{"number_samples":1,"species":"human","abstract":"We originally generated monoclonal antibodies which specifically react with Ser2 , Ser7 or Ser5 residue of the C-terminal domain of RNA polymerase 2. The genome wide recruitment of Ser5P, S7P and/or Ser2P RNA polymerase 2 was evaluated in Hela cells.","project":"DRP000366"}
{"number_samples":4,"species":"human","abstract":"Using next generation sequencer, we investigated the global transcriptional change against glucose deprivation in T24 bladder carcinoma cell line, which produced N-GlcNAc2-modifed proteins under glucose deprivation.","project":"DRP000425"}
{"number_samples":8,"species":"human","abstract":"Recent research hints at an underappreciated complexity in pre-miRNA processing and regulation. Global profiling of pre-miRNA and its potential to increase understanding of the pre-miRNA landscape is impeded by overlap with highly-expressed classes of other non-coding RNA. Here we present a dataset excluding these RNA before sequencing through locked nucleic acids (LNA), greatly increasing pre-miRNA sequence counts with no discernable effects on pre-miRNA or mature miRNA sequencing. Analysis of profiles generated in total, nuclear, and cytoplasmic cell fractions reveals pre-miRNAs are subject to a wide range of regulatory processes involving loci-specific 3’- and 5’-end variation entailing complex cleavage patterns with co-occurring polyuridylation. Additionally, examination of nuclear-enriched flanking sequences of pre-miRNA, particularly those derived from polycistronic miRNA transcripts, provides insight into miRNA and miRNA-offset (moRNA) production. Our findings point to particularly intricate regulation of the let-7 family, introduce novel and unify known forms of pre-miRNA regulation and processing, and shed new light on the byproducts of miRNA processing pathways.","project":"DRP000464"}
{"number_samples":21,"species":"human","project":"DRP000499"}
{"number_samples":4,"species":"human","abstract":"To appreciate the biological/biochemical impact of these splicing mutations, we expressed the wild-type and the mutant (S34F) U2AF35 in HeLa cells and performed RNA sequencing.","project":"DRP000527"}
{"number_samples":5,"species":"human","abstract":"In order to identify the transcripts those are regulated by UPF1 in human cells, we have measured the decay rates and the steady-state abundances of whole transcriptome in UPF1-depleted HeLa cells using BRIC-seq and RNA-seq.","project":"DRP000622"}
{"number_samples":7,"species":"human","abstract":"Using next generation sequencer, we investigated the global transcriptional time course-change against glucose deprivation in T24 bladder carcinoma cell line cells, which produced N-GlcNAc2-modified protein under glucose deprivation.","project":"DRP000665"}
{"number_samples":4,"species":"human","abstract":"Human inactive X chromosome (Xi) forms a compact structure called the Barr body, which is enriched in repressive histone modifications such as trimethylation of histone H3 Lys9 (H3K9me3) and Lys27 (H3K27me3). These two histone marks are distributed in distinct domains, and XIST (X inactive specific transcript) preferentially colocalizes with H3K27me3 domains. Here, we show that Xi compaction requires HBiX1, a heterochromatin protein 1 (HP1)–binding protein, and SMCHD1 (structural maintenance of chromosomes hinge domain containing 1), both of which are enriched throughout the Xi chromosome. HBiX1 localization to H3K9me3 and XIST-associated H3K27me3 (XIST-H3K27me3) domains was mediated through interactions with HP1 and SMCHD1, respectively. Furthermore, HBiX1 was required for SMCHD1 localization to H3K9me3 domains. Depletion of HBiX1 or SMCHD1, but not Polycomb repressive complex 2 (PRC2), resulted in Xi decompaction, similarly to XIST depletion. Thus, the molecular network involving HBiX1 and SMCHD1 links the H3K9me3 and XIST-H3K27me3 domains to organize the compact Xi structure.","project":"DRP000929"}
{"number_samples":122,"species":"human","abstract":"To understand the mutual association of the transcriptomes between host humans and infecting malaria parasites, Plasmodium falciparum (Pf), in vivo, we conducted the RNA-Seq analysis of 116 Indonesian malaria patients. Using RNAs extracted from peripheral blood of the patients as the mixture of the human and Pf transcripts, we generated a total of 3 billion RNA-Seq tags. Analysis of these tags allowed us to identify genes and pathways which were associated with clinical symptoms both on human and parasite sides. Particularly, we observed characteristic expression changes in the human innate immune response pathway genes depending on severity of malaria. We also identified a group of transcription regulatory factors and signaling molecule genes which have positive or negative correlations between humans and Pfs. These genes may change their expression patterns to accommodate the respective organisms to mutually conflicting environments. Furthermore, it was possible to utilize the RNA-Seq data for genotyping of the parasites to identify possible drug resistant genetic variations. By bypassing technical difficulty in preparing samples in field, this approach should provide a practically useful mean to describe mutually interacting transcritpomes of humans and parasites, which should eventually explain diverse malaria symptoms.","project":"DRP000987"}
{"number_samples":6,"species":"human","abstract":"MCF-7 human breast cancer cells acquire estrogen independent cell proliferation ability, when they are grown in the medium deprived of estrogen for an extended period (long term estrogen deprivation; LTED). This represents a cellular model for hormonal therapy resistance. On the other hand, phytoestrogen resveratrol (RES) inhibits the growth of breast cancer cells. In this study, we analyzed transcriptome of MCF-7 cells under LTED and RES treatment.","project":"DRP001048"}
{"number_samples":4,"species":"human","abstract":"A cohesin component RAD21 is recurrently mutated in myeloid neoplasms. To investigate the role of RAD21 mutation in leukemogenesis, we performed RNA sequencing of mock, wild-type or mutant RAD21 transduced Kasumi-1 cells and compared their effect on global gene expression.","project":"DRP001055"}
{"number_samples":12,"species":"human","abstract":"Comparison of CAGE and RNA-seq transcriptome profiling using a clonally amplified and single molecule next generation sequencing","project":"DRP001149"}
{"number_samples":19,"species":"human","abstract":"The FANTOM5 promoter expression atlas provides a rich source of expression and functional annotation of human and mouse cell-type specific transcriptomes with wide applications in biomedical research.","project":"DRP001150"}
{"number_samples":9,"species":"human","abstract":"5’ cap structure plays a very important role in the biological system, as it for instance regulates RNA stability and localization.Here we report an intensive study of the cap structure of short RNAs (sRNAs), using three different methods: thin layer chromatography, mass spectrometry, and sequencing of small RNA libraries.","project":"DRP001194"}
{"number_samples":1,"species":"human","abstract":"In addition to BCR, various rare fusion partners for the ABL1 gene have been reported in leukemia.  We have identified the fusion gene SNX2-ABL1 in a pediatric case of acute lymphoblastic leukemia (ALL), which has only once previously been reported in an adult patient.  Cytogenetic analysis detected this fusion gene arising from a t(5;9)(q22;q34) translocation.  ALL cells carrying a SNX2-ABL1 fusion exhibited a BCR-ABL1+ALL-like gene expression profile.  The patient poorly responded to dasatinib but partially responded to imatinib.  Treatment using tyrosine kinase inhibitors requires further investigation to optimize the genotype-based treatment stratification for patients with SNX2-ABL1 fusion.","project":"DRP001219"}
{"number_samples":1,"species":"human","abstract":"In addition to BCR, various rare fusion partners for the ABL1 gene have been reported in leukemia.  We have identified the fusion gene SNX2-ABL1 in a pediatric case of acute lymphoblastic leukemia (ALL), which has only once previously been reported in an adult patient.  Cytogenetic analysis detected this fusion gene arising from a t(5;9)(q22;q34) translocation.  ALL cells carrying a SNX2-ABL1 fusion exhibited a BCR-ABL1+ALL-like gene expression profile.  The patient poorly responded to dasatinib but partially responded to imatinib.  Treatment using tyrosine kinase inhibitors requires further investigation to optimize the genotype-based treatment stratification for patients with SNX2-ABL1 fusion.","project":"DRP001220"}
{"number_samples":7,"species":"human","abstract":"By integrative analysis of ChIP-seq, RNA-seq and our BRIC-seq, we found that transcriptional regulation and RNA degradation are independently regulated. In addition, we showed that RNA stability serves as an important determinant of eventual transcript levels. We also suggest that RNA binding proteins, such as UPF1, STAU1 and EXOSC5, play active roles in such controls.","project":"DRP001280"}
{"number_samples":337,"species":"human","abstract":"To understand heterogeneous behaviors of individual cancer cells, it should be important to investigate gene expression levels as well as their divergences between individual cells. Here we conducted a single cell RNA Seq analysis of a series of lung adenocarcinoma cell lines. We analyzed a total of 337 RNA Seq libraries of single cells and found gene expression diversities between individual cells are characteristic depending on genes. The genes showing highly variable expressions were enriched in particular pathways. Particularly, the EGFR pathway genes originally showed wider variations and further diversity was induced in response to an anticancer-drug stimulation. We also observed that cancer-related genes, which were identified from recent clinical cancer sequencing projects, showed potential expression variations, which firstly became apparent on the anti-cancer drug stimulation.","project":"DRP001358"}
{"number_samples":324,"species":"human","abstract":"Comprehensive epigenomic profiling for a set of vascular endothelial cells has been performed. The epigenomic profile contains epigenomic signals captured with ChIP-seq (H3K4me1, H3K4me3, H3K27ac, H3K36me3, H3K9me3, H3K27me3, H3K9-oligo, H3K27-oligo, control) and FAIRE-seq. The vascular endothelial cells (AoEC, HPAEC, HCAEC, HUVEC, HUAEC, encodardial celll, HBCAEC, HPAEC, and HBAEC) have been cultured from donors belong to multiple ethnisity, gender, age, and medical background.","project":"DRP001797"}
{"number_samples":26,"species":"human","abstract":"Cancer cells carry various types of aberrations at the levels of genome, epigenome and transcriptome. For future studies on the biological relevance of the aberrations and their mutual relations, we generated a multi-omics catalogue of 26 lung adenocarcinoma cell lines. By whole-genome sequencing, we detected SNVs and indels, copy number aberrations and chromosome rearrangements. We used RNA-Seq to examine whether those mutation-harboring genes are expressed in the corresponding cell lines or whether their expression levels are deviated between the cell lines. Also, we could detect known and novel abnormal transcripts, such as RET- and ALK-fusion transcripts and transcripts caused by splice site mutations. Similarly, target-captured bisulfite sequencing and ChIP-Seq analyses revealed epigenomic status in each cell line and their deviations. We integrated these multi-omics data to explain possible causes of the observed aberrant gene expressions, particular for the representative cancer-related genes. We unexpectedly found that the patterns of aberrations were highly diverse between genes; namely irregular chromatin status revealed by ChIP-Seq was the characteristic to the EGFR, while a large genomic deletion and hyper-DNA methylation in the promoter region were the most frequent for the CDKN2A gene. Our datasets, by complementing current whole-exome or whole-genome sequencing of clinical cancers, should lay valuable base for interpreting how various types of genomic and epigenomic aberration lead to aberrant transcriptomic appearances in cancers.","project":"DRP001919"}
{"number_samples":1,"species":"human","abstract":"Cancer cells carry various types of aberrations at the levels of genome, epigenome and transcriptome. For future studies on the biological relevance of the aberrations and their mutual relations, we generated a multi-omics catalogue of 26 lung adenocarcinoma cell lines. By whole-genome sequencing, we detected SNVs and indels, copy number aberrations and chromosome rearrangements. We used RNA-Seq to examine whether those mutation-harboring genes are expressed in the corresponding cell lines or whether their expression levels are deviated between the cell lines. Also, we could detect known and novel abnormal transcripts, such as RET- and ALK-fusion transcripts and transcripts caused by splice site mutations. Similarly, target-captured bisulfite sequencing and ChIP-Seq analyses revealed epigenomic status in each cell line and their deviations. We integrated these multi-omics data to explain possible causes of the observed aberrant gene expressions, particular for the representative cancer-related genes. We unexpectedly found that the patterns of aberrations were highly diverse between genes; namely irregular chromatin status revealed by ChIP-Seq was the characteristic to the EGFR, while a large genomic deletion and hyper-DNA methylation in the promoter region were the most frequent for the CDKN2A gene. Our datasets, by complementing current whole-exome or whole-genome sequencing of clinical cancers, should lay valuable base for interpreting how various types of genomic and epigenomic aberration lead to aberrant transcriptomic appearances in cancers.","project":"DRP002380"}
{"number_samples":477,"species":"human","abstract":"The goal of these libraries is to find transcriptome signatures of the different phases of the cell cycle.  Single HeLa cells expressing fluorescent reporters that peak in G1 and G2/M phases were captured in C1 Single-Cell Auto Prep systems (Fluidigm  recording red and green fluorescence for each cell individually, each cell was lysed and their RNAs converted to cDNAs in the C1 systems using SMARTer kits (Clontech).  The cDNAs were then collected and converted to RNA-seq libraries by tagmentation using Illumina/Nextera kits.  The cells used in this experiment are unpublished as of today, and precise details will be given later, together with the fluorescence values.  In the meantime, this dataset can be used to assess reproducibility of single-cell RNA-seq over 5 runs in the Fluidigm system.","project":"DRP002435"}
{"number_samples":255,"species":"human","abstract":"To understand heterogeneous behaviors of individual cancer cells, it should be important to investigate gene expression levels as well as their divergences between individual cells. Here we conducted a single cell RNA Seq analysis of a series of lung adenocarcinoma cell lines. We analyzed a total of 337 RNA Seq libraries of single cells and found gene expression diversities between individual cells are characteristic depending on genes. The genes showing highly variable expressions were enriched in particular pathways. Particularly, the EGFR pathway genes originally showed wider variations and further diversity was induced in response to an anticancer-drug stimulation. We also observed that cancer-related genes, which were identified from recent clinical cancer sequencing projects, showed potential expression variations, which firstly became apparent on the anti-cancer drug stimulation.","project":"DRP002586"}
{"number_samples":23,"species":"human","abstract":"Potential Small Guide RNAs for tRNase ZL from Human Plasma, Peripheral Blood Mononuclear Cells, and Cultured Cell Lines","project":"DRP002623"}
{"number_samples":3,"species":"human","abstract":"Exosomes are small membrane vesicles (30-100 nm in diameter) secrted by numerous cells. Exosome have been shown to contain mRNA and DNA. We found at least two types of salivary exosomes (exosome I and exosome II) that are different in size and have different proteome. In previous study, we performed small RNA transcriptome analysis by next generation sequencing technology and reported that many types of small RNA, such as miRNA, piwi-interacting RNA (piRNA), small nucleolar RNA (snoRNA), other small RNAs and genomic repeats were contained in exosome I, II and whole saliva. In this study, we performed whole-transcriptome analysis using cDNA libraries constructed from total RNA except small RNAs. We found that 11-32% of reads from salivary exosomes and whole saliva were mapped to pseudogene.  The Data Access Committee of the National Bioscience Database Center (NBDC) approved that this personal genetic data were publicated according to NBDC data sharing guidelines (http://humandbs.biosciencedbc.jp/).","project":"DRP002625"}
{"number_samples":4,"species":"human","abstract":"Gene expression in response to inflammatory stimuli is controlled by both transcriptional and posttranscriptional mechanisms in immune cells. Posttranscriptional regulation that modifies mRNA stability and translation provides rapid and flexible control of gene expression and control of mRNAs stability is important for coordinating the initiation and resolution of inflammation. However, the posttranscriptional mechanisms in innate immunity remain to be clarified. This study is aiming to investigate the posttranscriptional mechanisms in innate immunity based on the roles of RNA binding proteins and RNase we identified.","project":"DRP002667"}
{"number_samples":4,"species":"human","abstract":"Gene expression in response to inflammatory stimuli is controlled by both transcriptional and posttranscriptional mechanisms in immune cells. Posttranscriptional regulation that modifies mRNA stability and translation provides rapid and flexible control of gene expression and control of mRNAs stability is important for coordinating the initiation and resolution of inflammation. However, the posttranscriptional mechanisms in innate immunity remain to be clarified. This study is aiming to investigate the posttranscriptional mechanisms in innate immunity based on the roles of RNA binding proteins and RNase we identified.","project":"DRP002672"}
{"number_samples":1,"species":"human","abstract":"Pulmonary fibrosisinvolves the process known as epithelial-to-mesenchymal transition (EMT) in which alveolar epithelial cells become mesenchymal cells, as well as lung fibroblast changes to abnormal myofibroblasts, and the deposition of extracellular matrix molecules such as collagen. These changes are irreversible, so current treatment aims to halt disease progression. Pulmonary fibrosis can be drug-induced or caused by an infectious disease or occupational dust inhalation, but the cause is unknown in idiopathic pulmonary fibrosis (IPF). This project analyzes gene expression during EMT or fibrosis, using EMTmodel induced in human alveolar epithelial cells (A549) by transforming growth factor (TGF)-b1 stimulation or pulmonary fibrosis model mouse.","project":"DRP002712"}
{"number_samples":1,"species":"human","abstract":"The project aims to reveal biological function of histone variants. The samples are the HeLa cells that exogenously produce the GFP-tagged histone variants. We performed ChIP-Seq for the tags and epigenetic markers. We also performed mRNA-Seq to profile the gene expression in the cells.","project":"DRP002721"}
{"number_samples":2,"species":"human","abstract":"To determine the factors which might promote metastasis at downstream of ANGPTL2 in breast cancer cell, mRNA sequencing analyses of MDA-MB231 cells harboring ANGPTL2 knockdown (MB231/miANGPTL2) and relative to control (MB231/miLacZ) cells were carried out by using illumina GAIIx sequencer.","project":"DRP002835"}
{"number_samples":8,"species":"human","abstract":"1. To identify lncRNA regulating colorectal tumorigenesis, we performed RNA-seq analysis of cells with high and low tumorigenicity 2. To study the role of UPAT in colorectal cancer cells, we investigated the gene expression profiles of HCT116 cells in which UPAT expression had been suppressed by siRNA.","project":"DRP002851"}
{"number_samples":20,"species":"human","abstract":"The cell sheet engineering improved the function of the ocular surface, but the impairment of full recovery of visual acuity still remains. To address this problem, we transduced PAX6 isoforms, OCT4 and KLF4 into oral mucosal epithelial cells and investigated the molecular mechanism for induced corneal epithelium specific genes. This study will improve the clinical outcome of epithelial sheet transplantation.","project":"DRP002860"}
{"number_samples":48,"species":"human","project":"ERP000546"}
{"number_samples":10,"species":"human","project":"ERP000573"}
{"number_samples":12,"species":"human","project":"ERP000710"}
{"number_samples":3,"species":"human","project":"ERP000787"}
{"number_samples":1,"species":"human","project":"ERP000799"}
{"number_samples":2,"species":"human","abstract":"While the number and identity of proteins expressed in a single human                     cell type is currently unknown, this fundamental question can be addressed by                     advanced mass spectrometry (MS)-based proteomics. On-line liquid chromatography                     coupled to high resolution MS and MS/MS yielded more than 150,000 unique                     peptides that identified more than 10,000 different human proteins encoded by                     more than 9,000 human genes. Deep transcriptome sequencing revealed transcripts                     for nearly all detected proteins. We show that the abundances of more than 90%                     of proteins and transcripts fall within a 10,000-fold range, and allocate the                     proteome to different compartments, complexes and functions. Comparisons of the                     proteome and the transcriptome, and analysis of protein complex databases and GO                     categories, suggest that we achieved almost complete coverage of the functional                     transcriptome and the proteome of a single cell type.","project":"ERP000959"}
{"number_samples":18,"species":"human","project":"ERP000992"}
{"number_samples":1,"species":"human","project":"ERP001115"}
{"number_samples":18,"species":"human","project":"ERP001304"}
{"number_samples":14,"species":"human","abstract":"Sequencing of small transcript processed RNAs from T-cells that are involved in gene expression regulation and control of the immune response.","project":"ERP001317"}
{"number_samples":40,"species":"human","abstract":"Detection of unknown viral hepatitis agents in transplanted liver tissue","project":"ERP001344"}
{"number_samples":8,"species":"human","project":"ERP001458"}
{"number_samples":3,"species":"human","project":"ERP001574"}
{"number_samples":3,"species":"human","abstract":"A timecourse of adenovirus infection of human HeLa cells, using RNA-seq","project":"ERP001782"}
{"number_samples":7,"species":"human","abstract":"The aim of this experiment was to obtain a detailed transcriptional landscape of human and mouse islet-cells, including strand information, as well an epigenetic map of transcriptionally active chromatin.","project":"ERP001828"}
{"number_samples":2,"species":"human","abstract":"Investigation of the upregulation of lncRNA genes in 13q14.3 and ist correlation with downregulation in cis of a gene cluster regulating NF-kB","project":"ERP001895"}
{"number_samples":63,"species":"human","abstract":"A Cartes d'Identite des Tumeurs (CIT) project from the french Ligue Nationale Contre le Cancer (http://cit.ligue-cancer.net); Integrative study based on 89 patients with Head and Neck Squamous Cell Carcinoma (HNSCC); 89 Affymetrix HG-U133 Plus 2.0 GeneChips arrays;  88 samples on HumanCNV370-Quad GeneChips arrays; 84 samples on Illumina Human Methylation 27K arrays;Micro-RNA sequencing of  64 tumors. Please note: Characteristics[MetastasisFreeSurvivalEvent] indicates if there is a metastasis-free survival for a particular sample / patient (it is like an event-free survival variable where the event considered is a metastasis).  Characteristics[MetastasisFreeSurvivalDelay] - whenever the MetastasisFreeSurvivalEvent is 1, gives the duration of the metastasis-free survival period in months.","project":"ERP001908"}
{"number_samples":667,"species":"human","abstract":"The data is under embargo until the first publication by the investigators in early 2013. This RNA sequencing data set of 465 human lymphoblastoid cell line samples from the CEU, FIN, GBR, TSI and YRI populations from the 1000 Genomes sample collection was created by the Geuvadis consortium.","project":"ERP001942"}
{"number_samples":4,"species":"human","abstract":"We here present a comparative genome, transcriptome and functional network analysis of three human cancer cell lines (A431, U251MG and U2OS), and investigate their relation to protein expression. Gene copy numbers significantly influenced corresponding transcript levels; their effect on protein levels was less pronounced. We focused on genes with altered mRNA and/or protein levels to identify those active in tumor maintenance. We provide comprehensive information for the three genomes and demonstrate the advantage of integrative analysis for identifying tumor-related genes amidst numerous background mutations by relating genomic variation to expression/protein abundance data and use gene networks to reveal implicated pathways.","project":"ERP001948"}
{"number_samples":6,"species":"human","abstract":"Acute megakaryoblastic leukemia (AMKL) is a heterogeneous disease generally associated with poor prognosis. Gene expression profiles indicate the existence of distinct molecular subgroups and several genetic alterations have been characterized in the past years, including the t(1;22)(p13;q13) and the trisomy 21 associated with GATA1 mutations. However, the majority of patients do not present with known mutations and the limited access to primary patient leukemic cells also prevents efficient development of novel therapeutic strategies. Using a xenotransplantation approach, we have modeled human pediatric AMKL in immunodeficient mice. High-throughput sequencing of engrafted cases identified recurrent fusions genes that define new molecular subgroups. A group of patients present with MLL or NUP98 fusion genes leading to upregulation of the HOX A cluster genes as described in other subtypes of AML. Also, a novel CBFA2T3-GLIS2 fusion gene resulting from a cytogenetically invisible inversion of chromosome 16 was identified and observed in 31% of non Down syndrome AMKL. These data provide new markers that will be useful for the diagnosis and follow-up of patients. Finally, we show that AMKL xenograft models constitute a relevant preclinical screening platform to validate the efficacy of novel therapies such as dimethylfasudil, a novel small molecule ploidy inducer.","project":"ERP001971"}
{"number_samples":27,"species":"human","abstract":"To characterize the epigenetic programs that underlie pancreas differentiation, we have generated genome-scale maps of H3K4me and H3K27me3 patterns by ChIP-seq and determined expression profiles by RNA-seq from undifferentiated human ESCs, four intermediate differentiated stages (definitive endoderm, primitive gut tube, posterior foregut, and pancreatic endoderm), and in vitro-differentiated polyhormonal cells. Antibodies against CD142 and CD200 were used to select for targeted pancreatic and endocrine populations at the end of the culture. Cells at the end of culture were implanted into mice for further differentiation into mature insulin-producing beta-cells and compared to sorted polyhormonal cells by RNA-seq and ChIP-seq analysis. For the experimental factor values the time points are along a differentiation protocol from stem cells to tissue. Values in the CD antibody column refer to antibodies used for cell sorting. In this column 'not applicable' refers to samples that were not sorted.  'none' refers to samples that were sorted, but did not bind to either of the two antibodies, CD200 and CD142. Values in the histone antibody column are about the ChIP process.","project":"ERP002045"}
{"number_samples":7,"species":"human","abstract":"From altogether 220 million paired-end sequence reads from seven CRC cell lines we identified 3391 candidate fused transcripts, each with at least five-fold coverage across the fusion partners and at least three sequence-reads spanning the actual chimeric breakpoint. Stringent requirements left a set of eleven candidate fusion transcripts nominated for further experimental validation, of which ten were positive by RT-PCR and Sanger sequencing. Six of the fusions were intra-chromosomal transcripts and, interestingly, three of these, AKAP13-PDE8A, COMMD10-AP3S1, and CTB-35F21.1-PSD2, were present in respectively 18, 18 and 20 of 21 analyzed cell lines, and in respectively 18, 61 and 48 (17 to 58 percent) of 106 primary cancer tissues. These three fusion transcripts were also detected in two to four of 14 normal colonic mucosa samples (14 to 28 percent).","project":"ERP002049"}
{"number_samples":5,"species":"human","abstract":"RNA sequences are generally identical to the underlying DNA sequences but there are known exceptions.  These RNA-DNA differences (RDDs) have been found in the nuclear genomes of human cells and in the mitochondria of plants and animals but not in human mitochondria.  Here by deep sequencing of DNA and RNA of human mitochondria, we identified 3 RDD sites including an A-to-U and an A-to-G RDD at position 2617.  Examination of the precursor polycistronic mitochondrial transcripts shows that the RDD formation occurs post-transcriptionally.  Phylogenetic analysis shows that the ancestral allele at position 2617 was a thymine or a guanine.  Thus the RDD formation recapitulates the ancestral form of 16S rRNA. Our findings show that RDD formation like other RNA processing steps is conserved across species and likely has functional significance.","project":"ERP002075"}
{"number_samples":4,"species":"human","abstract":"Comprehensive transcriptome analysis of four distinct human limbal compartments, including basal limbal crypts (BLCs), superficial limbal crypts (SLCs), cornea, and the supporting stroma, with the aid of laser capture microdissection and deep RNA sequencing.","project":"ERP002232"}
{"number_samples":9,"species":"human","abstract":"We adapted the NextGen sequencing technology to obtain more accurate spectra of mRNA and miRNA in circulation, specifically to explore the plasma-miRNA association with colorectal cancer and ulcerative colitis.","project":"ERP002414"}
{"number_samples":7,"species":"human","abstract":"RNA-seq data for cell lines in the haematopoietic lineages, hematological malignancy","project":"ERP002588"}
{"number_samples":12,"species":"human","abstract":"Human embryonic stem cells (hESCs) harbor the ability to undergo lineage-specific differentiation into clinically relevant cell types. Transcription factors and epigenetic modifiers are known to play important roles in the maintenance of pluripotency of hESCs. However, little is known about regulation of pluripotency through splicing. In this study, we identify the spliceosome-associated factor SON as a novel factor essential for the maintenance of hESCs. Depletion of SON in hESCs results in the loss of pluripotency and cell death. Using genome-wide RNA profiling, we identified transcripts that are regulated by SON. Importantly, we confirmed that SON regulates the proper splicing of transcripts encoding for pluripotency regulators such as PRDM14, OCT4, E4F1 and MED24. Furthermore, we show that SON is bound to these transcripts in vivo. In summary, we connect a splicing-regulatory network for accurate transcription production to the maintenance of pluripotency and self-renewal of hESCs.","project":"ERP003259"}
{"number_samples":12,"species":"human","abstract":"TBA","project":"ERP003460"}
{"number_samples":4,"species":"human","abstract":"To assess the performance of computational methods for exon identification, transcript reconstruction and expression level quantification from RNA-seq data, 24 protocol variants of 14 independent software programs (AUGUSTUS, Cufflinks, Exonerate, GSTRUCT, iReckon, mGene, mTim, NextGeneid, Oases, SLIDE, Transomics, Trembly, Tromer and Velvet) were evaluated against transcriptome data from human cells and two model organisms.","project":"ERP003471"}
{"number_samples":8,"species":"human","abstract":"Cells were cultured in RPMI 1640 (Gibco), supplemented with 10% fetal bovine serum (FBS) (Gibco), 2mM L-glutamine (Gibco) and 1% Penicillin-Streptomycin solution (Gibco). IMR32 cells were treated as follows; untreated control, 24h 1uM azakenpaullone, 24h 1uM BIO or 24h 28mM LiCl with biological duplicates.","project":"ERP003495"}
{"number_samples":18,"species":"human","abstract":"To assess the performance of read mapping software for RNA-seq, 26 spliced alignment protocols based on 11 programs and pipelines (BAGET, GEM, GSNAP, GSTRUCT, MapSplice, PALMapper, PASS, ReadsMap, SMALT, STAR and TopHat) were applied to four human and mouse transcriptome data sets.","project":"ERP003536"}
{"number_samples":171,"species":"human","abstract":"RNA-seq was performed of tissue samples from 95 human individuals representing 27 different tissues in order to determine tissue-specificity of all protein-coding genes.","project":"ERP003613"}
{"number_samples":4,"species":"human","abstract":"Bone development and repair depends on the differentiation of mesenchymal stem cells (MSCs) into osteoblasts. MSCs can be differentiated towards osteoblasts in vitro, making these cells a convenient tool for investigation of osteogenesis. Molecular characterization of this process is relevant for the application of MSCs in skeletal regenerative medicine, and for understanding the deregulation of osteogenesis in bone disease. Cellular  differentiation is driven by highly regulated changes in gene expression, which at the level of transcription is associated with DNA binding of transcriptional regulators and local changes in chromatin landscape. By sequencing of RNA (RNA-Seq) and immunoprecipitated chromatin (ChIP-Seq) we have characterized gene expression, histone modification changes and DNA binding of the bone master regulator RUNX2 in osteogenic differentiation.  Data from the RNA-Seq experiment has also been deposited at ArrayExpress under accession number E-MTAB-1829 (https://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-1829/).","project":"ERP003789"}
{"number_samples":21,"species":"human","abstract":"Chronic lymphocytic leukemia (CLL), the most frequent adult leukemia in western countries, is a clonal accumulation of mature B-lymphocytes and its natural history is yet unclear. By using sequencing and cellular biology approaches on a cohort of CLL patient samples, we show here that acquired CLL mutations are observed in hematopoietic multipotent progenitor fractions in the majority of patients. These early CLL mutations include recurrent inactivating mutations in NFKBIE (10.7%) and missense mutations in BRAF (3.6%) and EGR2 (8.3%). Functional analyses demonstrated that BRAF-G469R affects lymphoid differentiation and transforms the T-cell lineage in vivo. In addition, the EGR2 recurrent mutations were associated with transcriptional activation of EGR2 target genes in patients and cell cycle abnormality in cellular model. Our findings indicate that CLL may develop from an initial infra-clinic, pre-leukemic phase affecting immature hematopoietic cells. We perform RNA-seq experiments on three EGR2-E356K samples, one EGR2-H384N sample and seven EGR2 wildtype patients. The RNA was extracted from B cells. The cDNA libraries were prepared using the ScriptSeq Complete Kit (Epicentre) and we performed paired-end sequencing (100 bp) using HiSeq2000 sequencing instruments.","project":"ERP003791"}
{"number_samples":3,"species":"human","abstract":"RNA-sequencing analysis of human platelet rRNA-depleted total RNA from two normal males and one female. The goal of this experiment was to identify genes expressed in unstimulated circulating platelets. Data from platelet polyA-mRNA has also been deposited at ArrayExpress, under accession number E-MTAB-715, https://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-715/ .","project":"ERP003815"}
{"number_samples":102,"species":"human","abstract":"We overexpress RUNX2 in ten human cell lines and identify genes that are affected by RUNX2 expression. These target genes provide a valuable resource into pathways regulated by RUNX2","project":"ERP003917"}
{"number_samples":34,"species":"human","abstract":"Analysis of the role of KAT2A in  maintenance vs. differentiation of human leukaemia.","project":"ERP003933"}
{"number_samples":16,"species":"human","abstract":"Micro-organisms inhabiting teeth surfaces grow on biofilms where a specific and complex succession of bacteria has been described by co-aggregation tests and DNA-based studies. Although the composition of oral biofilms is well established, the active portion of the bacterial community and the patterns of gene expression in vivo have not been studied. Using RNA-sequencing technologies, we present the first metatranscriptomic study of human dental plaque, performed by two different approaches: (1) A short-reads, high-coverage approach by Illumina sequencing to characterize the gene activity repertoire of the microbial community during biofilm development; (2) A long-reads, lower-coverage approach by pyrosequencing to determine the taxonomic identity of the active microbiome before and after a meal ingestion. The high-coverage approach allowed us to analyze over 398 million reads, revealing that microbial communities are individual-specific and no bacterial species was detected as key player at any time during biofilm formation. We could identify some gene expression patterns characteristic for early and mature oral biofilms. The transcriptomic profile of several adhesion genes was confirmed through qPCR by measuring expression of fimbriae-associated genes. In addition to the specific set of gene functions overexpressed in early and mature oral biofilms, as detected through the short-reads dataset, the long-reads approach detected specific changes when comparing the metatranscriptome of the same individual before and after a meal, which can narrow down the list of organisms responsible for acid production and therefore potentially involved in dental caries.","project":"ERP003984"}
{"number_samples":8,"species":"human","abstract":"We isolated and analyzed at single nucleotide resolution cancer-associated neochromosomes from well-/de-differentiated liposarcomas. Neochromosomes, which can exceed 600Mb in size, initially arise as circular structures following chromothripsis involving chromosome 12. The core of the neochromosome is amplified, rearranged and corroded through hundreds of breakage-fusion-bridge cycles. Under selective pressure, amplified oncogenes are over-expressed, while co-amplified passenger genes may be silenced epigenetically. New material may be captured during punctuated chromothriptic events. Centromeric corrosion leads to crisis, which is resolved through neocentromere formation or native centromere capture. Finally, amplification terminates and the neochromosome core is stabilized in linear form by telomere capture. This study provides insight into the dynamic mutational processes underlying the life history of a special form of cancer mutation.","project":"ERP004006"}
{"number_samples":34,"species":"human","abstract":"Comprehensive sequencing of human cancers has identified recurrent mutations in genes encoding chromatin regulatory proteins. For clear cell renal cell carcinoma (ccRCC), three of the five commonly mutated genes encode the chromatin regulators PBRM1, SETD2, and BAP1. How these mutations alter the chromatin landscape and transcriptional program in ccRCC or other cancers is not understood. Here, we identified alterations in chromatin organization and transcript profiles associated with mutations in chromatin regulators in a large cohort of primary human kidney tumors. By associating variation in chromatin organization with mutations in SETD2, which encodes the enzyme responsible for H3K36 trimethylation, we found that changes in chromatin accessibility occurred primarily within actively transcribed genes. This increase in chromatin accessibility was linked with widespread alterations in RNA processing, including intron retention and aberrant splicing, affecting approximately 25% of all expressed genes. Further, decreased nucleosome occupancy proximal to misspliced exons was observed in tumors lacking H3K36me3. These results directly link mutations in SETD2 to chromatin accessibility changes and RNA processing defects in cancer. Detecting the functional consequences of specific mutations in chromatin regulatory proteins in primary human samples could ultimately inform the therapeutic application of an emerging class of chromatin-targeted compounds.","project":"ERP004043"}
{"number_samples":2,"species":"human","abstract":"Bisphenol A (BPA) is an environmental endocrine disruptor which has been detected in almost all human bodies. Many studies have implied that BPA exposure is harmful to human health. Previous studies mainly focused on BPA effects on ER-positive cells. Genome-wide impact of BPA on gene regulation in ER-negative cells is unclear. In the present study, we performed RNA-seq to characterize BPA-induced gene regulation on ER-negative HEK 293 cells.","project":"ERP004062"}
{"number_samples":14,"species":"human","abstract":"Transcriptome sequencing (RNA-seq) experiment of 14 human lymphoblastoid cell line samples from the 1000 Genomes sample set (http://www.1000genomes.org/). Dataset includes two parent-daughter trios (CEU and YRI populations) and additional eight unrelated individuals (CEU population). This accession contains raw and mapped RNA-seq read data, other assays in this study are available under accession XX (ChIP-seq) and YY (GRO-seq).","project":"ERP004078"}
{"number_samples":40,"species":"human","abstract":"A human volunteer challenge model for typhoid fever has recently been established at the Oxford Biomedical Research Centre, University of Oxford. More than forty individuals have been infected with 1x10(4) CFU S. Typhi and the transcriptional response in peripheral blood of ten volunteers before infection and on development of fever or on day 14 id no fever was diagnosed has been determined using Illuminaâ„¢ Human HT-12_V4 Beadchips (Illumina, San Diego, CA, USA). A strong transcriptional signature was associated with typhoid diagnosis.We propose to determine the peripheral blood trancriptional response at additional time points. Further we will determine th peripheral blood trancriptional response to vaccination with an established live oral typhoid vaccine Ty21a, a novel live oral vaccine and a placebo control.","project":"ERP004094"}
{"number_samples":4,"species":"human","abstract":"To improve our understanding of the effect of differential methylation on gene expression between normal and tumor breast cells, we carried out RNA sequencing evaluate mRNA expression changes in normal breast tissue and MCF7 breast cancer cell line. We identified 6632 protein-coding genes to be significantly differentially expressed (FDR threshold of 0.05) in the MCF7 cells. IPA Bio-Function enrichment analysis showed the 'cell death and survival' and 'cellular growth and proliferation' are the top 2 terms associated with differentially expressed genes. Cancer versus normal analysis of The Cancer Genome Atlas (TCGA) Breast data revealed genes that were differentially up and down-regulated in MCF7 are mostly respectively over- and under-expressed in clinical specimens.","project":"ERP004151"}
{"number_samples":6,"species":"human","abstract":"Human embryonic stem cells (hESCs) are isolated from the inner cell mass of the blastocysts.  The pluripotent properties of hESCs enable the derivation of cell-types or tissues of different lineages for potential applications such as therapeutics discovery and regenerative medicine.  Even though hESCs are pluripotent, differences have been observed when compared to the native pluripotent epiblast cells of the blastocyst. We use a chemical approach (3iL: 3 small molecule inhibitor and cytokine) to induce an expression signature that more closely resembles native pluripotent cells. To better understand the difference between this hESC state and the conventional hESC state, we generated the global expression profiles using RNA-seq.","project":"ERP004211"}
{"number_samples":10,"species":"human","abstract":"The DNA damage response (DDR) is a vast signaling network that is robustly activated by DNA double-strand breaks, the critical lesion induced by ionizing radiation (IR). Although much of this response operates at the protein level, a critical component of the network sustains many DDR branches by modulating the cellular transcriptome. Using deep sequencing, we delineated three layers in the transcriptional response to IR in human breast cancer cells: changes in the expression of genes encoding proteins or long noncoding RNAs, alterations in genomic binding by key transcription factors, and dynamics of epigenetic markers of active promoters and enhancers. We identified protein-coding and previously unidentified noncoding genes that were responsive to IR, and demonstrated that IR-induced transcriptional dynamics was mediated largely by the transcription factors p53 and nuclear factor ?B (NF-?B) and was primarily dependent on the kinase ataxia-telangiectasia mutated (ATM). The resultant data set provides a rich resource for understanding a basic, underlying component of a critical cellular stress response.","project":"ERP004219"}
{"number_samples":12,"species":"human","abstract":"Comprehensive analysis of coding and non -coding transcriptome using ribo-depleted total RNA-seq and poly A selected RNA-seq of MCF-7 cells grown in hypoxia and normoxia. Breast cancer cell line (MCF-7) is cultured in normoxic condition (21% O2) and hypoxic condition (1%O2) for 24 hours.  Expression of HIF-1alpha and/or HIF-2alpha subunits was suppressed using siRNAs in hypoxic MCF-7 cells. Total RNA was isolated from both hypoxia and normoxia conditions were subjected for ribosomal depleted stand specific RNA-seq and poly A selected RNA-seq","project":"ERP004269"}
{"number_samples":6,"species":"human","abstract":"DICER_Ex5 cells were created by disrupting exon 5 of the human Dicer gene using an AAV targeting construct, thereby interrupting a well conserved segment of the N-terminal helicase domain while sparing the RNase III domains. In DICER_Ex5 cells, Dicer is expressed at a level lower than the wild type. This experiment started with RIP-anti-Ago2 pull-down, followed by high throughput sequencing analysis in wild type and Dicer-hypomorphic HCT116 cell lines.","project":"ERP004270"}
{"number_samples":4,"species":"human","abstract":"Human endogenous retrovirus subfamily H (HERVH) is a class of transposable elements whose members are expressed preferentially in human pluripotent stem cells. Here, we report that the long-terminal repeat regions of HERVH function as enhancers which are regulated by OCT4. Strikingly, we found that HERVH is a nuclear-localized long non-coding RNA that is required to maintain the undifferentiated state of human embryonic stem cells. Furthermore, we showed that HERVH is associated with OCT4 and co-activators such as P300, CBP and mediator subunits. Together, these results uncover a new and unexpected role of species-specific transposable elements in specifying the pluripotency of human cells.","project":"ERP004298"}
{"number_samples":3,"species":"human","abstract":"Characterization of AGO2 bound short RNAs from nuclear extracts of Jurkat cell line. Nuclei were isolated from cells; a fraction of Nuclear lysates was harvested and sequenced as the \"input\" samples. The rest of nuclear lysates were Immunoprecipitated using AGO2 monoclonal antibodies or Isotype matched IgG. Following immunoprecipitation, RNA was extracted from AGO2 or control IgG IP for short RNA sequencing. Two independent biological replicates were carried out.","project":"ERP004352"}
{"number_samples":194,"species":"human","abstract":"High-throughput cellular phenotyping to understand the etiology and pathology of host-pathogen interaction.","project":"ERP004375"}
{"number_samples":4,"species":"human","abstract":"Identifying miRNA-regulated genes is key to understanding miRNA function. However, many miRNA recognition elements (MREs) do not follow canonical \"seed\" base-pairing rules, making identification of bona fide targets challenging. Here, we apply an unbiased sequencing-based systems approach to characterize miR-522, a member of the oncogenic primate-specific chromosome 19 miRNA cluster, highly expressed in poorly differentiated cancers. To identify miRNA targets, we sequenced full-length transcripts captured by a biotinylated miRNA mimic. Within these targets, mostly non-canonical MREs were identified by sequencing RNase-resistant fragments (this dataset).","project":"ERP004399"}
{"number_samples":6,"species":"human","abstract":"In order to assess the effect of AGO2 on the transcriptional and post-transcriptional regulation of gene expression, RNA expression was profiled in untreated, CTRL siRNA transfected and AGO2 siRNA transfected HeLaS3 cells","project":"ERP004402"}
{"number_samples":9,"species":"human","abstract":"S. aureus infection stimulates de novo synthesis and secretion of Nerve Growth Factor beta (NGF beta). The release of NGF beta does not depend on phagocytosis and can be stimulated by exo-products from S. aureus growth conditioned media. Mutants of the SaeR/SaeS two-component regulatory system that modulate exoprotein release in S. aureus do not elicit the same response, suggesting that specific S. aureus proteins are involved in the induction of NGF beta. In this study, we aim to perform gene expression profiling of human monocytic cell line THP1 cells that have been incubated with live wild type S. aureus, a saeS- mutant, and dead wild type S. aureus.  The aims of the experiment is to identify a set of genes expressed in the wild type S. aureus but not in cells lacking either the response regulator or sensor kinase components of the sae global regulator.","project":"ERP004573"}
{"number_samples":3,"species":"human","abstract":"Next-generation sequencing (NGS) technologies have dramatically expanded the breadth of genomics. Genome-scale data, once restricted to a small number of biomedical model organisms with significant resources, can now be generated for virtually any species at remarkable speed and low cost. Yet non-model organisms often lack a suitable reference to map sequence reads against, making alignment-based quality control (QC) of NGS data more challenging than cases where a well-assembled genome is already available. Here we show that by generating a rapid, non-optimized draft assembly of raw reads, it is possible to obtain reliable QC metrics without the need for a high quality reference genome. We use benchmark datasets generated from control samples across a range of genome sizes to illustrate that QC inferences made using draft assemblies are broadly equivalent to those made using a well-established reference, and describe QC tools routinely used in our production facility to assess the quality of NGS data from non-model organisms.","project":"ERP004578"}
{"number_samples":23,"species":"human","abstract":"To identify genome-wide differentially expressed microRNAs (miRNAs) between Huntington's disease (HD) and control Human pre-frontal cortex samples","project":"ERP004592"}
{"number_samples":7,"species":"human","abstract":"Using RNA deep sequencing we determined gene expression patterns and expression genotypes in Acute Myeloid Leukemia (AML) cell lines harboring a 3q-aberration. These cell lines specifically overexpress the gene EVI1 which plays a significant part in leukomogenesis. Additionally we RNA sequenced AML cell lines that overexpress EVI1 resulting from different genetic causes or do not express EVI1 at all.","project":"ERP004617"}
{"number_samples":6,"species":"human","abstract":"RNAseq to assess changes in transcript abundance upon knockdown of the RNA-binding protein Unkempt in human SH-SY5Y cell lines.","project":"ERP004682"}
{"number_samples":5,"species":"human","abstract":"Ribosome profiling to assess changes in ribosome occpancy upon overexpression of the RNA-binding protein Unkempt (most likely acts as a translational regulator) in human HeLa cell lines.","project":"ERP004683"}
{"number_samples":7,"species":"human","abstract":"iCLIP experiments tomap the RNA binding sites of the RNA-binding protein Unkempt across the transcriptome in SH-SY5Y cells, HeLa cells with ectopic Unk expression and mouse E15 embryonic brain samples. Expression of Unk is normally largely restricted to the nervous system. We therefore mapped the binding sites in human SH-SY5Y and mouse E15 brain to detect its physiological binding sites (in SH-SY5Y, we also performed the RNAseq experiment upon Unk knockdown). HeLa cells on the other hand normally don't express Unk, but convert to neuron-like shape when the protein is ectopically expressed. So, here we hoped to identify those binding events (and hence target transcripts) that are critical for this morphological transformation.","project":"ERP004684"}
{"number_samples":8,"species":"human","abstract":"Systematic survey of gene and isoform allele-specific expression in human brain and liver tissues, and description of optimised bioinformatic and statistical methods to accurately measure allele-specific expression. DNA-seq data for the same set of samples have been deposited at the European Nucleotide Archive under project accessino PRJEB5279 ( http://www.ebi.ac.uk/ena/data/view/PRJEB5279 ).","project":"ERP004697"}
{"number_samples":6,"species":"human","abstract":"Atmospheric pressure cold plasma (APCP) is a novel tool in medicine for tissue disinfection. It is  produced by a device that ionizes a flow of helium gas partially mixed with air. We recently reported that 2-minute of APCP application significantly reduce the microbial viability without evident damage on the ocular cells and tissues. To validate its use as antimicrobial for biomedical purposes such as ophthalmic applications, with this study we verified the functional changes in human corneas exposed to APCP by means of global gene expression analysis. To our knowledge, this is the first analysis of the effects of cold plasma on gene expression.","project":"ERP005189"}
{"number_samples":2,"species":"human","abstract":"We are human embryologists from center for reproductive medicinel of Anhui Provincial Hospital Affiliated to Anhui Medical University, and we have the expertise to do all that properly in humans. By deep sequencing, the current experiment determined the miRNA profile of two intrafollicular somatic cell types: CRCs and COCs, isolated from women undergoing controlled ovarian stimulation and in vitro fertilization treatment. Ovarian follicles, which are a densely-packed shell of granulosa cells that contains an immature or mature oocyte, are above all responsible for the development, maturation, and release of mature egg for fertilization. They are also responsible for synthesizing and secreting hormones that are essential for follicular development, menstrual and estrous cycle, maintenance of the reproductive tracts and their functions, development of female secondary sex characteristics, and metabolism. During folliculogenesis, ovarian granulosa cells surrounding the oocyte differentiate into mural granulosa cells, involved in gonadal steroidogenesis, and into cumulus cells, which are ovulated with the oocyte at ovulation. These cumulus cells derive from the same population of early follicles, but differentiate into two distinct groups of cells: 1) Those directly lie on the zona pellucida are composed of the so called \"corona radiata cells\".(CRCs)  2) The other group surrounds the CRCs and consists of more numerous cells, forming the so called \"cumulus oophorus cells (COCs)\". In the present study, we described the miRNA expression profile to characterize the ensemble of both known and novel miRNAs expressed in CRCs, as well as in COCs, by using high-throughput Solexa technology.","project":"ERP005274"}
{"number_samples":8,"species":"human","abstract":"Investigating the response of long non-coding RNA expression to LPS stimulation in human monocytes.  This experiment is related to the microarray study E-MTAB-2408.","project":"ERP005301"}
{"number_samples":6,"species":"human","abstract":"RNA-seq analysis of poly-A RNA from untreated U2OS cells, and from U2OS cells exposed to heat stress (43 C) for 4 hours","project":"ERP005426"}
{"number_samples":4,"species":"human","abstract":"Overexpression of the transcription factor RHOXF2 in the HEK293_M2 cells confers resistance to several DNA damaging agents. To understand that resistance mechanism, we cultured HEK293_M2 with or without overexpression of RHOXF2 in the presence of 40nM of mitomycin C or DMSO (control) and profiled their transcriptional response by RNA-seq.","project":"ERP005583"}
{"number_samples":256,"species":"human","abstract":"The zygote and blastomeres of a cleavage stage mouse embryo have the capacity to differentiate to all lineages in the embryo proper and extraembryonic tissues and are considered totipotent. Mouse ESCs can differentiate to embryonic lineages but have restricted potential to trophoblasts. We report here that cultures of a new type of stem cells with expanded potential (EPSCs) can be established from in vitro cultured mouse preimplantation embryos and individual blastomeres, from directly converting mouse ESCs, or from genetically reprogramming somatic cells. EPSCs differentiate to trophoblasts in vitro, and a single EPSC contributes in chimeras to both extraembryonic lineages including the placenta trophoblasts and the embryo proper. Molecular analyses reveal that EPSCs express core pluripotency factors similar to ESCs, but have enriched signature of preimplantation blastomeres. The data described here suggest that similar cell lines from other mammalian species might also be established in culture and stably maintained.","project":"ERP005641"}
{"number_samples":18,"species":"human","abstract":"Brown adipose tissue (BAT) evolved in mammals as a natural defence system against hypothermia and obesity. While existence of BAT in adult humans has been recently appreciated, its cellular origin and molecular identity remain elusive due in large to high cellular heterogeneity within adipose tissues.  Here we isolated clonal adipocytes from adult human BAT as well as WAT (control) and critically analyzed their transcriptome to identify bona fide BAT markers and its new functions.","project":"ERP005953"}
{"number_samples":8,"species":"human","abstract":"SLC35F2 was identified as a transporter of DNA damage inducing agent YM155. We then did a comparative transcriptomics experiment comparing wild-type and SLC35F2 knock-out KBM7 cells.","project":"ERP006028"}
{"number_samples":85,"species":"human","abstract":"Background: Genomic analyses of hundreds of prostate tumors have defined a diverse landscape of mutations and genome rearrangements, but the transcriptomic effect of this complexity is less well understood, particularly at the individual tumor level. We selected a cohort of 25 high-risk prostate tumors, representing the lethal phenotype, and applied deep RNA-sequencing and matched whole genome sequencing, followed by detailed molecular characterization.   Results: Ten tumors were exposed to neo-adjuvant hormone therapy and expressed marked evidence of therapy response in all except one extreme case, which demonstrated early resistance via apparent neuroendocrine transdifferentiation. We observe high inter-tumor heterogeneity, including unique sets of outlier transcripts in each tumor. Interestingly, outlier expression converged on druggable cellular pathways associated with cell cycle progression, translational control or immune regulation, suggesting distinct contemporary pathway affinity and a mechanism of tumor stratification. We characterize hundreds of novel fusion transcripts, including a high frequency of ETS fusions associated with complex genome rearrangements and the disruption of tumor suppressors. Remarkably, several tumors express unique but potentially-oncogenic non-ETS fusions, which may contribute to the phenotype of individual tumors, and have significance for disease progression. Finally, one ETS-negative tumor has a striking tandem duplication genotype which appears to be highly aggressive and present at low recurrence in ETS-negative prostate cancer, suggestive of a novel molecular subtype.   Conclusions: The multitude of rare genomic and transcriptomic events detected in a high-risk tumors cohort offer novel opportunities for personalized oncology and their convergence on key pathways and functions has broad implications for precision medicine.","project":"ERP006077"}
{"number_samples":2,"species":"human","abstract":"We infected the human macrophage-like cell line THP-1 with the attenuated M. tuberculosis surrogate M. bovis Bacillus Calmette–Guérin (M. bovis BCG). We employed microbial enrichment combined with specific ribosomal RNA depletion to simultaneously analyze the transcriptional responses of host and pathogen during infection by dual RNA sequencing.","project":"ERP006097"}
{"number_samples":12,"species":"human","abstract":"The effect of the GSK3 inhibitor Azakenpaullone (Azak) and the differentiation agent retinoic acid (RA) was studied in low and high MYCN levels in the MYCN inducible neuroblastoma cell line SH-SY5Y/6TR(EU)/pTrex-Dest-30/MYCN (SY5Y-MYCN).  Azak was dissolved in DMSO in order to apply it to the cells. Therefore a vehicle control consisting of SY5Y-MYCN cells treated with 24h 1µl/ml DMSO only was used in duplicates. Doxycycline (Sigma) dissolved in water was used at a final concentration of 1µg/ml to induce MYCN expression in SY5Y-MYCN. A co-treatment study with Dox and Azak was conducted. SY5Y-MYCN cells were treated with 24h Azak, 24h Azak & 48h Dox and 48h Dox, with biological duplicates. 1 µM RA (dissolved in DMSO) and 1 µg/mL Doxycycline were given individually and in combination. SY5Y-MYCN cells were treated with 24h RA, 24h RA & 48h Dox, and 48h Dox and RNA was extracted in biological duplicates. For the 24h RA & 48h Dox co-treatment cells were treated with Dox for 24h and then with RA and fresh Dox for a further 24h.","project":"ERP006185"}
{"number_samples":16,"species":"human","abstract":"RNA-Seq data of U2AF1 isoform depletion experiments.","project":"ERP006200"}
{"number_samples":35,"species":"human","abstract":"We are generating macrophage and intestinal organoids from primary mouse and human tissues or induced pluripotent stem cells (iPSCs), these macrophages and organoids are challenged with live or dead pathogens/pathogen components in combination with various immune regulatory cytokines and growth factors.","project":"ERP006215"}
{"number_samples":35,"species":"human","abstract":"In this study, we aim to perform gene expression profiling of undifferentiated human pluripotent stem cells (hIPSC)  and macrophages and their progenitors derived from hIPSC.  The aims of the experiment is to identify a set of genes uniquely expressed in hIPSC but not in hIPSC derived monocytes/macrophages (hIPSdM) and define them as \"stemness\" genes. We will also identify genes upregulated in hIPSC-derived monocytes/macrophages but not hIPSC to provide insights into the regulatory switches of hIPSC differentiation.  We established protocols to generate monocytes and macrophages from hIPSC CRL1 line in vitro. Briefly,  hIPSC were grown in bacteriological dishes in the presence of hIPSC base medium to generate embryoid bodies (EBs). Subsequently, EBs were transferred to gelatinized tissue culture dish in medium supplemented with M-CSF and IL-3. Supernatants containing macrophage progenitors were collected, spun down and plated onto bacteriological dishes  to differentiate into macrophages. Two or three biological replicates of undifferentiated hIPSCs, macrophage progenitors and macrophages were collected and their RNA were isolated via the Qiagen RNA Isolation Kit. The RNA were treated with RNAse free DNase and eluted in RNase-free water. THe RNA integrity (RIN >8) were obtained for all samples by Agilent 2100 Bioanalyzer. We aim to sequence 5GBp per replicate per lane on an Illumina platform.","project":"ERP006216"}
{"number_samples":12,"species":"human","abstract":"APOBEC3G and APOBEC3F are cell-encoded restriction factors that have evolved to counteract virus infections. When packaged into HIV-1 particles, A3G and A3F impair reverse transcription and induce the hypermutation of viral DNA. We employed a next generation sequencing approach to identify the RNAs that these proteins bind in HIV-1 infected cells and HIV-1 virions. We then analysed the mechanism of packaging of APOBEC3 in detail.","project":"ERP006368"}
{"number_samples":8,"species":"human","abstract":"Untreated neuroblastoma cell lines SMS-KCN, SMS-KCNR, Kelly, and SH-SY5Y were sequenced on an Illumina GA IIx sequencer.","project":"ERP006533"}
{"number_samples":18,"species":"human","abstract":"MYCN overexpression in the doxycline MYCN inducible cell line SH-SY5Y/6TR(EU)/pTrex-Dest-30/MYCN (SY5Y-MYCN) using the dynamic transcriptome analysis (DTA) protocol. Samples at different time-points after MYCN over-expression and uninduced controls (0h) were sequenced in duplicates. The experimental setup consists of [A] total mRNA at 0h, 1h, 4h, 24h, which corresponds to the standard mRNA-seq protocol, [B] 4-thioUridine (4sU) labelled mRNA 30 min before extraction at time-points 0h, 1h, 4h, which is the freshly transcribed mRNA within the last 30 min before the time-point and [C] the counter-part, 4sU unlabelled mRNA 30 min before extraction which corresponds to the mRNA from before 30 min of extraction. The samples were sequenced on an Illumina GA IIx using the Illumina protocols.","project":"ERP006538"}
{"number_samples":10,"species":"human","abstract":"Neuroblastoma is one of the most common extracranial solid tumors in children. By sequencing transcriptomes of low- and high-risk neuroblastomas, we have identified differentially expressed annotated and novel long noncoding RNAs (lncRNAs). Our analysis identifies a lncRNA neuroblastoma associated transcript-1 (NBAT-1) as a biomarker for predicting the clinical outcome of the neuroblastoma patients. CpG methylation and a high-risk neuroblastoma associated SNP on chromosome 6p22 functionally contribute to differential expression of NBAT-1. Loss of NBAT-1 results in increased proliferation and invasion of cells. It controls these processes via epigenetic silencing of target genes. Its loss affects neuronal differentiation through activation of the neuronal-specific transcription factor NRSF/REST. Thus, loss of NBAT-1 contributes to aggressive neuroblastoma by increasing proliferation and impairing differentiation of neuronal precursors.","project":"ERP006577"}
{"number_samples":29,"species":"human","abstract":"RNA-seq was performed of tissue samples from 127 human individuals representing 32 different tissues in order to study the human tissue transcriptome. This submission contains 32 samples and the remaining data is located at E-MTAB-1733.","project":"ERP006650"}
{"number_samples":32,"species":"human","abstract":"The aim of the study is to analise the transcriptional profile of single leukemic cells in order to understand the early stage of leukaemogenesis. Oligoclonal haemopoetic and progenitor cell populations from several leukemic mutants were flow sorted into single cells and processed for RNAseq.","project":"ERP006662"}
{"number_samples":15,"species":"human","abstract":"Identification of Wig-1-associated RNAs by RNA-immunoprecipitation followed by high-throughput sequencing in HCT116 and Saos2 cancer cell lines","project":"ERP006686"}
{"number_samples":12,"species":"human","abstract":"In this study RNA was extracted from the olfactory mucosa of three individuals of four mammalian species: cow, macaque, marmoset and humans. The RNA was sequenced using the Illumina Hiseq2500 platform.","project":"ERP006696"}
{"number_samples":6,"species":"human","abstract":"Human pluripotent cells were reset to ground state pluripotency by transient overexpression of NANOG and KLF2 and subsequent inhibition of ERK and protein kinase C. Transcriptional profiling of reset cells and conventional pluripotent stem cell cultures was carried out by RNA-seq, in tandem with mouse embryonic stem cells propagated under similar conditions to assess the combinatorial effects of MEK inhibitor PD0325901, GSK3 inhibitor CHIR99021 and PKC inhibitor Go6983.","project":"ERP006823"}
{"number_samples":2,"species":"human","abstract":"Human embryonic stem cells (WA01) were differentiated in a step-wise manner into three-dimensional human gastric organoids (hGOs). At day 34 of differentiation, the hGOs were collected and analyzed by RNA-sequencing.","project":"ERP006872"}
{"number_samples":6,"species":"human","abstract":"The epigenetic dysregulation of tumor suppressor genes is a major driver of human carcinogenesis. We have combined genome-wide methylation analyses with functional screening to identify novel candidate tumor suppressor genes in diffuse large B-cell lymphoma (DLBCL). We find that the dual-specificity phosphatase DUSP4 is aberrantly silenced in nodal and extranodal DLBCL due to promoter hypermethylation; ectopic expression of wild type DUSP4, but not of a phosphatase-deficient mutant, dephosphorylates c-JUN N-terminal kinase (JNK) and induces apoptosis in DLBCL cells. JNK inhibition prevents DLBCL survival in vitro and in vivo, and synergizes strongly with inhibitors of chronic active B-cell receptor signaling. Our results provide a mechanistic basis for the clinical development of JNK inhibitors in DLBCL, alone or in synthetic lethal combinations. A methylation profiling data set related to this experiment was also deposited at ArrayExpress under accession number E-MTAB-2926: http://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-2926/","project":"ERP006964"}
{"number_samples":3,"species":"human","abstract":"RNAseq was prepared in parallel with ribosome profiling, using the same protocol for cDNA library preparation.","project":"ERP007008"}
{"number_samples":3,"species":"human","abstract":"Long noncoding RNAs (lncRNAs) regulate gene expression.","project":"ERP007014"}
{"number_samples":1,"species":"human","abstract":"Background: The human neuroblastoma cell line, SH-SY5Y, is a commonly used cell line in studies related to neurotoxicity, oxidative stress, and neurodegenerative diseases. Although this cell line is often used as a cellular model for Parkinson’s disease, the relevance of this cellular model in the context of Parkinson’s disease (PD) and other neurodegenerative diseases has not yet been systematically evaluated.  Results: We have used a systems genomics approach to characterize the SH-SY5Y cell line using whole-genome sequencing to determine the genetic content of the cell line and used transcriptomics and proteomics data to determine molecular correlations. Further, we integrated genomic variants using a network analysis approach to evaluate the suitability of the SH-SY5Y cell line for perturbation experiments in the context of neurodegenerative diseases, including PD.  Conclusions: The systems genomics approach showed consistency across different biological levels (DNA, RNA and protein concentrations). Most of the genes belonging to the major Parkinson’s disease pathways and modules were intact in the SH-SY5Y genome. Specifically, each analysed gene related to PD has at least one intact copy in SH-SY5Y. The disease-specific network analysis approach ranked the genetic integrity of SH-SY5Y as higher for PD than for Alzheimer’s disease but lower than for Huntington’s disease and Amyotrophic Lateral Sclerosis for loss of function perturbation experiments.   Keywords (3-10): SH-SY5Y, cell line, whole genome sequencing, RNA-seq, proteomics, cell line suitability evaluation, Parkinson’s Disease","project":"ERP007022"}
{"number_samples":16,"species":"human","abstract":"RNA-seq was performed for transcriptional analysis of MCF10A cells, an epithelial mammary cell line. MCF10A cells were cultured in 3D acinus forming conditions (in Matrigel). Timepoints analysed were 24h, 34h, 36h, 38h and 48h into acinus formation. Control cells were monoloayer.","project":"ERP007109"}
{"number_samples":134,"species":"human","abstract":"The HipSci project brings together diverse constituents in genomics, proteomics, cell biology and clinical genetics to create a UK national iPS cell resource and use it to carry out cellular genetic studies.  In this sub-study we perform RNAseq on iPS cells generated from skin biopsies from healthy volunteers.","project":"ERP007111"}
{"number_samples":38,"species":"human","abstract":"Single-cell RNA-seq of DDX4 FACS sorted cells from ovarian cortical tissue. Regarding singe cell quality, genes were filtered based on a RPKM greater than or equal to 1 in at least one cell, where the RPKM cutoff was determined from histograms of number of expressed transcripts versus RPKM, and it was also sensible with respect to the number of molecules per RPKM estimated from the ERCC spike-in. Three clear outliers were observed in a principal component analysis and therefore removed from further analysis.","project":"ERP007170"}
{"number_samples":12,"species":"human","abstract":"ES cells grown in culture are heterogeneous with respect to their transcriptional profiles, their methylation landscape and, functionally, their potency. This study is investigating overall changes in transcriptional landscape of ES cells upon overexpression of epigenetic regulators which can indicate a shift in the distribution of subpopulations.","project":"ERP007180"}
{"number_samples":57,"species":"human","abstract":"Whole transcriptome sequencing of 19 matched patient sample trios: normal oral epithelium, oral dysplasia and oral squamous cell carcinoma from the same patent. All material is from FFPE blocks that have been stained and sections and had sample histology confirmed by 2 independent pathologists. A ScriptSeq library kit was used to create strand specific 100bp paired-end sequencing libraries following RiboZero treatment to deplete rRNA. Both coding and non-coding RNA <200bp is sequenced on an Illumina HiSeq2500 with an average of 4 samples per lane.","project":"ERP007185"}
{"number_samples":18,"species":"human","abstract":"We have develop a simple fact method to isolate transcription factories and analyzed their RNA content as compared to total poly(A)+ and chromatin associated RNA fractions in presence or absence of pro-inflammatory cytokine.","project":"ERP008445"}
{"number_samples":7,"species":"human","abstract":"RNA-sequencing experiments performed on control siRNA treated and PRPF8 depleted cells","project":"ERP008458"}
{"number_samples":15,"species":"human","abstract":"Technological advances have enabled the analysis of cellular protein and RNA levels with unprecedented depth and sensitivity, allowing for an unbiased re?evaluation of gene regulation during fundamental biological processes. Here, we have chronicled the dynamics of protein and mRNA expression levels across a minimally perturbed cell cycle in human myeloid leukemia cells using centrifugal elutriation combined with mass spectrometry?based proteomics and RNA?Seq, avoiding artificial synchronization procedures.","project":"ERP008483"}
{"number_samples":11,"species":"human","abstract":"RNA-seq of 11 osteosarcoma cell lines","project":"ERP008492"}
{"number_samples":4,"species":"human","abstract":"We use a novel approach (an analysis of metatranscriptomics data) to study antibiotic resistance, a topic that poses major health care concerns. We investigate resistance gene expression in microbial communities in a diverse range of environments.","project":"ERP008520"}
{"number_samples":87,"species":"human","abstract":"RNA-Seq analysis of human fibroblasts, induced neurons and cortex from donors of several ages","project":"ERP008608"}
{"number_samples":4,"species":"human","abstract":"The genomic regulatory programs that underlie human organogenesis are poorly understood. Human pancreas development, in particular, has pivotal implications for pancreatic regeneration, cancer, and diabetes. We have now created maps of transcripts, active enhancers, and transcription factor networks in pancreatic multipotent progenitors obtained from human embryos, or derived in vitro from human embryonic stem cells. This revealed that artificial progenitors recapitulate salient transcriptional and epigenomic features of their natural counterparts. Using this resource, we show that TEAD1, a transcription factor controlled by Hippo signaling, is a core component of the combinatorial code of pancreatic progenitor enhancers. TEAD thus activates genes encoding regulators of signaling pathways and stage-specific transcription factors that are essential for normal pancreas development. Accordingly, chemical and genetic perturbations of TEAD and its coactivator YAP inhibited expression of known regulators such as FGFR2 and SOX9, and suppressed the proliferation and expansion of mouse and zebrafish pancreatic progenitors. These findings provide a resource of active enhancers and transcripts in human pancreatic multipotent progenitors, and uncover a central role of TEAD and YAP as signal-responsive regulators of the transcriptional program of early pancreas development.","project":"ERP008682"}
{"number_samples":6,"species":"human","abstract":"PPARb/d agonist and antagonist response was characterized in (human) monocyte derived macrophages.","project":"ERP008803"}
{"number_samples":4,"species":"human","abstract":"RNA-Seq was carried out in order to obtain the expression profile of Solute Carrier Family (SLC) of proteins in two commonly used celllines. We were specifically interested in the subset of SLCs that are capable of transporting amino acids.","project":"ERP008834"}
{"number_samples":108,"species":"human","abstract":"The activation of receptors displayed on the surface of cells typically initiates cytoplasmic signalling cascades that lead to changes in cellular transcriptional responses so that a cell expresses proteins that enable it to behave appropriately as a function of their position within an organism. Understanding these signals and learning how to either prevent or ectopically activate them could have many therapeutic benefits and applications. In some cases these signals are highly localised, being delivered by directly neighbouring cells that express ligands which specifically bind to and activate receptors; for example, stem cells display receptors for ligands provided by cells within their niche to both ensure continual production of cells, and that their cellular progeny are directed towards appropriate fates. While for some cell types we know something of these signals enabling the in vitro differentiation of stem cells towards certain cellular fates, the differentiation processes is often not complete and so “immature” cell types which are not fully functional are produced, implying that some important signals are missing. Here, we propose to systematically identify receptor-triggered signalling pathways by stimulating receptors on the surface of human stem cells by using a library of soluble recombinant oligomerised protein ligands and measuring the consequent transcriptional perturbations using RNA-seq. We propose to screen an existing library of ~50 oligomerised human ligands previously used in receptor protein interaction screens and first test them for their ability to bind to human stem cells to identify those ligands for which the stem cells express a cell surface receptor. For those ligands which interact with the stem cells (we estimate between 2 and 5), they will be added to stem cell cultures and their effects on transcription quantified. The budget requested is sufficient for us to include controls and test important parameters such different time points after addition of the proteins.","project":"ERP008967"}
{"number_samples":9,"species":"human","abstract":"RNAseq was carried out to compare global transcriptional profiles of human pluripotent H9 stem cells with tissues derived from these cells through directed differentiation, including endoderm and human intestinal organoids.","project":"ERP008992"}
{"number_samples":13,"species":"human","abstract":"The transcriptome of serous ovarianc cancer tumor associated macrophages was characterized. Additionally, their transcription response to PPARD agonists and inverse agonists in cell culture was analyzed.","project":"ERP009022"}
{"number_samples":4,"species":"human","abstract":"Transfer RNAs (tRNAs) are key adaptor molecules in the protein translation machinery. In order to become fully active, they are heavily modified posttranscriptionally. Inosine at the first residue of the anticodon (position 34; I34) is an essential tRNA modification that has been poorly studied thus far. The modification in eukaryotes results from a deamination reaction of adenine that is catalysed by the heterodimeric Adenosine Deaminase Acting on tRNA (hetADAT), composed of two subunits: ADAT2 and ADAT3. Using high-throughput small RNA sequencing, we show that this modification takes place in human tRNAs at the precursor tRNA level and as this precursor maturates. We also functionally validated the human genes encoding for hetADAT and show that in cells, HsADAT2 and HsADAT3 co-localize in nucleus. Lastly, we knocked down HsADAT2 and show for the first time that it is possible to modulate the levels of I34 modification on all human hetADAT tRNA substrates. These findings give mechanistic insights on the biology of tRNA modifications and the tools reported in this study will prove useful in the future to address the exact molecular role of I34 modification on protein translation.","project":"ERP009050"}
{"number_samples":2,"species":"human","abstract":"Polycomb repressive complexes (PRC) play a critical role during tumorigenesis and development. The histone methyltransferase Enhancer of Zeste homologue 2 (EZH2), as a core component of PRC2, is frequently overexpressed in a wide variety of cancers. EZH2-mediated gene silencing contributes to carcinogenesis. To further the understanding of the EZH2 biology in gastric cancer, here we performed transcriptome analyses and identified EZH2-responsive genes upon EZH2 knockdown by RNA-seq.","project":"ERP009263"}
{"number_samples":18,"species":"human","project":"ERP009282"}
{"number_samples":6,"species":"human","abstract":"Circular RNAs (circRNAs), formed by non-sequential back-splicing of pre-mRNA transcripts, are a widespread form of non-coding RNA in animal cells. However, it is unclear whether the majority of circRNAs represent splicing by-products without function, or are produced in a regulated manner to carry out specific cellular functions. We show that hundreds of circRNAs are regulated during human epithelial-mesenchymal transition (EMT) and find that the production of over one-third of abundant circRNAs is dynamically regulated by the alternative splicing factor, Quaking (QKI), which itself is regulated during EMT. Furthermore, by modulating QKI levels we show the effect on circRNA abundance is dependent on intronic QKI binding motifs. Critically, the addition of QKI motifs is sufficient to induce de novo circRNA formation from transcripts that are normally linearly spliced. These findings demonstrate circRNAs are both purposefully synthesised and regulated by cell-type specific mechanisms, suggesting they play specific biological roles in EMT.","project":"ERP009283"}
{"number_samples":7,"species":"human","abstract":"The goal of this project is to investigate genes regulating definitive endodermal differentiation of human iPSC using genetic screening. The differentiation of human iPSC into definitive endoderm is going to be examined by global RNA expression patterns.","project":"ERP009295"}
{"number_samples":384,"species":"human","abstract":"DCM,dilated cardiomyopathy, is a major cause of chronic heartfailure.  In this study,128 samples from DCM patients (left ventricle) have been sequenced by RNAseq. We obtained cardiac tissue from 128 subjects with end-stage heart failure.  All subjects were transplanted at the Royal Brompton and Harefield NHS Foundation Trust (London, United Kingdom).  On this occasion, cardiac samples were obtained and immediately frozen in liquid nitrogen.","project":"ERP009437"}
{"number_samples":23,"species":"human","abstract":"HepG2 cells (human liver hepatocarcinoma) were exposed for 6 different time points (6,12,18,24,36 and 48h) to Benzo[a]pyrene (BaP) in duplicate. Total RNAs were ribo-depleted and sequenced on an Illumina Hiseq2000 in pair-end 100 bp reads. One of the duplicates for BaP-treated sample at the 24 hour time point was found to have a minor fastq file integrity issue and was removed from this data set as a precaution.","project":"ERP009514"}
{"number_samples":6,"species":"human","abstract":"Human lung organoids (HLOs) were derived from human embryonic stem cell line H9 NIH registry #0062. Sequencing of HLOs was performed by the UM DNA Sequencing Core, using Illumina Hi-Seq platform and single-end 50 bp reads. The UM Bioinformatics Core downloaded the reads files from the Sequencing Core storage, and concatenated those into a single .fastq file for each sample. 3 HLOs were cultured for 65 days and 3 HLOs were cultured in vitro for 110 days.  HLO Culture Protocol 4-day Activin A (R&D systems) differentiation protocol was used to derive definitive endoderm from human pluripotent stem cells (hPSCs). Cells were treated with Activin A (100?ng?ml-1) for three consecutive days in RPMI 1640 media (Life Technologies) with increasing concentrations of 0%, 0.2% and 2% HyClone defined fetal bovine serum (dFBS, Thermo Scientific). The fourth day was a repeat of day 3 media (2% HyClone FBS).   After differentiation into definitive endoderm, cells were incubated in foregut media (advanced DMEM/F12 plus N-2 and B27 supplement, 10mM Hepes, 1x L-Glutamine (200mM), 1x Penicillin-streptomycin (5,000 U/mL, all from Life Technologies)) with 200ng/mL Noggin (NOG, R&D Systems) and 10µM SB431542 (SB, Stemgent) for 4 days, SB, 500ng/mL FGF4 (R&D Systems), and 2 µM CHIR99021 (Chiron, Stemgent), and 1µM SAG (Enzo Life Sciences) for 4-6 days. After 4 days with treatment of growth factors, three-dimensional floating spheroids were present in the culture. Three-dimensional spheroids were transferred into Matrigel to support 3D growth. Spheroids were embedded in a droplet of Matrigel (BD Bioscience  #356237) in one well of a 24 well plate, and incubated at room temperature for 10 minutes. After the Matrigel solidified, foregut media with 1% Fetal bovine serum (FBS, CAT#: 16000-044, Life Technologies) and 500ng/mL FGF10 (R&D Systems) were overlaid and replaced every 4 days. Organoids were transferred into new Matrigel droplets every 10 to 15 days.","project":"ERP009612"}
{"number_samples":58,"species":"human","abstract":"In this study we used human primary granulocytes to assess natural variability of lncRNA expression. We used polyA+ granulocyte RNA-seq from 10 healthy individuals to define granulocyte lncRNA transcriptome, which was not available before. We annotated 6,249 lncRNA transcripts forming 1,591 lncRNA loci, contributing 268 novel loci to the human genome annotation. We show that focusing on granulocytes allows identification of less well expressed, less efficiently spliced and more granulocyte specific lncRNAs. We used Ribosomal depleted granulocyte RNA-seq from 7 healthy individuals sampled in 3 replicates to estimate reproducibility and variability of lncRNA expression and found that though being as well reproducible between replicates, lncRNAs are significantly more variable in their expression than mRNAs. We confirmed this finding in publicly available Geuvadis lymphoblastoid cell line (LCL) RNA-seq dataset from 462 individuals and also used this dataset to show that the number of identified lncRNA increases with the number of donors analyzed. Overall, we showed that high variability of lncRNAs is an important feature that distinguishes them from mRNAs and influences their identification strategy, making it necessary to include as many individuals as possible into the identification effort for every given cell type in order to comprehensively annotate all the lncRNAs in the human genome.","project":"ERP009768"}
{"number_samples":6,"species":"human","abstract":"In this study RNA was extracted from the olfactory mucosa of three human individuals. The RNA was sequenced using the Illumina Hiseq2500 platform.","project":"ERP009841"}
{"number_samples":6,"species":"human","abstract":"The aim of this experiment was to determine the amount of RNA-seq signal degradation that results from MARIS and to test how well 4 different RNA-seq library construction techniques perform on partially degraded RNA.","project":"ERP009913"}
{"number_samples":7,"species":"human","abstract":"The aim of this experiment was to observe the transcriptional profile of Ins+ cells in human cadaveric islets and at the terminal stage of our in vitro beta-cell differentiation protocol across both the Hues8 and H1 cell lines. One polyhormonal sample (Ins+/Gcg+) was also sorted  for additional comparison.","project":"ERP009935"}
{"number_samples":24,"species":"human","abstract":"Ataxin-2 (ATXN2) polyglutamine domain expansions of large size result in an autosomal dominantly inherited multi-system-atrophy of the nervous system named Spinocerebellar ataxia type 2 (SCA2), while expansions of intermediate size act as polygenic risk factors for levodopa-responsive Parkinson’s disease (PD) and for motor neuron disease (ALS and FTLD). In view of the established role of ATXN2 for RNA processing and the expression of ATXN2 in blood cells such as platelets, we investigated whether global deep RNA sequencing of whole blood from SCA2 patients identifies a molecular profile which might serve as diagnostic biomarker. The bioinformatic analysis of the SCA2 blood transcriptome revealed various significant effects, prominently on the pathways of Huntington’s disease and PD where mitochondrial dysfunction is crucial. Notably, an induction of PINK1 and PARK7 expression was observed.  Conversely, PINK1 expression was severely decreased upon global transcriptome profiling of Atxn2-knockout mouse cerebellum and liver, in parallel to strong effects on OPA1 and GHITM, which encode known mitochondrial dynamics regulators.  These results were validated by quantitative PCR and immunoblots. ATXN2 expression was induced by nutrient deprivation of human SH-SY5Y neuroblastoma cells, with a time course in parallel to PINK1 expression, and PINK1 levels appeared to depend on ATXN2 knockdown. Thus, ATXN2 may be upstream of or parallel to PINK1 in mitochondrial quality control and autophagy. Given that PINK1 is responsible for autosomal recessive juvenile Parkinson’s disease (PD), this genetic interaction offers an explanation how the degeneration of nigrostriatal dopaminergic neurons and the Parkinson phenotype are triggered by ATXN2 mutations.","project":"ERP009992"}
{"number_samples":40,"species":"human","abstract":"Parkinson’s disease (PD) is a frequent neurodegenerative process at old age. Accumulation and aggregation of the lipid-binding SNARE complex component alpha-synuclein (SNCA) underlies this vulnerability and defines stages of disease progression. Determinants of SNCA levels and mechanisms of SNCA neurotoxicity are intensely investigated. In view of physiological SNCA roles in blood to modulate vesicle release, we studied blood samples from a new large pedigree with SNCA gene duplication (PARK4 mutation), to identify effects of SNCA gain-of-function as potential disease biomarkers. The expression of other Parkinson’s disease gene was not, but CPLX1 mRNA downregulation was correlated with genotype. In global RNAseq profiling of blood from presymptomatic PARK4, bioinformatics detected significant upregulations for platelet activation, hemostasis, lipoproteins, endocytosis, lysosome, cytokine, toll like receptor signalling and extracellular pathways. In PARK4 platelets, stimulus-triggered degranulation was impaired. Strong SPP1, GZMH, and PLTP mRNA upregulations were validated in PARK4. Analysing prodromal stages of general PD, only blood CPLX1 levels were altered in cases with REM sleep behaviour disorder (RBD). Validation experiments confirmed an inverse mutual regulation of SNCA and CPLX1 mRNA levels. In the 3’-UTR of the CPLX1 gene we identified a SNP that is significantly associated with PD risk. In summary, our data provide functional insights into the role and regulation of blood alpha-synuclein levels. The novel blood biomarkers of synucleinopathy may become useful for PD prediction.","project":"ERP010003"}
{"number_samples":8,"species":"human","abstract":"Aim: To determine the baseline gene expression profile of human gallbladder and cystic duct cells at various stages from isolation in fresh biopsy samples to early and later passages in culture.","project":"ERP010065"}
{"number_samples":16,"species":"human","abstract":"Aim: To determine the baseline miRNA expression profile of human gall bladder and cystic duct cells at various stages from isolation in fresh biopsy samples to early and later passages in culture.","project":"ERP010082"}
{"number_samples":3,"species":"human","abstract":"The aim of this experiment was to identify novel, small (<600bp), capped, polyA-tailed RNA transcripts in mouse E14.5 pancreas, liver, and brain and at stages S1, S2, S3 within our human in vitro beta-cell differentiation protocol. Low-molecular-weight (LMW) and high-molecular-weight (HMW) fractions were separated.","project":"ERP010136"}
{"number_samples":78,"species":"human","abstract":"Despite the explosion in numbers of cancer genomic studies, metastasis is still the major cause of cancer mortality. In breast cancer, approximately one-fifth of metastatic patients survive 5 years. Therefore, detecting the patients at a high risk of developing distant metastasis at first diagnosis is critical for effective treatment strategy. We conducted comprehensive genetic analyses to discover driver mutations escalating the risk of metastasis, using Exome and RNA sequencing of the samples of 22 HRM (high-risk for distant metastasis) and 56 LRM (low-risk for distant metastasis) breast cancer patients. We hypothesized that the genetic mutations responsible for the metastasis may be associated with TFs (transcription factors) to perturb the downstream processes which result in DEGs (differentially expressed genes). Based on this, we developed a new integrated systematic approach to predict candidate driver mutations by measuring the sample coverage and the weight of the pathways linking mutations to TFs. The analysis led to the discovery of HRM-specific mutations in ADPGK, NUP93, PCGF6, PKP2, and SLC22A5 which significantly enhanced cancer cell migration and metastatic signaling pathways. The discovered somatic mutations may be useful for identifying patients who are likely to develop distant metastasis. Furthermore, our integrative systematic analyses suggest a new way to discover and understand driver mutations without relying on simple frequency-based analyses.","project":"ERP010142"}
{"number_samples":14,"species":"human","abstract":"We have isolated, analyzed and compared the RNA content of various cell compartments including total RNA, nucleolplasmic RNA and nucleolar RNA.  In addition, cells have been subjected to various treatment to specifically inhibit each of the RNA polymerases I, II and III, as well as protein synthesis. Cells were also treated with antisense oligos to induce the degradation of specific RNA transcripts. The whole RNA content of the cells after these specific treatments was isolated and sequenced.","project":"ERP010177"}
{"number_samples":48,"species":"human","abstract":"The IFNL4 gene is negatively associated with spontaneous and treatment-induced clearance of hepatitis C virus infection. The activity of IFN?4 plays an important causal role in the pathogenesis, but the molecular details are not fully understood. One possible reason for the detrimental effect of IFN?4 could be a tissue specific regulation of an unknown subset of genes. To address both tissue- and subtype subtype-specificity in the interferon response, we treated primary human hepatocytes and airway epithelial cells with IFNa, IFN?3 or IFN?4 and assessed interferon mediated gene regulation using transcriptome sequencing. Our data show a surprisingly similar response to all three subtypes of interferon. We also addressed the tissue tissue-specificity of the response, and identified a subset of tissue tissue-specific genes. However, the interferon response is robust in both tissues with the majority of the identified genes being regulated in hepatocytes as well as airway epithelial cells. Thus we provide an in-depth analysis of the liver interferon response seen over an array of interferon subtypes and compare it to the response in the lung epithelia.","project":"ERP010227"}
{"number_samples":32,"species":"human","abstract":"In cystic fibrosis (CF), chronic Pseudomonas aeruginosa infection of the lower respiratory tract is associated with epithelial inflammation that eventually causes tissue damage. To identify the molecular determinants of an effective response to P. aeruginosa infection, we performed a transcriptomic analysis of primary human bronchial epithelial cells from healthy donors (CTRL) 2, 4, and 6 h after induced P. aeruginosa infection.","project":"ERP010372"}
{"number_samples":11,"species":"human","abstract":"The long (2.2Mb) human dystrophin transcript (DMD) takes 16 hours to be transcribed and is co-transcriptional spliced. The presence of long introns (24 over 10kb long, 5 over 100kb long) poses challenges to the splicing machinery. Additionally, the heterogeneity in intron size suggests that intron removal not always takes place consecutively. We explored the order of intron removal and potential multi-step splicing for the DMD transcripts in human skeletal muscle cell lines. A customized library of probes (120bp) covering all exons and introns, has been generated capturing pre-mRNA and targeting DMD, followed by Hiseq sequencing. SplicePie pipeline has been used to analyze capture-pre-mRNA-sequencing data. Analysis showed that DMD introns can be removed non-sequentially generating exon blocks – joined blocks of exons flanked by unspliced introns. Exon blocks were detected by analysis of the coverage of paired-end reads and validated experimentally using PCR and Sanger sequencing. No correlation between intron length and speed of intron removal was observed. Computational analysis and experimental validation revealed that intron removal takes place in several steps for the majority of dystrophin introns. We found two mechanisms of multi-step intron removal in DMD– recursive and nested splicing. Non-sequential and multi-step splicing events were found throughout the DMD gene across three cell lines. We believe that our findings of non-sequential and multi-step splicing provide insight in the splicing mechanism and will be useful to optimize therapeutic strategies that interfere with the splicing process.","project":"ERP010500"}
{"number_samples":6,"species":"human","abstract":"RNASeq of the human epithelial cell line LoVo following Listeria monocytogenes infection","project":"ERP010786"}
{"number_samples":6,"species":"human","abstract":"More potent targeting of the androgen receptor (AR) in advanced prostate cancer is driving an increased incidence of neuroendocrine prostate cancer (NEPC), an aggressive and treatment-resistant AR-negative variant. Its molecular pathogenesis remains poorly understood but appears to require TP53 and RB1 aberration. We modeled the development of NEPC from conventional prostatic adenocarcinoma using a patient-derived xenograft and found that the placental gene PEG10 is de-repressed during the adaptive response to AR interference and subsequently highly upregulated in clinical NEPC. We found that the AR and the E2F/RB pathway dynamically regulate distinct post-transcriptional and post-translational isoforms of PEG10 at distinct stages of NEPC development. In vitro, PEG10 promoted cell cycle progression from G0/G1 in the context of TP53 loss, and regulated Snail expression via TGF-ß signaling to promote invasion. Taken together, these findings show the mechanistic relevance of RB1 and TP53 loss in NEPC and suggest PEG10 as an NEPC-specific target.","project":"ERP010791"}
{"number_samples":30,"species":"human","abstract":"In this study we performed RNA-seq analysis of gastric corpus biopsies to study transcriptional changes during early gastric carcinogenesis. We included patients with non-atrophic gastritis, with intermediate and extensive corpus atrophy, and patients with intestinal metaplasia. We also included a control group of H. pylori-negative subjects. All subjects were from Nicaragua, which has a population of high gastric cancer risk.  Total RNA samples were treated with RiboZero Magnetic (Human/Mouse/Rat) Kit (Epicentre Biotechnologies) to deplete rRNA and the cDNA libraries were prepared using ScriptSeqTM v2 RNA-Seq Library Preparation Kit (Epicentre). Sequencing was performed on the Illumina HiScan2500 platform, 2*100bp reads.","project":"ERP010889"}
{"number_samples":2,"species":"human","abstract":"The two subunits of IL-35, EBI3 and P35, were fused together and transfected into Panc-1 cell via lentivirus. The sequence of the fused gene is identical to that of a commercial IL-35-overexpessed plasmid (InvivoGEN, pORF9-hIL35elasti). An empty vector was used as the control. The two cell lines were subjected to a genome-wide RNA sequencing.","project":"ERP010929"}
{"number_samples":49,"species":"human","abstract":"Low grade gliomas (LGG; WHO grade 2 astrocytomas, oligodendrogliomas and oligoastrocytomas) account for about 25% of diffuse gliomas. Most occur in young adults between the ages of 30 and 45 years, and are usually only diagnosed after a seizure. In general, they can be characterised by a long period of continuous slow growth, followed by malignant transformation that will be the cause of death up to 25 years after onset. However, there is a significant number of patients for whom malignant progression is more rapid, with mortality observed within 5 years. This suggests that, as with other tumour types, there may be different subtypes of LGG with specific prognosis. It follows that being able to identify these subtypes may permit better patient stratification and aid targeted treatments. Until recently, our understanding of the variables involved in patient prognosis included the type of tumour â?? oligodendroglial tumours indicate better prognosis than oligoastrocytic or astrocytic â?? and presence of the 1p-19q co-deletion. In addition, the recent discovery of mutations in IDH1&2 in the majority of LGGs provided another means of stratifying patients, while offering an important insight into their biology. However, we still understand very little of the biology behind the genesis and progression of the 70-80% of LGG that bear IDH1&2 mutations, let alone the remaining IDH wild-type tumours.","project":"ERP010930"}
{"number_samples":6,"species":"human","abstract":"Elucidation of FOXP1 target genes by RNA-Seq of DLBCL cell lines transfected with siRNAs targeting FOXP1 expression","project":"ERP011000"}
{"number_samples":8,"species":"human","abstract":"Recently, a novel subset of TCRaß+ CD4- CD8- double-negative (DN) T cells has been described to suppress immune responses in both mice and humans. Moreover, in murine models infusion and/or activation of DN T cells specifically suppressed alloreactive T cells and prevented development of Graft-versus-Host disease (GvHD) after allogeneic hematopoietic stem cell transplantation (HSCT). We have demonstrated that human DN T cells like their murine counterparts are highly potent suppressor cells of both, CD4+ and CD8+ T-cell responses. After HSCT and other lymphopenic conditions, Interleukin (IL)-7 plays an important role in the reconstitution, survival, and homeostasis of the T-cell compartment. Since IL-7 was shown to interfere with T-cell functionality, we asked whether IL-7 affects the functionality of human DN T cells. Intriguingly, IL-7 diminished the suppressive activity of DN T cells towards allogeneic CD4+ effector T cells.  Of interest our studies revealed that IL-7 activates the Akt/mTOR pathway in human DN T cells. Importantly, selective inhibition of the protein kinases Akt or mTOR was able to reverse the IL-7 effect thereby restoring the functionality of DN T cells, whereas inhibition of other central T-cell signaling pathways did not. Further analyses suggest that the IL-7/Akt/mTOR signaling cascade downregulates anergy-associated genes and upregulates activation- and proliferation-associated factors which may be crucial for DN T cell functionality. These findings indicate that IL-7 and Akt/mTOR signaling are critical factors for the suppressive capacity of DN T cells. Targeting of these pathways by pharmacological agents may restore and/or enhance DN T-cell functionality in GvHD.","project":"ERP011264"}
{"number_samples":6,"species":"human","abstract":"Quiescent and activated human skeletal muscle stem cells are analyzed by whole-transcriptome RNA-sequencing. These cells are prospectively isolated from latissimus dorsi muscle. Quiescent cells are analyzed immediately following prospective isolation. Activated cells are analyzed after 1 week in culture. Additionally, activated cells (again cultured for one week) and treated with a p38 MAPK inhibitor are analyzed.","project":"ERP011282"}
{"number_samples":6,"species":"human","project":"ERP011301"}
{"number_samples":148,"species":"human","abstract":"RNA sequencing of pancreatic ductal adenocarcinoma (PDAC) primary cultures from five different patient-derived xenograft models grown either in adherent conditions selecting for non-CSCs or in CSC-enriching anchorage-independent sphere conditions. A and B are two biological duplicates, from the same patient-derived xenograft model of PDAC.","project":"ERP011411"}
{"number_samples":8,"species":"human","abstract":"GalNAc-transferase (GalNAc-T) isoforms modify distinct subsets of the O-glycoproteome and GalNAc-type O-glycosylation is found on most proteins trafficking through the secretory pathway in metazoan cells. The O-glycoproteome is regulated by up to 20 polypeptide GalNAc-Ts and the contributions and biological functions of individual GalNAc-Ts are poorly understood. Here, we used a zinc-finger nuclease (ZFN)-directed knockout strategy to probe the contributions of the major GalNAc-Ts (GalNAc-T1 and T2) in liver cells, and explore how the GalNAc-T repertoire quantitatively affects the O-glycoproteome. We demonstrate that the majority of the O-glycoproteome is covered by redundancy, whereas distinct subsets of substrates are modified by non-redundant functions of GalNAc-T1 and T2. Differential transcriptomic analysis indicates that loss of function of a GalNAc-T induces specific transcriptional response. The non-redundant O-glycoproteome subsets for and the transcriptional responses for each isoform appeared to be related to different cellular processes, and for the GalNAc-T2 isoform supporting a role in lipid metabolism. The results demonstrate that GalNAc-Ts have non-redundant glycosylation functions, and that these may affect distinct cellular processes. The data provides a comprehensive resource for unique substrates for individual GalNAc-Ts. Our study provides a new view on the regulation of the O-glycoproteome, suggesting that the plurality of GalNAc-Ts arose to regulate distinct protein functions and cellular processes.","project":"ERP012063"}
{"number_samples":18,"species":"human","abstract":"We identified cohort of 18 glioma patients and performed Next Generation Sequencing (Exome, RNA, and Whole Genome Sequencing) and methylation profiling to identify genetic, epigenetic, and transcriptomic differences between long-term survivors (LTS, OS > 18 months) and short-term survivors (STS, OS < 12 months).","project":"ERP012180"}
{"number_samples":10,"species":"human","abstract":"The project aims to identify functional heterogeneity of cancer-associated stromal fibroblasts (CAF) which show specific gene expression profile in Chondrosarcoma (CS). The current knowledge of the role of CAF in the tumour microenvironment in CS regulating the expression of tumour cells is limited. In this study, we have determined the capability of primary CAFs to promote tumorigenesis of CS cancer cells. In order to confirm whether the primary fibroblast have activated features, we performed indirect co- culture (Trans- well migration assay) with CS tumour cell lines. On the basis of this migration assay we have identified that each primary fibroblast exhibited different induction of migration of CS tumour cells.   This differential pro- migratory capacity of CAFs will help identifying a gene expression signature that allows classification of patients with CS into high and low risk groups by RNA- sequencing","project":"ERP012188"}
{"number_samples":9,"species":"human","abstract":"Strand-specific polyA-selected mRNA libraries were generated using Illumina TruSeq RNA Library prep kits and protocols. Targeted fragment size was 300 bp. Libraries were multiplexed, pooled, and sequenced on multiple lanes of an Illumina HiSeq2500 to an average depth > 50M paired-end 50-cycle reads for each sample. Each library also includes 1 ul of a 1:10 dilution of ERCC RNA Control Spike-Ins (Ambion) that were added from one of two mixes, each containing the same 92 synthetic RNA standards of known concentration and sequence.","project":"ERP012633"}
{"number_samples":4,"species":"human","abstract":"This study involves characterization of four head and neck cancer cell lines -- NT8e, OT9, AW13516 and AW8507, established from Indian head and neck cancer patients, using SNP arrays, whole exome and whole transcriptome sequencing.","project":"ERP012768"}
{"number_samples":8,"species":"human","abstract":"We performed RNA-seq analyses of both estrogen receptor-positive and negative breast cancer cells over-expressing miR-515-5p which revealed down-regulation of several transcripts coding for migration promoting proteins. Modulating the expression of such proteins and miR-515 we found that miR-515 reduced migration and metastatic behaviour through direct inhibition of such pro-metastatic genes.","project":"ERP013303"}
{"number_samples":6,"species":"human","abstract":"The  KBM-7  reference  clone  B  (product  no.  P00174E07)  and  the  KBM-7-NuKO  derivative (P01289H04) in which the NUDT2 gene has been inactivated by retroviral gene-trap insertion were  obtained  from  Haplogen and  maintained. Three independent samples of total RNA were prepared from both KBM-7 and NuKO cells and sequenced using Illumina HiSeq2000.","project":"ERP013422"}
{"number_samples":25,"species":"human","abstract":"Based on the ability of FGF and/or WNT signaling to control posterior fate and intestinal lineage commitment, several groups have reported that treating mouse or human Pluripotent Stem Cell (PSC) derived definitive endoderm (DE) with small molecules or ligands that activate WNT signaling, or a combination of WNT and FGF signaling can induce an intestinal fate in human DE.  In this current study, we leverage hESC derived human intestinal organoids (HIOs) to test the hypothesis that the duration of exposure to high levels of FGF and WNT signaling controls regional intestinal identity, with shorter durations generating intestine similar to the proximal duodenum, and longer durations distalizing HIOs to become similar to jejunum/ileum. Our results demonstrate that exposing human definitive endoderm (DE) cultures to short or long incubations of media that activate WNT and FGF siganling results in gene and protein expression profiles that are consistent with tissue that has been patterned into proximal (duodenum) or distal (ileum) small intestine, respectively.","project":"ERP013713"}
{"number_samples":1,"species":"human","abstract":"Whole genome sequencing of (YRI) Yoruba HapMap population","project":"SRP000542"}
{"number_samples":1,"species":"human","abstract":"Whole genome sequencing of (LWK) Luhya in Webuye, Kenya HapMap population","project":"SRP000543"}
{"number_samples":1,"species":"human","abstract":"Whole genome sequencing of (CHB) Han Chinese in Beijing, China HapMap population","project":"SRP000546"}
{"number_samples":40,"species":"human","abstract":"We report an applicaton of small RNA sequencing using high throughput next generation sequencing to identify the small RNA content of cell lines. By sequencing over 30 million reads we could identify a new class of small RNAs previousy observed with tiling arrays and mapping to promoter regions of coding genes. We also identified a large number of small RNAs corresponding to internal exons of coding genes. By using different enzymatic treatments and immunoprecipitation experiments, we have determined that both the promoter associated small RNAs as well as ones within the body of the genes bear 5' cap structures. Overall design: Examination of the expression of small RNAs (<200nt).","project":"SRP000599"}
{"number_samples":8,"species":"human","abstract":"Adenosine-to-Inosine (A-to-I) RNA editing leads to transcriptome diversity and is important for normal brain function. To date, only a handful of functional sites have been identified in mammals. We developed an unbiased assay to screen >36,000 computationally predicted non-repetitive A-to-I sites, utilizing massively parallel target capture and DNA sequencing. The first comprehensive set of hundreds of human RNA editing sites were detected by comparing genomic DNA with RNAs from seven tissues of a single individual. Specificity of our profiling was supported by observations of enrichment with known features of ADAR targets and validation by capillary sequencing. Many sites are unique to the primate lineage. This expended repertoire of editing substrates will provide additional targets for understanding the role that RNA editing plays in normal brain function.","project":"SRP000617"}
{"number_samples":23,"species":"human","abstract":"Profiling of miRNA transcriptome in Human mature B cells from similar populations Overall design: Profiling of miRNA transcriptome in Human Normal and Malignant B cells, 8 normal samples corresponding to Naïve, Germinal Center, Plasma  and Memory B cells and 23 samples derived from B cell malignancy","project":"SRP000729"}
{"number_samples":23,"species":"human","abstract":"Cortisol is a glucocorticoid steroid hormone released by the adrenal glands in response to acute stress and as a messenger in circadian rhythms.  Transcriptional responses to this hormonal signal are mediated by the glucocorticoid receptor (GR).  Here, using massively parallel DNA sequencing, we determine GR binding across the human genome (using ChIP-seq) and measure related changes in gene expression (using mRNA-seq) in response to the glucocorticoid dexamethasone (DEX).  We identify 4,392 genomic positions occupied by the GR and 234 genes with significant changes in expression in response to DEX.  This genomic census reveals striking differences between activating and repressing activity of the GR.  First, genes activated by DEX have GR occupancy within a median distance of 10 kb from the transcriptional start site (TSS) while, for the genes repressed by DEX, the nearest GR binding site is a median of 150 kb from the TSS.  Furthermore, a reduction of GR levels by siRNA significantly and substantially reduces activation of GR target genes, but repression of transcription by DEX was largely unaffected.  These results suggest that DEX-mediated repression occurs independently of promoter proximal GR binding and is less sensitive to a reduction in the level of GR.  We also find that the GR can distinguish between different levels of corticosteroids in a gene-specific manner by, for example, selectively inducing PER1, a major transcription factor in circadian rhythms, at low doses of DEX.  Overall, our work both provides a genome-wide analysis of GR:DNA binding and transcriptional response and also reveals substantial new insights into the gene regulatory activities managed by GR.","project":"SRP000762"}
{"number_samples":14,"species":"human","abstract":"Paired end sequencing of cDNA isolated from individual melanoma samples via the Illumina sequencing platform to identify genetic aberrations that may play a role in melanoma genesis.","project":"SRP000931"}
{"number_samples":345,"species":"human","abstract":"The human embryonic stem cells (hESCs) are a unique model system for investigating the mechanisms of human development due to their ability to replicate indefinitely while retaining the capacity to differentiate into a host of functionally distinct cell types. In addition, these cells could be potentially used as therapeutic agents in regenerative medicine. Differentiation of hESCs involves selective activation or silencing of genes, a process controlled in part by the epigenetic state of the cell. In order to gain a better understanding of the epigenetic mechanisms regulating differentiation of hESCs, and produce general reference epigenome maps of the human cells, we propose to establish an Epigenome Center in San Diego. Our center will be focused on both undifferentiated hESC and four hESC-derived early embryonic cell lineages including extraembryonic endoderm, trophoblast, mesendoderm (a common precursor to mesodermal and endodermal lineages), and mesenchymal cells (a specific mesoderm derivative). We have developed and validated high throughput technologies for mapping the state of DNA methylation and chromatin modifications throughout the genome, and will use these methods to generate high-resolution maps of the reference epigenomes. Specifically, we will grow and differentiate hESCs into multiple lineages, and map DNA methylation sites using a newly developed technology that combines bisulfite conversion and whole genome shotgun sequencing. We will also determine the histone modification status in the genome by performing both ChlP-chip and ChlP-Seq analysis. We will develop advanced statistical and algorithmic solutions to facilitate high-throughput sequencing data analysis, and establish an informatics pipeline for collecting, storage, and distribution of epigenome maps. Finally, we will perform integrated data analysis to identify new epigenetic patterns in the genome that could provide insights in mechanisms of epigenetic regulation.","project":"SRP000941"}
{"number_samples":32,"species":"human","abstract":"The NIH Roadmap Epigenomics Mapping Consortium aims to produce a public resource of epigenomic maps for stem cells and primary ex vivo tissues selected to represent the normal counterparts of tissues and organ systems frequently involved in human disease. Characterization of the reference epigenome in humans by use of ChIP-Seq in a diverse panel of ES cells, tissue stem cells, reprogrammed stem cells, primary cells and tissues.","project":"SRP000996"}
{"number_samples":9,"species":"human","abstract":"miRNA expression profile Keywords: small RNA profiling by high throughput sequencing Overall design: The small RNA fraction (18-30nt) was cloned and sequenced from total RNA of each cell line","project":"SRP001313"}
{"number_samples":2,"species":"human","abstract":"Pre-mRNA splicing is regulated through combinatorial activity of RNA motifs including splice sites and splicing regulatory elements (SREs).  Here, we show that the activity of a major class of mammalian SREs is highly sensitive to the strength of the adjacent 5' splice site (5'ss) sequence, and that this has important functional and evolutionary implications.  Activity of G-run SREs was higher for intermediate strength 5'ss by ~4-fold relative to weak 5'ss, and by ~1.3-fold relative to strong 5'ss.  The dependence on 5'ss strength was supported both by comparative genomics and by microarray and Illumina mRNA-Seq analyses of splicing changes following RNAi against the splicing factor heterogeneous nuclear ribonucleoprotein (hnRNP) H, which binds G-runs.  This dependence implies that the responsiveness of exons to changes in hnRNP H levels is a bivariate function of both SRE abundance and 5'ss strength; this relationship may hold also for other splicing factors.  This pattern of activity enables G-runs and hnRNP H to buffer the effects of 5'ss mutations, augmenting both the frequency of 5'ss polymorphism and the evolution of new splicing patterns. Overall design: Examine mRNA expression in 293T cells following hnRNP H or control siRNA knockdown","project":"SRP001349"}
{"number_samples":167,"species":"human","abstract":"Study of chromatin accessibility","project":"SRP001371"}
{"number_samples":3,"species":"human","abstract":"In flies, small interfering RNAs (siRNAs) direct Argonaute2 (Ago2) to defend cells from viruses and transposons. In contrast, micro RNAs (miRNAs) mainly guide Ago1 to repress target mRNAs. A key step in the production of a functional Ago2/siRNA but not an Ago1/miRNA complex is Hen1-mediated methylation of the small RNA guide at the 2* hydroxyl of the 3*-terminal ribose. The function of this modification in animals is unknown. Here we show that highly complementary target RNAs decrease the steady-state level of Ago1-bound miRNAs. In vitro, the decline in miRNA levels requires high, but not complete complementarity between the target RNA and the small RNA; is accompanied by 3*-terminal nucleotide addition; and is blocked by Hen1-catalyzed 2*-O-methylation of small RNAs. The absence of Hen1 results in 3*-terminal uridylation and destabilization of Ago2-associated small RNAs in vivo. Our results suggest that small RNA-target RNA complementarity has shaped the coevolution of miRNAs and their target RNAs in flies and we present human cultured cell experiments that the target RNA-dependent small RNA tailing and degradation machinery may be conserved in mammals.","project":"SRP001381"}
{"number_samples":4,"species":"human","abstract":"Next-generation sequencing has become an important tool for genome-wide quantification of DNA and RNA.  However, a major technical hurdle lies in the need to map short sequence reads back to their correct locations in a reference genome.  Here we investigate the impact of SNP variation on the reliability of read-mapping in the context of detecting allele-specific expression (ASE).We generated sixteen million 35 bp reads from mRNA of each of two HapMap Yoruba individuals.  When we mapped these reads to the human genome we found that, at heterozygous SNPs, there was a significant bias towards higher mapping rates of the allele in the reference sequence, compared to the alternative allele.  Masking known SNP positions in the genome sequence eliminated the reference bias but, surprisingly, did not lead to more reliable results overall. We find that even after masking, $\\sim$5-10\\% of SNPs still have an inherent bias towards more effective mapping of one allele. Filtering out inherently biased SNPs removes 40\\% of the top signals of ASE.  The remaining SNPs showing ASE are enriched in genes previously known to harbor cis-regulatory variation or known to show uniparental imprinting.  Our results have implications for a variety of applications involving detection of alternate alleles from short-read sequence data. Scripts, written in Perl and R, for simulating short reads, masking SNP variation in a reference genome, and analyzing the simulation output are available upon request from JFD. Overall design: RNA-Seq on two YRI Hapmap cell lines. Each individual sequenced on two lanes of the Illumina Genome Analyzer","project":"SRP001462"}
{"number_samples":2,"species":"human","abstract":"Whole genome sequencing of (CLM) Colombia from Medellin, Colombia HapMap population","project":"SRP001523"}
{"number_samples":161,"species":"human","abstract":"Understanding the genetic mechanisms underlying natural variation in gene expression is a central goal of both medical and evolutionary genetics, and studies of expression quantitative trait loci (eQTLs) have become an important tool for achieving this goal. While all eQTL studies to date have assayed mRNA levels using expression microarrays, recent advances in RNA sequencing enable the analysis of transcript variation at unprecedented resolution. We sequenced RNA from 69 lymphoblastoid cell lines (LCLs) derived from unrelated Nigerian individuals that have been extensively genotyped by the International HapMap Project. Pooling data from all individuals, we generated a map of the transcriptional landscape of these cells, identifying extensive use of unannotated polyadenylation sites and over 100 novel putative protein-coding exons.  Using the genotypes from the HapMap project, we identified over a thousand genes at which genetic variation influences overall expression levels or splicing. We demonstrate that eQTLs near genes generally act via a mechanism involving allele-specific expression, and that variation that influences the inclusion of an exon is enriched within or near the consensus splice sites. Our results illustrate the power of high-throughput sequencing for the joint analysis of variation in transcription, splicing, and allele-specific expression across individuals. Overall design: RNA-Seq in 69 lymphoblastoid cell lines from multiple Yoruban HapMap individuals in at least two replicate lanes per individual","project":"SRP001540"}
{"number_samples":12,"species":"human","abstract":"Comparative studies of gene regulation suggest an important role for natural selection in shaping gene expression patterns within and between species. Most of these studies, however, estimated gene expression levels using microarray probes designed to hybridize to only a small proportion of each gene. Here we used recently-developed RNA sequencing protocols, which side-step this limitation, to assess intra- and inter-species variation in gene regulatory processes in considerably more detail than was previously possible. Specifically, we used RNAseq to study transcript levels in humans, chimpanzees, and rhesus macaques, using liver RNA samples from three males and three females from each species. Our approach allowed us to identify a large number of genes whose expression levels likely evolve under natural selection in primates. These include a subset of genes with conserved sexually dimorphic expression patterns across the three species, which we found to be enriched for genes involved in lipid metabolism. Our data also suggest that while alternative splicing is tightly regulated within and between species, sex-specific and lineage-specific changes in the expression of different splice forms are also frequent. Intriguingly, among genes in which a change in exon usage occurred exclusively in the human lineage, we found an enrichment of genes involved in anatomical structure and morphogenesis, raising the possibility that differences in the regulation of alternative splicing have been an important force in human evolution. Keywords: Gene Regulation Study Overall design: Examination of gene expression levels in livers from three primate species (human, chimpanzee, and rhesus macaque), using 3 male and 3 female samples from each species.","project":"SRP001558"}
{"number_samples":45,"species":"human","abstract":"Expression levels of human genes vary extensive among individuals.  Gene expression determines cell function and characteristics thus this variation likely contributes to phenotypic variation.  Genetic studies have shown that there is a heritable component to gene expression variation, and have identified genomic regions that contain polymorphic regulators.  However, most of these regions are quite large, and few regulators have been identified.  In this genetic of gene expression study, we used a large sample to search the genome for polymorphic regulators that influence gene expression, and followed up the results with deep sequencing of transcriptomes and molecular analyses. Key word(s): Transcriptome Analysis Overall design: genetics of gene expression study, 41 Coriell cell line samples examined.","project":"SRP001563"}
{"number_samples":4,"species":"human","abstract":"Recent transcriptome analysis indicates that >90% of human genes undergoes alternative splicing, underscoring the contribution of differential RNA processing to diverse proteomes in higher eukaryotic cells. The polypyrimidine tract binding protein PTB is a well-characterized splicing repressor, but PTB knockdown causes both exon inclusion and skipping. Genome-wide mapping of PTB-RNA interactions and construction of a functional RNA map now revealed that dominant PTB binding near a competing constitutive splice site generally induces exon inclusion whereas prevalent binding close to an alternative site often causes exon skipping. This positional effect was further demonstrated by disrupting or creating a PTB binding site on minigene constructs and testing their responses to PTB knockdown or overexpression. These findings suggest a mechanism for PTB to modulate splice site competition to produce opposite functional consequences, which may be generally applicable to RNA binding splicing factors to positively or negatively regulate alternative splicing in mammalian cells. Overall design: Examination of PTB-RNA binding in Hela cells using CLIP-seq (Cross-Linking ImmunoPrecipitation coupled with high-throughput sequencing) method. Peaks: The four alignment files (linked as supplementary files on Sample records) were combined together for peak finding, as we found that most of the monomeric and dimeric tags are similarly distributed in the genome with high pearson correlation coefficient. The method to detect the peaks above gene-specific randomized background was similar to (Yeo et al., 2009) and described in the paper (Xue et al., 2009).","project":"SRP001758"}
{"number_samples":12,"species":"human","abstract":"Alternative splicing of pre-mRNA is a prominent mechanism to generate protein diversity, yet its regulation is poorly understood. Here, we demonstrate a direct role for histone modifications in alternative splicing. We find distinctive histone modification signatures which correlate with splicing outcome in a set of human genes. Modulation of histone modifications causes splice site switching. The mechanism for histone-mediated splice site selection involves a histone mark which is read by a chromatin protein, which in turn recruits a splicing regulator. These results outline an adaptor system for reading of histone marks by the pre-mRNA splicing machinery. Overall design: To obtain an estimate of how many PTB-dependent alternative splicing events are regulated by SET2/MRG15-mediated recruitment of PTB, we carried out a genomewide comparative analysis of alternative splicing in hMSC cells depleted of either SETD2, MRG15 or PTB using specific siRNAs, or mock-depleted using a control siRNA.","project":"SRP001893"}
{"number_samples":8,"species":"human","abstract":"The p53-family member p73 functions in various cellular signaling pathways and can have tumor suppressor properties. Several isoforms of p73 exist that differ considerably in their function. Whereas the functions of the N-terminal isoforms (TA and ?Np73) and their opposing pro- and anti-apoptotic roles became evident, the functional differences of the distinct C-terminal spliceforms of TAp73 have remained unclear. Here, we characterized the genomic binding sites for TAp73a and TAp73ß and identified a specific p73 consensus binding-motif. Furthermore, an AP1 motif is strongly enriched close to binding sites for TAp73a. These AP1 motif-containing target genes are selectively upregulated by TAp73a, while their mRNA expression is repressed upon TAp73ß induction. Recruitment of c-Jun to the respective AP1 sites was impaired upon TAp73ß expression in part due to downregulation of c-Jun. We show that several of these AP1-site containing TAp73a-induced genes reduce on apoptosis-induction suggesting an underlying molecular mechanism for the observed functional differences between TAp73a and TAp73ß. Overall design: ChIP-seq and RNA-seq profiles of TAp73alpha, TAp73beta and p53 stably transfected in human osteosarcoma Saos cells","project":"SRP001997"}
{"number_samples":1,"species":"human","abstract":"RNA polymerase (Pol) III transcribes many noncoding RNAs (for example, transfer RNAs) important for translational capacity and other functions. We localized Pol III, alternative TFIIIB complexes (BRF1 or BRF2) and TFIIIC in HeLa cells to determine the Pol III transcriptome, define gene classes and reveal 'TFIIIC-only' sites. Pol III localization in other transformed and primary cell lines reveals previously uncharacterized and cell type–specific Pol III loci as well as one microRNA. Notably, only a fraction of the in silico–predicted Pol III loci are occupied. Many occupied Pol III genes reside within an annotated Pol II promoter. Outside of Pol II promoters, occupied Pol III genes overlap with enhancer-like chromatin and enhancer-binding proteins such as ETS1 and STAT1. Moreover, Pol III occupancy scales with the levels of nearby Pol II, active chromatin and CpG content. These results suggest that active chromatin gates Pol III accessibility to the genome. Overall design: Use of ChIP-seq to identify genomic regions bound by RNA Polymerase III machinery in multiple cell types as well as RNA-seq in HeLa for gene expression analysis.  See GSE20609 for whole human genome raw Pol III ChIP-array data.  See link below for supplementary methods and analysis.","project":"SRP002001"}
{"number_samples":62,"species":"human","abstract":"Dynamic transcriptomes during neural differentiation of human embryonic stem cells revealed by short long, and paired-end sequencing In order to examine the fundamental mechanisms governing neural differentiation, we analyzed the transcriptome changes that occur during the differentiation of human embryonic stem cells (hESCs) into the neural lineage. Undifferentiated hESCs as well as cells at three stages of early neural differentiation, N1 (early initiation), N2 (neural progenitor), and N3 (early glial-like) were analyzed using a combination of single read, paired-end read, and long read RNA sequencing. The results revealed enormous complexity in gene transcription and splicing dynamics during neural cell differentiation. We found previously unannotated transcripts and spliced isoforms specific for each stage of differentiation. Interestingly, splicing isoform diversity is highest in undifferentiated hESCs and decreases upon differentiation, a phenomenon we call “isoform specialization.” During neural differentiation, we observed differential expression of many types of genes including those involved in key signaling pathways, and a large number of extracellular receptors exhibit stage-specific regulation. These results provide a valuable resource for studying neural differentiation and reveal insights into the mechanisms underlying in vitro neural differentiation of hESCs, such as neural fate specification, NPC identity maintenance and the transition from a predominantly neuronal state into one with increased gliogenic potential Overall design: Characterization of changes in the transcriptome profiles during early stages of human neural differentiation from H1 !Series_overall_design=hESCs. 454 SFF files are missing. hESC_B_s9.fastq--> SRR037192 hESC_B_s8.fastq--> SRR037191 hESC_B_s7.fastq--> SRR037190 hESC_B_s6.fastq--> SRR037189 hESC_B_s5.fastq--> SRR037188 hESC_B_s4.fastq--> SRR037187 hESC_B_s3.fastq--> SRR037186 hESC_B_s2.fastq--> SRR037185 hESC_B_s16.fastq--> SRR037184 hESC_B_s15.fastq--> SRR037183 hESC_B_s14.fastq--> SRR037182 hESC_B_s13.fastq--> SRR037181 hESC_B_s12.fastq--> SRR037180 hESC_B_s11.fastq--> SRR037179 hESC_B_s10.fastq--> SRR037178 hESC_B_s1.fastq--> SRR037177 hESC_B_p6_2.fastq--> SRR037176 hESC_B_p6_1.fastq--> SRR037176 hESC_B_p5_2.fastq--> SRR037175 hESC_B_p5_1.fastq--> SRR037175 hESC_B_p4_2.fastq--> SRR037174 hESC_B_p4_1.fastq--> SRR037174 hESC_B_p3_2.fastq--> SRR037173 hESC_B_p3_1.fastq--> SRR037173 hESC_B_p2_2.fastq--> SRR037172 hESC_B_p2_1.fastq--> SRR037172 hESC_B_p1_2.fastq--> SRR037171 hESC_B_p1_1.fastq--> SRR037171 hESC_A_s3.fastq--> SRR037170 hESC_A_s2.fastq--> SRR037169 hESC_A_s1.fastq--> SRR037168 hESC_A_p3_2.fastq--> SRR037167 hESC_A_p3_1.fastq--> SRR037167 hESC_A_p2_2.fastq--> SRR037166 hESC_A_p2_1.fastq--> SRR037166 hESC_A_p1_2.fastq--> SRR037165 hESC_A_p1_1.fastq--> SRR037165 N3_A_s4.fastq--> SRR037226 N3_A_s3.fastq--> SRR037225 N3_A_s2.fastq--> SRR037224 N3_A_s1.fastq--> SRR037223 N3_A_p3_2.fastq--> SRR037222 N3_A_p3_1.fastq--> SRR037222 N3_A_p2_2.fastq--> SRR037221 N3_A_p2_1.fastq--> SRR037221 N3_A_p1_2.fastq--> SRR037220 N3_A_p1_1.fastq--> SRR037220 N2_B_s8.fastq--> SRR037219 N2_B_s7.fastq--> SRR037218 N2_B_s6.fastq--> SRR037217 N2_B_s5.fastq--> SRR037216 N2_B_s4.fastq--> SRR037215 N2_B_s3.fastq--> SRR037214 N2_B_s2.fastq--> SRR037213 N2_B_s1.fastq--> SRR037212 N2_B_p6_2.fastq--> SRR037211 N2_B_p6_1.fastq--> SRR037211 N2_B_p5_2.fastq--> SRR037210 N2_B_p5_1.fastq--> SRR037210 N2_B_p4_2.fastq--> SRR037209 N2_B_p4_1.fastq--> SRR037209 N2_B_p3_2.fastq--> SRR037208 N2_B_p3_1.fastq--> SRR037208 N2_B_p2_2.fastq--> SRR037207 N2_B_p2_1.fastq--> SRR037207 N2_B_p1_2.fastq--> SRR037206 N2_B_p1_1.fastq--> SRR037206 N2_A_s4.fastq--> SRR037205 N2_A_s3.fastq--> SRR037204 N2_A_s2.fastq--> SRR037203 N2_A_s1.fastq--> SRR037202 N2_A_p3_2.fastq--> SRR037201 N2_A_p3_1.fastq--> SRR037201 N2_A_p2_2.fastq--> SRR037200 N2_A_p2_1.fastq--> SRR037200 N2_A_p1_2.fastq--> SRR037199 N2_A_p1_1.fastq--> SRR037199 N1_A_s3.fastq--> SRR037198 N1_A_s2.fastq--> SRR037197 N1_A_s1.fastq--> SRR037196 N1_A_p3_2.fastq--> SRR037195 N1_A_p3_1.fastq--> SRR037195 N1_A_p2_2.fastq--> SRR037194 N1_A_p2_1.fastq--> SRR037194 N1_A_p1_2.fastq--> SRR037193 N1_A_p1_1.fastq--> SRR037193","project":"SRP002079"}
{"number_samples":15,"species":"human","abstract":"Epigenetic control is an important aspect of gene regulation.  Despite detailed understanding of many examples, the transcription of non-coding RNA genes by RNA polymerase (pol) III is less well characterized.  Here we profile the epigenetic features of pol III target genes  throughout the human genome.  This reveals that the chromatin landscape of pol III-transcribed genes resembles that of pol II templates in many ways, although there are also clear differences.  Our analysis also discovered an entirely unexpected phenomenon, namely that pol II co-localizes with the majority of genomic loci that are bound by pol III. Overall design: RNA-seq experiment for total RNA in CD4+ cells.","project":"SRP002105"}
{"number_samples":14,"species":"human","abstract":"We examined genome-wide variation in transcription factor binding in different individuals and a chimpanzee using chromatin immunoprecipitation followed by massively-parallel sequencing (ChIP-Seq). The binding sites of RNA Polymerase II (Pol II) as well as a key regulator of immune responses, NFkB, were mapped in ten HapMap lymphoblastoid cell lines derived from individuals of African, European, and Asian ancestry, including a parent-offspring trio. We also mapped gene expression in all ten human cell lines for two treatment conditions: a) no treatment and b) following induction by TNF-alpha. Overall design: Genome-wide comparison of Pol II and NF-KappaB binding in ten individuals. RNA-seq study with no treatment.","project":"SRP002126"}
{"number_samples":10,"species":"human","abstract":"We examined genome-wide variation in transcription factor binding in different individuals and a chimpanzee using chromatin immunoprecipitation followed by massively-parallel sequencing (ChIP-Seq). The binding sites of RNA Polymerase II (Pol II) as well as a key regulator of immune responses, NFkB, were mapped in ten HapMap lymphoblastoid cell lines derived from individuals of African, European, and Asian ancestry, including a parent-offspring trio. We also mapped gene expression in all ten human cell lines for two treatment conditions: a) no treatment and b) following induction by TNF-alpha. Overall design: Genome-wide comparison of Pol II and NF-KappaB binding in ten individuals. RNA-seq study with TNF-alpha treatment.","project":"SRP002128"}
{"number_samples":6,"species":"human","abstract":"Epidermal growth factor (EGF) is a key regulatory growth factor activating a myriad of processes affecting cell proliferation and survival that are relevant to normal development and disease. Here we have used a combined approach to study the EGF dependent transcriptome of HeLa cells. We obtained mRNA expression profiles using multiple long oligonucleotide based microarray platforms (from Agilent, Operon, Febit, and Illumina) in combination with digital gene expression profiling (DGE) with the Illumina Genome Analyzer I (GA-I). By applying a procedure for cross-platform data meta-analysis based on rank product and global ancova tests, we establish a well validated gene set with transcript levels altered after EGF treatment. We used this robust gene list to build higher order networks of gene interaction by interconnecting associated networks, supporting and extending the important role of the EGF signaling pathway in cancer. In addition, we found a whole new set of genes previously unrelated to the currently accepted EGF associated cellular functions, among which are metallothionein genes. We propose the use of global genomic cross-validation to generate more reliable datasets derived from high content technologies (microarrays or deep sequencing). This approach should help to improve the confidence of downstream in silico functional inference analyses based on high content data. Keywords: treated vs. untreated comparison, time course Overall design: Time course experiment comparing HeLa gene expression in response to EGF analyzed on different microarray platforms (Agilent, IMPPC, Illumina, and Operon) and by digital gene expression using short read high throughput tag sequencing. Three independent experiments were performed where HeLa cells were serum deprived for 24 hours and were either left untreated or treated with EGF for 6, and 24 h and harvested for RNA extraction. Technical dye swap duplicates were performed for each of the three biological replicates in both time points. Comparative genomic hybridization of HeLa cell genomic DNA versus  poooled genomic DNA from blood obtained from human females conducted on commercial oligonucleotide microarrays (Human Genome CGH Microarray Kit 244A, Agilent Technologies) in order to assess DNA dosage dependence of gene expression levels and response to EGF. Digital gene expression using short read high throughput tag sequencing data submitted to NCBI's SRA","project":"SRP002184"}
{"number_samples":4,"species":"human","abstract":"MicroRNAs are a class of small non-coding RNAs that regulate mRNA expression at the post-transcriptional level and thereby many fundamental biological processes. A number of methods, such as multiplex polymerase chain reaction, microarrays have been developed for profiling levels of known miRNAs. These methods lack ability to identify novel miRNAs and accurately determine expression at a range of concentration. Deep or massively parallel sequencing methods are providing suitable platforms for genome wide transcriptome analysis and have the ability to identify novel transcripts. The results of analysis of small RNA sequences obtained by Solexa technology of normal peripheral blood mononuclear cells, tumor cell lines K562 (chronic myelogenous leukemia) and HL60 (acute promyelogenous leukemia) are presented. Custom computation pipelines were used to generate expression profiles of known and for identification of novel miRNAs. Some of the highly expressed miRNAs in the leukocytes include several members of the let 7 family, mir-21, 103, 185, 191 and 320a. Comparison of the miRNA profiles of normal versus K562 cells or HL60 revealed a specific set of differentially expressed molecules. Correlation of the miRNA with that of mRNA expression profiles, obtained by microarray, revealed a set of target genes showing inverse correlation with miRNA levels. Our computational pipeline also predicted a number of novel miRNAs. Some of the predictions were validated by realtime RT-PCR and or RNAase protection assay. Overall design: The small RNA population from four samples - Two Normal Peripheral blood mononuclear cells (PBMC) samples, K562 cell line (This cell line is used as a model to study Chronic Myelogenous Leukemia), HL60 (This cell line is used to study acute promyelogenous leukemia) was sequenced using Solexa technology.","project":"SRP002220"}
{"number_samples":1,"species":"human","abstract":"We deep sequenced chromatin-associated RNAs (CARs) from human fibroblast (HF) cells. This resulted in the identification of 141 intronic regions and 74 intergenic regions harbouring CARS. Overall design: We purified CARs from normal HFs by isolating soluble chromatin after MNase treatment, followed by separation of chromatin fragments of different lengths on a sucrose gradient. CARs were converted into double-stranded cDNAs and sequenced using the Illumina Genome Analyzer I.","project":"SRP002245"}
{"number_samples":15,"species":"human","abstract":"We applied small RNA Solexa sequencing technology to identify microRNA expression in human liver samples from surgically removed liver tissues including three normal liver tissues (distal normal liver tissue of liver hemangioma), an hepatitis B virus (HBV)-infected liver, a severe chronic hepatitis B liver, two HBV-related hepatocellular carcinoma (HCC), an hepatitis C virus (HCV)-related HCC, and an HCC without HBV or HCV infection. All samples were collected with the informed consent of the patients and the experiments were approved by the ethics committee of Second Military Medical University, Shanghai, China. We investigated the miRNome in human normal liver and suggested some deregulated abundantly expressed microRNAs in HCC. center_name: National Key Laboratory of Medical Immunology & Institute of Immunology, Second Military Medical University, Shanghai, China. Overall design: Examination of miRNome in human liver samples from surgically removed liver tissues including three normal liver tissues (distal normal liver tissue of liver hemangioma), an hepatitis B virus (HBV)-infected liver tissue, a severe chronic hepatitis B liver tissue, an HBV-related hepatocellular carcinoma (HCC) tissue and adjacent liver tissues of different regions,an HBV-related HCC tissue and adjacent liver tissue, an hepatitis C virus (HCV)-related HCC tissue and adjacent liver tissue, and an HCC without HBV or HCV infection and adjacent liver tissue. All 15 human liver tissue samples.","project":"SRP002272"}
{"number_samples":6,"species":"human","abstract":"Apply the Illumina next generaton sequencing technology to obtain 22 millions of 50-bp paired-end reads Overall design: High-throughput RNA-seq in human brain tissues. Mapping results on human genome (hg18) by SeqMap.","project":"SRP002274"}
{"number_samples":1,"species":"human","abstract":"We have recently shown that transcription initiation RNAs (tiRNAs) are derived from sequences downstream of transcription start sites. Here we report the identification of a second class of nuclear-specific ~17-18 nucleotide small RNA whose 3’ ends map precisely to the splice donor site of internal exons in animals. These splice-site RNAs (spliRNAs) are associated with highly expressed genes, and show evidence of developmental stage- and region-specific expression. We also confirm that tiRNAs are nuclear localized, enriched at chromatin marks associated with transcription initiation, and possess a 3’ nucleotide bias. Additionally, we find that microRNA-offset RNAs (moRNAs), the oncogenic miR-15/16 cluster and most snoRNA-derived small RNAs (sdRNAs) are enriched in the nucleus, whereas most miRNAs and two H/ACA sdRNAs are cytoplasmically enriched. We propose that nuclear localized tiny RNAs are involved in epigenetic regulation of gene expression. Overall design: Discovery and characterization of small RNA species through high-througput deep sequencing of nuclear, cytoplasmic and total small RNA fractions from THP-1 cells, a monocytic leukemia cell line. Additional files and information are available at http://matticklab.com/index.php?title=NuclearTinyRNAs","project":"SRP002278"}
{"number_samples":3,"species":"human","abstract":"Most tumors of the upper gastrointestinal tract are known to depend on an excessive expression of Shh. It was recently discovered that this Shh does not signal on the tumor cells, but rather the stromal cells that in turn produce an unknown set of reciprocal signals that act as growth- or survival cues for the tumor cells. This sequencing effort aims to identify the reciprocal signals. Overall design: Mouse fibroblasts and Shh-expressing human pancreatic adenocarcinoma cells were either grown alone, or in a non-adherent coculture system. To be able to assess the effect of Shh on fibroblast gene expression (being the reciprocal signal), 5E1 blocking antibody was added to the cells and after 5 days, RNA was isolated. n=1","project":"SRP002306"}
{"number_samples":38,"species":"human","abstract":"We prepared small RNA libraries from 29 tumor/normal pairs of human cervical tissue samples. Analysis of the resulting sequences (42 million in total) defined 64 new human microRNA (miRNA) genes. Both arms of the hairpin precursor were observed in twenty-three of the newly identified miRNA candidates. We tested several computational approaches for analysis of class differences between high throughput sequencing datasets, and describe a novel application of log linear model that has provided the most datasets, and describe a novel application of log linear model that has provided the most effective analysis for this data. This method resulted in the identification of 67 miRNAs that were differentially-expressed between the tumor and normal samples at a false discovery rate less than 0.001. Overall design: A total of 29 tumor/normal pairs of human cervical tissue samples were analyzed. Two samples (G699N_2 and G761T_2) were performed in duplicates. No Fastq files for GSM532871 to GSM532889, GSM532929, and GSM532930. Sequence files are provided as text files for these 22 Sample records in GSE20592_RAW.tar.  38 samples with quality scores are available from SRA as SRP002/SRP002326 (see Supplementary file below).","project":"SRP002326"}
{"number_samples":38,"species":"human","abstract":"We applied 4-thiouridine to cultured cells expressing the FLAG/HA-tagged RNA-binding proteins (RBPs) followed by UV 365 nm irradiation. The crosslinked RNA-protein complexes were isolated by immunoprecipitation, and the covalently bound RNA was partially digested with RNase T1 and radiolabeled. The radiolabeled RNPs were subsequenctly separated by SDS-PAGE, the crosslinked RNA segments recovered and converted into a cDNA library and sequenced. Overall design: RBPs were UV-crosslinked to their RNA targets containing 4-thiouridine. The RNA segments were recovered after immunoprecipitation and seqeunced by Solexa.","project":"SRP002487"}
{"number_samples":7,"species":"human","abstract":"Messenger RNAs are regulated by a variety of degradation mechanisms in mammalian cells.  In the canonical animal microRNA pathway, microRNAs in complex with Argonaute proteins bind to many mRNA targets with imperfect complementarity, leading to degradation of the mRNA through the regular decay machinery.  The ancestral “slicer” endonuclease activity of Argonaute2 itself, which requires more extensive complementarity with the target RNA, is not used in this pathway, and has only been observed in two microRNA-guided cases.  Nevertheless, the cleavage capacity of mammalian Ago2 is conserved and essential for viability.  Here, we assess the endonucleolytic function of Ago2 and other nucleases by identifying cleavage products retaining 5`-phosphate groups in mouse ES cells on a transcriptome-wide scale.  We detect a significant signature of Ago2-dependent cleavage events and validate several targets.  Unexpectedly, a broader class of Ago2-independent cleavage sites is also observed, indicating participation of additional nucleases in this mode of mRNA regulation.  Within this class, we identify a cohort of Drosha-dependent mRNA cleavage events, including one in the Dgcr8 mRNA, that functionally regulate mRNA levels in mES cells.  Together, these results highlight the underappreciated role of endonucleolytic cleavage in controlling mRNA fates in mammals. Overall design: Global 5`-phosphate-dependent RACE in WT, Ago2-KO and Drosha-excised mouse ES cells and human 293S cells","project":"SRP002543"}
{"number_samples":24,"species":"human","abstract":"MicroRNAs (miRNAs) are endogenous ~22-nucleotide RNAs that mediate important gene-regulatory events by pairing to the mRNAs of protein-coding genes to direct their repression. Repression of these regulatory targets leads to decreased translational efficiency and/or decreased mRNA levels, but the relative contributions of these two outcomes have been largely unknown, particularly for endogenous targets expressed at low-to-moderate levels. Here, we use ribosome profiling to measure the overall effects on protein production and compare these to simultaneously measured effects on mRNA levels. For both ectopic and endogenous miRNA regulatory interactions, lowered mRNA levels account for most (=84%) of the decreased protein production. These results show that changes in mRNA levels closely reflect the impact of miRNAs on gene expression and indicate that destabilization of target mRNAs is the predominant reason for reduced protein output. Overall design: Examine ribosome footprints and mRNA abundance of HeLa cells transfected with miR-1 or miR-155, versus mock-transfected cells, at two different time points post-transfection. Supplementary processed data files linked below. mir155_summaryTable.txt: log2 fold changes (miR-155-transfected versus mock-transfected HeLa cells, 32hr). mir1_summaryTable.txt: log2 fold changes (miR-1-transfected versus mock-transfected HeLa cells, 32hr).","project":"SRP002605"}
{"number_samples":8,"species":"human","abstract":"to be edited","project":"SRP002621"}
{"number_samples":30,"species":"human","abstract":"We used RNA-seq to interrogate prostate cancer specific gene fusions, alternative splicings, somatic mutations and novel transcripts. Overall design: We sequenced the transcriptome (polyA+) of 20 prostate cancer tumors and 10 matched normal tissues using Illumina GAII platform. Then we used bioinformatic approaches to identify prostate cancer specific aberrations which include gene fusion, alternative splicing, somatic mutation, etc.","project":"SRP002628"}
{"number_samples":8,"species":"human","abstract":"We present “centered sites,” a class of microRNA target sites that lacks both perfect seed pairing and 3'-compensatory pairing and instead has 11–12 contiguous Watson–Crick pairs to the center of the microRNA.  In elevated Mg2+, centered sites impart mRNA cleavage, but in cells, centered sites repress protein output without consequential Agronaute-catalyzed cleavage.  Our study also identified novel extensively paired sites that are cleavage substrates in cultured cells and human brain.  This expanded repertoire of cleavage targets and the identification of the centered site type help explain why central regions of many microRNAs are evolutionarily conserved. Overall design: To study centered sites and identify miRNA cleavage targets, mRNA degradomes were sequenced from human brain and HeLa cells, and smallRNAs were sequenced from human brain and zebrafish embryo at 24 hours post fertilization (hpf). Replicates were combined before the analysis. Fastq files are not available for GSM548638 and GSM548639.","project":"SRP002640"}
{"number_samples":4,"species":"human","abstract":"We have used deep sequencing to explore the repertoire of both poly(A)+ and poly(A)- RNAs from two standard cell lines, HeLa cells and human embryonic stem cell (hESC) H9 cells. Overall design: Examination of nonpolyadenylated and polyadenylated RNA in 2 cell types.","project":"SRP002789"}
{"number_samples":2,"species":"human","abstract":"Background: MicroRNAs (miRNAs) are 22-nt small non-coding regulatory RNAs that have generally been considered to regulate gene expression at the post-transcriptional level in the cytoplasm. However, recent studies have reported that some miRNAs localize to and function in the nucleus. Methodology/Principal Findings: To determine the number of miRNAs localized to the nucleus, we systematically investigated the subcellular distribution of small RNAs (sRNAs) by independent deep sequencing the nuclear and cytoplasmic pools of 18- to 30-nucleotide sRNAs from human cells. We identified 339 nuclear and 324 cytoplasmic known miRNAs, 300 of which overlap, suggesting that the majority of miRNAs are imported into the nucleus. With the exception of a few miRNAs evidently enriched in the nuclear pool, such as the mir-29b, the ratio of miRNA abundances in the nuclear fraction versus in the cytoplasmic fraction vary to some extent. Moreover, our results revealed that a large number of tRNA 3' trailers are exported from the nucleus and accumulate in the cytoplasm. These tRNA 3' trailers accumulate in a variety of cell types, implying that the biogenesis of tRNA 3' trailers is conserved and that they have a potential functional role in vertebrate cells. Conclusion/Significance: Our results provide the first comprehensive view of the subcellular distribution of diverse sRNAs and new insights into the roles of miRNAs and tRNA 3' trailers in the cell Overall design: Discovery and characterization of small RNA species through deep sequencing of nuclear and cytoplasmic small RNA fractions from 5-8F cells, a nasopharyngeal carcinoma cell line.","project":"SRP002861"}
{"number_samples":1,"species":"human","abstract":"We generate this 130-nt dataset to benchmark SplicePL, TopHat and SpliceMap. Overall design: Retina sample from one human donor.","project":"SRP002881"}
{"number_samples":27,"species":"human","abstract":"To investigate potential links between Stat3 transcriptional activity and other signaling pathways in breast cancer, we determined the gene expression profiles of three breast cancer cell lines treated with JAK, PTGIS, PFKFB3, CXCR2, HAS1, or NQO1 inhibitor (all of which decreased Stat3 transcriptional activity in Hs 578T cells except for the NQO1 inhibitor), inhibitor treatment vehicle alone (DMSO), STAT3 siRNAs, or non-targeting siRNAs. Using the resulting data, we identified a gene signature that was significantly regulated by STAT3 siRNAs and similarly affected by the JAK and at least 3 other inhibitors (or by 4 other inhibitors) but not by the NQO1 inhibitor in Hs 578T cells that was enriched in genes involved in development and correlated with shorter distant metastasis-free survival in primary lymph-node-negative invasive breast tumors. These results emphasize the central importance of Stat3 in CD44+/CD24- stem-cell-like breast cancer cells. Overall design: This study includes SAGE-Seq libraries obtained using 27 samples that each underwent individual protocols. There are nine samples of each of three cell lines - Hs 578T, MCF7, and SUM159PT. The samples in each group were treated with either one of the six inhibitors, DMSO only (the background control for inhibitor-treated cells), STAT3 siRNAs, or non-targeting siRNAs (the background control for STAT3 siRNAs).","project":"SRP002915"}
{"number_samples":6,"species":"human","abstract":"small RNA profiles of 6 human tonsillar B cell populatios (naive B cells, pre-germinal center B cells, centrocytes, centroblasts, memory B cells, and plasma cells) were determined by deep sequencing.  These samples were compared to mouse developing lymphocytes, various hematopoietic cell lineages, and tissues. Overall design: small RNA expression profiles of 6 well defined B cell populations isolated from human tonsils.","project":"SRP002958"}
{"number_samples":2,"species":"human","abstract":"We used immunoprecipitation to pulldown Lin28 associated RNA targets and used genome-wide high throughput deep sequencing to identified those Lin28-associated RNAs. Keywords: Lin28 IP-RNAseq Overall design: Examination of Lin28 immunoprecipitated RNA transcripts","project":"SRP003021"}
{"number_samples":1,"species":"human","abstract":"Reprogramming of the human fibroblast genome into the state of the hematopoietic progenitors Overall design: One sample analyzed.","project":"SRP003227"}
{"number_samples":2,"species":"human","abstract":"A comparison of RNA-Seq and high-density exon array for detecting differential gene expression between closely related species","project":"SRP003276"}
{"number_samples":53,"species":"human","abstract":"Prostate adenocarcinoma and matched adjacent normal samples were profiled by deep transcriptional sequencing to analyze transcription-induced chimeras and gene fusions.  Reference samples from the MAQC and brain and universal reference libraries were also sequenced. Overall design: Two-condition experiment plus reference samples: Prostate adenocarcinoma versus matched normal from three separate patients, plus brain and universal reference samples from the MAQC project.","project":"SRP003611"}
{"number_samples":3,"species":"human","abstract":"Changes in gene regulation are thought to play an important role in speciation and adaptation, especially in primates. However, we still know relatively little about the mechanisms underlying regulatory evolution. In particular, the extent to which epigenetic modifications underlie gene expression differences between primates is not yet known. Our study focuses on an epigenetic histone modification, H3K4me3, which is thought to promote transcription. To investigate the contribution of H3K4me3 to regulatory differences between species, we collected gene expression data and identified H3K4me3-associated genomic regions in lymphoblastoid cell lines (LCLs) from humans, chimpanzees, and rhesus macaques, using three cell lines from each species. We found strong evidence for conservation of H3K4me3 localization in primates. Moreover, regardless of species, H3K4me3 is consistently enriched near annotated transcription start sites (TSS), and highly expressed genes are more likely than lowly expressed genes to have the histone modification near their TSS. Interestingly, we observed an enrichment of interspecies differences in H3K4me3 at the TSS of genes that are differentially expressed between species. We estimate that as much as 7% of gene expression differences between the LCLs of humans, chimpanzees, and rhesus macaques may be explained, at least in part, by changes in the status of H3K4me3 histone modifications. Our results suggest a modest, yet important role for epigenetic changes in gene expression differences between primates. Overall design: Examination of H3K4me3 profiles in lymphoblastoid cell lines (LCLs) from three primate species (human, chimpanzee, and rhesus macaque), using 3 samples from each species, compared to a pooled input control from each species. Expression profiling was also done in the same LCL samples.","project":"SRP003637"}
{"number_samples":12,"species":"human","abstract":"We have used deep sequencing to explore the repertoire of both poly(A)+ and poly(A)- RNAs from two standard cell lines, HeLa cells and human embryonic stem cell (hESC) H9 cells. Overall design: Examination of nonpolyadenylated and polyadenylated in 2 cell types.","project":"SRP003672"}
{"number_samples":14,"species":"human","abstract":"We present a powerful application of ultra high-throughput sequencing, SAGE-Seq, for the accurate quantification of normal and neoplastic mammary epithelial cell transcriptomes. We develop data analysis pipelines that allow the mapping of sense and antisense strands of mitochondrial and RefSeq genes, the normalization between libraries, and the identification of differentially expressed genes. We find that the diversity of cancer transcriptomes is significantly higher than that of normal cells. Our analysis indicates that transcript discovery plateaus at 10 million reads/sample, and suggests a minimum desired sequencing depth around 5 million reads. Comparison of SAGE-Seq and traditional SAGE on normal and cancerous breast tissues reveals higher sensitivity of SAGE-Seq to detect less abundant genes including those encoding for known breast cancer-related transcription factors and G protein-coupled receptors (GPCRs). SAGE-Seq is able to identify genes and pathways abnormally activated in breast cancer that traditional SAGE failed to call. SAGE-Seq is a powerful method for the identification of biomarkers and therapeutic targets in human disease. Overall design: Global transcriptome profilings of 7 samples in normal and 7 samples in cancer from clinical breast tissue using Sage-Seq","project":"SRP003726"}
{"number_samples":156,"species":"human","abstract":".","project":"SRP003754"}
{"number_samples":20,"species":"human","abstract":"we combined their genome-wide profiles of tumors and normal adjacent tissues in 10 ccRCC patients. The results showed that 283 miRNAs were down-regulated and 187 up-regulated, meanwhile 7473 mRNAs were up-regulated and 3439 down-regulated in ccRCC. Expressions of 12 miRNAs and genes were validated in 10 patients we studied by RT-qPCR (validation rate from 60% to 100%). Differentially expressed gene analysis showed the collective change of miR-200 and SLC22A gene family, and pathway analysis revealed down-regulation of multiple metabolic pathways and up-regulation of focal adhesion, ECM-receptor interaction, etc. Moreover, A miRNA cluster located in chromosome Xq27.3 were significantly down-regulated in ccRCC, which were verified in 53 ccRCC patients by RT-qPCR (validation rate from 63.8% to 88.6%).56 and 586 potentially novel miRNAs and mRNA we detected according to statistical analyses. In addition, 14 miRNA candidates were randomly selected to validate using qRT-PCR and Sanger sequcencing technology, 10 of out 14 could be successfully amplified,.Plus, the sequences of all 5 randomly selected amplified products were confirmed by cloned Sanger sequencing.Our data demonstrated a combined profiling of miRNAs and mRNAs in ccRCC, which showed the decreased metabolism and the perturbation of renal cell function. Moreover, this study identified the expression alteration of a miRNA cluster on Xq27.3, which suggested its potential function in carcinogenesis of ccRCC Overall design: Examination of 10 pairs of kidney normal cells and cancer cells .","project":"SRP003901"}
{"number_samples":20,"species":"human","abstract":"we combined their genome-wide profiles of tumors and normal adjacent tissues in 10 ccRCC patients. The results showed that 283 miRNAs were down-regulated and 187 up-regulated, meanwhile 7473 mRNAs were up-regulated and 3439 down-regulated in ccRCC. Expressions of 12 miRNAs and genes were validated in 10 patients we studied by RT-qPCR (validation rate from 60% to 100%). Differentially expressed gene analysis showed the collective change of miR-200 and SLC22A gene family, and pathway analysis revealed down-regulation of multiple metabolic pathways and up-regulation of focal adhesion, ECM-receptor interaction, etc. Moreover, A miRNA cluster located in chromosome Xq27.3 were significantly down-regulated in ccRCC, which were verified in 53 ccRCC patients by RT-qPCR (validation rate from 63.8% to 88.6%).56 and 586 potentially novel miRNAs and mRNA we detected according to statistical analyses. In addition, 14 miRNA candidates were randomly selected to validate using qRT-PCR and Sanger sequcencing technology, 10 of out 14 could be successfully amplified,.Plus, the sequences of all 5 randomly selected amplified products were confirmed by cloned Sanger sequencing.Our data demonstrated a combined profiling of miRNAs and mRNAs in ccRCC, which showed the decreased metabolism and the perturbation of renal cell function. Moreover, this study identified the expression alteration of a miRNA cluster on Xq27.3, which suggested its potential function in carcinogenesis of ccRCC Overall design: Examination of 10 pairs of kidney normal cells and cancer cells .","project":"SRP003902"}
{"number_samples":3,"species":"human","abstract":"The stromal microenvironment plays key roles in prostate development and cancer. Cancer associated fibroblasts (CAFs) and other stromal cells stimulate tumourigenesis via several mechanisms including the expression of pro-tumourigenic factors. Mesenchyme (embryonic stroma) controls prostate organogenesis, and in some circumstances can re-differentiate prostate tumours. Epithelia are regulated by powerful paracrine signalling from the stroma in both development and disease, and identification of these stromal signals is important.  We have applied next-generation Tag profiling to fetal human prostate, normal human prostate fibroblasts (NPFs) and CAFs to identify molecules expressed in prostatic stroma Overall design: Each sample was used for Tag library construction, by Solexa Inc","project":"SRP004042"}
{"number_samples":58,"species":"human","abstract":"Noncoding RNAs (ncRNAs) are emerging as key molecules in human cancer, with the potential to serve as novel markers of disease and to reveal uncharacterized aspects of tumor biology. Here we discover 121 unannotated prostate cancer–associated ncRNA transcripts (PCATs) by ab initio assembly of high-throughput sequencing of polyA+ RNA (RNA-Seq) from a cohort of 102 prostate tissues and cells lines. We characterized one ncRNA, PCAT-1, as a prostate-specific regulator of cell proliferation and show that it is a target of the polycomb repressive complex 2 (PRC2). We further found that patterns of PCAT-1 and PRC2 expression stratified patient tissues into molecular subtypes distinguished by expression signatures of PCAT-1–repressed target genes. Taken together, our findings suggest that PCAT-1 is a transcriptional repressor implicated in a subset of prostate cancer patients. These findings establish the utility of RNA-Seq to identify disease-associated ncRNAs that may improve the stratification of cancer subtypes. Overall design: 21 prostate cell lines sequenced on the Illumina Genome Analyzer and GAII.  Variable number of replicates per sample.  RNA-Seq data from prostate cancer tissues used in this study will be made available on dbGAP.","project":"SRP004637"}
{"number_samples":3,"species":"human","abstract":"Transcriptome profiling studies suggest that a large fraction of the genome is transcribed and many transcripts function independent of their protein coding potential. The relevance of noncoding RNAs (ncRNAs) in normal physiological processes and in tumorigenesis is increasingly recognized. Here, we describe consistent and significant differences in the distribution of sense and antisense transcripts between normal and neoplastic breast tissues. Many of the differentially expressed antisense transcripts likely represent long ncRNAs. A subset of genes that mainly generate antisense transcripts in normal but not cancer cells is involved in essential metabolic processes. These findings suggest fundamental differences in global RNA regulation between normal and cancer cells that might play a role in tumorigenesis. Overall design: Global strand-specific transcriptome profilings of 2 samples in cancer and 1 sample in normal from clinical breast tissue using asymmetrical strand-specific analysis of gene expression (ASSAGE).","project":"SRP004837"}
{"number_samples":8,"species":"human","abstract":"Transcriptome profiling studies suggest that a large fraction of the genome is transcribed and many transcripts function independent of their protein coding potential. The relevance of noncoding RNAs (ncRNAs) in normal physiological processes and in tumorigenesis is increasingly recognized. Here, we describe consistent and significant differences in the distribution of sense and antisense transcripts between normal and neoplastic breast tissues. Many of the differentially expressed antisense transcripts likely represent long ncRNAs. A subset of genes that mainly generate antisense transcripts in normal but not cancer cells is involved in essential metabolic processes. These findings suggest fundamental differences in global RNA regulation between normal and cancer cells that might play a role in tumorigenesis. Overall design: Global transcriptome profilings of 12 samples in normal and 9 samples in cancer from clinical breast tissue using Sage-Seq.","project":"SRP004847"}
{"number_samples":3,"species":"human","abstract":"Here we describe the application of high-throughput sequencing technology for profiling histone and DNA methylation, and gene expression patterns of normal human mammary progenitor-enriched and luminal lineage-committed cells. We observed significant differences in histone H3 lysine 27 tri-methylation (H3K27me3) enrichment and DNA methylation of genes expressed in a cell type-specific manner, suggesting their regulation by epigenetic mechanisms and a dynamic interplay between the two processes that together define developmental potential. The technologies we developed and the epigenetically regulated genes we identified will accelerate the characterization of primary cell epigenomes and the dissection of human mammary epithelial lineage-commitment and luminal differentiation. Overall design: Global transcriptome profilings of 15 samples in normal breast tissue using Sage-Seq.","project":"SRP004965"}
{"number_samples":3,"species":"human","abstract":"H2B mono-ubiquitylation is required for multiple methylations of both H3K4 and H3K79 and has been implicated in gene expression from yeast to human. However, molecular crosstalk between H2BUb1 and other modifications, especially H3K4 and H3K79 methylations, remains unclear in vertebrates. To understand the functional role of H2BUb1, genome-wide histone modification patterns were measured in human cells. This study proposes dual roles of H2BUb1 that are both H3 methylation dependent and independent. First, H2BUb1 is a 5'-enriched active transcription mark and is co-occupied with H3K79 methylations in actively transcribed regions. Importantly, this study found a unique role of H2BUb1 in chromatin architecture independent of histone H3 methylations. H2BUb1 is well positioned in exon-intron boundaries of highly expressed exons and is specifically enriched in 5'-biased exons. Furthermore, H2BUb1 demonstrates increased occupancy in skipped exons compared to flanking exons for the human and mouse genome. Our findings suggest that a potentiating mechanism links H2BUb1 to both H3K79 methylations in actively transcribed regions and the exon-intron structure of highly expressed exons through the regulation of nucleosome dynamics during transcription elongation. Overall design: We generated high-throughput sequencing (ChIP-seq) data for genome-wide occupancy of H2BUb1, nucleosome, H3K4me3, H3K36me3, H379me1/2/3, H3Ac, and mRNA in human embryonic carcinoma cells. We performed ChIP-seq for seven different histone modifications, MNase-seq, mRNA-seq (two replications), and inputDNA-seq in NCCIT cell lines (human embryonic carcinoma cell lines). We also performed mRNA-seq for RNF20-siRNA transfected NCCIT cells, where H2BUb1 signals decreased.","project":"SRP005129"}
{"number_samples":1,"species":"human","abstract":"The human tumor antigen PRAME (Preferentially expressed antigen of melanoma) is frequently overexpressed in tumors. High PRAME levels correlate with poor clinical outcome of several cancers, but the mechanisms by which PRAME could be involved in tumorigenesis remain largely elusive. We applied protein-complex purification strategies and identified PRAME as a substrate recognition subunit of a Cullin2-based E3 ubiquitin ligase. Genome-wide chromatin immunoprecipitation experiments revealed that PRAME is specifically enriched at NF-Y promoters that are transcriptionally active, suggesting a role in gene activation. Our results are consistent with the involvement of the PRAME ubiquitin ligase complex in NF-Y-mediated transcriptional regulation. Overall design: ChIP-seq binding profiles of PRAME (ChIP-seq using the preimmune serum was used as negative control), NFYA, and NFYB, and expression analysis by RNA-seq in K562 human leukemia cell line","project":"SRP005174"}
{"number_samples":1,"species":"human","abstract":"Alternative polyadenylation (APA) of mRNAs has emerged as an important mechanism for post-transcriptional gene regulation in higher eukaryotes. Although microarrays have recently been used to characterize APA globally, they have a number of serious limitations that prevents comprehensive and highly quantitative analysis. To better characterize APA and its regulation, we have developed a deep sequencing-based method called Poly(A) Site Sequencing (PAS-Seq) for quantitatively profiling RNA polyadenylation at the transcriptome level. PAS-Seq not only accurately and comprehensively identifies poly(A) junctions in mRNAs and noncoding RNAs, but also provides quantitative information on the relative abundance of polyadenylated RNAs. Overall design: We first analyzed HeLa cell transcriptome using PAS-Seq to demonstrate that PAS-Seq not only accurately and comprehensively identifies poly(A) junctions, but also provide quantitative information on the relativel abundance of APA isoforms. Next we analyzed the mouse embryonic stem cells, neural stem/progenitor cells, and neurons by PAS-Seq to characterize the dynamic changes of mRNA polyadenylation during stem cell differentiation.","project":"SRP005177"}
{"number_samples":10,"species":"human","abstract":"The second wave of next generation sequencing technologies, referred to as single-molecule sequencing (SMS), carries the promise of profiling samples directly without employing polymerase chain reaction steps used by amplification-based sequencing (AS) methods. To examine the merits of both technologies, we examine mRNA sequencing results from single-molecule and amplification-based sequencing in a set of human cancer cell lines and tissues. We observe a characteristic coverage bias towards high abundance transcripts in amplification-based sequencing. A larger fraction of AS reads cover highly expressed genes, such as those associated with translational processes and housekeeping genes, resulting in relatively lower coverage of genes at low and mid-level abundance. In contrast, the coverage of high abundance transcripts plateaus off using SMS; Consequently, SMS is able to sequence lower- abundance transcripts more thoroughly, including some that are undetected by AS methods; however, these include many more mapping artifacts. A better understanding of the technical and analytical factors introducing platform specific biases in high throughput transcriptome sequencing applications will be critical in cross platform meta-analytic studies.","project":"SRP005242"}
{"number_samples":36,"species":"human","abstract":"We determined the genome-wide digital gene expression (DGE) profiles of primary acute lymphoblastic leukemia (ALL) cells from 28 patients and fractionated blood cells from healthy blood donors taking advantage of “second generation” sequencing technology. The patients included in the study represent distinct subtypes of B-cell precursor (BCP) ALL and T-cell lineage ALL (T-ALL) and the controls are fractionated CD19+ and CD3+ cells. Overall design: Gene expression analysis of 28 ALL patient samples with different  immunophenotypic backgrounds including T-ALL (n=4) and patients with BCP ALL with diverse cytogenetic backgrounds: High Hyperploidy (HeH) (n=10), t(9;22) BCR-ABL1 (n=3), t(12;21) ETV6-RUNX1 (n=4), dic(9;20) (n=3), t(1;19)TCF3-PBX1, MLL/11q23 (n=1) and undefined/non-recurrent aberrations (n=1). Fractionated b-cells (CD19+) and t-cells (CD3+) isolated from peripheral blood of healthy donors were used as controls. Sequencing libraries were prepared from 1 µg of total RNA using reagents from the NlaIII Digital Gene Expression Tag Profiling kit (Illumina Inc, San Diego, CA, USA). mRNA was captured on magnetic oligo(dT) beads and reverse transcribed into double-stranded cDNA. The cDNA was cleaved using the restriction enzyme NlaIII. An adapter sequence containing the recognition sequence for the restriction enzyme MmeI was ligated to the NlaIII cleavage sites. The adapter-ligated cDNA was digested with MmeI to release the cDNA from the magnetic bead, while leaving 17 base-pairs of sequence in the fragment. The fragments were dephosphorylated and purified by phenol-chloroform. A second adapter was ligated at the MmeI cleavage sites. The adapter-ligated cDNA fragments were amplified by PCR, and the PCR products were purified on a 6% polyacrylamide gel. The ~96 base pair PCR products were excised from the gel and eluted overnight, followed by ethanol precipitation and re-suspension. Purified libraries were quality controlled and quantified on a Bioanalyzer using DNA 1000 series or High Sensitivity chips.  The DGE libraries were diluted to a 10 nM concentration and sequenced on one lane of an Illumina GAII or GAIIx for 18 cycles.","project":"SRP005279"}
{"number_samples":7,"species":"human","abstract":"Enhancers of transcription activate transcription via binding of sequence-specific transcription factors to their target sites in chromatin. In this report, we identify GATA1-bound enhancers genome-wide and find a global reorganization of the nucleosomes at these enhancers during differentiation of hematopoietic stem cells (HSCs) to erythrocytes. We show that the catalytic subunit BRG1 of BAF complexes localizes to these enhancers during differentiation and generates a longer nucleosome repeat length surrounding the GATA1 sites by shifting the flanking nucleosomes away. Intriguing, we find that the nucleosome shifting specifically facilitates binding of TAL1 but not GATA1, which subsequently activates transcription of target genes. Overall design: Human hematopoietic stem cells (referred as CD34+ cells) were differentiated into erythroid lineage expressing the cell surface marker CD36 (referred as CD36+ cells). To study the function of ATP-dependent chromatin remodeling BAF complexes in the establishment of erythroid specific enhancers during differentiation, we mapped the genome-wide binding sites of GATA1 and TAL1, the key transcription regulators of erythroid differentiation, in CD36+ cells, then determined the genomic positions of the catalytic subunit BRG1 of the BAF complexes and the nucleosome landscapes, by ChIP-Seq and Mnase-Seq, respectively, for both HSCs and the differentiated cells. We compared changes in nucleosome landscapes with BRG1 binding at GATA1 and TAL1 binding sites and found that BRG1 binding is correlated with nucleosome shifting away from the binding sites, which is critical for TAL1 binding. To confirm that BRG1 is responsible for the nucleosome shift, we knocked down BRG1 in CD36+ cells and analyzed the nucleosome changes in the knockdown cells. The data indicated that inhibition of BRG1 caused a nucleosome shift towards the GATA1 or TAL1 sites, indicating that BRG1 is indeed responsible for the observed nucleosome shift.","project":"SRP005281"}
{"number_samples":4,"species":"human","abstract":"In this study, we took advantage of deep sequencing approaches to investigate the miRNA expression profiles of human endometrium on days LH+2 and LH+7 in natural cycles, and compare them with those on days hCG+4 and hCG+7 in stimulated cycles during IVF treatment. Overall design: Examination of miRNA expression in human endometrium during natural and stimulated cycles","project":"SRP005309"}
{"number_samples":12,"species":"human","abstract":"Abstract: Alternative splicing (AS) plays a major role in the generation of proteomic diversity and in gene regulation. However, the role of the basal splicing machinery in regulating AS remains poorly understood. Here we show that the core snRNP protein SmB/B’ self-regulates its expression by promoting the inclusion of a highly-conserved alternative exon in its own pre-mRNA that targets the spliced transcript for nonsense-mediated mRNA decay (NMD). Depletion of SmB/B’ in human cells results in reduced levels of snRNPs and in a striking reduction in the inclusion levels of hundreds of alternative exons, with comparatively few effects on constitutive exon splicing levels. The affected alternative exons are enriched in genes encoding RNA processing and other RNA binding factors, and a subset of these exons also regulate gene expression by activating NMD. Our results thus demonstrate a role for the core spliceosomal machinery in controlling an exon network that appears to modulate the levels of many RNA processing factors. Overall design: HeLa cells were transfected with a control non-targeting siRNA pool (siNT), or with siRNA pools designed to knockdown SmB/B' or SRSF1 (also known as SF2/ASF/SFRS1). Sequence reads were aligned to exon-exon junction sequences in a database of EST/cDNA-mined cassette-type alternative splicing events. Processed data files (.bed and .txt) provided as supplementary files on the Series record.  Processed data file build information: hg18.","project":"SRP005342"}
{"number_samples":1,"species":"human","abstract":"Second-generation sequencing is gradually becoming the method of choice for miRNA detection and expression profiling1. Given the relatively small number of miRNAs and improvements in DNA sequencing technology studying miRNA expression profiles of multiple samples in a single flow cell lane becomes feasible. Multiplexing strategies require marking each miRNA library with a DNA bar code. Here we report that bar-codes introduced through adapter ligation confer significant bias on miRNA expression profiles. This bias is much higher than expected Poisson noise and masks significant expression differences between miRNA libraries. This bias can be eliminated by adding bar-codes during PCR amplification of libraries. The accuracy of miRNA expression measurement in multiplexed experiments becomes a function of sample number.","project":"SRP005449"}
{"number_samples":4,"species":"human","abstract":"The Carboxy-terminal domain (CTD) of RNA Polymerase II (RNAPII) in mammals undergoes extensive post-translational modification, which is essential for transcriptional initiation and elongation. Here, we show that the CTD of RNAPII is methylated at a single arginine (R1810) by the transcriptional co-activator CARM1. Although methylation at R1810 is present on the hyper-phosphorylated form of RNAPII in vivo, Ser-2 or Ser-5 phosphorylation inhibit CARM1 activity towards this site in vitro, suggesting that methylation occurs before transcription initiation. Mutation of R1810 results in the mis-expression of a variety of snRNAs and snoRNAs, an effect that is also observed in Carm1-/- MEFs.  These results demonstrate that CTD methylation facilitates the expression of select RNAs, perhaps serving to discriminate the RNAPII-associated machinery recruited to distinct gene types. Overall design: To address the function of RNAPII methylation, we generated Raji cell lines expressing an RNA Polymerase II resistant to a-amanitin and carrying either wild-type R1810 or an arginine to alanine substitution at that same residue, abolishing R1810 methylation of the CTD. In cells cultured in a-amanitin, the a-amanitin-resistant mutants fully replaced the functions of endogenous RNAPII, allowing us to study if gene-expression is affected by the absence of R1810me","project":"SRP005846"}
{"number_samples":4,"species":"human","abstract":"Transcriptionally active and inactive chromatin domains tend to segregate into separate sub-nuclear compartments to maintain stable expression patterns. However, we have uncovered here an inter-chromosomal network connecting active loci enriched in circadian genes to repressed lamina-associated domains (LADs). The interactome is regulated by PARP1 and its co-factor, CTCF, which not only mediate chromatin fiber interactions, but also promote the recruitment of circadian genes to the lamina. Synchronization of circadian rhythm by serum shock induces oscillations in PARP1-CTCF interactions, which is accompanied by oscillating recruitment of circadian loci to the lamina, followed by the acquisition of repressive H3K9me2 marks and transcriptional attenuation. Furthermore, depletion of H3K9me2/3, inhibition of PARP activity by Olaparib or down-regulation of PARP1 or CTCF expression counteracts both recruitment to the envelope and circadian transcription. PARP1- and CTCF-regulated contacts between circadian loci and the repressive chromatin environment at the lamina thus mediate circadian transcriptional plasticity. Overall design: Examination of intra- and interchromosomal interactions in human embryonic stem cells (HESCs) and derived embryoid bodies (HEBs) generated by 4C-seq, as described in (Zhao et al., 2006), using Illumina Genome Analyzer II and Illumina MiSeq. Examination of mRNA profiles in HESCs and derived HEBs generated by deep sequencing, in duplicate, using Illumina HiSeq 2500.","project":"SRP005858"}
{"number_samples":3,"species":"human","abstract":"Mammalian genomes are populated with thousands of transcriptional enhancers that orchestrate cell type-specific gene expression programs; however, the potential that there are pre-established enhancers in different functional classes that permit alternative signal-dependent transcriptional responses has remained unexplored. Here we present evidence that cell lineage-specific factors, such as FoxA1, can simultaneously facilitate and restrict key regulated transcription factors, exemplified by the androgen receptor (AR), acting at structurally- and functionally-distinct classes of pre-established enhancers, thus licensing specific signal-activated responses while restricting others. Consequently, FoxA1 down-regulation, an unfavorable prognostic sign in advanced prostate tumors, causes a massive switch in AR binding from one functional class of enhancers to another, with a parallel switch in levels of enhancer-templated non-coding RNAs (eRNAs) revealed by the global run-on assay (GRO-seq), which documents the dramatic reprogramming of the hormonal response.  The molecular basis for this switch lies in the release of FoxA1-mediated restriction of AR binding to the new enhancer class with no apparent nucleosome remodeling, which is required for stimulating their eRNA transcription and/or enhancing enhancer:promoter looping and gene activation. Together, these findings reveal a large repository of pre-determined enhancers in the human genome that can be dynamically tuned to induce their transcription and activation of alternative gene expression programs, which may underlie many sequential gene expression events in development or during disease progression. Overall design: ChIP-Seq, Gro-Seq, and gene expression profiling was performed in LNCaP cells with hormone treatment and siRNA against FoxA1 ChIP-Seq and Gro-Seq data presented here. Supplementary file GroSeq.denovo.transcripts.hg18.bed represents analysis using GSM686948-GSM686950.","project":"SRP005967"}
{"number_samples":8,"species":"human","abstract":"HP1 alpha, beta, and gamma play an evolutionarily conserved role in the formation and maintenance of heterochromatin. In addition, some HP1 family members may also participate in transcriptional regulation of genes. Recently, HP1 gamma binding to the bodies of a subset of genes has been observed in human and murine cells. However, the generality of this phenomenon and the role HP1 gamma may play in this context are unknown. Genome-wide localization analysis reveals broad and general HP1 gamma binding at actively transcribed genes, which strongly correlates with gene activity across multiple cell types. Unexpectedly, we find HP1 gamma binding is necessary for efficient RNA processing of its target genes. Loss of HP1 gamma results in defective recruitment of splicing factors including U1-70K, and leads to accumulation of unspliced nascent transcripts genome-wide. Therefore, our data argue that the primary role of HP1 gamma is to ensure efficient RNA processing in mammalian cells. Overall design: Genome-wide localization and function of HP1 gamma in multiple cell types using ChIP-chip, ChIP-seq, and RNA-seq","project":"SRP006309"}
{"number_samples":13,"species":"human","abstract":"Crosslinking and immunoprecipitation (CLIP) is increasingly used to map transcriptome-wide binding sites of RNA-binding proteins (RBPs). We developed a method for CLIP data analysis and applied it to compare 254 nm CLIP with PAR-CLIP, which involves crosslinking of photoreactive nucleotides with 365 nm UV light. We found small differences in the accuracy of these methods in identifying binding sites of HuR, a protein that binds low-complexity sequences and Argonaute 2, which has a complex binding specificity. We show that crosslink-induced mutations lead to single-nucleotide resolution for both PAR-CLIP and CLIP. Our results confirm the expectation from original CLIP publications that RNA-binding proteins do not protect sufficiently their sites under the denaturing conditions used during the CLIP procedure, and we show that extensive digestion with sequence-specific ribonucleases strongly biases the set of recovered binding sites. We finally show that this bias can be substantially reduced by milder nuclease digestion conditions. Overall design: We performed duplicate experiments for each variant of the CLIP protocol (CLIP, PAR-CLIP), each protein (HuR, Ago2), and enzymatic digestion (complete T1 digestion, mild MNase digestion). In addition, we performed a single PAR-CLIP experiment with mild T1 digestion.","project":"SRP006474"}
{"number_samples":10,"species":"human","abstract":"Crosslinking and immunoprecipitation (CLIP) is increasingly used to map transcriptome-wide binding sites of RNA-binding proteins (RBPs). We developed a method for CLIP data analysis and applied it to compare 254 nm CLIP with PAR-CLIP, which involves crosslinking of photoreactive nucleotides with 365 nm UV light. We found small differences in the accuracy of these methods in identifying binding sites of HuR, a protein that binds low-complexity sequences and Argonaute 2, which has a complex binding specificity. We show that crosslink-induced mutations lead to single-nucleotide resolution for both PAR-CLIP and CLIP. Our results confirm the expectation from original CLIP publications that RNA-binding proteins do not protect sufficiently their sites under the denaturing conditions used during the CLIP procedure, and we show that extensive digestion with sequence-specific ribonucleases strongly biases the set of recovered binding sites. We finally show that this bias can be substantially reduced by milder nuclease digestion conditions. Overall design: We performed duplicate mRNA-Seq experiments for cells grown in the presence of modified nucleotides or in normal medium, and following crosslinking at 365nm, 254nm or without crosslinking. In addition, we performed single mRNA-Seq experiments of cells transfected with anti-GFP or anti-HuR siRNA.","project":"SRP006475"}
{"number_samples":1,"species":"human","abstract":"Combining an in vitro hNCC differentiation protocol with epigenomic profiling, we provide the first whole-genome characterization of cis-regulatory elements in this highly relevant cell type. With this data at hand, we have characterized the chromatin state and dynamics of all human gene promoters during the course of NCC in vitro differentiation. Most importantly, we have identified a large cohort of active and NCC-specific enhancers, which we showed to be functionally relevant in vivo, in the context of embryonic development. Finally, through sequence analysis of the identified NCC enhancers, we uncovered the orphan nuclear receptors NR2F1 and NR2F2 as novel hNCC transcriptional regulators both in vitro and in vivo. Overall design: RNA-seq experiments in human neural crest cells (hNCC)","project":"SRP006561"}
{"number_samples":245,"species":"human","project":"SRP006574"}
{"number_samples":114,"species":"human","abstract":"Molecular characterization of tumors has been critical for identifying important genes in cancer biology and for improving tumor classification and diagnosis. Long non-coding RNAs (lncRNAs), as a new, relatively unstudied class of transcripts, provide a rich opportunity to identify both functional drivers and cancer-type specific biomarkers. However, despite the potential importance of lncRNAs to the cancer field, no comprehensive survey of lncRNA expression across various cancers has been reported. We used 3'-End Sequencing for Expression Quantification (3SEQ) to quantify transcript abundance across 64 solid tumors representing 17 diagnostic subtypes of adenocarcinomas, squamous cell carcinomas, and sarcomas. We identified hundreds of transcripts from among the known 1,065 lncRNAs surveyed that show variability in transcript levels between the tumor types, and therefore, make potential biomarker candidates. We discovered 1,071 novel intergenic transcribed regions and demonstrate that these show similar patterns of variability between tumor types. We find that many of these differentially expressed cancer transcripts are also expressed in normal tissues. One such novel transcript specifically expressed in breast tissue was further evaluated using RNA in situ hybridization on a panel of breast tumors and shown to correlate with low tumor grade and estrogen receptor expression, thereby representing a potentially important new breast cancer biomarker. This study provides the first large survey of lncRNA expression within a panel of solid cancers and also identifies a number of novel transcribed regions differentially expressed across distinct cancer types that represent candidate biomarkers for future research. Overall design: 3SEQ was performed on 64 formalin-fixed, paraffin-embedded (FFPE) human tumors representing 17 diagnostic cancer subtypes. Duplicate libraries were prepared for two of the tumors (ESS STT5520 and LMS STT516). 3SEQ was also performed on 27 normal human tissue samples. RNA-Seq was performed on 6 breast cancer cell lines for the examination of the breast-specific transcript on chr10. Series supplementary files: 'GSE28866_raw_counts_54511_peaks_cancer_and_normal.txt': Raw_counts: Total 3SEQ reads in each peak for each sample. File includes read counts for 54,511 peaks for the 66 cancer libraries and the 27 normal libraries. 'GSE28866_36048_normalized_peaks_cancer_and_normal.txt': Normalized_peaks: Normalized 3SEQ expression data for the 36,048 filtered peaks for the 66 cancer libraries and the 27 normal libraries. Peaks were classified as coding, lncRNA (Rinn, Bartel, Hughes, Gencode_lnc), other known transcripts (ref_known_Gencode_nc, all_mrna, other known), downstream, intron, promoter, or novel_intergenic. Peaks determined to be differentially expressed in one of the 17 cancer subtypes using a series of 2-Class SAM analyses are noted along with the type of cancer showing upregulation. Expression data was normalized using the sequencing depth of each sample by scaling the data using the mean value of each sample. Data was further compressed to reduce outliers by taking the square root of each value.","project":"SRP006575"}
{"number_samples":4,"species":"human","abstract":"Regulatory T-cells (Treg) play an essential role in the negative regulation of immune answers by developing an attenuated cytokine response that allows suppressing proliferation and effector function of T-cells (CD4+ Th). The transcription factor FoxP3 is responsible for the regulation of many genes involved in the Treg gene signature. Its ablation leads to severe immune deficiencies in human and mice. Recent developments in sequencing technologies have revolutionized the possibilities to gain insights into transcription factor binding by ChiP-Seq and into transcriptome analysis by mRNA-Seq. We combine FoxP3 ChiP-Seq and mRNA-Seq in order to understand the transcriptional differences between primary human CD4+ T helper and regulatory T-cells, as well as to study the role of FoxP3 in generating those differences. We show, that mRNA-Seq allows analyzing the transcriptomal landscape of T-cells including the expression of specific splice variants at much greater depth than previous approaches, whereas 50% of transcriptional regulation events have not been described before by using diverse array technologies.","project":"SRP006674"}
{"number_samples":8,"species":"human","abstract":"mRNA expression was profiled from pooled bronchial airway epithelial cell brushings (n=3 patients/pool) obtained during bronchoscopy from healthy never (NS) and current smokers (S) and smokers with (C) and without (NC) lung cancer Overall design: 4 samples were sequened, each representing a pool of 3 patients.  The phenotypes of the 4 samples were as follows: healthy non-smoker, healthy smoker, smoker without lung cancer and smoker with lung cancer.","project":"SRP006676"}
{"number_samples":2,"species":"human","abstract":"Transcriptomic analyses have revealed an unexpected complexity of the human transcriptome, whose breadth and depth exceeds current RNA sequencing capacity 1-3. Here we combine tiling arrays and RNA sequencing technologies, in an approach we term RNA Capture-Seq, to enable the assembly and characterisation of transcripts whose expression level is below the detection limits of conventional sequencing approaches. By this technique we identify novel exons and splicing patterns to even well-studied genes, such as the p53 and Hox loci, and expose widespread, regulated and remarkably complex noncoding transcription in intergenic gene deserts. We show that unassigned tags observed in conventional RNA sequencing datasets are not noise but can derive from rare transcripts fully assembled by RNA Capture-Seq. We expect RNA Capture-Seq to prove an invaluable technique for future research into gene expression and its relationship to phenotypic variation. Overall design: Application of RNA Capture sequencing from human fibroblasts for identifying rare novel transcripts. Hybridization enhancing oligos are designed to cover the full sequencing adaptor and MID sequences during hybridisation. Enhancing Oligo A 5   CCATCTCATCCCTGCGTGTCTCCGACTCAG/3ddc/ Enhancing Oligo B 5'   CCTATCCCCTGTGTGCCTTGGCAGTCTCAG/3ddc/","project":"SRP006717"}
{"number_samples":3,"species":"human","abstract":"Next Generation Sequencing technologies have enabled de novo gene fusion discovery that could reveal candidates with therapeutic significance in cancer. Here we present an open-source software package, ChimeraScan, for the discovery of chimeric transcription between two independent transcripts. Overall design: Three cancer cell lines with known gene fusions","project":"SRP006719"}
{"number_samples":4,"species":"human","abstract":"While genetic mutation is a hallmark of cancer, many cancers also acquire epigenetic alterations during tumorigenesis including aberrant DNA hypermethylation of tumor suppressors as well as changes in chromatin modifications as caused by genetic mutations of the chromatin-modifying machinery. However, the extent of epigenetic alterations in cancer cells has not been fully characterized. Here, we describe the first complete methylome maps at single nucleotide resolution of a low-passage breast cancer cell line and primary human mammary epithelial cells. We find widespread DNA hypomethylation in the cancer cell, primarily at partially methylated domains (PMDs) in normal breast cells. Unexpectedly, genes within these regions are largely silenced in cancer cells. The loss of DNA methylation in these regions is accompanied by formation of repressive chromatin, with a significant fraction displaying allelic DNA methylation where one allele is DNA methylated while the other allele is occupied by histone modifications H3K9me3 or H3K27me3. Our results show a mutually exclusive and complementary relationship between DNA methylation and H3K9me3 or H3K27me3. These results suggest that global DNA hypomethylation in breast cancer is tightly linked to the formation of repressive chromatin domains and gene silencing, thus identifying a potential epigenetic pathway for gene regulation in cancer cells and suggesting a possible new approach toward the development of cancer therapeutics. Overall design: mRNA-Seq of polyA-selected RNA from breast cancer HCC1954 and normal breast HMEC. 36 cycles of sequencing on Illumina platform.","project":"SRP006726"}
{"number_samples":11,"species":"human","abstract":"Beginning with precursor lesions, aberrant DNA methylation marks the entire spectrum of prostate cancer progression. We mapped the global DNA methylation patterns in selected prostate tissues and cell lines using Methylplex-Next Generation Sequencing (M-NGS). Hidden Markov Model based next generation sequence analysis identified ~68,000 methylated regions per sample. While global CpG Island (CGI) methylation was not differential between benign adjacent and cancer samples, overall promoter CGI methylation increased from ~12.6% in benign samples to 19.3% and 21.8% in localized and metastatic cancer tissues, respectively. We found distinct patterns of promoter methylation around transcription start sites, where methylation occurred not only on the CGIs, but also on flanking regions and CGI sparse promoters. Among the 6,691 methylated promoters in prostate tissues, 2481 differentially methylated regions (DMRs) are cancer specific, including numerous novel DMRs. A novel cancer specific DMR in WFDC2 promoter showed heavy methylation in cancer (17/22 tissues, 6/6 cell lines), but not in the benign tissues (0/10) and normal PrEC cells. Integration of LNCaP DNA methylation and H3K4me3 data suggested an epigenetic mechanism for alternate transcription start site utilization and these modifications segregated into distinct regions when present on the same promoter. Finally, we observed differences in repeat element methylation, particularly LINE-1, between ERG gene fusion positive and negative cancers. This comprehensive methylome map will further our understanding of epigenetic regulation in prostate cancer progression. Overall design: Next generation Sequencing for Gene expression using the RNA-Seq methodology from LNCaP and PrEC cell lines","project":"SRP006731"}
{"number_samples":10,"species":"human","abstract":"We report how unique RNA seq profiles of ribosomal RNA (18S, 28S) present in sperm indicate specific cleavage sites.  Additionally, these sequencing results reveal how overall character of RNA (GC content) can significantly affect overall distribution of mapped reads. Overall design: Examination of rRNA population and distribution in spermatozoa","project":"SRP006769"}
{"number_samples":4,"species":"human","abstract":"The RNA from two paired normal and colorectal tumor tissue were sequenced by Illumina genome analyzer. 0.8 to 1.1 million reads were generated. Sequencing data was aligned with human RNA, human mitochondrial DNA and human genomic DNA by BWA.  Base calling and Phred-like score was calculated by SAMTools. Overall design: Whole transcriptome sequence of paired normal and tumor tissues obtained from two colorectal cancer patients","project":"SRP006900"}
{"number_samples":5,"species":"human","abstract":"Background: Breast cancers arising from germline BRCA1 mutations are usually high grade, clinically aggressive tumors that are associated with a poor prognosis. Alterations in a number of pathways contribute to this phenotype including genomic stability, cell cycle and cell proliferation. RNA splicing is also often perturbed in cancers but it is not known whether this contributes to tumorigenesis in BRCA1-related breast cancers.  Methods: Using next-generation mRNA sequencing (RNA-Seq), we compared the transcriptome of four breast cancer cell lines and three primary breast cancers from BRCA1 germline mutation carriers to five breast cancer cell lines and two breast cancers occurring in non-BRCA mutation carriers. We created a systematic computational pipeline to detect potentially abnormal novel splice junctions (i.e. absent from RefSeq and UCSC) with irregular expression levels in BRCA1¬-related breast cancers compared to the non-BRCA control cancers.   Results: We found that a greater number of common splice junctions not annotated in RefSeq or UCSC databases were either up- or down-regulated in the BRCA1-related breast cancers than expected by chance. We present a list of highly recurrent splicing events, with the most discordant expression patterns compared to other breast cancers. Most of these are predicted to disrupt protein domains, cause frameshifts or are located in important regulatory genes. However, we were not able to detect any abnormally spliced transcripts which clearly contribute to the phenotype of the BRCA1-related breast cancers.  Conclusions: RNA-Seq is a powerful tool for analyzing abnormal transcripts on a genome-wide scale. With RNA-Seq, several potentially abnormal transcripts were found to be more highly expressed in the BRCA1-related breast cancers than in the non-BRCA1 breast cancer controls. However, we did not find evidence that highly recurrent, highly expressed abnormal alternative splice variants of specific genes are predominant drivers of tumorigenesis in BRCA1-related breast cancers.","project":"SRP006908"}
{"number_samples":5,"species":"human","abstract":"TGF-beta1 is the major cytokine driver of fibrotic scarring observed in diabetic nephropathy and other fibrosis-related diseases. RNA-sequencing offers the potential for more sensitive assessment of the TGF-ß1-driven transcriptome. Overall design: There were two treatment groups: vehicle, 48 hr TGFb1. Each treatment was carried out in triplicate. Upon quality control assessment, one TGFß1 treated sample was excluded from further analyses, leaving 3 unstimulated and 2 TGFß1 samples.","project":"SRP006912"}
{"number_samples":4,"species":"human","abstract":"Establishing the functional roles of genetic variants remains a significant challenge in the post-genomic era. Here, we present a method, allele-specific alternative mRNA processing (ASARP), to identify genetically influenced mRNA processing events using transcriptome sequencing (RNA-Seq) data. The method examines RNA-Seq data at both single nucleotide and whole-gene/isoform levels to identify allele-specific expression (ASE) and existence of allele-specific regulation of mRNA processing. We applied the methods to data obtained from the human glioblastoma cell line U87MG and primary breast cancer tissues and found that 26–45% of all genes with sufficient read coverage demonstrated ASE, with significant overlap between the two cell types. Our methods predicted potential mechanisms underlying ASE due to regulations affecting either whole-gene-level expression or alternative mRNA processing, including alternative splicing, alternative polyadenylation and alternative transcriptional initiation. Allele-specific alternative splicing and alternative polyadenylation may explain ASE in hundreds of genes in each cell type. Reporter studies following these predictions identified the causal single nucleotide variants (SNVs) for several allele-specific alternative splicing events. Finally, many genes identified in our study were also reported as disease/phenotype-associated genes in genome-wide association studies. Future applications of our approach may provide ample insights for a better understanding of the genetic basis of gene regulation underlying phenotypic diversity and disease mechanisms. Overall design: Examine allele-specific gene expression and alternative RNA processing in U87MG cell line","project":"SRP006970"}
{"number_samples":6,"species":"human","abstract":"We performed the integrative transcriptome analysis of human esophageal squamous cell carcinoma (ESCC) using Illumina high-throughput sequencing. A total of 187 million 38bp sequencing reads were generated containing 7 billion bases for three pairs of matched patient-derived ESCC clinical specimens and their adjacent non-tumorous tissues. By investigating the digital gene expression profiling, we found 1425 genes significantly differentially expressed and detected more than 9000 SNPs across all six samples. We also identified protein tyrosine kinase 6 (PTK6) as a novel tumor suppressor gene, which is critical in ESCC development. Overall design: Analysis of whole transcriptome from 3 paired patient-derived ESCC clinical specimens and their adjacent non-tumorous tissues.","project":"SRP007169"}
{"number_samples":16,"species":"human","abstract":"Thousands of human genes contain introns ending in NAGNAG motifs (N any nucleotide), where both NAGs can function as 3' splice sites, yielding isoforms differing by inclusion/exclusion of just three bases. However, the functional importance of NAGNAG alternative splicing is highly controversial. Using very deep RNA-Seq data from sixteen human and eight mouse tissues, we found that approximately half of alternatively spliced NAGNAGs undergo tissue-specific regulation and that regulated events have been selectively retained: alternative splicing of strongly tissue-specific NAGNAGs was ten times as likely to be conserved between species as for non-tissue-specific events. Further, alternative splicing of human NAGNAGs was associated with an order of magnitude increase in the frequency of exon length changes at orthologous mouse/rat exon boundaries, suggesting that NAGNAGs accelerate exon evolution. Together, our analyses show that NAGNAG alternative splicing constitutes a major generator of tissue-specific proteome diversity and accelerates evolution of proteins at exon-exon boundaries. Overall design: mRNA-Seq of sixteen human and eight mouse tissues. Supplementary files: human.nagnag.junctions.gff and mouse.nagnag.junctions.gff are the annotation files (in GFF3 format) corresponding to the 'bwtout' mapped reads files linked to the Sample records. Raw data files provided for Samples GSM742937-GSM742952 only.","project":"SRP007338"}
{"number_samples":4,"species":"human","project":"SRP007351"}
{"number_samples":11,"species":"human","abstract":"We describe PolyA-Seq, a strand-specific method for high-throughput sequencing of the 3' ends of polyadenylated transcripts. PolyA-Seq is as accurate for digital gene expression as existing RNA sequencing approaches, and superior to microarrays. We used the approach to map polyadenylation (polyA) sites in 24 samples from normal tissues in human, rhesus, dog, mouse, and rat. Overall design: Detection of polyA sites in a mixture of 24 tissues in human, mouse, rat, dog and rhesus. Samples included two replicates each of MAQC Human Brain Reference and MAQC Universal Human Reference. Two additional human sets of reads are included that were used to distinguish true polyA sites from internal priming sites.","project":"SRP007359"}
{"number_samples":2,"species":"human","abstract":"Regulation of cell-cell junction formation and regulation of cell migration were enriched among EMT (Epithelial-Mesenchymal Transition)-associated alternatively splicing events. Our analysis suggested that most EMT-associated alternative splicing events are regulated by one or more members of the RBFOX, MBNL, CELF, hnRNP or ESRP classes of splicing factors. The EMT alternative splicing signature was confirmed in human breast cancer cell lines, which could be classified into basal and luminal subtypes based exclusively on their EMTassociated splicing pattern. Expression of EMT-associated alternative mRNA transcripts was also observed in primary breast cancer samples, indicating that EMT-dependent splicing changes occur commonly in human tumors. The functional significance of EMT-associated alternative splicing was tested by expression of the epithelial-specific splicing factor ESRP1 or depletion of RBFOX2 in mesenchymal cells, both of which elicited significant changes in cell morphology and motility towards an epithelial phenotype, suggesting that splicing regulation alone can drive critical aspects of EMT-associated phenotypic changes. The molecular description obtained here may aid in the development of new diagnostic and prognostic markers for analysis of breast cancer progression. Overall design: Examination of transcriptomes of HMLE/Twist-ER before and after induction of EMT by tamoxifen","project":"SRP007403"}
{"number_samples":21,"species":"human","abstract":"Changes in gene expression are thought to underlie many of the phenotypic differences between species. However, large-scale analyses of gene expression evolution were until recently prevented by technological limitations. Here we report the sequencing of polyadenylated RNA from six organs across ten species that represent all major mammalian lineages (placentals, marsupials and monotremes) and birds (the evolutionary outgroup), with the goal of understanding the dynamics of mammalian transcriptome evolution. We show that the rate of gene expression evolution varies among organs, lineages and chromosomes, owing to differences in selective pressures: transcriptome change was slow in nervous tissues and rapid in testes, slower in rodents than in apes and monotremes, and rapid for the X chromosome right after its formation. Although gene expression evolution in mammals was strongly shaped by purifying selection, we identify numerous potentially selectively driven expression switches, which occurred at different rates across lineages and tissues and which probably contributed to the specific organ biology of various mammals. Our transcriptome data provide a valuable resource for functional and evolutionary analyses of mammalian genomes. Overall design: To study mammalian transcriptome evolution at high resolution, we generated RNA-Seq data (~3.2 billion Illumina Genome Analyser IIx reads of 76 base pairs) for the polyadenylated RNA fraction of brain (cerebral cortex or whole brain without cerebellum), cerebellum, heart, kidney, liver and testis (usually from one male and one female per somatic tissue and two males for testis) from nine mammalian species: placental mammals (great apes, including humans; rhesus macaque; mouse), marsupials (gray short-tailed opossum) and monotremes (platypus). Corresponding data (~0.3 billion reads) were generated for a bird (red jungle fowl, a non-domesticated chicken) and used as an evolutionary outgroup. Important note: Fluorophore intensity files were processed with the Ibis base caller (version 1.11). As illustrated in Supplementary Note Table 1 and Supplementary Note Figure 1 of Brawand et al. (Nature 2011), we found that Ibis significantly increased the number of usable reads and drastically reduced the error rate. However, Ibis quality values are not comparable to those from Illumina values, which should be taken into account when using our data and interpreting their quality relative to data established using other base calling approaches.","project":"SRP007412"}
{"number_samples":4,"species":"human","abstract":".","project":"SRP007417"}
{"number_samples":205,"species":"human","project":"SRP007461"}
{"number_samples":2,"species":"human","abstract":"We report the gene expression profile of human endometriual stromal cells and armadillo endometrial samples Overall design: We report the gene expression profile of human endometriual stromal cells and armadillo endometrial samples generated with Illumina GA2","project":"SRP007481"}
{"number_samples":12,"species":"human","abstract":"Autism spectrum disorder (ASD) is a common, highly heritable neurodevelopmental condition characterized by marked genetic heterogeneity. Thus, a fundamental question is whether autism represents an aetiologically heterogeneous disorder in which the myriad genetic or environmental risk factors perturb common underlying molecular pathways in the brain. Here, we demonstrate consistent differences in transcriptome organization between autistic and normal brain by gene co-expression network analysis. Remarkably, regional patterns of gene expression that typically distinguish frontal and temporal cortex are significantly attenuated in the ASD brain, suggesting abnormalities in cortical patterning. We further identify discrete modules of co-expressed genes associated with autism: a neuronal module enriched for known autism susceptibility genes, including the neuronal specific splicing factor A2BP1 (also known as FOX1), and a module enriched for immune genes and glial markers. Using high-throughput RNA sequencing we demonstrate dysregulated splicing of A2BP1-dependent alternative exons in the ASD brain. Moreover, using a published autism genome-wide association study (GWAS) data set, we show that the neuronal module is enriched for genetically associated variants, providing independent support for the causal involvement of these genes in autism. In contrast, the immune-glial module showed no enrichment for autism GWAS signals, indicating a non-genetic aetiology for this process. Collectively, our results provide strong evidence for convergent molecular abnormalities in ASD, and implicate transcriptional and splicing dysregulation as underlying mechanisms of neuronal dysfunction in this disorder. Overall design: To identify potential A2BP1-dependent differential splicing events in ASD brain, we performed high-throughput RNA sequencing (RNA-Seq) on three autism samples with significant downregulation of A2BP1 (average fold change by quantitative RT-PCR = 5.9) and three control samples with average A2BP1 levels. The list of potential A2BP1-depending differential splicing events in ASD is given in the Supplementary file linked at the foot of this record.","project":"SRP007483"}
{"number_samples":9,"species":"human","abstract":"Large intergenic non-coding RNAs (lincRNAs) are emerging as key regulators of diverse cellular processes. Determining the function of individual lincRNAs remains a challenge. Recent advances in RNA sequencing (RNA-Seq) and computational methods allow for an unprecedented analysis of such transcripts. Here, we present an integrative approach to define a reference catalogue of over 8,000 human lincRNAs. Our catalogue unifies previously existing annotation sources with transcripts we assembled from RNA-Seq data collected from ~4 billion RNA-Seq reads across 24 tissues and cell types.  We characterize each lincRNA by a panorama of more than 30 properties, including sequence, structural, transcriptional, and orthology features. We find that lincRNA expression is strikingly tissue specific compared to coding genes, and that they are typically co-expressed with their neighboring genes, albeit to a similar extent to that of pairs of neighboring protein-coding genes. We distinguish an additional sub-set of transcripts that have high evolutionary conservation but may include short open reading frames, and may serve either as lincRNAs or as small peptides. Our integrated, comprehensive, yet conservative reference catalogue of human lincRNAs reveals the global properties of lincRNAs and will facilitate experimental studies and further functional classification of these genes. Overall design: We extracted profiled the transcriptome expression polyadenylated mRNA-Seq.  We then used these to reconstruct the transcriptome using de-novo assemblers and identify long non coding RNAs and their expression.","project":"SRP007494"}
{"number_samples":9,"species":"human","abstract":"Post-transcriptional gene regulation relies on hundreds of RNA binding proteins (RBPs) but the function of most RBPs is unknown. The human RBP HuR/ELAVL1 is a conserved mRNA stability regulator. We used PAR-CLIP, a method based on RNA-protein crosslinking, to identify transcriptome wide ~26,000 HuR binding sites. These sites were on average highly conserved, enriched for HuR binding motifs and mainly located in 3' untranslated regions. Surprisingly, many sites were intronic, implicating HuR in splicing. Upon HuR knock down, mRNA levels and protein synthesis of thousands of target genes was down regulated, validating functionality. HuR and miRNA binding sites tended to reside nearby but generally did not overlap. Additionally, HuR knock down triggered strong and specific up regulation of miR-7. In summary, we identified thousands of direct and functional HuR targets, found a human miRNA controlled by HuR, and propose a role for HuR in splicing. Overall design: PolyA mRNA was extracted from anti HuR siRNA treated and mock transfected HeLa cells to identify changes in mRNA expression and splicing. 2x100 paired end sequencing was performed according to the protocol on the Illumina HiSeq. PARCLIP was performed as in Hafner et. Al 2010 but with an antibody against endogenous HuR (3A2, Santa Cruz, sc-5261) in unstressed HeLa cells. We used, independently, 4-thiouridine (4SU) and 6-thioguanosine (6SG) to assess a possible nucleotide bias. As our proteomics measurements required labeling of cells in a special medium we also performed PAR-CLIP on cells grown in SILAC medium. Altogether we performed three PAR-CLIP experiments: 4SU labeling in standard DMEM medium, 4SU labeling in SILAC medium (as a replicate) and 6SG labeling in SILAC medium. Small RNA was extracted from anti HuR siRNA treated and mock transfected HeLa cells to identify changes in mRNA expression. Sequencing was performed on Illumina GAII using the standard sRNA 36cycle protocol.","project":"SRP007498"}
{"number_samples":4,"species":"human","abstract":"Transcriptional regulation of developmentally controlled genes is at the heart of differentiation and organogenesis. In this study, we have performed mRNA transcript abdundance analyses in human cells in response to serum activation signal by RNA-seq. Overall design: mRNA transcript abundance determined before and after serum activation signals using two biological replicates.","project":"SRP007506"}
{"number_samples":5,"species":"human","abstract":"The human mitochondrial genome comprises a distinct genetic system transcribed as precursor polycistronic transcripts that are subsequently cleaved to generate individual mRNAs, tRNAs and rRNAs. Here we provide a comprehensive analysis of the human mitochondrial transcriptome across multiple cell lines and tissues. Using directional deep sequencing and parallel analysis of RNA ends, we demonstrate wide variation in mitochondrial transcript abundance, and precisely resolve transcript processing and maturation events. We identify novel transcripts, including small RNAs, and observe the enrichment of several nuclear RNAs in mitochondria. Using high-throughput in vivo DNaseI footprinting, we establish the global profile of DNA-binding protein occupancy across the mitochondrial genome at single nucleotide resolution, revealing novel regulatory features at mitochondrial transcription initiation sites and functional insights into disease-associated variants. Together, this integrated analysis of the mitochondrial transcriptome reveals unexpected complexity in the regulation, expression, and processing of mitochondrial RNA, and it  provides a resource for the future study of mitochondrial function (accessed at mitochondria.matticklab.com). Overall design: Examination of the mitochondiral transcriptome by long and small RNA sequencing, PARE sequencing and DNAseI hypersensitivity mapping.","project":"SRP007508"}
{"number_samples":181,"species":"human","abstract":"BCL6 is crucial for B-cell activation and lymphomagenesis. We used integrative genomics to explore BCL6 mechanism in normal and malignant B-cells. Surprisingly, BCL6 assembled distinct complexes at enhancers vs. promoters. At enhancers BCL6 preferentially recruited SMRT, which mediated H3K27 deacetylation through HDAC3, antagonized p300 activity and repressed transcription, but without decommissioning enhancers. This provides a biochemical basis for toggling enhancers from the active to poised state. Virtually all SMRT was bound with BCL6 suggesting that in B-cells BCL6 uniquely sequesters SMRT from other factors. In promoters BCL6 preferentially recruited BCOR, but most potently repressed promoters where it formed a distinctive ternary complex with SMRT and BCOR. Promoter repression was associated with decreased H3K36me3, H3K79me2 and Pol II elongation, linking BCL6 to transcriptional pausing. Overall design: We identified the binding patterns of BCL6, SMRT, NCOR and BCOR corepressors in normal germinal center B cells and a DLBCL cell line (OCI-Ly1) using ChIP-seq. Additionally we treated lymphoma cells with siRNA against BCL6 and a non-targeted siRNA (NT control) and performed RNA-seq to identify the genes bound and repressed by BCL6. RNA-seq experiments were performed at 24h and 48h after siRNA treatments. Additional biological triplicate RNA-seq experiments were performed at 48h after BCL6 knockdown. Furthermore, a series of histone mark ChIP-seq and RNA polymerase ChIP-seq (total, Ser5-P and Ser2-P) were preformed to capture the chromatin states associated with the formation of BCL6 corepressor complexes.","project":"SRP007525"}
{"number_samples":4,"species":"human","abstract":"Transcriptome for human liver cancer","project":"SRP007560"}
{"number_samples":3,"species":"human","abstract":"Monocytes are a heterogeneous cell population with subset-specific functions and phenotypes. The differential expression of CD14 and CD16 distinguishes classical CD14++CD16-, intermediate CD14++CD16+ and non-classical CD14+CD16++ monocytes. However, CD14++CD16+ monocytes remain the most poorly characterized subset so far. Therefore we analyzed the transcriptomes of the three monocyte subsets using SuperSAGE in combination with high-throughput sequencing. Analysis of 5,487,603 tags revealed unique identifiers of CD14++CD16+ monocytes, delineating these cells from the two other monocyte subsets. CD14++CD16+ monocytes were linked to antigen processing and presentation (e.g. CD74, HLA-DR, IFI30, CTSB), to inflammation and monocyte activation (e.g. TGFB1, AIF1, PTPN6), and to angiogenesis (e.g. TIE2, CD105). Therefore we provide genetic evidence for a distinct role of CD14++CD16+ monocytes in human immunity. Overall design: Human monocyte subsets (CD14++CD16-, CD14++CD16+, CD14+CD16++) were isolated from 12 healthy volunteers based on MACS technology. Total RNA from monocyte subsets was isolated and same aliquots from each donor and monocyte subset were matched for SuperSAGE. Three SuperSAGE libraries (CD14++CD16-, CD14++CD16+ and CD14+CD16++) were generated.","project":"SRP007569"}
{"number_samples":24,"species":"human","abstract":"A strain of Fusobacterium nucleatum, which we named CC53, was cultured from a colon cancer tumor surgical section and its genome sequenced.","project":"SRP007584"}
{"number_samples":9,"species":"human","abstract":"We applied deep-sequencing based technique, 3'-Seq, to obtain comprehansive maps of poly-A sites in human cells. 3'-Seq was applied to two cell lines (U2OS and RPE-1), in control and PABPN1 knockdown cells Overall design: Examination of poly-A sites in control and PABPN1kd cells (in two different cell lines)","project":"SRP007596"}
{"number_samples":12,"species":"human","abstract":"RNA editing is a post-transcriptional event that recodes hereditary information. Here we describe a comprehensive profile of the RNA editome of a male Han Chinese individual based on analysis of ~767 million sequencing reads from poly(A)+, poly(A)- and small RNA samples. We developed a computational pipeline that carefully controls for false positives while calling RNA editing events from genome and whole-transcriptome data of the same individual. We identified 22,688 RNA editing events in noncoding genes and introns, untranslated regions and coding sequences of protein-coding genes. Most changes (~93%) converted A to I(G), consistent with known editing mechanisms based on adenosine deaminase acting on RNA (ADAR). We also found evidence of other types of nucleotide changes; however, these were validated at lower rates. We found 44 editing sites in microRNAs (miRNAs), suggesting a potential link between RNA editing and miRNA-mediated regulation. Our approach facilitates large-scale studies to profile and compare editomes across a wide range of samples.","project":"SRP007605"}
{"number_samples":4,"species":"human","abstract":"microRNAs (miRNAs) are a large class of small non-coding RNAs which post-transcriptionally regulate the expression of a large fraction of all animal genes and are important in a wide range of biological processes. Recent advances in high-throughput sequencing allow miRNA detection at unprecedented sensitivity, but the computational task of accurately identifying the miRNAs in the background of sequenced RNAs remains challenging. For this purpose we have designed miRDeep2, a substantially improved algorithm which identifies canonical and non-canonical miRNAs such as those derived from transposable elements and informs on high-confidence candidates that are detected in multiple independent samples. Analyzing data from seven animal species representing the major animal clades, miRDeep2 identified miRNAs with an accuracy of 98.6-99.9% and reported hundreds of novel miRNAs. To test the accuracy of miRDeep2, we knocked down the miRNA biogenesis pathway in a human cell line and sequenced small RNAs before and after. The vast majority of the >100 novel miRNAs expressed in this cell line were indeed specifically down-regulated, validating most miRDeep2 predictions.  Last, a new miRNA expression profiling routine, low time and memory usage and user-friendly interactive graphic output can make miRDeep2 useful to a wide range of researchers.\" Overall design: high-throughput sequencing was used to profile small RNA expression in a human MCF-7 cell line before and after Dicer knock-down","project":"SRP007641"}
{"number_samples":1,"species":"human","abstract":"Our previous study revealed that nucleosomal H3.3-H4 tetramers split at a ratio of about 8%-10% during each cell cycle. However, the genomic distribution pattern of H3.3 splitting events is unknown. Here we used an unbiased approach based on sequential ChIP and high throughput sequencing to investigate the splitting distribution in a dual inducible HeLa cell line.","project":"SRP007665"}
{"number_samples":5,"species":"human","abstract":"A-to-I editing modifies RNA transcripts from their genomic blueprint. A prerequisite for this process is a double-stranded RNA (dsRNA) structure. Such dsRNAs are formed as part of the micro-RNA (miRNA) maturation process, and it is therefore expected that miRNAs are affected by A-to-I editing. Editing of miRNAs has the potential to add another layer of complexity to gene regulation pathways, especially if editing occurs within the miRNA recognition site. Thus, it is of interest to study the extent of this phenomenon. Current reports in the literature disagree on its extent; while some reports claim that it may be widespread others deem the reported events as rare. Utilizing a next-generation sequencing approach supplemented by an extensive bioinformatic analysis, we were able to systematically identify A-to-I editing events in mature miRNAs derived from human brain tissue. Our algorithm successfully identified many of the known editing sites in mature miRNAs, and revealed additional novel sites, six of which are in the recognition sites of the miRNAs. The editing efficiency of most sites identified is very low. Thus, we find that A-to-I editing does alter several miRNAs but it is not widespread.","project":"SRP007819"}
{"number_samples":67,"species":"human","abstract":"We report the application of Illumina small RNA sequencing to normal human skin, as well as uninvolved and involved psoriatic skin. By obtaining over 600 million qualified reads from 20 healthy controls and 47 psoriasis biopsies (uninvolved/involved), we have generated a complete small RNA profile in normal and diseased human skin, with particular emphasis on miRNAs. We report the discovery of 284 putative novel miRNAs as well as 98 differentially expressed miRNAs in psoriatic skin. Overall design: miRNA discovery and expression profiling in 67 normal and psoriatic human skin biopsies","project":"SRP007825"}
{"number_samples":2,"species":"human","abstract":"We assessed whether human transcripts expressed in an aneuploid mouse that carries human chromosome 21 (HsChr21) are spliced in a human-specific or mouse-specific fashion. In almost all cases, human-specific alternative splicing is maintained in the mouse nucleus. Species-specific splicing therefore appears to be primarily directed by cis-acting elements, rather than changes in the levels or activities of trans-acting factors. Overall design: Approximately 485 million Illumina 50-nt sequence reads were generated for brain and liver tissues from normal human, Tc0 (wildtype) mouse and Tc1 mouse strains.  Sequence reads were mapped to splice junctions and %in levels were estimated.","project":"SRP007862"}
{"number_samples":8,"species":"human","abstract":"Prostate cancer is the second leading cancer that causes the death of American men. In this study, we  perform an integrative analysis of the whole-transcriptome of metastatic prostate cancer clinical samples. Specifically, we prepare our samples using an innovative protocol NuGEN, which captures the profile of the total RNA, rather than only the mRNA as the popular polyA selection protocol does. Unexpectedly high intronic expressions are observed including the gene AR (Androgen Receptor) and KLK3/PSA, a marker of prostate cancer. Further, certain amount of reads can be found to be across the intron-exon boundaries, suggesting that they come from unspliced pre-mRNAs.  Interestingly, we also observe high splicing on a non-coding RNA MALAT-1. We believe that the aberrant splicing patterns are associated with prostate cancer development. Overall design: Analysis of 8 clinical samples of metastatic prostate cancer patients","project":"SRP007881"}
{"number_samples":6,"species":"human","abstract":"The goal of this study was to investigate the role of intragenic CTCF in alternative pre-mRNA splicing through a combined CTCF-ChIP-seq and RNA-seq approach. CTCF depletion led to decreased inclusion of weak upstream exons. Overall design: CTCF ChIP-seq was performed in BJAB and BL41 B cell lines and normalized relative to Rabbit Ig control IP-seq reads. RNA-seq was performed in BJAB and BL41 cells transduced with shRNA against CTCF or RFP as a control, and in untransduced cells as well.","project":"SRP007885"}
{"number_samples":34,"species":"human","abstract":"Genome-wide gene expression profile using deep sequencing technologies can drive the discovery of cancer biomarkers and therapeutic targets. Such efforts are often limited to profiling the expression signature of either mRNA or microRNA (miRNA) in a single type of cancer.Here we provided an integrated analysis of the genome-wide mRNA and miRNA expression profiles of three different genitourinary cancers: carcinomas of the bladder, kidney and testis.Our results highlight the general or cancer-specific roles of several genes and miRNAs that may serve as candidate oncogenes or suppressors of tumor development. Further comparative analyses at the systems level revealed that significant aberrations of the cell adhesion process, p53 signaling, calcium signaling, the ECM-receptor and cell cycle pathways, the DNA repair and replication processes and the immune and inflammatory response processes were the common hallmarks of human cancers. Gene sets showing testicular cancer-specific deregulation patterns were mainly implicated in processes related to male reproductive function, and general disruptions of multiple metabolic pathways and processes related to cell migration were the characteristic molecular events for renal and bladder cancer, respectively. Furthermore, we also demonstrated that tumors with the same histological origins and genes with similar functions tended to group together in a clustering analysis. By assessing the correlation between the expression of each miRNA and its targets, we determined that deregulation of 'key' miRNAs may result in the global aberration of one or more pathways or processes as a whole.This systematic analysis deciphered the molecular phenotypes of three genitourinary cancers and investigated their variations at the miRNA level simultaneously. Our results provided a valuable source for future studies and highlighted some promising genes, miRNAs, pathways and processes that may be useful for diagnostic or therapeutic applications. Overall design: Examination of microRNA expression of cancer and matched adjacent tissues from 7 testicular germ cell tumors and 10 transitional cell carcinomas of bladder","project":"SRP007946"}
{"number_samples":34,"species":"human","abstract":"Genome-wide gene expression profile using deep sequencing technologies can drive the discovery of cancer biomarkers and therapeutic targets. Such efforts are often limited to profiling the expression signature of either mRNA or microRNA (miRNA) in a single type of cancer.Here we provided an integrated analysis of the genome-wide mRNA and miRNA expression profiles of three different genitourinary cancers: carcinomas of the bladder, kidney and testis.Our results highlight the general or cancer-specific roles of several genes and miRNAs that may serve as candidate oncogenes or suppressors of tumor development. Further comparative analyses at the systems level revealed that significant aberrations of the cell adhesion process, p53 signaling, calcium signaling, the ECM-receptor and cell cycle pathways, the DNA repair and replication processes and the immune and inflammatory response processes were the common hallmarks of human cancers. Gene sets showing testicular cancer-specific deregulation patterns were mainly implicated in processes related to male reproductive function, and general disruptions of multiple metabolic pathways and processes related to cell migration were the characteristic molecular events for renal and bladder cancer, respectively. Furthermore, we also demonstrated that tumors with the same histological origins and genes with similar functions tended to group together in a clustering analysis. By assessing the correlation between the expression of each miRNA and its targets, we determined that deregulation of 'key' miRNAs may result in the global aberration of one or more pathways or processes as a whole.This systematic analysis deciphered the molecular phenotypes of three genitourinary cancers and investigated their variations at the miRNA level simultaneously. Our results provided a valuable source for future studies and highlighted some promising genes, miRNAs, pathways and processes that may be useful for diagnostic or therapeutic applications. Overall design: Examination of mRNA expression of cancer and matched adjacent tissues of 7 testicular germ cell tumors and 10 transitional cell carcinomas of bladder","project":"SRP007947"}
{"number_samples":1,"species":"human","abstract":"The ubiquitously expressed RNA-binding protein Hu Antigen R (HuR) or ELAVL1 is implicated in a variety of biological processes as well as being linked with a number of diseases, including cancer. Despite a great deal of prior investigation into HuR, there is still much to learn about its function. We take an important step in this direction by conducting iCLIP (CrossLinking and ImmunoPreciptation) and RNA Sequencing experiments followed by an extensive computational analysis to determine the characteristics of the HuR binding site and impact on the transcriptome. We reveal that HuR targets predominantly uracil-rich single-stranded stretches of varying size, with a strong conservation of structure and sequence composition. Despite the fact that HuR sites are observed in intronic regions, our data does not support a role for HuR in regulating splicing. HuR sites in 3'UTRs overlap extensively with predicted miRNA target sites suggesting interplay between the functions of HuR and miRNAs. Network analysis showed that identified targets containing HuR binding sites in the 3' UTR are highly interconnected.","project":"SRP008069"}
{"number_samples":18,"species":"human","abstract":"We apply a massively parallel reporter assay (MPRA) that relies on mRNA and plasmid tag sequencing (Tag-Seq) to compare the regulatory activities of more than 27,000 distinct variants of two inducible enhancers in human cells: a synthetic cAMP-regulated enhancer and the virus-inducible interferon beta enhancer. The resulting data define accurate maps of functional transcription factor binding sites in both enhancers at single-nucleotide resolution and can be used the to train quantitative sequence-activity models (QSAMs). Overall design: Reporter Tag-Seq from HEK293 cells transfected with each of six MPRA plasmid pools, with and without stimulation (forskolin or Sendai virus). The reporter mRNAs contain unique 10 nucleotide tags that facilitates quantitation of their abundances. The same tags were also sequenced from each ransfected plasmid pool to facilitate normalization to plasmid copy numbers. The reporter constructs were designed according to two different mutagenesis strategies: 'single-hit scanning' and 'multi-hit sampling'. The specific variants are included in the processed data files.","project":"SRP008145"}
{"number_samples":2,"species":"human","abstract":"Primary effusion lymphoma (PEL) is caused by Kaposi's sarcoma-associated herpesvirus (KSHV) and frequently also harbors Epstein-Barr virus (EBV). The expression of KSHV- and, often, EBV-encoded microRNAs (miRNAs) in PELs suggests a role for these miRNAs in viral latency and lymphomagenesis. Here we report the direct and transcriptome-wide identification of miRNA target sites for all miRNAs expressed in PEL cell lines. The resulting dataset revealed that KSHV miRNAs directly target more than 2000 cellular mRNAs encoding proteins that function in pathways with relevance to KSHV pathogenesis. Moreover, ~50% of these mRNAs are also targeted by EBV miRNAs, via distinct binding sites. In addition to a known viral analog of miR-155, we show that KSHV encodes a viral miRNA that mimics cellular miR-142-3p function. In summary, these experiments identify an extensive list of mRNAs targeted by KSHV miRNAs and indicate that these are likely to strongly influence viral replication and pathogenesis. Overall design: small RNA sequencing, 3 samples Ago2 (EIF2C2) PAR-CLIP, 2 samples","project":"SRP008216"}
{"number_samples":20,"species":"human","abstract":"Gene expression, DNA and histone methylation profiles were performed on multiple cell types purified from normal human nulliparous and parous breast tissue using SAGE-seq, MSDK-seq and ChIP-seq [GSE26141]. Overall design: SAGE-seq, MSDK-seq and ChIP-seq libraries were made from individual cell types and integrated to find a comprehensive view of parity associated changes.","project":"SRP008218"}
{"number_samples":8,"species":"human","abstract":"Alternative splicing (AS) is a key process underlying the expansion of proteomic diversity and the regulation of gene expression. However, the contribution of AS to the control of embryonic stem cell (ESC) pluripotency is not well understood. Here, we identify an evolutionarily conserved ESC-specific AS event that changes the DNA binding preference of the forkhead family transcription factor FOXP1. We show that the ESC-specific isoform of FOXP1 stimulates the expression of transcription factor genes required for pluripotency including OCT4, NANOG, NR5A2 and GDF3, while concomitantly repressing genes required for ESC differentiation. Remarkably, this isoform also promotes the maintenance of ESC pluripotency and the efficient reprogramming of somatic cells to induced pluripotent stem cells. These results thus reveal that an AS switch plays a pivotal role in the regulation of pluripotency and functions by controlling critical ESC-specific transcriptional programs. Overall design: Exons 18 and 18b form a mutually exclusive splicing event. The FOXP1 (non-ES) isoform contains only exon 18 and not 18b, while the FOXP1-ES isoform contains only exon 18b and not 18. To investigate whether FOXP1 and FOXP1-ES control different sets of genes, we performed knockdowns using custom siRNA pools targeting FOXP1 exons 18 or 18b in undifferentiated H9 cells, followed by RNA-Seq profiling.","project":"SRP008225"}
{"number_samples":2,"species":"human","abstract":"We report here that human mitochondria contain small RNA including microRNA, piRNA, tRNA, rRNA, and RNA repeats. Mitochondria from human cells were purified and RNA isolated. Small RNAs were purified, library generated and analyzed by Illumina Hiseq 2000 system. The sequencing generated 19.5 and 17.7 million reads from HEK-293 and HeLa respectively. 91% and 97% sequences of HEK293 and HeLa respectively were annotated to various classes of small RNA. The total percentage of 4.21 and 2.58 sequences from HEK293 and HeLa respectively was found to be of miRNA. Further, we found only 1.2 % sequences from both the libraries aligned to mitochondrial genome. These results suggest that there is efficient transport of nuclear encoded small RNA to mitochondria. The small RNA in mitochondria may regulate critical cellular processes. Overall design: Analyzing the smallRNA in human mitochondria from two human cell lines (HEK-293 and HeLa).","project":"SRP008258"}
{"number_samples":9,"species":"human","abstract":"We have analyzed publicly available K562 Hi-C data, which enables genome-wide unbiased capturing of chromatin interactions, using a Mixture Poisson Regression Model to define a highly specific set of interacting genomic regions. We integrated multiple ENCODE Consortium resources with the Hi-C data, using DNase-seq data and ChIP-seq data for 46 transcription factors and 8 histone modifications. We classified 12 different sets (clusters) of interacting loci that can be distinguished by their chromatin modifications and which can be categorized into three types of chromatin hubs. The different clusters of loci display very different relationships with transcription factor binding sites. As expected, many of the transcription factors show binding patterns specific to clusters composed of interacting loci that encompass promoters or enhancers. However, cluster 6, which is distinguished by marks of open chromatin but not by marks of active enhancers or promoters, was not bound by most transcription factors but was highly enriched for 3 transcription factors (GATA1, GATA2, and c-Jun) and 3 chromatin modifiers (BRG1, INI1, and SIRT6). To validate the identification of the clusters and to dissect the impact of chromatin organization on gene regulation, we performed RNA-seq analyses before and after knockdown of GATA1 or GATA2. We found that knockdown of the GATA factors greatly alters the expression of genes within cluster 6. Our work, in combination with previous studies linking regulation by GATA factors with c-Jun and BRG1, provide genome-wide evidence that Hi-C data identifies sets of biologically relevant interacting loci. Overall design: RNA-seq of control, siGATA1 and siGATA2 K562 cells","project":"SRP008280"}
{"number_samples":15,"species":"human","abstract":"We have developed two methods for efficiently consructing RNA-seq libraries using transposition.  Each method constructs high quality RNA-seq libraries when compared to standard approaches. One of the methods (Directional Tn-RNA-seq) maintains strand-of-origin information and exhibits strand specificity comparable to current approaches. Overall design: RNA-seq libraries were constructed from ECC-1, a human cell line, and Universal Human Reference RNA using transposon-based and standard RNA-seq library construciton methods.","project":"SRP008331"}
{"number_samples":4,"species":"human","abstract":"Breast milk is a complex liquid that enriched in immunological components and affect the development of the infant immune system. Exosomes, the membranous vesicles of endocytic origin, are ubiquitously in various body fluids which can mediate intercellular communication. MicroRNAs (miRNAs), a well-defined group of non-coding small RNAs, in human breast milk are packaged inside exosomes. Here, we present the identification of miRNAs in human breast milk exosomes using deep sequencing technology. We found that the immune-related miRNAs are enriched in breast milk exosomes, and are resistant to the general harsh conditions. Overall design: Four small RNA libraries in human breast milk exosomes from four healthy women (30 +/- 0.9 years old, primiparity) when the infant were aged at 60 days were sequenced.","project":"SRP008339"}
{"number_samples":1,"species":"human","abstract":"NGS of IGIB-NIV 01 Hologenome","project":"SRP008479"}
{"number_samples":5,"species":"human","abstract":"The regulation of endothelial functions is essential for maintaining circulatory homeostasis and the physiological function of different organs. Thrombin can stimulate endothelial cells and regulate the expression, release and activation of a number of biological mediators. However, transcriptional regulation of vascular endothelial cells by thrombin is not completely understood. In the present study, Illumina RNA-seq was used to profile the transcriptome in human pulmonary microvascular endothelial cells (HMVEC-L) treated with thrombin to gain insight into thrombin's direct effects on the endothelial function. Out of 100 million total reads from a paired end sequencing assay, 91-94% of the reads were aligned to over 16,000 genes in the reference human genome. Thrombin upregulated 150 known genes and 480 known isoforms, and downregulated 2,190 known genes and 3,574 known isoforms by at least 2 fold. Of note, thrombin upregulated 1,775 unknown isoforms and downregulated 12,202 isoforms by at least 2 fold. Many genes displayed isoform specific differential expression levels and different usage of transcriptional start sites after the thrombin treatment. Since the dysregulation of vascular endothelial cells by different agonists including thrombin has been implicated in the development of a number of pathologic disorders, such as inflammatory conditions, cancer, diabetes, coronary heart disease, further in-depth follow-up analysis of the transcriptional regulation reported in this study may shed light on molecular pathogenic mechanisms underlying thrombin mediated endothelial dysfunction in those processes and provide new leads of potential relevant therapeutic targets.","project":"SRP008482"}
{"number_samples":12,"species":"human","abstract":"Twelve clinical samples from human esophageal squamous cell carcinoma (ESCC) (seven tumors and five non-tumors) were sequenced using Illumina high-throughput sequencing and the RNA-Seq profiling was investigated with 1730 genes significantly differentially expressed. The gene Rab25 was found to be down-regulated in tumors (p-value < 1E-20) and identified as a novel candidate tumor suppressor gene. The down-regulation of Rab25 was examined in a large cohort of ESCC and non-tumor cases by qPCR and immunohistochemistry analyses. Aberrant methylation in the promoter region of Rab25 was studied by demethylation treatment with 5-aza-dC and bisulfite genomic sequencing (BGS). In order to assess the effect of Rab25 on tumor growth and angiogenesis, in vitro and in vivo functional studies in ESCC cell lines using lentiviral-based overexpression or knockdown models were also performed. Overall design: 7 tumor and 5 normal samples","project":"SRP008496"}
{"number_samples":6,"species":"human","abstract":"In this study, we employed massively parallel sequencing technology to identify miRNAs expressed in B-cells from Ashkenazi Jewish centenarians, i.e., those living to a hundred and a human model of exceptional longevity, and younger controls without a family history of longevity. With data from 26.7 million reads comprising 9.4x108 bp from 3 centenarian and 3 control individuals, we discovered a total of 276 known miRNAs and 8 unknown miRNAs ranging several orders of magnitude in expression levels. A total of 22 miRNAs were found to be significantly upregulated, with only 2 miRNAs downregulated, in centenarians as compared to controls. Overall design: Examination of miRNA profile of two different ages","project":"SRP008552"}
{"number_samples":2,"species":"human","abstract":"Adenovirus small e1a causes ~70% reduction in cellular levels of histone H3 lysine 18 acetylation (H3K18ac). It is unclear, however, where this dramatic reduction occurs genome-wide. ChIP-seq revealed that e1a erases 95% of H3K18ac peaks in normal fibroblasts and replaces them with one-third as many at new genomic locations. H3K18ac at promoters and intergenic regions of genes with fibroblast-related functions are relocalized after infection to promoters of highly-induced genes that regulate cell cycling and to new putative enhancers. Strikingly, a significant fraction of the post-infection H3K18ac peaks occurs precisely at regions bound by RB1 in uninfected cells, but not by p107 or p130 without RB1. In contrast, over half of H3K9ac peaks are similarly distributed before and after infection, independently of RB1. The strategic redistribution of H3K18ac by e1a highlights the importance of this modification for transcriptional activation and cellular transformation. Overall design: Examination of two histone acetylations and RB family members binding. mRNA-Seq RPKM file linked as supplementary file on Series record.","project":"SRP008554"}
{"number_samples":2,"species":"human","abstract":"As a preliminary step towards applying next generation sequencing using neurons derived from patient-specific iPSCs, we have carried out an RNA-Seq analysis on control human neurons. Dramatic changes in the expression of coding genes, long non-coding RNAs (lncRNAs), pseudogenes, and splice isoforms were seen during the transition from pluripotent stem cells to early differentiating neurons. A number of genes that undergo radical changes in expression during this transition include candidates for schizophrenia (SZ), bipolar disorder (BD) and autism spectrum disorders (ASD) that function as transcription factors and chromatin modifiers, such as POU3F2 and ZNF804A, and genes coding for cell adhesion proteins implicated in these conditions including NRXN1 and NLGN1. In addition, a number of novel lncRNAs were found to undergo dramatic changes in expression, one of which is HOTAIRM1, a regulator of several HOXA genes during myelopoiesis. The increase we observed in differentiating neurons suggests a role in neurogenesis as well. Finally, several lncRNAs that map near SNPs associated with SZ in genome wide association studies also increase during neuronal differentiation, suggesting that these novel transcripts may be abnormally regulated in a subgroup of patients. Overall design: Total RNAs were isolated from both iPS cells and differentiating neurons, and the sequenced material was pre-amplified using the NuGen Ovation amplification system.  RNA-Seq libraries were prepared and carried out on Illumina HiSeq200.","project":"SRP008681"}
{"number_samples":4,"species":"human","abstract":"RNA Sequencing of 12 primates and 4 mammals.","project":"SRP008743"}
{"number_samples":6,"species":"human","abstract":"RNA sequencing was performed on ten breast cancer transcriptomes (five BRCA1-mutated breast cancer cell lines, three BRCA1-mutated primary tumours, two secretory breast cancer primary tumours and one non-tumorigenic breast epithelial cell line) using the Illumina Genome Analyzer. We sought to identify putative gene fusions using a bioinformatics approach. As a proof of concept, we were able to identify two previously characterized gene fusions in our samples using both single-end and paired-end approaches. In addition, we identified three novel in-frame fusions, but none were recurrent. Two of the candidates, WWC1-ADRBK2 in HCC3153 cell line and ADNP-C20orf132 in a primary tumour, were confirmed by Sanger sequencing and RT-PCR. RNA-Seq expression profiling of these two fusions showed a distinct overexpression of the 3’ partner genes, suggesting that its expression may be under the control of the 5’ partner gene’s regulatory elements.","project":"SRP008746"}
{"number_samples":8,"species":"human","abstract":"Clinicians need additional metrics for predicting quality of human oocytes for IVF procedures. Human polar bodies reflect the oocyte transcript profile. Quantitation of polar body mRNAs could allow for both oocyte ranking and embryo preferences in IVF applications. The transcriptome of a polar body has never been reported, in any organism. Overall design: Eight total samples. There are 2 biological replicates of the following four conditions: pooled oocytes and their sister polar bodies and a single oocyte and its sister polar body.","project":"SRP008775"}
{"number_samples":4,"species":"human","abstract":"The goal of this study was to investigate the role of hnRNP L-like in alternative pre-mRNA splicing in human B-cells through an RNA-Seq approach. Overall design: RNA-Seq was performed in DG75 cell line with over expression of hnRNP L-like or GFP as control.","project":"SRP008930"}
{"number_samples":25,"species":"human","abstract":"We have determined the whole genome sequence of an individual at high accuracy and performed an integrated analysis of omics profiles over a 1.5 year period that included healthy and two virally infected states. Omics profiling of transcriptomes, proteomes, cytokines, metabolomes and autoantibodyomes from blood components have revealed extensive, dynamic and broad changes in diverse molecular components and biological pathways that occurred during healthy and disease states. Many changes were associated with allele- and edit-specific expression at the RNA and protein levels, which may contribute to personalized responses. Importantly, genomic information was also used to predict medical risks, including Type II Diabetes (T2D), whose onset was observed during the course of our study using standard clinical tests and molecular profiles, and whose disease progression was monitored and subsequently partially managed. Our study demonstrates that longitudinal personal omics profiling can relate genomic information to global functional omics activity for physiological and medical interpretation of healthy and disease states. Overall design: Examination of blood component in 20 different time points over 1.5 years which includes 2 disease state and 18 healty state Related exome studies at: SRX083314 SRX083313 SRX083312 SRX083311","project":"SRP008976"}
{"number_samples":4,"species":"human","abstract":"Most cancer genomics papers to date have focused on aberrations in genomic DNA and protein-coding transcripts. However, around 50% of transcripts have no coding potential and may exist as non-coding RNA. We performed RNA-seq in BRAFv600e melanoma skin cancer and on melanocytes over-expressing oncogenic BRAF to catalog transcriptome remodeling. We discovered that BRAF regulates expression of 1027 protein coding transcripts, 39 annotated lncRNAs and 70 novel transcripts. Many of the novel transcripts are lncRNAs. We used an indepenedent dataset to interrogate our novel transcripts and found that the novel lncRNA BANCR is a BRAF-regulated lncRNA recurrently upregulated in melanoma. Knockdown of BANCR impairs melanoma cell migration. Overall design: Whole transcriptome RNA-seq followed assembly to Refseq/Gencode and de novo transcript assembly. RNA-seq performed on: 2 BRAFv600e melanomas, melanocytes+RFP control and melanocytes + BRAFv600e. We discovered protein-coding transcripts, annotated ncRNAs and novel transcripts (including lncRNAs) regulated by BRAFv600e also coordinately expressed in melanoma.","project":"SRP009029"}
{"number_samples":12,"species":"human","abstract":"We set out to investigate genomic mechanisms by which alternative polyadenylation impacts gene expression and its variation across genetically distinct human individuals. We used 3’-end RNA-seq to maximize genome coverage and precision of transcript end positions, and to measure quantitative expression levels of transcript forms. The results shed light on the architecture of transcript ends and regulatory elements in human 3’ UTRs and the principles of genetic variation in 3’ length form usage. Overall design: Survey of transcript 3' end positions in B-lymphocyte cell lines derived from six human individuals (two biological replicates each)","project":"SRP009067"}
{"number_samples":2,"species":"human","abstract":"Identification of the all RNA species associated with DNMT1. Using a comparative genome-scale approach we identified and correlated the RNA species physically associated with DNMT1 and proximal to the annotated genes to the methylation status of the corresponding loci and expression levels of the respective genes. This comparative approach delineated the first -DNMT1 centered- 'epitranscriptome' map, a comprehensive map cross-referencing DNMT1-interacting transcripts to (i) DNA methylation and (ii) gene expression profile. Overall design: Relationship between DNMT1-RNA interactions, DNA methylation and gene expression","project":"SRP009094"}
{"number_samples":2,"species":"human","abstract":"Transcriptional elongation by RNA polymerase II (Pol II) is regulated by positive transcription elongation factor b (P-TEFb) in association with Bromodomain-containing protein 4 (BRD4). We used genome-wide chromatin immunoprecipitation sequencing in primary human CD4+ T cells to reveal that BRD4 co-localizes with Ser2-phosphorylated Pol II (Pol II Ser2) at both enhancers and promoters of active genes. Disruption of bromodomain:histone acetylation interactions by JQ1, a small-molecule bromodomain inhibitor, resulted in decreased BRD4 binding, reduced Pol II Ser2, and reduced expression of lineage-specific genes in primary human CD4+ T cells. A large number of JQ1-disrupted BRD4 binding regions exhibited di-acetylated H4 (lysine-5 and -8) and H3K27 acetylation (H3K27ac), which correlated with the presence of histone acetyltransferases and deacetylases. Genes associated with BRD4/H3K27ac co-occupancy exhibited significantly higher activity than those associated with H3K27ac or BRD4 binding alone. Comparison of BRD4 binding in T cells and in human embryonic stem cells revealed that enhancer BRD4 binding sites were predominantly lineage-specific. Our findings suggest that BRD4-driven Pol II phosphorylation at serine 2 plays an important role in regulating lineage-specific gene transcription in human CD4+ T cells. Overall design: Examination of BRD4, total Pol II, serine 2 phosphorylated Pol II and serine 5 phosphorylated Pol II binding in CD4+ T cells (with and without JQ1 treatment) and BRD4 binding in human embryonic  stems cell; PolyA RNA expression in CD4+ T cells( with and without JQ1 treatment) using RNA-seq","project":"SRP009105"}
{"number_samples":6,"species":"human","abstract":"Deep high-throughput transcriptome sequencing (RNA-seq) performed on 3 pairs of matched tumor and adjacent non-tumorours (NT) tissues from HCC patients of Chinese origin generated 183.6-million reads that could be aligned. We discovered a number of differentially expressed genes and multiple types of somatic single nucleotide variations (SNVs) in expressed genes.  After the removal of the error alignments, high-quality reads were mapped to the human reference sequence (GRCh37/hg19) using three different softwares TopHat, Burrows-Wheeler Aligner (BWA) and CLC Genomics Workbench (CLC).  The high-quality variants were identified using VarScan with the following parameters: minimum coverage depth of 10, variation frequency of more than 30% and base quality of more than 15.  A total of 568, 545 and 494 potential somatic single nucleotide variants (SNVs), including 94, 89 and 101 coding somatic SNVs (cSNVs), were identified in 3 tumor samples HCC448T, HCC473T and HCC510T, respectively. Validation analysis was carried out for 10 of the intersected cSNVs (all are non-synonymous substitutions) within selected genes of interests with the majority confirmed. Overall design: Examination of 3 paired human hepatocellular carcinoma and matched non-tumor tissues","project":"SRP009123"}
{"number_samples":2,"species":"human","abstract":"To investigate differential gene expression, we analyzed the entire transcriptomes of tumor and matched normal brain tissues obtained from a patient who had glioblastoma multiforme.  We extracted and sequenced the mRNA using Illumina GA2 platform. The raw data was analyzed using our recently developed program called RNASEQR, as well as ERANGE, MapSplice, SpliceMap, and TopHat. Overall design: Tumor and matched control brain tissues were obtained from a Han-Chinese patient.","project":"SRP009144"}
{"number_samples":4,"species":"human","abstract":"Small RNA deep sequencing analysis was conducted on primary human fibroblasts infected with human cytomegalovirus (HCMV). HCMV-encoded miRNAs accumulated to ~20% of the total smRNA population at late stages of infection, and our analysis led to improvements in viral miRNA annotations and identification of novel HCMV miRNAs. Through crosslinking and immunoprecipitation of Argonaute-bound RNAs from infected cells, followed by high-throughput sequencing (Ago CLIP-seq), we obtained direct evidence for incorporation of all HCMV miRNAs into the endogenous host silencing machinery. Additionally, significant upregulation was observed during infection for a host miRNA cluster containing miR-96, miR-182 and miR-183. We also identified novel non-miRNA forms of virus-derived smRNAs, revealing greater complexity within the smRNA population during HCMV infection. Overall design: High-throughput profiling of smRNAs,  Ago1-, and Ago2-associated miRNAs from HCMV-infected fibroblast cells. Wild-type HCMV Towne (Genbank FJ616285.1) was used for these studies.","project":"SRP009246"}
{"number_samples":17,"species":"human","abstract":"We identified human-specific gene expression patterns in the brain by comparing expression with chimpanzee and rhesus macaque. Overall design: DpnII tag-based libraries were generated from human, chimpanzee, and rhesus macaque brain (frontal pole, hippocampus, caudate nucleus), and sequenced using an Illumina Genome Analyzer.","project":"SRP009247"}
{"number_samples":10,"species":"human","abstract":"We evaluated the trancriptome of primary cutaneous leisions caused by infection with Leishmania braziliensis. mRNA-seq technique was used to study the trancriptome of both host and parasite. A total of 10 samples was obtained from primary skin ulcers of two extreme clinical forms of American tegumentary leishmaniasis: (i) individuals that after antimonial treatment cured completely (localized cutaneous leishmaniasis - LCL, n=5) and (ii) individuals that developed mucosal lesions in naso and oropharynx areas long after initial healing of the cutaneous lesion (mucosal leishmaniasis - ML, n=5).  The sequencing generated an average of 13+ 5 million reads per samples. The reads were aligned to Homo sapiens (USCS - hg19) and to Leishmania braziliensis (Wellcome Trust Sanger Institute - V2_29072008) genomes. Approximately, 15,000 human genes could be detected in the samples. Low amount of L. braziliensis reads did not allow the evaluation of parasite gene expression. LCL and ML samples showed different patterns of gene expression, indicating a more robust immune response in LCL individuals. In summary, this study demonstrated that next-generation sequencing can be used for identification of potentially important biological pathways and drug targets in the host-response to L. braziliensis infection and for characterization of a gene expression signature that could be used to predict the disease outcome. Moreover, we also showed the ability of this technique in, simultaneously, sequence host and pathogen mRNA. Overall design: Examination of 10 fragments of cutaneous lesions: 5 from localized cutaneous leishmaniasis patients and 5 from mucosal leishmaniasis patients.","project":"SRP009251"}
{"number_samples":5,"species":"human","abstract":"Using an enhanced RNA-seq pipeline to analyze Epstein-Barr virus transcriptomes, we have  investigated viral and cellular gene expression in the Akata cell line following B-cell receptor mediated reactivation. Robust induction of EBV gene expression was observed, with most viral genes induced more than 200-fold and with EBV transcripts accounting for 7% of all mapped reads within the cell. Following induction, hundreds of candidate splicing events were detected using the junction mapper TopHat, including a novel non-productive splicing event at the gp350/gp220 locus and several alternative splicing events at the LMP2 locus. A more detailed analysis of lytic LMP2 transcripts showed an overall lack of the prototypical type III latency splicing events. Analysis of nuclear versus cytoplasmic RNA-seq data showed that the lytic forms of LMP2, EBNA-2, EBNA-LP, and EBNA-3A, B, and C have higher nuclear-to-cytoplasmic accumulation ratios than most lytic genes, including classic late genes. This data raises the possibility that at least some lytic transcripts derived from these latency gene loci may have unique, non-coding nuclear functions during reactivation. Our analysis has also identified two previously unknown genes, BCLT1 and BCRT2, that map to the BamHI C-region of the EBV genome. Pathway analysis of cellular gene expression changes following BCR activation identified inflammatory response as the top predicted function, and ILK and TREM1 as the top predicted canonical pathways.","project":"SRP009262"}
{"number_samples":42,"species":"human","project":"SRP009266"}
{"number_samples":1,"species":"human","abstract":"Cross-talk between DNA methylation and histone modifications drives the establishment of composite epigenetic signatures and is traditionally studied using correlative rather than direct approaches. Here we present sequential ChIP-bisulfite-sequencing (ChIP- BS-seq) as an approach to quantitatively assess DNA methylation patterns associated with chromatin modifications or chromatin-associated factors directly. A chromatin- immunoprecipitation (ChIP)-capturing step is used to obtain a restricted representation of the genome occupied by the epigenetic feature of interest, for which a single-base resolution DNA methylation map is then generated. When applied to H3 lysine 27 tri- methylation (H3K27me3), we found that H3K27me3 and DNA methylation are compatible throughout most of the genome, except for CpG islands, where these two marks are mutually exclusive. Further ChIP-BS-seq-based analysis in Dnmt triple- knock-out (TKO) embryonic stem cells revealed that total loss of CpG methylation is associated with alteration of H3K27me3 levels throughout the genome: H3K27me3 in localized peaks is decreased while broad local enrichments (BLOCs) of H3K27me3 are formed. At an even broader scale, these BLOCs correspond to regions of high DNA methylation in wild-type ES cells, suggesting that DNA methylation prevents H3K27me3 deposition locally and at megabase scale. Our strategy provides an unique way of investigating global interdependencies between DNA methylation and other chromatin features. Overall design: ChIP (chromatin immunoprecipitation) is followed by bisulfite conversion and deep sequencing to directly assess DNA methylation levels in captured chromatin fragments (ChIP-BS-seq). We used ChIP-BS-seq to study the potential global cross-talk between H3K27me3 and DNA methylation, which are both linked to repression. First, we used capturing of methylated DNA, followed by bisulfite-deep sequencing (MethylCap-BS-seq). Genomic DNA isolated from normal and tumor colon tissues was used for MethylCap-BS-seq as well as for conventional MethylCap-seq experiments. Second, we performed ChIP-BS-seq on H3K27me3, using HCT116 colon carcinoma cells. Third, to further study the relevance of the observations, we generated genome-wide profiles for H3K27me3 and DNA methylation by conventional ChIP-seq and MethylCap-seq, and RNA-seq, respectively. Finally, we performed H3K27me3-ChIP-BS-seq and MethylCap-seq on wild-type mouse ES cells as well as Dnmt-triple-knockout (TKO) mouse ES cells.","project":"SRP009269"}
{"number_samples":10,"species":"human","abstract":"We mapped and quantified poly(A) sites in BJ and MCF10A cells under proliferative, arrested and transformed states Overall design: Two human cell lines (BJ and MCF10A) examined under proliferative, arrested and transformed states","project":"SRP009276"}
{"number_samples":30,"species":"human","abstract":"Basal cell carcinomas (BCCs) are the most common cancers in the US.  Histologic appearance distinguishes several subtypes, each of which can have a different biologic behavior.  In this study global miRNA expression was quantified by high throughput sequencing in nodular BCCs, a subtype that is slow growing, and infiltrative BCCs, aggressive tumors that extend through the dermis and invade structures such as cutaneous nerves.  Principal components analysis correctly classified seven out of eight infiltrative tumors on the basis of miRNA expression.  The remaining tumor, on pathology review, contained a mixture of nodular and infiltrative elements.  Nodular tumors did not cluster tightly, likely reflecting broader histopathologic diversity in this class, but trended toward forming a group separate from infiltrative BCCs.  qPCR assays were developed for six of the miRNAs that showed significant differences between the BCC subtypes, and five of these six were validated in a replication set of four infiltrative and three nodular tumors.  The expression level of miR-183, a miRNA that inhibits invasion and metastasis in several types of malignancies, was consistently lower in infiltrative than nodular tumors and could be one element underlying the difference in invasiveness.  These results represent the first miRNA profiling study in BCCs and demonstrate that miRNA gene expression may be involved in tumor pathogenesis and particularly in determining the aggressiveness of these malignancies. Overall design: Total RNA was extracted from eight nodular and eight infiltrative basal cell carcinomas. miRNA was then processed into libraries using the Illumina Small RNA Ver 1.5 protocol. Samples were then sequenced on the Illumina GAII system","project":"SRP009310"}
{"number_samples":41,"species":"human","abstract":"Burkitt lymphoma (BL) can often be cured by intensive chemotherapy, but the toxicity of such therapy precludes its use in the elderly and in patients with endemic BL in developing countries, necessitating new strategies.  We used high throughput RNA sequencing and RNA interference screening to discover essential regulatory pathways that cooperate with MYC, the defining oncogene of this cancer.  In 38% of sporadic BL (sBL) cases, oncogenic CCND3 mutations produced highly stable cyclin D3 isoforms that drive cell cycle progression.  In 70% of sBL cases, mutations affecting the transcription factor TCF3 or its negative regulator ID3 fostered TCF3 dependency.  TCF3 activated the pro-survival PI(3) kinase pathway in BL, in part by augmenting constitutive B cell receptor signaling.  These findings suggest opportunities to improve therapy for patients with BL.","project":"SRP009316"}
{"number_samples":6,"species":"human","abstract":"Most human pre-mRNAs are spliced into linear molecules that retain the exon order defined by the genomic sequence.  By deep sequencing of RNA from a variety of normal and malignant human cells, we found RNA transcripts from many human genes in which the exons were arranged in a non-canonical order.  Statistical estimates and biochemical assays provided strong evidence that a substantial fraction of the spliced transcripts from hundreds of genes are circular RNAs.  Our results suggest that a non-canonical mode of RNA splicing, resulting in a circular RNA isoform, is a widespread and perhaps general feature of the gene expression program in human cells. Overall design: 3 samples of non-malignant primary human leukocytes, one replicate each","project":"SRP009373"}
{"number_samples":2,"species":"human","abstract":"Examination of Pin1-regulated Myc target genes in a human breast epithelial cell line. Overall design: Two samples: control GFP-expressing MCF10A-Myc cells and Pin1-expressing MCF10A-Myc cells.","project":"SRP009374"}
{"number_samples":3,"species":"human","abstract":"To support our research of colon cancer in human genome, we conducted massively parallel pyrosequencing of mRNAs (RNA-seq) using normal,paracancerouse and cancerous human colon tissues. We obtained a total of 29.9M reads from normal,33.0M reads from paracancerous and 36.5M reads from cancerous.The RNA-seq data derived from the sample illustrated the differencially expreesion genes among normal,paracancerous and cancerous colon tissues of human. Overall design: 3 samples examined: normal tissue, paracancerous tissue, cancerous tissue.","project":"SRP009386"}
{"number_samples":62,"species":"human","abstract":"Background: MicroRNAs (miRNAs) are small noncoding RNAs about 22 nucleotides in length and regulate mRNA expression by binding to 3?UTR of mRNAs and causing translation repression or degradation of mRNAs. Recent studies have revealed that microRNAs play important regulatory roles in carcinogenesis. However, the complement of miRNAs involved in lung carcinogenesis and tumor biology is generally unknown. We used high-throughput sequencing technology to investigate miRNA expression profiles in a study of paired lung non-small cell carcinomas and remote noncancerous lung tissues. Methods: Total RNA was isolated by Trizol from patient lung tissues, including 1 non-pair of macroscopic lung adenocarcinoma tissue, and 20 pairs of macroscopic lung adenocarcinoma tissues and corresponding paired noncancerous lung tissues from the same patients, 1 non-pair of noncancerous lung tissues of squamous cell carcinoma patient, and 10 pairs of squamous cell carcinoma and corresponding paired noncancerous lung tissues from the same patients. Small RNA library of each total RNA was constructed with Illumina?s Small RNA Sample Prep Kit, followed by high-throughput sequencing. Statistical differences in microRNA expression between groups were analyzed by GenePattern software. Results: Using 2-fold threshold with FDR<0.05, there were ~90 miRNAs differentiated the lung cancers from nontumor lung tissues, both histologies.  (a) The sets of microRNAs differentiating adenocarcinoma from paired nontumor tissues are largely different from those differentiating squamous tumors. There were 40 upregulated and 23 downregulated microRNAs differentiating adenocarcinoma Tvs.NT .  For squamous cell, the numbers were 19 and 6, respectively.  (b) In common between the two tumor histologies, there were six common upregulated microRNAs (miR-21, 135b, 200a, 494, 708. 1259), and four common down-regulated microRNAs (miR-144, 184, 516a, 1251). (c) Comparing our 22 miRNAs in common with Yanaihara/Harris Cancer Cell 2006 using spotted microarrays, we found similar direction of change in 19/22.  (d) As a group, the top over-expressed miRNA in lung tumors (determined as fold change > 2, and comprising at least .01% of total miRNA) are able to predict mRNAs that are differentially expressed in those same samples (p=4.6x10^-6). Conclusions: This effort represents one of the more comprehensive surveys of the lung tumor micronome. (i) Tumor microRNA complement differs from far-adjacent lung; (ii) Histologies differ in their overall, and tumor-distinctive microRNA complement; (iii) Statistical correlations of individual microRNA:mRNA (putative) pairing from the same tissue sets are pending.  (iv) Experimental confirmation of microRNA:mRNA targeting, using our microRNA affinity pull down assay is also pending. Overall design: Examination of different microRNA expression profiles between lung cancer tissues and adjacent lung tissues. Our lung tissues were from 32 patients, including 21 adenocarcinoma patient (20 pairs of tumor and non-tumor and 1 non-pair tumor) and 11 squamous cell carcinoma patients (10 pairs of tumor and non-tumor and 1 non-pair non-tumor).","project":"SRP009408"}
{"number_samples":1,"species":"human","abstract":"The epididymis in male reproductive system plays a crucial role in regulating the development of sperm motility and fertilizing capacity. Small non-coding RNAs (sncRNAs), especially microRNAs (miRNAs), can participate in the regulation of various physiological pathways. However, their abundance and whether they are involved in the regulation of gene expression in the human epididymis are unknown. By adopting the Solexa deep sequencing approach, we systematically investigated the sncRNAs in the adult human epididymis. A total of 4,903 unique reads representing 527 known miRNA were discovered. Eighteen novel miRNA genes encoding 23 mature miRNAs were also identified, suggesting a complicated miRNA regulatory network. The expression of some of the novel miRNA was confirmed by qRT-PCR. The presence of Piwi-interacting RNAs (piRNAs) in the library also adds to the diversity of sncRNA population in the human epididymis. This research lays the foundation for the functional study of sncRNAs in huamn reproductive system. Overall design: The epididymis from a  26-year-old male who is healthy and fertile was deep-sequenced.","project":"SRP009455"}
{"number_samples":3,"species":"human","abstract":"In this study, we combined a genome-wide analysis of the Polycomb protein Bmi1 and an in vivo RNAi screening to identify critical targets whose repression in neural progenitor and Malignant Glioma cells enables normal and aberrant self-renewal. Overall design: Bmi1 ChIP-sequencing generated in 2 primary adult mouse Neural Progenitor cell lines, 1 mouse astrocytic cell line, 1 mouse Glioma initiating cell line, and 1 mouse Glioma cell line. In human cell lines, Bmi1 ChIP-seq was performed for foetal Neural Progenitor Cells, and 2 Glioblastoma Stem-like cells. ChIP-seq for AP-1 in one Glioblastoma stem-like cell line. ChIP-sequencing were generated using either Illumina GxII or HiSeq. We have further performed RNA-seq of  adult Ink4a/Arf -/-;BMI1or GFP  dox-inducible shRNA mNPC with a doxycycline inducible shRNA targeting BMI1 or GFP (as control). These cells were either treated with doxyclyine for 48h to ablate BMI1 and GFP expression or further subjected to short-term differentiation using either BMP4 (10ng/ml or 50ng/ml for 3 or 6hrs), or FBS (1 or 10% for 3 or 6hrs). Additional mouse RNA-seq were performed in two biological replica of adult mouse brains (littermate) with either wild-type FVB background or BMI1 -/-. Finally, we performed RNA-seq in human Glioblastoma stem-like cells (most likely belonging to the mesenchymal GBM subtype) treated with BMP7, Arvanil and doxycycline or  EtOH treated as ctrl.","project":"SRP009474"}
{"number_samples":10,"species":"human","abstract":"The elongation stage of transcription is a highly regulated in metazoan. We previously purified the AFF1/AFF4-containing Super Elongation Complex (SEC) as a major regulator of development and cancer pathogenesis. Here, we report the biochemical isolation of SEC-like 2 (SEC-L2) and SEC-like 3 (SEC-L3) containing AFF2 and AFF3 in association with P-TEFb, ENL, and AF9. The SEC family members demonstrate high levels of Pol II CTD kinase activity, however, only SEC is required for the proper induction of the HSP70 gene upon stress. Genome-wide mRNA-seq analyses demonstrate that SEC-L2-3 control the expression of different subsets of genes, while AFF4/SEC plays a more dominant role in rapid gene expression in cells. MYC is one of the direct targets of AFF4/SEC, and the SEC requirement to the MYC gene regulates its expression in different cancer cells bearing either acute myeloid or lymphoid leukemia. These findings suggest that AFF4/SEC could be a potential therapeutic target for the treatment of leukemia or other cancers associated with MYC overexpression. Overall design: RNA-seq in human embyonic kidney 293T cells of wild-type and after RNAi of AFF2, AFF3, AFF4.  ChIP-seq of AFF4 and Pol2 in human 293T cells.","project":"SRP009568"}
{"number_samples":12,"species":"human","abstract":"Summary: K562-shX cells are made in an effort to validate TFBS data and ChIP-seq antibodies in Myers lab (GSE32465). K562 cells are transduced with lentiviral vector having Tet-inducible shRNA targeting a transcription factor gene. Cells with stable integration of shRNA constructs are selected using puromycin in growth media. Doxycycline is added to the growth media to induce the expression of shRNA and a red fluorescent protein marker.  A successful shRNA cell line shows at least a 70% reduction in expression of the target transcription factor as measured by qPCR. For identification, we designated these cell lines as K562-shX, where X is the transcription factor targeted by shRNA and K562 denotes the parent cell line.  For example, K562-shATF3 cells are K562 derived cells selected for stable integration of shRNA targeting the transcription factor ATF3 gene and showed at least a 70% reduction in the expression of ATF3 gene when measured by qPCR. Cells growing without doxycycline (uninduced) are used as a control to measure the change in expression of target transcription factor gene after induction of shRNA using doxycycline. For detailed growth and culturing protocols for these cells please refer to http://hudsonalpha.org/myers-lab/protocols . To identify the potential downstream targets of the candidate transcription factor, analyze the mRNA expression profile of the uninduced and induced K562-shX using RNA-seq.    For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf    Overall Design: Make K562-shX cells as described in the http://hudsonalpha.org/myers-lab/protocols . Measure the mRNA expression levels in uninduced K562-shX and induced K562-shX cells in two biological replicates using RNA-seq. Identify the potential downstream targets of the candidate transcription factor.","project":"SRP009615"}
{"number_samples":4,"species":"human","abstract":"RNA editing enhances the diversity of gene products at the post-transcriptional level. Approaches for genome-wide identification of RNA editing face two main challenges: separating true editing sites from false discoveries and accurate estimation of editing levels. We developed an approach to analyze transcriptome sequencing data (RNA-Seq) for global identification of RNA editing in cells for which whole-genome sequencing data are available. We applied the method to analyze RNA-Seq data of a human glioblastoma cell line, U87MG. Around 10,000 DNA-RNA differences were identified, the majority being putative A-to-I editing sites. These predicted A-to-I events were associated with a low false discovery rate (~5%). Moreover, the estimated editing levels from RNA-Seq correlated well with those based on traditional clonal sequencing. Our results further facilitated unbiased characterization of the sequence and evolutionary features flanking predicted A-to-I editing sites and discovery of a conserved RNA structural motif that may be functionally relevant to editing. Genes with predicted A-to-I editing were significantly enriched with those known to be involved in cancer, supporting the potential importance of cancer-specific RNA editing. A similar profile of DNA-RNA differences as in U87MG was predicted for another RNA-Seq data set obtained from primary breast cancer samples. Remarkably, significant overlap exists between the putative editing sites of the two transcriptomes despite their difference in cell type, cancer type and genomic backgrounds. Our approach enabled de novo identification of the RNA editome, which sets the stage for further mechanistic studies of this important step of post-transcriptional regulation. Overall design: Examine mRNA expression in U87MG cells following ADAR1 or control siRNA knockdown","project":"SRP009659"}
{"number_samples":8,"species":"human","abstract":"Nuclear factor kappaB (NF-kB) is a transcription factor that regulates various aspects of immune response, cell death and differentiation as well as cancer. In this study we introduce the Py-Im polyamide 1 that binds preferentially to the sequences 5'-WGGWWW-3' and 5' GGGWWW-3'. The compound is capable of binding to kB sites and reducing the expression of various NF-kB driven genes including IL6 and IL8 by qRT-PCR. Chromatin immunoprecipitation experiments demonstrate a reduction of p65 occupancy within the proximal promoters of those genes. Genome-wide expression analysis by RNA-seq compares the DNA-binding polyamide with the well-characterized NF-kB inhibitor PS1145, identifying overlaps and differences in affected gene groups and showing that both affect comparable numbers of TNFa inducible genes. Inhibition of NF-kB DNA binding via direct displacement of the transcription factor is a potential alternative to the existing antagonists. Overall design: A549 cells were treated with either a Py-Im polyamide or PS1145, subsequently induced with TNFa and compared with the untreated TNFa induced and the basal state by RNAseq. The NF-kB binding motif was derived by ChIP-seq","project":"SRP009790"}
{"number_samples":20,"species":"human","abstract":"We profiled the transcriptome of matched diagnosis and relapse samples from 10 pediatric B precursor Acute Lymphoblastic Leukemia (ALL) patients using massively parallel sequencing (RNA-Seq) technology to identify novel mutations specific at disease recurrence.","project":"SRP009840"}
{"number_samples":2,"species":"human","abstract":"The nuclear matrix associated hnRNP U/SAF-A protein has been implicated in diverse  pathways from transcriptional regulation to telomere length control to X inactivation, but the precise  mechanism underlying each of these processes has remained elusive. Here, we report hnRNP U as a  regulator of SMN2 splicing from a custom RNAi screen. Genome-wide analysis by CLIP-seq reveals that  hnRNP U binds virtually to all classes of regulatory non-coding RNAs, including all snRNAs required for  splicing of both major and minor classes of introns, leading to the discovery that hnRNP U regulates U2  snRNP maturation and Cajal body morphology in the nucleus. Global analysis of hnRNP U-dependent  splicing by RNA-seq coupled with bioinformatic analysis of associated splicing signals suggests a  general rule for splice site selection through modulating the core splicing machinery. These findings  exemplify hnRNP U/SAF-A as a potent regulator of nuclear ribonucleoprotein particles in diverse gene  expression pathways. Overall design: Examination of hnRNP U regulated splicing in Hela cells with CLIP-seq (two biological replicates) and paired-end RNA-seq (control and hnRNP U knockdown)","project":"SRP009861"}
{"number_samples":72,"species":"human","project":"SRP009862"}
{"number_samples":1,"species":"human","abstract":"In humans, there are four Ago proteins (Ago1–4) and AGO1- and 2 were previously implicated in TGS induced by exogenous siRNAs and microRNAs (miRs) directed against gene promoter transcripts via promotion of changes in histone covalent modifications and DNA methylation. Not-with-standing, many mechanistic details of this process remain poorly defined in human cells, and very little is known about the identity of possible endogenous signals, which may drive this process in human cells. Given the evolutionary conserved role of siRNAs and AGO proteins in TGS and heterochromatin formation, we set out to analyse their possible involvement in senesence-associated repression of E2F target genes.   To obtain a detailed picture of AGO-immunoprecipitating miRs (RIP) in senescent cells, we used next-generation sequencing (NGS)(RIP-Seq). We also included histone H3 dimethylated on lysine 9 (H3K9me2) in this analysis to assign potential AGO2-interacting miRs to a repressive chromatin state and unfractionated, cellular RNA from senescent cells for normalisation. Overall design: Determination of AGO AGO-immunoprecipitating miRs in WI-38 senescent human fibroblast","project":"SRP009864"}
{"number_samples":40,"species":"human","abstract":"A total of 332 genes were identified which modified their expression in a dose-dependent manner upon exposure to doxycycline. The more represented cellular pathways included all mitochondrial genes, the endoplasmic reticulum stress response, integrins and extracellular matrix components, and growth factors. Overall design: Examination of 4 different doses of doxycycline in ten human pterygium samples.","project":"SRP010038"}
{"number_samples":6,"species":"human","abstract":"Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease of unknown cause that lacks a proven therapy for altering its high mortality rate. Microarrays have been employed to investigate the pathogenesis of IPF, but are presented mostly at the gene-expression level due to technologic limitations. In as much as, alternative RNA splicing isoforms are increasingly identified as potential regulators of human diseases, including IPF, we propose a new approach with the capacity to detect splicing variants using RNA-Seq data. We conducted a joint analysis of differential expression and differential splicing on annotated human genes and isoforms, and 333 non-differentially expressed genes were identified with a high degree of “switch” between major and minor isoforms. Three cases with variant mechanisms for alternative splicing were validated using qRT-PCR, among the group of genes in which expression was not significantly changed at the gene level. We also identified 37 genes related to IPF specific novel transcripts using exon-exon junction evidence, and selected a representative for qRT-PCR validation. The results of our study are likely to provide new insight into the pathogenesis of pulmonary fibrosis and may eventuate in new treatment targets. Moreover, a similar approach can be applied to discovering novel splicing variants in other diseases.","project":"SRP010041"}
{"number_samples":11,"species":"human","abstract":"Using DNase-seq, mRNA-seq and publicly available ChIP-seq data sets, we examined the role of chromatin accessibility (DNase-seq) in androgen receptor binding to the genome (ChIP-seq) and AR-mediated transcriptional changes (mRNA-seq). Our data reveals genome-wide changes in chromatin structure that correspond to AR binding and  differential gene expression.  A focused examination of DNase-seq data around androgen receptor motifs within androgen receptor ChIP-seq peaks reveals distinct patterns of protection from DNaseI cleavage. Overall design: Examination of chromatin accessibility (DNase-seq), AR binding (AR ChIP-seq), and transcription (mRNA-seq) in LNCaP cells before and after 12 hours of 1 nM R1881 treatment This Series represents the RNA-Seq data only. Exon microarray data generated under the same conditions is available through GSE15805.  The DNase-seq data is publicly available through GSE32970 as well as the UCSC genome browser (genome.ucsc.edu) under Regulation::ENCODE DNase/FAIRE::Duke DNaseI HS:LNCaP and LNCaP + Andro.  The accession numbers for the ChIP-seq experiments used are GSE14097 and GSE28126.","project":"SRP010054"}
{"number_samples":9,"species":"human","abstract":"RNA-Seq reads and TopHat (Trapnell et al. Bioinformatics 2009) alignments of K562 cell-line transcriptome.  These were used to validate the expression of short peptides idenitified by Mass-Spectrometry in K562 cells. Overall design: K562 polyA+ RNA (Batch 1) and total RNA (batch 2) was purchased from Ambion. We used oligo (dT)-selected polyA+ RNA to construct libraries for RNA-Seq.We then profiled the transcriptome of polyadenylated mRNA-Seq using Illumina sequencing platforms. We then used the sequenced reads to reconstruct the transcriptome using the Cufflinks de-novo assembler (Trapnell et al. Nat.Bio.Tech. 2010). Recent computational and ribosome profiling analyses suggest that many short open reading frames (sORFs) in eukaryotic genomes are translated. However, evidence that these sORFs produce stable polypeptides is lacking. Here we develop a strategy to discover and validate novel sORF-encoded polypeptides (SEPs) in human cells. In total, we detect 117 SEPs, 114 of which are novel, varying in length from 15 to 149 amino acids. Of these, 10 SEPs (0.5%) are derived from long intergenic non-coding RNAs (lincRNAs). We also observe the presence of polycistronic genes and the widespread use of non-AUG start codons, which is a phenomenon historically thought to be rare in the mammalian genome. Quantitative measurements reveal that SEPs can be found at concentrations between ~10-2000 copies per cell, which is within the range of typical cellular proteins. We confirm the translation of a number of these SEPs through heterologous expression of their encoding cDNAs. We also discover that several SEPs possess properties characteristic of functional proteins. These results demonstrate that human sORFs produce numerous stable polypeptides, revealing that the human proteome is larger and more diverse than previously appreciated.","project":"SRP010061"}
{"number_samples":1,"species":"human","abstract":"The ability to chronicle transcription factor binding events throughout the development of an organism would facilitate mapping of transcriptional networks that control cell fate decisions. We describe a method for permanently recording protein-DNA interactions in mammalian cells.  We endow transcription factors with the ability to deposit a transposon into the genome near to where they bind. The transposon becomes a “Calling Card” the transcription factor leaves behind to record its visit to the genome. The locations of the Calling Cards can be determined by massively-parallel DNA sequencing. We show that the transcription factor SP1 fused to the piggyBac transposase directs insertion of the piggyBac transposon near SP1 binding sites. The locations of transposon insertions are highly reproducible, and agree with sites of SP1-binding determined by ChIP-seq. Genes bound by SP1 are more likely to be expressed in the HCT116 cell line we used, and SP1-bound CpG islands show a strong preference to be unmethylated. This method has the potential to trace transcription factor binding throughout cellular and organismal development in a way that has heretofore not been possible. Overall design: These data contain mapped insertion sites from the piggyBac (PB) transposon into HCT116 cell line and sequenced using an Illumina GAII analyzer.  The first set contains the insertion sites of transposons mapped from the wild-type PB transposase and the second set contains the insertion sites of transposons mapped with the PB transposase fused to the transcription factor SP1.  Other sequencing data present are ChIP-seq data for SP1 in the HCT116 cell line and RNA-seq data.","project":"SRP010064"}
{"number_samples":40,"species":"human","abstract":"We profiled and quantitated miRNAs in two skin tumors (Basal cell carcinoma and Merkel cell carcinoma) and identified tumor-specific miRNAs. We used these tumor-specific miRNAs to guide development of miRNA fluorescence in situ hybridization. Overall design: 2 barcoded sequencing runs, including 40 unique samples (36 used in manuscript). The details of each sample can be found in Supplementary Tables S1 and S2.","project":"SRP010129"}
{"number_samples":16,"species":"human","abstract":"In this manuscript, we have used RNA-Sequencing experiment to obtain and integrate a variety of genomic features in order to identify signaling pathways that are associated to mutant KRAS lung tumors. Overall design: 8 lung adenocarcinomas with mutant KRAS in lung tumors and 8 lung adenocarcarcinomas without KRAS mutation of which one sample is ran twice for QC purposes.","project":"SRP010166"}
{"number_samples":520,"species":"human","abstract":"The human leukocyte antigen (HLA) is key to many aspects of human physiology and medicine. All current sequence-based HLA typing methodologies are targeted approaches requiring the amplification of specific HLA gene segments. Whole genome, exome and transcriptome shotgun sequencing can generate prodigious data but due to the complexity of HLA loci these data have not been immediately informative regarding HLA genotype. We describe HLAminer, a computational method for identifying HLA alleles directly from shotgun sequence datasets (http://www.bcgsc.ca/platform/bioinfo/software/hlaminer). This approach circumvents the additional time and cost of generating HLA-specific data and capitalizes on the increasing accessibility and affordability of massively parallel sequencing.","project":"SRP010181"}
{"number_samples":15,"species":"human","abstract":"Understanding how RNA binding proteins control the splicing code is fundamental to human biology and disease. Here we present a comprehensive study to elucidate how heterogeneous nuclear ribonucleoparticle (hnRNP) proteins, among the most abundant RNA binding proteins, coordinate to regulate alternative pre-mRNA splicing (AS) in human cells. Using splicing-sensitive microarrays, cross-linking and immunoprecipitation coupled with high-throughput sequencing, and cDNA sequencing, we find that more than half of all AS events are regulated by multiple hnRNP proteins, and that some combinations of hnRNP proteins exhibit significant synergy, whereas others act antagonistically. Our analyses reveal position-dependent RNA splicing maps, in vivo consensus binding sites, a surprising level of cross- and auto-regulation among hnRNP proteins, and the coordinated regulation by hnRNP proteins of dozens of other RNA binding proteins and genes associated with cancer. Our findings define an unprecedented degree of complexity and compensatory relationships among hnRNP proteins and their splicing targets that likely confer robustness to cells. Overall design: CLIPseq for hnRNP A1, hnRNP A2/B1, hnRNP F, hnRNP M, and hnRNP U in human 293T cells","project":"SRP010279"}
{"number_samples":31,"species":"human","abstract":"Understanding how RNA binding proteins control the splicing code is fundamental to human biology and disease. Here we present a comprehensive study to elucidate how heterogeneous nuclear ribonucleoparticle (hnRNP) proteins, among the most abundant RNA binding proteins, coordinate to regulate alternative pre-mRNA splicing (AS) in human cells. Using splicing-sensitive microarrays, cross-linking and immunoprecipitation coupled with high-throughput sequencing, and cDNA sequencing, we find that more than half of all AS events are regulated by multiple hnRNP proteins, and that some combinations of hnRNP proteins exhibit significant synergy, whereas others act antagonistically. Our analyses reveal position-dependent RNA splicing maps, in vivo consensus binding sites, a surprising level of cross- and auto-regulation among hnRNP proteins, and the coordinated regulation by hnRNP proteins of dozens of other RNA binding proteins and genes associated with cancer. Our findings define an unprecedented degree of complexity and compensatory relationships among hnRNP proteins and their splicing targets that likely confer robustness to cells. Overall design: RNAseq for control, hnRNP A1, hnRNP A2/B1, hnRNP H1, hnRNP F, hnRNP M, and hnRNP U siRNA treated human 293T cells","project":"SRP010280"}
{"number_samples":18,"species":"human","abstract":"Prostate cancer is the most common cancer in men and cardiac glycosides inhibit prostate cancer cell proliferation. In order to investigate the mechanism by which cardiac glycosides inhibit  prostate cancer cells, we observed genome-wide RNA expression in prostate cancer LNCaP-abl cells, hormone resistant cells, after the cardiac glycoside treatment using RNA-Seq. In addition, we profiled LNCaP-abl cells after androgen receptor (AR) knockdown to observe whether cardiac glycoside effect on RNA expression is similar to that of  AR knockdown. Overall design: Observation of three cardioglycosides, Digoxin, Peruvoside and Strophanthidin,  and AR knockdown regulated RNA expression in LNCaP-abl with RNA-Seq (each triplicates)","project":"SRP010350"}
{"number_samples":5,"species":"human","abstract":"In the universal genetic code, most amino acids can be encoded by multiple trinucleotide codons, and the choice among available codons can influence position-specific translation elongation rates. By using sequence-based ribosome profiling, we obtained transcriptome-wide profiles of in vivo ribosome occupancy as a function of codon identity in Caenorhabditis elegans and human cells. Particularly striking in these profiles was a universal trend of higher ribosome occupancy for codons translated via G:U wobble base-pairing compared with synonymous codons that pair with the same tRNA family using G:C base-pairing. These data support a model in which ribosomal translocation is slowed at wobble codon positions.","project":"SRP010374"}
{"number_samples":3,"species":"human","abstract":"In order to support our research of bladder cancer in human genome, we conducted massively parallel pyrosequencing of mRNAs (RNA-Seq) using normal, paracancerouse and cancerous human bladder tissues. We obtained a total of 30.0 million read pairs from normal, 33.1 million read pairs from paracancerous and 36.5 million read pairs from cancerous. The RNA-Seq data derived from the sample illustrated the differencially expression genes among normal, paracancerous and cancerous bladder tissues of human. Overall design: 3 samples examined: normal tissue, paracancerous tissue, cancerous tissue.","project":"SRP010384"}
{"number_samples":1,"species":"human","abstract":"DNA methylation (5mC) is an epigenetic modification classified by the addition of a methyl group (Ch3-) catalysed by DNA methyltransferases (DNMTs) and S-adenosyl methionine (SAM). Recently hydroxymethylation at the same carbon-5 residue of the cytosine base catalysed by the TET (Ten–Eleven Translocation) family of proteins has been shown occur in mouse embryonic stem cells and the brain. 5-hydroxymethylcytosine (5hmC) has been shown to be distributed at a lesser extent in most mammalian tissues with an increased occurrence in Purkinje cells of the mouse brain. Here we assess genome-wide 5hmC distribution relative to 5mC enrichment in the human cerebellum and peripheral blood lymphocytes (PBLs) using high-resolution sequencing by synthesis. We found a significant increase and change in 5hmC distribution in human cerebellum at loci highly expressed in the human brain and postsynaptic density (PSD) compared to PBLs. Minimal changes in 5mC distribution were observed at these same loci and genome-wide in both human cerebellum and PBLs. Using high-throughput RNA-Seq of total RNA from human brain, cerebellum and PBLs we quantified elevated expression of gene products implicated in neuronal activity and having significantly increased 5hmC distribution across their gene bodies only in brain tissue compared to PBLs. Furthermore we show elevated expression of gene loci in the cerebellum to co-localise with increased 5hmC in Purkinje neurons. Our data provide new evidence for 5hmC mediated regulation of gene loci highly expressed in the human brain and PSD and implicated in neuronal activity and development. Overall design: High-throughput RNA-Seq analysis of total RNA isolated from human cerebellum (Capital Biosciences)","project":"SRP010427"}
{"number_samples":4,"species":"human","abstract":"The association between hyper-inflammatory states and numerous diseases is widely recognized, but our understanding of the molecular strategies that have evolved to prevent uncontrolled activation of inflammatory responses remains incomplete. Here, we report a critical, non-transcriptional role of GPS2 as a guardian against hyperstimulation of TNFA-induced gene program. GPS2 cytoplasmic actions are required to specifically modulate RIP1 ubiquitylation and JNK activation by inhibiting TRAF2/Ubc13 enzymatic activity. In vivo relevance of GPS2 anti-inflammatory role is confirmed by inhibition of TNFA target genes in macrophages and by improved insulin signaling in the adipose tissue of aP2-GPS2 transgenic mice. As the non-transcriptional role is complemented by GPS2 functioning as positive and negative cofactor for nuclear receptors, in vivo overexpression also results in elevated circulating level of resistin and development of hepatic steatosis. Together, these studies define GPS2 as a molecular guardian required for precise control of inflammatory responses involved in immunity and homeostasis. Overall design: RNA-sequencing of polyA selected RNA molecules in 293T cells and ChIP-seq of GPS2, TBL1, and NCOR.","project":"SRP010430"}
{"number_samples":10,"species":"human","abstract":"We have used RNA-seq to identify transcripts, including splice variants, expressed in human islets of Langerhans under control condition or following exposure to the pro-inflammatory cytokines interleukin-1ß (IL-1ß) and interferon-? (IFN-?). A total of 29,776 transcripts were identified as expressed in human islets. Expression of around 20% of these transcripts was modified by pro-inflammatory cytokines, including apoptosis- and inflammation-related genes. Chemokines were among the transcripts most modified by cytokines. Interestingly, 35% of the genes expressed in human islets undergo alternative splicing as annotated in RefSeq, and cytokines caused substantial changes in spliced transcripts. Nova1, previously considered a brain-specific regulator of mRNA splicing, is expressed in islets. 25/41 of the candidate genes for type 1 diabetes are expressed in islets, and cytokines modified expression of several of these transcripts. Overall design: 5 human islet of Langerhans preparations examined under 2 conditions (control and cytokine treatment)","project":"SRP010483"}
{"number_samples":17,"species":"human","abstract":"Genome-wide gene expression was measured in peripheral blood mononuclear cells (PBMCs) from patients with cystic fibrosis (CF) after treatment in vitro with the flagellin protein fliC, and/or synthetic peptide IDR-1018 to assess patterns of gene expression. The patterns of gene expression suggest that CF cells have a hyperinflammatory phenotype including dysfunctional autophagy processes. The synthetic peptide IDR-1018 attentuates this hyperinflammatory phenotype. Overall design: Total RNA was obtained from PBMCs obtained from CF patients after treatment with the fliC flagellin protein (that is known to play a role in CF lung inflammation), and/or the peptide IDR-1018 that has anti-inflammatory properties. Comparison of genes and pathways affected by these treatments indicated the role of autophagy process in CF disease.","project":"SRP010644"}
{"number_samples":2,"species":"human","abstract":"Nonsense-mediated mRNA decay (NMD) functions to degrade transcripts bearing premature stop codon (PTC) and is a crucial regulator of gene expression. NMD and the UPF3B gene have been implicated as the cause of various forms of intellectual disability (ID) and other neurological symptoms. Here, we reports three patients with global developmental delay carrying hemizygous deletions of the UPF2 gene, another important member of the NMD pathway and direct interacting partner of UPF3B. Overall design: Using RNA-SEQ on lymphoblastoid cells from UPF2 deletion patients, we identified 1009 differently expressed genes (DEGs). 38% of these DEGs overlapped with DEGs identified in UPF3B patients. More importantly, 95% of all DEGs in either UPF2 or UPF3B patients share the same trend of de-regulation. This demonstrates that the transcriptome deregulation in these two patient groups is similar and that UPF2 should be considered as a new candidate gene for ID in man. We expanded our inq`uiries and performed a comprehensive search for copy number variations (CNVs) encompassing all NMD genes in cohorts of ID patients and controls. We found that UPF2, UPF3A, Y14, SMG6 and EIF4A3 are frequently deleted and/or duplicated in ID patients. These CNVs are likely to be the root of the problems or to act as predisposing factors. Our results suggest that dosage imbalance of NMD factors is associated with ID and further emphasize the importance of NMD in normal learning and memory processes.","project":"SRP010647"}
{"number_samples":2,"species":"human","abstract":"Hepatitis B virus (HBV) is a hepatotropic virus that can regulate many host cellular gene expressions participating in the HBV life cycle, liver inflammation and hepatocellular injury. However, the underlying mechanism of differential gene expression is not understood. We report here a genome-wide analysis of histone methylation on two histone H3 lysine residues (H3K4me3 and H3K27me3) and gene expression profiles in HepG2 and HepG2.2.15 cells. We found that specific correlation exists between gene expression and the amounts of H3K4me3 (positive correlation) and H3K27me3 (negative correlation) across the gene body. These correlations displayed three distinct modes (repressive, active and poised), reflecting different functions of these genes in the HBV life cycle, liver inflammation and hepatocellular injury. Furthermore, a permissive chromatin state of each gene was established by a combination of different histone modifications. Our findings reveal a complex regulation by histone methylation in differential gene expression and suggest that histone methylation may be responsible for the HBV life cycle, liver inflammation and hepatocellular injury induced by HBV. Overall design: A large-scale analysis of gene expression of 2 different cell types (HepG2, HepG2.2.15) using a digital gene expression (DGE) tag profiling approach.","project":"SRP010670"}
{"number_samples":6,"species":"human","abstract":"Dicer plays a key role in RNA silencing. By processing pre-miRNAs and dsRNAs, Dicer generates miRNAs and siRNAs that act as post-transcriptional regulators of gene expression. Dicer is also implicated in heterochromatin formation and toxic RNA degradation. Here we report that Dicer is controlled in cell cycle. The Dicer protein level drastically rises at late G1 phase and falls at early S phase. Interestingly, the protein stability of Dicer is decreased specifically at early S phase. MG132, an inhibitor of proteasome, increases Dicer protein level, suggesting that the stability of Dicer is controlled via ubiquitination-dependent proteasome pathway.","project":"SRP010675"}
{"number_samples":3,"species":"human","abstract":"The aim of this study was to establish a deeply sequenced transcriptome at multiple timepoints during the differentiation of human epidermal keratinocytes from the progenitor state (d0).  These transcriptomes were then assembled in order to discover novel genes and transcriptional events that are dynamically regulated during terminal differentiation of a human somatic tissue. Overall design: Paired-end RNA sequencing was performed on primary human keratinocytes at three timepoints during calcium-induced epidermal differentiation.","project":"SRP010678"}
{"number_samples":12,"species":"human","abstract":"The mammalian target of rapamycin (mTOR) kinase is a master regulator of protein synthesis that couples nutrient sensing to cell growth and cancer. However, the downstream translationally regulated nodes of gene expression that may direct cancer development are poorly characterized. Employing ribosome profiling we uncover specialized translation of the prostate cancer genome by oncogenic mTOR signaling, revealing a remarkably specific repertoire of genes involved in cell proliferation, metabolism, and invasion. We extend these findings by functionally characterizing a class of translationally controlled pro-invasion mRNAs that we show direct prostate cancer invasion and metastasis downstream of oncogenic mTOR signaling. We further develop a clinically relevant ATP site inhibitor of mTOR, INK128, which reprograms this gene expression signature with therapeutic benefit for prostate cancer metastasis, for which there is presently no cure. Together, these findings significantly extend our understanding of how the “cancerous” translation machinery steers specific cancer cell behaviors and may be therapeutically targeted. Overall design: Examination of mRNA translation in human prostate cancer upon differential inhibition of the mTOR signaling pathway.","project":"SRP010679"}
{"number_samples":2,"species":"human","abstract":"Two normal frontal white matter samples, obtained from two pediatric patients (A boy of 3 year old and a girl of 3 mount old) undergoing focal brain resection for head injury, were obtained. miRVana miRNA Isolation Kit (Ambion, USA) was used following the manufacturing instruction to isolate miRNA from these two samples. Brain samples were collected during neurosurgery of the patients at the Policlinico Gemelli in Rome. The study was approved by the local ethical committee. The miRNA was sequenced using Illumina HiSeq2000 machine.","project":"SRP010712"}
{"number_samples":5,"species":"human","abstract":"Viral infection is commonly associated with virus-driven hijacking of host proteins. We describe a novel mechanism by which influenza virus impacts host cells through the interaction of influenza NS1 protein with the infected cell epigenome. We show that the NS1 protein of influenza A H3N2 target the transcription elongation PAF1 complex (hPAF1C). We demonstrate that binding of NS1 to hPAF1C results in suppression of hPAF1C-mediated transcriptional elongation.  In the following data sets, we show that NS1 colocalizes with hPAF1 on the chromatin of infected cells and that siRNA-mediated reduction of hPAF1 expression results in reduced recruitment of NS1 to the chromatin. Overall design: Examination of  different histone modifications in infected cells and RNA-Seq and GRO-Seq transcript measurements.","project":"SRP010907"}
{"number_samples":5,"species":"human","abstract":"The precise control of microRNA (miRNA) biogenesis is important for various cellular functions, and its dysregulation is often associated with human diseases. We previously reported that Terminal uridylyl transferase 4 (TUT4) down-regulates let-7 miRNA biogenesis by oligo-uridylating let-7 precursor (pre-let-7) in mouse embryonic stem cells and that a pluripotency marker Lin28 promotes a processivity of TUT4. Here we find that TUT4 positively controls let-7 biogenesis by adding a uridine residue to the 3’ end of pre-let-7 in the absence of Lin28. Such mono-uridylation enhances Dicer processing by generating an optimal end structure of pre-let-7 for Dicer recognition and may protect pre-miRNA from trimming. Moreover, TUT7, TUT4 and TUT2 redundantly regulate pre-let-7 processing and simultaneous knock down of these TUTs leads to the decrease of mature let-7 and the accumulation of pre-let-7 in cells. This study provides a novel regulation mechanism of miRNA biogenesis, which may function in development and tumorigenesis. Overall design: HeLa cells were transfected with siRNA two times over a 4~5 day period.","project":"SRP010943"}
{"number_samples":4,"species":"human","abstract":"Background: Sm proteins are multimeric RNA-binding factors, found in all three domains of life. Eukaryotic Sm proteins, together with their associated RNAs, form small ribonucleoprotein (RNP) complexes important in multiple aspects of gene regulation. Comprehensive knowledge of the RNA components of Sm RNPs is critical for understanding their functions. Results: We developed a multi-targeting RNA-immunoprecipitation sequencing (RIP-seq) strategy to reliably identify Sm-associated RNAs from Drosophila ovaries and cultured human cells. Using this method, we discovered three major categories of Sm-associated transcripts: small nuclear (sn)RNAs, small Cajal body (sca)RNAs and mRNAs. Additional RIP-PCR analysis showed both ubiquitous and tissue-specific interactions. We provide evidence that the mRNA-Sm interactions are mediated by snRNPs, and that one of the mechanisms of interaction is via base pairing. Moreover, the Sm-associated mRNAs are mature, indicating a splicing-independent function for Sm RNPs. Conclusions: This study represents the first comprehensive analysis of eukaryotic Sm-containing RNPs, and provides a basis for additional functional analyses of Sm proteins and their associated snRNPs outside of the context of pre-mRNA splicing. Our findings expand the repertoire of eukaryotic Sm-containing RNPs and suggest new functions for snRNPs in mRNA metabolism. Overall design: RNA-Immunoprecipitation sequencing of RNA-Sm protein complexes.","project":"SRP010944"}
{"number_samples":6,"species":"human","abstract":"The formation and execution of a productive immune response requires, among many things, the maturation of competent T cells and a robust change in cellular activity upon antigen challenge. Such changes in cellular function require regulated alterations of protein expression. Much work has previously gone into defining the transcriptional changes that regulate protein expression during T cell development and antigen stimulation. Here we describe a parallel pathway of gene regulation that occurs during T cell stimulation, namely alternative splicing. Specifically, we use RNA-Seq to identify 178 exons in 168 genes that exhibit robust changes in inclusion in response to a stimulation of a human T cell line. Interestingly, these signal-responsive genes are enriched for functions related to immune response including, cell trafficking, inflammatory and immune response, immunologic disease and several cell signaling pathways. The vast majority of these genes also exhibit different isoform expression in naive and activated primary T cells. Comparison of the responsiveness of splicing to various stimuli in the cultured and primary T cells reveal important insight into the diversity of signaling pathways that control splicing. Using this data we are able to classify signal-responsive exons into at least three distinct networks. Importantly, we find that each regulatory network is characterized by distinct sequence hallmarks, further suggesting independent regulatory mechanisms. Overall design: We utilize high-throughput RNA sequencing (RNA-Seq) to investigate global changes in alternative splicing in a cultured T cell line and in primary human T cells. We identify 178 genes that are predicted to exhibit robust signal-induced changes in isoform expression in cultured T cells.","project":"SRP010961"}
{"number_samples":2,"species":"human","abstract":"Genome-wide determination of a broad ESRP-regulated post-transcriptional network by high throughput sequencing.","project":"SRP011008"}
{"number_samples":2,"species":"human","abstract":"RNAs directly interacting with EZH2 were isolated from human colorectal HCT116 cells using an in vivo crosslinking and immunoprecipitation strategy (iCLIP, König J et al, Nat Struct Mol Biol 2010) coupled to an ultrasequencing approach. Overall design: RIP sequencing for 1 Human sample","project":"SRP011054"}
{"number_samples":39,"species":"human","abstract":"Candida albicans is the major invasive fungal pathogen of humans, causing diseases ranging from superficial mucosal infections to disseminated, systemic infections which are often life threatening. Hematogenously disseminated candidiasis (HDC) has a 47% mortality rate despite current antifungal therapy. An increase in prevalence, as well as an increasing resistance to most of the clinically important antifungal therapies, provides a strong impetus to understand the molecular mechanisms of pathogenesis and the acquisition of drug resistance. This information holds promise to identify novel therapeutic targets. A complete and accurate characterization of how the transcriptome of C. albicans responds to its interaction with cells from the host is an absolute necessity to accomplish this goal. RNA-seq (deep-sequencing of cDNA) provides an unbiased method to define comprehensively and systematically the transcriptome of an organism. We propose a comprehensive characterization of the C. albicans transcriptome in two different human tissue culture models, using RNA-seq. Such a characterization will shed unprecedented light on how fungal pathogens sense and respond to the host environment and how the host tissue responds to fungal invasion. A major problem in the design of antifungal agents stems from the fact that fungi and mammals are both eukaryotic organisms. Pharmaceutical compounds that can kill fungal cells will often kill human cells resulting in a drug having toxic side effects. Since a successful antifungal drug must inhibit the function of a gene that is expressed during the course of an infection, this work will provide a list of potential novel therapeutic targets to treat candidiasis, as well as mycoses caused by other serious invasive fungal pathogens, in the clinic. This project is led by Vincent M. Bruno, Ph.D., Institute for Genome Sciences, in collaboration with Dr. Scott Filler, David Geffen School of Medicine at UCLA.","project":"SRP011085"}
{"number_samples":2,"species":"human","abstract":"Identify YB1 associated RNAs","project":"SRP011107"}
{"number_samples":4,"species":"human","abstract":"We examine the concordance between miRNA-seq and cDNA-mediated annealing, selection, extension, and ligation (DASL) miRNA assays. Overall design: We profiled two technically replicated FFPE specimens on both DASL and RNA-seq. This submission represents the RNA-seq component of study. The DASL samples used in the study are GSM876557, GSM876558, GSM876564, and GSM876565.","project":"SRP011130"}
{"number_samples":1,"species":"human","abstract":"5'-complete cDNA sequencing on ribosome-depleted total RNA from the human K562 cell line. Provides high-quality, genome-wide single-base resolution profiling of transcription start sites and their expression levels. Overall design: This dataset represents a whole-genome, single-base resolution profiling of transcription start site (TSS) expression in the human K562 cell line. These profiles were established using RAMPAGE, a high-throughput, high-accuracy 5'-complete cDNA sequencing method implemented on the Illumina platform. The data was analyzed using custom scripts and algorithms that are all available upon request.","project":"SRP011185"}
{"number_samples":4,"species":"human","abstract":"While the importance of random sequencing errors decreases at higher DNA or RNA sequencing depths, systematic sequencing errors (SSEs) dominate at high sequencing depths and can be difficult to distinguish from biological variants.  These SSEs can cause base quality scores to underestimate the probability of error at certain genomic positions, resulting in false positive variant calls, particularly in mixtures such as samples with RNA editing, tumors, circulating tumor cells, bacteria, mitochondrial heteroplasmy, or pooled DNA.  Most algorithms proposed for correction of SSEs require a training data set, which is typically either from a part of the data set being “recalibrated” (Genome Analysis ToolKit, or GATK) or from a separate data set with special characteristics (SysCall).  Here, we combine the advantages of these approaches by adding synthetic RNA spike-in standards to human RNA, and use GATK to recalibrate base quality scores with reads mapped to the spike-in standards. Compared to conventional GATK recalibration that uses reads mapped to the genome, spike-ins improve the accuracy of Illumina base quality scores by a mean of 5 units, and by as much as 13 units  at CpG sites.  In addition, since reads mapping to the genome are not used for recalibration, our method allows run-specific recalibration even for the many species without a comprehensive and accurate SNP database.  We also use GATK with the spike-in standards to demonstrate that the Illumina RNA sequencing runs overestimate quality scores for AC, CC, GC, GG, and TC dinucleotides, while SOLiD has less dinucleotide SSEs but more SSEs for certain cycles.  We conclude that using these DNA and RNA spike-in standards with GATK improves base quality score recalibration. Overall design: Four human RNA samples with equimolar ERCC spike-in standards were sequenced on Illumina.  Two human brain/liver/muscle RNA mixtures with dynamic range of ERCC spike-in standards were sequenced on SOLiD.","project":"SRP011192"}
{"number_samples":8,"species":"human","abstract":"Whole-genome transcriptome measurements are pivotal for characterizing carcinogenic mechanisms of chemicals and predicting toxic classes, such as genotoxicity, from in vitro and in vivo assays. DNA microarrays have evolved as the gold standard for this purpose. In recent years deep sequencing technologies have been developed that hold the promise of measuring the transcriptome with RNA-seq in a more accurate and unbiased manner than microarrays. So far, however, few applications have been published that assess the performance of RNA-seq within a toxicogenomics context. Here, we applied RNA-seq for the characterization of the in vitro transcriptomic responses in HepG2 cells upon exposure to benzo[a]pyrene (BaP), a well-known DNA damaging carcinogen. We demonstrate the performance of RNA-seq with respect to the identification of differentially expressed genes and associated pathways, in comparison with microarray technology. RNA-seq data generates more complete and thus accurate data on differentially expressed genes and affected pathways than microarrays. Additionally, we highlight the potential of RNA-seq for characterizing mechanisms related to alternative splicing and thereby gathering new information. Exposure to BaP alters the isoform distribution for many genes, including regulators of cell death and DNA repair such as TP53, BCL2 and XPA, which are relevant for genotoxic responses. Finally, we demonstrate that RNA-seq enables to investigate allele-specific gene expression, although no changes for that could be observed. Our results provide evidence that RNA-seq is a powerful tool for toxicology which, compared to microarrays, is capable of adding valuable information at the transcriptome level for characterizing toxic effects caused by chemicals. Overall design: Examination of 2 biological replicates at 2 different timepoints","project":"SRP011233"}
{"number_samples":6,"species":"human","abstract":"Deficiencies in the ATM gene are the underlying cause for ataxia telangiectasia, a congenital syndrome characterized by neurological, motor and immunological defects, as well as a predisposition to cancer risks. MicroRNAs (miRNAs) are small regulators of post-transcriptional gene expression and a useful tool for cancer diagnosis, staging, and prediction of therapeutic responses to clinical regimens. In particular, miRNAs have been used to develop signatures for breast cancer profiling.  We are interested in the consequences of ATM deficiency on miRNA expression in breast epithelial cells and the potential contribution to cancer predisposition. In this study we investigate the effects of ATM loss on the miRNA expression and related gene expression changes in normal human mammary epithelial cells (HME-CC). We have identified 81 significantly differently expressed miRNAs in the ATM-deficient HME-CCs using small RNA sequencing. Many of these differentially expressed miRNAs have been described and implicated in tumorigenesis and proliferation. These changes include down-regulation of tumor suppressor miRNAs, such as hsa-miR-29c and hsa-miR-16, as well as the over-expression of pro-oncogenic miRNAs hsa-miR-93 and hsa-mir-221. All 81 miRNAs were combined with genome wide gene expression profiles to investigate possible targets of miRNA regulation. We identified messenger RNA (mRNA) targets of these miRNAs that were also significantly regulated after the depletion of ATM. Predicted targets included many genes implicated in cancer formation and progression, including SOCS1 and the proto-oncogene MAF. Integrated analysis of miRNA and mRNA expression allows us to build a more complete understanding of the pathways and networks involved in the breast cancer predisposition observed in individuals deficient in ATM. This study highlights miRNA and predicted mRNA target expression changes in ATM-deficient HME-CCs and suggests a mechanism for the breast cancer-prone phenotype seen in ATM deficient cells and patients. Additionally, this study provides preliminary data for defining miRNA profiles that may be used prognostic biomarkers for breast cancer predisposition. Overall design: Examination of small RNA population in human mammary epithelial cell lines. Each condition was preformed in triplicate.","project":"SRP011278"}
{"number_samples":15,"species":"human","abstract":"Here we identify a novel class of small RNAs that are ~21-nucleotide in length and are produced from the sequences in the vicinity of DNA double strand break (DSB) sites in Arabidopsis and humans. We named them diRNAs for DSB-induced small RNAs. In Arabidopsis, the biogenesis of diRNAs requires the PI3 kinase ATR, RNA polymerase IV (Pol IV), and Dicer-like proteins. Mutations in these proteins as well as in Pol V cause significant reduction in DSB repair efficiency. diRNAs are recruited by Argonaute 2 (AGO2) to mediate DSB repair. In humans, knocking down Dicer or Ago2 causes a significant reduction in DSB repair. Our findings reveal a novel biological function for small RNAs in the DSB repair pathway. We propose that diRNAs may function as guide molecules for chromatin modifications or recruitment of repair complexes at DSB sites to facilitate repair. Overall design: 28 samples from Arabidopsis thaliana in various genetic backgrounds and 5 samples from the human cells, small RNA sequencing","project":"SRP011369"}
{"number_samples":3,"species":"human","abstract":"In adult cancers, epigenetic changes and aberrant splicing of the DNMT3B is commonly observed, and the pattern of gene methylation and expression has been shown to be modified by DNMT3B7, a truncated protein of DNMT3B.  Much less is known about the mechanism of epigenetic changes in the pediatric cancer neuroblastoma.  To investigate if aberrant DNMT3B transcripts alter DNA methylation, gene expression and tumor phenotype in neuroblastoma, we measured DNMT3B isoform expression in primary tumors and cell lines. Higher levels of DNMT3B7 were detected in differentiated ganglioneuroblastomas compared to undifferentiated neuroblastomas, suggesting that expression of DNMT3B7 may induce a less clinically aggressive tumor phenotype.  To test this hypothesis, we investigated the effects of forced DNMT3B7 in neuroblastoma cells.  We found that DNMT3B7 expression significantly inhibited neuroblastoma cell proliferation in vitro, and in neuroblastoma xenografts, DNMT3B7 decreased angiogenesis and tumor growth. DNMT3B7-positive cells had higher levels of total genomic methylation, and RNA-sequencing revealed a dramatic decrease in expression of FOS and JUN family members, AP1 complex components.  Consistent with the established antagonistic relationship between AP1 expression and retinoic acid receptor activity, decreased proliferation and increased differentiation was seen in the DNMT3B7-expressing neuroblastoma cells following treatment with all trans retinoic acid (ATRA) compared to controls. Our results demonstrate that high levels of DNMT3B7 modify the epigenome in neuroblastoma cells, induce changes in gene expression, inhibit tumor growth, and increase sensitivity to ATRA.    Further knowledge regarding mechanisms by which DNMT3B7 regulates gene methylation may ultimately lead to the development of therapeutic strategies that reverse the epigenetic aberrations that drive neuroblastoma pathogenesis. Overall design: DNMT3B7, a truncated DNMT3B isoform, was stably transfected into an N-type neuroblastoma cell line (LA1-55n) using a Tet-off inducible system. DNMT3B7 expressing cells were compared to vector control cells after 21 days of induction.","project":"SRP011378"}
{"number_samples":31,"species":"human","abstract":"We report results on microRNA profiles revealing the distribution of \"isomirs\" of microRNA in a cancerous state. Deep sequencing was conducted at Stanford University and data analysis was conducted at the University of Connecticut Health Center. Overall design: Small RNA profiling to deduce differential microRNA expression levels along various stages of melanoma. Deep sequencing was performed on formalin-fixed paraffin-embedded (FFPE) archived tissue samples, fresh frozen samples from melanomas, and cell lines.","project":"SRP011422"}
{"number_samples":1,"species":"human","abstract":"In order to support our research of chronic myeloid leukemia in human genome, we conducted massively parallel pyrosequencing of mRNAs (RNA-seq) using chronic myeloid leukemia blood in early disease. We obtained a total of 17.74 million read pairs from blood in early disease.The RNA-seq data derived from the sample illustrated the expreesion genes in chronic myeloid leukemia blood in early disease of human. Overall design: 1 sample examined: blood in early disease.","project":"SRP011486"}
{"number_samples":124,"species":"human","abstract":"Find the casual relationship between gene expression network and cellular phenotype at single cell resolution. We collected donated human pre-implatation embryos, and the embryonic stem cells derived from them, isolate individual cells, prepared single cell cDNAs, and sequenced them by HiSeq2000. Then we analyzed the expression of known RefSeq genes. Overall design: We get transcriptome of 124 individual cells from human pre-implantation embryos and human embryonic stem cells by applying single cell RNA-seq technique we recently developed[1][2][3][4]. We did in-depth bioinformatic analysis to these data and found very dynamic expression of protein-coding genes. [1] Tang, F. et al. (2010a) Tracing the Derivation of Embryonic Stem Cells from the Inner Cell Mass by Single-Cell RNA-Seq Analysis. Cell Stem Cell 6, 468-478. [2] Tang, F. et al. (2010b) RNA-Seq analysis to capture the transcriptome landscape of a single cell. Nat Protocols 5, 516-535. [3] Tang, F. et al. (2009) mRNA-Seq whole-transcriptome analysis of a single cell. Nat Meth 6, 377-382. [4] Tang, F. et al. (2011) Development and applications of single-cell transcriptome analysis. Nat Meth 8, S6-S11.","project":"SRP011546"}
{"number_samples":2,"species":"human","abstract":"Recent studies demonstrated that cancer stem cells (CSCs) have higher tumorigenesis properties than those of differentiated cancer cells and that transcriptional factor-SOX2 plays a vital role in maintaining the unique properties of CSCs; however, the function and underlying mechanism of SOX2 in carcinogenesis of lung cancer are still elusive. This study applied immunohistochemistry to analyze the expression of SOX2 in human lung tissues of normal individuals as well as patients with adenocarcinoma, squamous cell carcinoma, large cell and small cell carcinoma and demonstrated specific overexpression of SOX2 in all types of lung cancer tissues. This finding supports the notion that SOX2 contributes to the tumorigenesis of lung cancer cells and can be used as a diagnostic probe. In addition, obviously higher expression of oncogenes c-MYC, WNT1, WNT2 and NOTCH1 was detected in side population (SP) cells than in none side population (NSP) cells of human lung adenocarcinoma cell line-A549, revealing a possible mechanism for the tenacious tumorigenic potential of CSCs. To further elucidate the function of SOX2 in tumorigenesis of cancer cells, A549 cells were established with expression of luciferase and doxycycline inducible shRNA targeting SOX2. We found silencing of SOX2 gene reduces the tumorigenic property of A549 cells with attenuated expression of c-MYC, WNT1, WNT2 and NOTCH1 in xenografted NOD/SCID mice. By RNA-Seq method, additional 246 target cancer genes of SOX2 were revealed. These results present evidence that SOX2 may regulate the expression of oncogenes in CSCs to promote the development of human lung cancer. Overall design: Examination of mRNA profiles in A549 cells with SOX2 silencing","project":"SRP011578"}
{"number_samples":12,"species":"human","abstract":"Retinal Pigment Epithelium (RPE) derived from two human embryonic stem cell lines, H1 and H9, were compared with human fetal RPE (hfRPE) using RNA-seq. Nominally, the transcriptome of H1-derived RPE showed greater overlap with hfRPE. For cells maintained in the medium used to differentiate RPE, 6.2% (H1-RPE) and 4.2% (H9-RPE) of the transcripts were expressed in amounts that were statistically different from hfRPE (false discovery rate: 5%).  After adaptation to the serum-free medium, SFM-1, only 1.0 % (H1-RPE) and 1.9% (H9-RPE) were expressed in amounts that were statistically different. For RPE signature genes, statistical differences were observed for 1.0 % (H1-RPE) and 1.9% (H9-RPE) of the transcripts. For some barrier-function related mRNAs the statistical differences were greater than these small differences would predict. For adhesion proteins and plasma membrane transporters, the differences were as great as 6.9% and 4.3%, respectively. After adaptation to SFM-1, the statistical differences between H1- and H9-RPE were only 0.4% for all transcripts, 1.4% for signature genes, and 0.7% for membrane transporters.  No statistical differences were observed for the transcripts related to tight junctions, adhesion junctions or ion channels. In summary, SFM-1 promoted the maturation of stem cell-derived RPE, and the statistical difference between two stem cell lines was minimal. Overall design: RNA sequencing of hfRPE from three fetuses (reference sample) and three independent isolates of RPE derived from the H1, and three from the H9, human embryonic stem cell (hESC) lines. The cultures were maintained in a serum-free medium, SFM-1.  Comparisons were also made to cultures maintained in the medium used to differentiate the cells, KSR.","project":"SRP011895"}
{"number_samples":15,"species":"human","abstract":"We used RNA sequencing to identify the RBFOX1 splicing network at a genome-wide level in primary human neural stem cells during differentiation. We observe that RBFOX1 regulates a large set of alternative splicing events implicated in neurogenesis and cell maintenance. Subsequent alterations in gene expression define an additional transcriptional network regulated by RBFOX1 involved in neurodevelopmental pathways remarkably parallel to those affected by splicing. Overall design: RNA sequencing at a 75bp single-end read scale was performed using polyA-enriched RNA from 5 biological replicates of primary human neural progenitor cell lines generated by lentiviral-mediated knockdown of GFP (control) or RBFOX1 and differentiated for 4 weeks.","project":"SRP011903"}
{"number_samples":29,"species":"human","abstract":"Right ventricular mRNA profiles from 22 patients with Tetralogy of Fallot (TOF) and mRNA-seq profiles from the left and right ventricle (LV and RV, respectively) of 4 healthy unaffected individuals (NH) were generated. The total RNA was isolated from the 30 human heart samples using TRIzol. mRNAs were isolated from total RNA and prepared for sequencing using Illumina Kit RS-100-0801 according to the manufacturer's protocol (Preparing Samples for Sequencing of mRNA Sept 2008). The sequencing libraries were generated using a non-strand-specific library construction method. The purified DNA fragments were used directly for cluster generation, and 36 cycles of single-end read sequencing were performed using the Illumina Genome Analyzer. Sequencing reads were extracted from the image files using the open source Firecrest and Bustard applications (Illumina Genome Analyzer pipeline 1.3.2, 1.4.0 and 1.5.0). Overall design: Right ventricular mRNA-seq profiles from 22 patients with Tetralogy of Fallot and mRNA-seq profiles from the left and right ventricle of 4 healthy unaffected individuals. Supplementary processed data files: GSE36761_gene_expression_levels.txt: Gene expression profiling data for all samples. GSE36761_gene_expression_levels_normalized.txt: Normalized gene expression profiling data for all samples. GSE36761_transcript_expression_levels.txt: Transcript expression profiling data for all samples. GSE36761_transcript_expression_levels_normalized.txt: Normalized transcript expression profiling data for all samples. Genome build: hg18","project":"SRP011924"}
{"number_samples":4,"species":"human","abstract":"Cornelia de Lange syndrome (CdLS) is a dominantly inherited congenital malformation disorder caused by mutations in the cohesin-loading protein NIPBL1,2 for nearly 60% of individuals with classical CdLS 3-5 and in the core cohesin components SMC1A (~5%) and SMC3 (<1%) for a smaller fraction of probands6,7. In humans, the multi-subunit complex cohesin is comprised of SMC1, SMC3, RAD21 and a STAG protein to form a ring structure proposed to encircle sister chromatids to mediate sister chromatid cohesion (SCC)8 as well as play key roles in gene regulation9. SMC3 is acetylated during S-phase to establish cohesiveness of chromatin-loaded cohesin10-13 and in yeast, HOS1, a class I histone deacetylase, deacetylates SMC3 during anaphase14-16. Here we report HDAC8 as the vertebrate SMC3 deacetylase and the identification of loss-of-function HDAC8 mutations in six CdLS probands. Loss of HDAC8 activity results in increased SMC3 acetylation (SMC3-ac) and inefficient dissolution of the “used” cohesin complex released from chromatin in both prophase and anaphase. While SMC3 with retained acetylation is loaded onto chromatin, ChIP-Seq analysis demonstrates decreased occupancy of cohesin localization sites to result in a consistent pattern of altered transcription seen in CdLS cell lines with either NIPBL or HDAC8 mutations.","project":"SRP011927"}
{"number_samples":8,"species":"human","abstract":"We attempted to identify alterations in gene expression that occur during the progression from normal breast to ductal carcinoma in situ (DCIS) with the aim to elucidate significant genes and pathways underlying the premalignant transformation.  To determine the expression changes that are common to multiple DCIS models (MCF10.DCIS, SUM102 and SUM225) and normal mammary epithelial cells (MCF10A), we grew the cells in three dimensional overlay culture with reconstituted basement membrane and used the extracted RNA for 76 cycles of deep sequencing (mRNA-Seq) using Illumina Genome Analyzer GAIIx.  Analysis of mRNA-Seq results showed 295 consistently differentially expressed transcripts in DCIS models as compared to MCF10A.  These differentially expressed genes are associated with a number of signaling pathways such as integrin, fibroblast growth factor and TGFß signaling.  Many differentially expressed transcripts in DCIS were found to be involved in cell-cell signaling, cell-cell adhesion and cell proliferation.  We further investigated ALDH5A1 gene that encodes for the enzyme, aldehyde dehydrogenase 5A1, which is involved in glutamate metabolism. Further, inhibition of ALDH5A1 with different pharmacological drugs resulted in significant inhibition of cell growth and proliferation in the DCIS models. Overall design: Four cell lines examined: normal mammary epithelial cell line (one sample) and three ductal carcinoma in situ cell lines (three samples). Each sample has two duplicates","project":"SRP011974"}
{"number_samples":2,"species":"human","abstract":"In order to detect the transcriptomic differences during chemotherapy treatment of de novo AML, we adopted massively parallel pyrosequencing of mRNAs (RNA-seq) using blood tissues of an patient with AML (FAB subtype M2) in tumor stage and remission stage. We obtained a total of 34.6 and 30.8 million paired reads from the two samples. The RNA-seq data derived from the sample illustrated the differentially expression genes between the two stages. Overall design: Blood samples of an AML-M2 patient in two stages examined: primary tumor and chemotherapy induced remission.","project":"SRP011977"}
{"number_samples":6,"species":"human","abstract":"Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived human M1- and M2-like macrophage profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to establish a high resolution transcriptome of human macrophages. Total RNA was isolated from classically and aternative activated human macrophages.mRNA profiles were generated by deep sequencing og M1 and M2 macrophages from 3 donors using Illumina HiSeqSQ. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Casava1.8 and TopHat followed by Cufflinks. qRT–PCR validation was performed using LightCycler and SYBR Green assays. Overall design: mRNA profiles were generated by deep sequencing og M1 and M2 macrophages from 3 donors using Illumina HiSeqSQ.","project":"SRP012015"}
{"number_samples":4,"species":"human","abstract":"Through alternative polyadenylation, human mRNAs acquire longer or shorter 3' untranslated regions, the latter typically associated with higher transcript stability and increased protein production. To understand the dynamics of polyadenylation site usage, we mapped transcriptome-wide both binding sites of 3' end processing factors CPSF-160, CPSF-100, CPSF-73, CPSF-30, Fip1, CstF-64, CstF-64tau, CF Im25, CF Im59, and CF Im68 and 3' end processing sites in HEK293 cells. We found that although binding sites of these factors generally cluster around the poly(A) sites most frequently used in cleavage, CstF-64/CstF-64tau and CF Im proteins have much higher positional specificity compared to CPSF components. Knockdown of CF Im68 induced a systematic use of proximal polyadenylation sites, indicating that changes in relative abundance of a single 3' end processing factor can modulate the length of 3' untranslated regions transcriptome-wide, and suggesting a mechanism behind the previously observed increase in tumor cell invasiveness upon CF Im68 knockdown. Overall design: 3' ends of transcripts were profiled by high-throughput sequencing in HEK 293 cells under normal conditions, and in HEK 293 cells depleted of 3' end processing factors CF Im 68 and CstF-64.","project":"SRP012056"}
{"number_samples":4,"species":"human","abstract":"To better understand the mechanisms of blockage of myeloid differentiation and apoptosis and induction of proliferation by miR-125b, we proceeded to identify miR-125b target genes involved in these pathways. We analyzed the total cellular gene expression pattern by RNA-sequencing of the parental NB4 myeloid cell line and that transiently transfected with miR-125b. We generated four cDNA libraries corresponding to duplicates of miR-125b and control cells. Overall design: Compare the gene expression levels in miR control transfected cells with that in miR-125b transfected NB4 cells. ","project":"SRP012062"}
{"number_samples":4,"species":"human","abstract":"To gain insight into possible processes that require m6A for their function, METTL3 was knocked down (KD) in HepG2 cells by siRNA transfections Overall design: Differential expression analysis of METTL3 KD versus mock-transfected HepG2 cells, in 2 biological replicates","project":"SRP012096"}
{"number_samples":8,"species":"human","abstract":"We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing. Overall design: Identification of m6A modified sequences in HepG2 cells. HepG2 cells were incubated with either IFNg (200ng/ml) or HGF/SF (10 ng/ml) over night. Stress effects were tested in HepG2 cells by either 30 minutes incubation at 43ºC (heat shock) or UV irradiation of 0.04 J/cm2 followed by 4 hours of recovery in normal growing conditions prior to harvesting using Trypsin.","project":"SRP012098"}
{"number_samples":7,"species":"human","abstract":"We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing. Overall design: Identification of m6A modified sequences in HepG2 cells.","project":"SRP012099"}
{"number_samples":2,"species":"human","abstract":"We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing Overall design: Identification of m6A modified sequences in mouse liver and human brain","project":"SRP012100"}
{"number_samples":27,"species":"human","abstract":"Stimulation of estrogen receptor beta in PHPT, genetic changes after 24 and 48h of treatments vs. Control Overall design: Treatment of parathyroid adenomas (4 patients, 4 adenomas) with DPN 24h (4 samples), DPN 48h (4 samples), OHT 24h (4 samples), OHT 48h (4 samples), control 24h (3 samples), control 48h (4 samples). Omission of 1 sample based on low RNA quality.","project":"SRP012167"}
{"number_samples":1,"species":"human","abstract":"The hair of all mammals consists of terminally differentiated cells that undergo a specialized form of apoptosis called cornification. While DNA is destroyed during cornification, the extent to which RNA is lost is unknown. Here we find that multiple types of RNA are incompletely degraded after hair shaft formation in both mouse and human. Notably, mRNAs and short regulatory microRNAs (miRNAs) are stable in the hair as far as 10 cm from the scalp. To better characterize the post-apoptotic RNAs that escape degradation in the hair, we performed sequencing (RNA-seq) on RNA isolated from hair shafts pooled from several individuals. This hair shaft RNA library, which encompasses different hair types, genders, and populations, revealed 7,193 mRNAs, 449 miRNAs and thousands of unannotated transcripts that remain in the post-apoptotic hair. A comparison of the hair shaft RNA library to that of viable keratinocytes revealed surprisingly similar patterns of gene coverage and indicates that degradation of RNA is highly inefficient during apoptosis of hair lineages. The generation of a hair shaft RNA library could be used as months of accumulated transcriptional history useful for retrospective detection of disease, drug response and environmental exposure.","project":"SRP012289"}
{"number_samples":6,"species":"human","abstract":"HDAC inhibitors are thought to regulate gene expression by post-translational modification of histone as well as non-histone proteins. Often studied at single loci, increased histone acetylation is the paradigmatic mechanism of action, however, little is known of the extent of genome-wide changes of the mammalian genome when stimulated by the hydroxamic acids, TSA and SAHA. In primary human vascular endothelial cells we map the chromatin modifications, histone H3 acetylation of lysine 9 and 14 (H3K9/14ac) using chromatin immunoprecipitation (ChIP) coupled with massive parallel sequencing (ChIP-seq). Since acetylation mediated gene expression is often associated with modification of other lysine residues we also examined H3K4me3 and H3K9me3 as well as changes in CpG methylation (CpG-seq). Genome-wide mRNA sequencing indicates the differential expression of about 30% of genes, with almost equal numbers being up- and down- regulated. We observe deacetylation conferred by TSA and SAHA that are associated with decreased gene expression. Histone deacetylation is associated with the loss of p300/CBP binding at gene promoters. This study provides an important framework for HDAC inhibitor function in vascular biology and a comprehensive description of genome-wide deacetylation. Overall design: HAEC mRNA profiles of TSA treated and control samples were generated by deep sequencing, in triplicate, using Illumina GAIIx.","project":"SRP012295"}
{"number_samples":384,"species":"human","abstract":"Background: Tissue regeneration and recovery in the adult body depends on self-renewal and differentiation   of stem and progenitor cells. Mesenchymal stem cells (MSCs) that have the ability to   differentiate into various cell types, have been isolated from the stromal fraction of virtually all   tissues. However, little is known about the true identity of MSCs. MSC populations exhibit great   tissue-, location- and patient-specific variation in gene expression and are heterogeneous in cell   composition.   Methodology/Principal findings: Our aim was to analyze the dynamics of differentiation of two closely related stromal cell types,   adipose tissue-derived MSCs and dermal fibroblasts (FBs) along adipogenic, osteogenic and   chondrogenic lineages using multiplex RNA-seq technology. We found that undifferentiated   donor-matched MSCs and FBs are distinct populations that stay different upon differentiation   into adipocytes, osteoblasts and chondrocytes. The changes in lineage-specific gene expression   occur early in differentiation and persist over time in both MSCs and FBs. Further, MSCs and   FBs exhibit similar dynamics of adipogenic and osteogenic differentiation but different dynamics   of chondrogenic differentiation.  Conclusion: Our findings suggest that stromal stem cells including adipose-derived MSCs and dermal FBs   exploit different molecular mechanisms of differentiation to reach a common cell fate. The early   mechanisms of differentiation are lineage-specific and are similar for adipogenic and osteogenic   differentiation but are distinct for chondrogenic differentiation between MSCs and FBs. Overall design: A total of 91 samples were analyzed by multiplex RNA-seq. Samples represented replicates from two patients, two cell types and three differentiation protocols, as indicated by the sample annotation. 5 barcodes were unused, but the corresponding FASTQ files are included for completeness.","project":"SRP012461"}
{"number_samples":2,"species":"human","abstract":"The Photo-Activatable Ribonucleoside-enhanced CrossLinking and ImmunoPrecipitation (PAR-CLIP) method was recently developed for global identification of RNAs interacting with proteins. The strength of this versatile method results from induction of specific T to C transitions at sites of interaction. However, current analytical tools do not distinguish between non-experimentally and experimentally induced transitions. Furthermore, geometric properties at potential binding sites are not taken into account. To surmount these shortcomings, we developed a two-step algorithm consisting of a non-parametric two-component mixture model and a wavelet-based peak calling procedure. Our algorithm can reduce the number of false positives up to 24% thereby identifying high confidence interaction sites. We successfully employed this approach in conjunction with a modified PAR-CLIP protocol to study the functional role of nuclear MOV10, a putative RNA helicase interacting with Argonaute2 and Polycomb. Our method, available as the R package wavClusteR, is generally applicable to any substitution-based inference problem in genomics. Overall design: The data comprises one MOV10 PAR-CLIP data file and one nuclear RNA-seq file","project":"SRP012463"}
{"number_samples":4,"species":"human","project":"SRP012540"}
{"number_samples":35,"species":"human","abstract":"MicroRNAs (miRNAs), non-coding RNAs regulating gene expression, are frequently aberrantly expressed in human cancers. Next-generation deep sequencing technology enables genome-wide expression profiling of known miRNAs and discovery of novel miRNAs at unprecedented quantitative and qualitative accuracy.   Deep sequencing was performed on 22 fresh frozen clear cell renal cell carcinoma (ccRCC), 11 non-tumoral renal cortex (NRC) samples and 2 ccRCC cell lines (n=35). The 22 ccRCCs patients belonged to 3 prognostic sub-groups, i.e. Those without disease recurrence, with recurrence and with metastatic disease at diagnosis Overall design: Deep sequencing was performed on 22 fresh frozen clear cell renal cell carcinoma (ccRCC), 11 non-tumoral renal cortex (NRC) samples and 2 ccRCC cell lines (n=35). The 22 ccRCCs patients belonged to 3 prognostic sub-groups, i.e. Those without disease recurrence, with recurrence and with metastatic disease at diagnosis.","project":"SRP012546"}
{"number_samples":42,"species":"human","project":"SRP012557"}
{"number_samples":3,"species":"human","abstract":"RNA-seq analysis of U251 glioblastoma transcriptome after Msi1 knockdown.","project":"SRP012568"}
{"number_samples":10,"species":"human","abstract":"Retinal pigment epithelium (RPE) cells can be obtained through in vitro differentiation of both embryonic stem cell (ESC) and induced pluripotent stem cells (iPSC) for cell replacement therapy. We have previously identified 87 signature genes relevant to RPE cell differentiation and function through transcriptome analysis of both human ESC- and iPSC-derived RPE as well as normal fetal RPE. Here, we profiled miRNA expression through small RNA-seq in human ESCs and their RPE derivatives. Much like conclusions drawn from our previous transcriptome analysis, we found that the overall miRNA landscape in RPE is distinct from ESCs and other differentiated somatic tissues. We also profiled miRNA expression during intermediate stages of RPE differentiation and identified unique subsets of miRNAs that are gradually up- or downregulated, suggesting dynamic regulation of these miRNAs is associated with the RPE differentiation process. Indeed, the down-regulation of a subset of miRNAs during RPE differentiation is associated with up-regulation of RPE-specific genes, such as RPE65, which is exclusively expressed in RPE. We conclude that miRNA signatures can be used to classify different degrees of in vitro differentiation of RPE from human pluripotent stem cells. We suggest that RPE-specific miRNAs likely contribute to the functional maturation of RPE in vitro, similar to the regulation of RPE-specific mRNA expression. Overall design: Study miRNA in ESC-derived RPE","project":"SRP012585"}
{"number_samples":12,"species":"human","abstract":"High-throughput RNA sequencing (RNA-Seq) has enabled accurate gene discovery and expression estimation, but robust differential analysis of gene and transcript abundances has proven difficult. We present a new algorithm, implemented in the freely available tool Cuffdiff, which integrates transcript-level expression estimation with a method to control for variability across replicate samples. Cuffdiff robustly identifies differentially regulated isoforms and genes and reveals differential splicing or promoter-preference changes. We demonstrate the accuracy of our approach through differential analysis of lung fibroblasts in response to loss of the developmental transcription factor HOXA1 and uncover a critical role for this gene in the maintenance of adult cells. We show that HOXA1 is required for lung fibroblast and HeLa cell cycle progression, and loss of HOXA1 results in significant expression level changes in thousands of individual transcripts, along with isoform switching events in key regulators of the cell cycle. Overall design: Lung Fibroblasts were transfected with either a HOXA1 directed siRNA pool or a scramble non-targeting siRNA control. RNA was collected 48 hours after transfection and changes in gene expression were assayed for using Agilent microarrays.","project":"SRP012607"}
{"number_samples":3,"species":"human","abstract":"Background & Aims: MicroRNAs have been shown to offer great potential in the diagnosis of cancer. We aimed to identify microRNAs in peripheral blood mononuclear cells (PBMCs) for diagnosing pancreatic cancer (PC). Methods: PBMCs microRNA expression was investigated in three independent cohorts including 352 participants (healthy, benign pancreatic/peripancreatic diseases (BPD), and PC). First, we used sequencing technology to identify differentially expressed microRNAs in 60 PBMCs samples for diagnosing PC. Quantitative reverse-transcriptase polymerase chain reaction assay was then applied to evaluate the expression of selected microRNAs. A logistic regression model was constructed using an independent cohort. Area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. Results: We found that PBMCs miR-27a-3p could efficiently discriminate PC from BPD (AUC=0.840; 95% CI, 0.787 to 0.885; sensitivity=82.2%, specificity=76.7%). A panel composed of PBMCs miR-27a-3p and serum CA19-9 provided a high diagnostic accuracy in differentiating PC from BPD in the clinical setting (AUC=0.886; 95% CI, 0.837 to 0.923; sensitivity=85.3%, specificity=81.6%). The satisfactory diagnostic performance of the panel persisted regardless of disease status (AUCs for tumour-node-metastasis stages?,?, and ? were 0.881, 0.884, and 0.893, respectively).  Conclusion: PBMCs miR-27a-3p could be a potential marker for PC screening. A panel composed of PBMCs miR-27a-3p and serum CA19-9 has considerable clinical value in diagnosing early-stage PC. Therefore, patients who would have otherwise missed the curative treatment window can benefit from optimal therapy. Overall design: Examination of different MicroRNA profiles in 3 types of PBMCs samples","project":"SRP012608"}
{"number_samples":1,"species":"human","abstract":"We report a HSV-1 encoded miRNA present during lytic infection that influences replication efficiency in a tissue culture model.  miRNAs were identified using high-throughput sequencing of HSV-1 infected epithelial cells.  Functional assays were performed by mutating the miRNA coding region and testing grow of the mutant in vitro. Overall design: Construction of a miRNA library and sequencing to reveal novel viral miRNAs from lytically infected cells","project":"SRP012612"}
{"number_samples":2,"species":"human","abstract":"Emerging evidence suggests that chromatin adopts a non-random three-dimensional (3D) topology and that the organization of genes into structural hubs and domains affects their transcriptional status. How chromatin conformation changes in diseases such as cancer is poorly understood. Moreover, how oncogenic transcription factors, which bind to thousands of sites across the genome, influence gene regulation by globally altering the topology of chromatin requires further investigation. To address these questions, we performed unbiased high-resolution mapping of intra- and inter-chromosome interactions upon over-expression of ERG, an oncogenic transcription factor frequently over-expressed in prostate cancer as a result of a gene fusion. By integrating data from genome-wide chromosome conformation capture (Hi-C), ERG binding and gene expression, we demonstrate that oncogenic transcription factor over-expression is associated with global, reproducible and functionally coherent changes in chromatin organization. The results presented here have broader implications, as genomic alterations in other cancer types frequently give rise to aberrant transcription factor expression e.g., EWS-FLI1, c-Myc, n-Myc, PML-RARa. Overall design: We used stable isogenic, normal benign prostate epithelial cell lines (RWPE1) that differ with respect to ERG3 over-expression. To test whether ERG over-expression is associated with global changes in chromatin structure, we performed unbiased chromatin interaction mapping using the Hi-C technique from both RWPE1-ERG and RWPE1-GFP cells, with biological replicates. Successful fill-in and ligation were determined as previously reported by testing for a known interaction between two distant genomic loci located on chromosome 6. The Hi-C libraries were paired-end sequenced using an Illumina GAIIx platform. Following alignment to the human genome (hg18) and filtering to remove un-ligated and self-ligated DNA, we identified intra-chromosomal (or cis-) and inter-chromosomal (or trans-) interactions in both RWPE1 cell lines. To characterize ERG binding and ERG-mediated gene expression changes in these cells, we performed chromatin-immunoprecipitation combined with high-throughput sequencing (ChIP-seq) and RNA sequencing (RNA-seq). ERG was bound to 6,398 sites in RPWE1-ERG cells. Based on paired-end RNA-seq data from both cell lines, 1,266 genes were differentially expressed between RWPE1-ERG and RWPE1-GFP cell lines.","project":"SRP012651"}
{"number_samples":36,"species":"human","abstract":"One of the most fertile applications of next generation sequencing will be in the field of cancer genomics. Here, we report a high-throughput multi-dimensional sequencing study of primary non-small cell lung adenocarcinoma tumors and adjacent normal tissues of 6 never-smoker Korean female patients. Our data encompass results from exome-seq, RNA-seq, small RNA-seq, and MeDIP-seq. We identified and validated novel genetic aberrations including 47 somatic mutations and 20 fusion transcripts. We also characterized gene expression profiles which we sought to integrate with genomic aberrations and epigenetic regulations into functional networks. Importantly, among others the gene network module governing G2/M cell check point emerged as the primary source of disturbance in these patients. In addition, our study strongly suggests that microRNAs make key regulatory inputs into this gene network module. Our study offers a paradigm for integrative genomics analysis and proposes potential target pathways for the control of non-small cell lung adenocarcinoma. Overall design: Study of primary non-small cell lung adenocarcinoma tumors and normal tissues of 6 patients.","project":"SRP012656"}
{"number_samples":3,"species":"human","abstract":"Analysis of mRNA in THP1 (human monocytic leukemia) cell line in order to correlate miRNA activity with target abundance. Overall design: THP1 mRNA profiles were generated in triplicates by deep-sequencing in Illumina HiSeq2000.","project":"SRP012695"}
{"number_samples":20,"species":"human","abstract":"Complex genome rearrangements are frequently observed in cancer, but their impact on tumour molecular biology is largely unknown. Recent studies have identified a new phenomenon involving the simultaneous generation of tens to hundreds of genomic rearrangements, called chromothripsis. To understand the molecular consequences of these events, we sequenced the genomes and transcriptomes of two prostate tumours exhibiting evidence of chromothripsis. We identified several complex fusion transcripts, each containing sequence from three different genes, originating from different parts of the genome. One such poly-gene fusion transcript appeared to be expressed from a chain of small genomic fragments. Furthermore, we detected poly-gene fusion transcripts in the prostate cancer cell line LNCaP, suggesting they may represent a common phenomenon. Finally in one tumour with chromothripsis, we identified multiple mutations in the p53 signaling pathway, expanding on recent work associating aberrant DNA damage response mechanisms with chromothripsis. Overall, our data shows that chromothripsis can manifest as massively rearranged transcriptomes. The implication that multigenic changes can give rise to poly-gene fusion transcripts is potentially of great significance to cancer genetics.","project":"SRP013021"}
{"number_samples":4,"species":"human","abstract":"RNA sequencing technology has been carried out in order to evaluate mRNA expression changes after manipulation of miR-23b in both MCF-7 and MDA-MB-231 breast cancer cell lines Overall design: Our study implicates miR-23b in cytoskeletal remodeling in breast cancer. To evaluate the entire set of genes modulated by miR-23b we performed RNA-seq after ectopic manipulation of this miRNA in breast cancer cell lines. We over-expressed this miRNA in MCF-7 epithelial cancer cell lines and also reduced its activity by stably transfecting MDA-MB-231 mesenchymal-like cancer cell lines with a specific sponge vector. RNA-seq analysis revealed a number of candidate targets of this miRNA.","project":"SRP013022"}
{"number_samples":11,"species":"human","abstract":"We sequenced the total mRNA from infected cells and detected differences in the expression of both host mRNA. We detected a small but significant suppression of T cell activation-related genes at 12 hpi. This suppression persisted and expanded by 24 hpi providing new possible markers of virus-induced T cell cytopathology. By 24 hpi the expression of over 50% of detectable host loci was also altered indicating widespread alteration of host processes including RNA processing, splicing, and transport to an extent not previously reported. In addition next-generation sequencing provided insights into the expression of non-coding RNAs including microRNA host genes. Overall design: We isolated polyadenylated RNA from SUPT1 cells infected with HIV-1 strain LAI at 12 and 24 hours post-infection (3 replicates for each time point). As controls we isolated polyadenylated RNA from mock-infected cells at 12 and 24 hours post-infection (2 replicates at 12 hours post-infection, 3 replicates at 24 hours post-infection).","project":"SRP013224"}
{"number_samples":2,"species":"human","abstract":"RNA-Seq data of two bladder cancer cell lines--5637 and T24. The data can be used to do some analysis , such as expression, alternative splicing, gene fusion.","project":"SRP013232"}
{"number_samples":20,"species":"human","abstract":"We report the immediate effects of estrogen signaling on the transcriptome of breast cancer cells using Global Run-On and sequencing (GRO-seq). We found that estrogen signaling directly regulates a strikingly large fraction of the transcriptome in a rapid, robust, and unexpectedly transient manner. In addition to protein-coding genes, estrogen regulates the distribution and activity of all three RNA polymerases, and virtually every class of non-coding RNA that has been described to date. This data submission covers >95% of mapped reads comprising nearly all transcript classes described.  Reads mapping to intergenic and enhancer transcripts were removed from this data submission and will be reported separately (manuscripts in preparation). Bed files are tab-separated text files in which columns represent: chrom, chromStart (5' End of the read), chromEnd (chromStart+1), name (unused always 'n'), score (the number of mismatches), and strand.  Note that because of the inclusion of reads mapping to the rRNA chromosome, bed files cannot be uploaded to the UCSC genome browser directly.  Instead, use the wiggle files (coming soon!) for this purpose. Overall design: Using GRO-seq over a time course (0, 10, 40, 160 min) of estrogen signaling in ER-alpha positive MCF-7 human breast cancer cells.","project":"SRP013239"}
{"number_samples":2,"species":"human","abstract":"Posttranslational histone modifications play important roles in regulating chromatin structure and function. Histone H2B ubiquitination and deubiquitination have been implicated in transcriptional regulation, but the function of H2B deubiquitination is not well defined, particularly in higher eukaryotes. Here we report the purification of USP49 as a histone H2B specific deubiquitinase and demonstrate that H2B deubiquitination by USP49 is required for efficient co-transcriptional splicing of a large set of exons. USP49 forms a complex with RVB1 and SUG1, and specifically deubiquitinates histone H2B in vitro and in vivo. USP49 knockdown results in small changes in gene expression, but affects the abundance of over 9,000 isoforms.  Exons down-regulated in USP49 knockdown cells show both elevated levels of alternative splicing and a general decrease in splicing efficiency.  Importantly, USP49 is relatively enriched at this set of exons.  USP49 knockdown increased uH2B levels at these exons as well as upstream 3’ and downstream 5’ intronic splicing elements.  Change in H2B ubiquitination level, as modulated by USP49, regulates U1A and U2B association with chromatin and binding to nascent pre-mRNA. Although H3 levels are relatively stable after USP49 depletion, H2B levels at these exons are dramatically increased, suggesting that uH2B may enhance nucleosome stability. Therefore, this study identifies USP49 as a histone H2B specific deubiquitinase and uncovers a critical role for H2B deubiquitination in co-transcriptional pre-mRNA processing events. Overall design: Examination of gene expression in wild type and USP49 knockdown cells [RNA-Seq]","project":"SRP013288"}
{"number_samples":19,"species":"human","abstract":"Preimplantation human embryonic development is a highly dynamic cellular and molecular process, the correct execution of which is critical for successful development to occur. However the mechanism of this event remains largely unknown due to the scarcity of the material, legal restraints and limited techniques to  faithfully analyze small amounts of samples. The development of single cell RNA-seq technology to measure gene expression at single cell levels opens new avenues for investigations into preimplantation human embryonic development.","project":"SRP013299"}
{"number_samples":10,"species":"human","abstract":"Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing. Application to a human embryonic kidney cell line identified close to 800 proteins. Close to one third of these proteins, were neither previously annotated nor could be functionally predicted to bind RNA. Protein occupancy profiling provides a transcriptome-wide catalog of potential cis-regulatory regions on mammalian mRNAs and showed that large stretches in 3' UTRs can be contacted by the mRNA-bound proteome, with numerous putative binding sites in regions harboring disease-associated nucleotide polymorphisms. Our observations indicate the presence of a large number of unexpected mRNA-binders with novel molecular functions participating in combinatorial post-transcriptional gene-expression networks. Overall design: PARCLIP was performed as in Hafner et al. 2010 Cell 141, 129–141, with HEK293 cell lines stably expressing HIS/FLAG/HA-tagged ALKBH5, C17orf85, C22orf28, CAPRIN1, and ZC3H7B. We used 4-thiouridine (4SU) to enhance the crosslink and generated two PAR-CLIP cDNA libraries per protein.","project":"SRP013363"}
{"number_samples":16,"species":"human","abstract":"To obtain an integrated view of gene regulation in response to environmental and endogenous estrogens on a genome-wide scale, we performed ChIP-seq, to identify estrogen receptor 1 (ER) binding sites, and RNA-seq in endometrial cancer cells exposed to bisphenol A (BPA; found in plastics), genistein (GEN; found in soybean), or 17ß-estradiol (E2; an endogenous estrogen).  GEN and BPA treatment induces thousands of ER binding sites and >50 gene expression changes, representing a subset of E2-induced gene regulation changes.  Genes affected by E2 were highly enriched for ribosome-associated proteins; however, GEN and BPA failed to regulate most ribosome-associated proteins and instead enriched for transporters of carboxylic acids.  Treatment-dependent changes in gene expression were associated with treatment-dependent ER binding sites, with the exception that many genes up-regulated by E2 harbored a BPA-induced ER binding site, but failed to show any expression change after BPA treatment.   GEN and BPA exhibited a similar relationship to E2 in the breast cancer line T-47D, where cell type specificity played a much larger role than treatment specificity.  Overall, both environmental estrogens clearly regulate gene expression through ER on a genome-wide scale, although with lower potency resulting in less ER binding sites and less gene expression changes compared to the endogenous estrogen, E2. Overall design: RNA-seq of human cancer cell lines treated with estradiol, bisphenol A, genistein or DMSO (control)","project":"SRP013389"}
{"number_samples":6,"species":"human","abstract":"We generated iPS cells with a synthetic self-replicative RNA that expresses four independent reprogramming factors (OCT4, KLF4, SOX2 and either c-MYC or GLIS1).  We performed whole genome RNA sequencing (RNA-seq) of iPS cell clones, parental BJ and HUES9 ES cell controls. All iPS cell clones analyzed by RNA-seq showed unsupervised hierarchical clustering and expression signatures characteristic of human HUES9 ES cells that were highly divergent from parental human fibroblasts. Overall design: RNA-seq in two OKS-iM iPS clones (generated from OCT4, KLF4, SOX2 and cMYC expressing RNA replicon), two OKS-iG clones (generated from OCT4, KLF4, SOX2 and GLIS1 expressing RNA replicon), HUES9 and BJ cells.","project":"SRP013402"}
{"number_samples":20,"species":"human","abstract":"Adenosine deaminases, RNA specific (ADAR) are proteins that deaminate adenosine to inosine which is then recognized in translation as guanosine.  To study the roles of ADAR proteins in RNA editing and gene regulation, we carried out DNA and RNA sequencing, RNA interference and RNA-immunoprecipitation in human B-cells.  We also characterized the ADAR protein complex by mass spectrometry.  The results uncovered over 60,000 sites where the adenosines (A) are edited to guanosine (G) and several thousand genes whose expression levels are influenced by ADAR.  We also identified more than 100 proteins in the ADAR protein complex; these include splicing factors, heterogeneous ribonucleoproteins and several members of the dynactin protein family.  Our findings show that in human B-cells, ADAR proteins are involved in two independent functions: A-to-G editing and gene expression regulation.  In addition, we showed that other types of RNA-DNA sequence differences are not mediated by ADAR proteins, and thus there are co- or post-transcriptional mechanisms yet to be determined. Overall design: Here we studied human B-cells where ADAR proteins (ADAR1 and ADAR2) are expressed but APOBECs are not.  We identified the sequence differences between DNA and the corresponding RNA in B-cells from two individuals.  Then, we carried out RNA interference, RNA-immunoprecipitation and next generation sequencing to determine the contribution of ADAR proteins in mediating A-to-G editing and other types of RNA-DNA sequence differences.","project":"SRP013450"}
{"number_samples":6,"species":"human","abstract":"Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing. Application to a human embryonic kidney cell line identified close to 800 proteins. Close to one third of these proteins, were neither previously annotated nor could be functionally predicted to bind RNA. Protein occupancy profiling provides a transcriptome-wide catalog of potential cis-regulatory regions on mammalian mRNAs and showed that large stretches in 3' UTRs can be contacted by the mRNA-bound proteome, with numerous putative binding sites in regions harboring disease-associated nucleotide polymorphisms. Our observations indicate the presence of a large number of unexpected mRNA-binders with novel molecular functions participating in combinatorial post-transcriptional gene-expression networks. Overall design: We generated protein occupancy cDNA libraries for two biological replicates. Briefly, we crosslinked 4SU-labeled cells and purified protein-mRNA complexes using oligo(dT)-beads. The precipitate was treated with RNAse I to reduce the protein-crosslinked RNA fragments to a length of about 30-60 nt. To remove non-crosslinked RNA, protein-RNA complexes were precipitated with ammonium sulfate and blotted onto nitrocellulose. The RNA was recovered by Proteinase K treatment, ligated to cloning adapters, and reverse transcribed. The resulting cDNA libraries were PCR-amplified and next-generation sequenced","project":"SRP013456"}
{"number_samples":4,"species":"human","abstract":"Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing. Application to a human embryonic kidney cell line identified close to 800 proteins. Close to one third of these proteins, were neither previously annotated nor could be functionally predicted to bind RNA. Protein occupancy profiling provides a transcriptome-wide catalog of potential cis-regulatory regions on mammalian mRNAs and showed that large stretches in 3' UTRs can be contacted by the mRNA-bound proteome, with numerous putative binding sites in regions harboring disease-associated nucleotide polymorphisms. Our observations indicate the presence of a large number of unexpected mRNA-binders with novel molecular functions participating in combinatorial post-transcriptional gene-expression networks. Overall design: To obtain a more detailed picture of the RNA present in the pooled precipitates of four consecutive oligo(dT)-purifications, we constructed a cDNA library by random priming of 4-thiouridine (4SU)- and 6-thioguanosine (6SG)-labeled RNA derived from UV-irradiated (365 nm)and non-irradiated cells. Digital gene expression analysis of the cDNA library of non-irradiated cells, labeled with 4SU and 6SG, was performed.  To monitor the incorporation of photoreactive nucleotides into mRNA, we isolated 4SU- and 6SG-labeled RNA from the oligo(dT) precipitate of non-crosslinked cells by biotinylation and streptavidin purification (Dolken et al., 2008).","project":"SRP013463"}
{"number_samples":4,"species":"human","abstract":"We obtained 1,367 GATA2 up-regulated and 759 GATA2 down-regulated genes (p< 0.001). Overall design: mRNA profiles of LNCaP cell treated with or without SiGATA2 were generated by deep sequencing  in duplicate using Hi-Seq 2000.","project":"SRP013473"}
{"number_samples":3,"species":"human","abstract":"The TCF7L2 transcription factor is linked to a variety of human diseases, including type 2 diabetes and cancer. One mechanism by which TCF7L2 could influence expression of genes involved in diverse diseases is by binding  to distinct regulatory regions in different tissues. To test this hypothesis, we performed ChIP-seq for TCF7L2 in 6 human cell lines. We identified 116,000 non-redundant TCF7L2 binding sites, with only 1,864  sites common to the 6 cell lines. Using ChIP-seq, we showed that many genomic regions that are marked by both H3K4me1 and H3K27Ac are also bound by TCF7L2, suggesting that TCF7L2 plays a critical role in enhancer  activity. Bioinformatic analysis of the cell type-specific TCF7L2 binding sites revealed enrichment for multiple transcription factors, including HNF4alpha and FOXA2 motifs in HepG2 cells and the GATA3 motif in MCF7 cells. ChIP-seq analysis revealed that TCF7L2 co-localizes with HNF4alpha and FOXA2 in HepG2 cells and with GATA3 in MCF7 cells. Interestingly, in MCF7 cells the TCF7L2 motif is enriched in most TCF7L2 sites but is not enriched in the sites bound by both GATA3 and TCF7L2. This analysis suggested that GATA3 might tether TCF7L2 to the genome at these sites. To test this hypothesis, we depleted GATA3 in MCF7 cells and showed that TCF7L2 binding was lost at a subset of sites. RNA-seq analysis suggested that TCF7L2 represses transcription when tethered to the genome via GATA3. Our studies demonstrate a novel relationship between GATA3 and TCF7L2,  and reveal important insights into TCF7L2-mediated gene regulation. Overall design: RNAseq analysis of MCF7 cells transfected with siCONTROL, siTCF7L2 or siGATA3. ChIP-seq analysis of H3K27ac, H3K4me1, H3K27me3, H3K9me3 in MCF7 cells; H3K4me1 and H3K27ac in HCT116 cells.","project":"SRP013504"}
{"number_samples":3,"species":"human","abstract":"RNA-seq was performed on human PBMCs subjected to 0.004% and 0.04% cigarette smoke condensate to examine changes in gene expression levels.","project":"SRP013619"}
{"number_samples":2,"species":"human","abstract":"RNA-Seq uncovers transcriptomic variations associated with the lethal phenotype conversion on LNCaP progression cell model","project":"SRP013621"}
{"number_samples":2,"species":"human","abstract":"Strand-specific RNA sequencing was done on a normal and a cancer cell line to examine how isoforms are used differently between these two states. Overall design: One PrEC sample, a normal cell line. One LNCaP sample, a cancer cell line.","project":"SRP013724"}
{"number_samples":2,"species":"human","abstract":"Capped analysis of gene expression (CAGE) sequencing was done on a normal and a cancer cell line to examine how promoter usage changes between these two states. Overall design: One PrEC sample, a normal cell line. One LNCaP sample, a cancer cell line.","project":"SRP013725"}
{"number_samples":6,"species":"human","abstract":"Variation in chromatin composition and organization often reflects differences in genome function. Histone variants, for example, replace canonical histones to contribute to regulation of numerous nuclear processes including transcription, DNA repair and chromosome segregation. Here we focus on H2A.Bbd, a rapidly evolving variant found in mammals but not in invertebrates. We report that in human cells, nucleosomes bearing H2A.Bbd form unconventional chromatin structures enriched within actively transcribed genes and characterized by shorter DNA protection and nucleosome spacing. Analysis of transcriptional profiles from cells depleted for H2A.Bbd demonstrated widespread changes in gene expression with a net down-regulation of transcription and disruption of normal mRNA splicing patterns. In particular, we observed changes in exon inclusion rates and increased presence of intronic sequences in mRNA products upon H2A.Bbd depletion. Taken together, our results indicate that H2A.Bbd is involved in formation of a specific chromatin structure that facilitates both transcription and initial mRNA processing. Overall design: RNA-seq was used to examine changes in gene expression upon shRNA-assisted depletion of H2A.Bbd and H2A.Z histone variants in HeLa cells. The cells treated with shRNA with no homology to the human genome were used as control.","project":"SRP013773"}
{"number_samples":24,"species":"human","abstract":"The expansion of the neocortex during mammalian brain evolution results primarily from an increase in neural progenitor cell divisions in its two principal germinal zones during development, the ventricular zone (VZ) and the subventricular zone (SVZ). Using mRNA sequencing, we analyzed the transcriptomes of fetal human and embryonic mouse VZ, SVZ and cortical plate (CP). We describe sets of genes that are up- or down-regulated in each germinal zone. These data suggest that cell adhesion and cell-extracellular matrix (ECM) interactions promote the proliferation and self-renewal of neural progenitors in the developing human neocortex. Notably, relevant ECM-associated genes include distinct sets of collagens, laminins, proteoglycans and integrins, along with specific sets of growth factors and morphogens. Our data establish a basis for identifying novel cell-type markers and open up avenues to unravel the molecular basis of neocortex expansion during evolution. Overall design: Total RNA was isolated from the VZ, inner SVZ (ISVZ), outer SVZ (OSVZ) and CP of six 13-16 weeks post-conception (w.p.c.) human fetuses and from the VZ, SVZ and CP of five E14.5 mouse embryos using laser capture microdissection of Nissl-stained cryosections of dorsolateral telencephalon. Poly A+ RNA was used as template for the preparation of cDNA which were then subjected to single-end 76-bp RNA-Seq.","project":"SRP013825"}
{"number_samples":1,"species":"human","abstract":"This project aims to discover canonical gene fusion events from mixed human tissue cell lines.","project":"SRP013842"}
{"number_samples":4,"species":"human","abstract":"MicroRNAs (miRNAs) are a class of small RNA molecules that regulate expression of specific mRNA targets. They can be released from cells, often encapsulated within extracellular vesicles (EVs), and therefore have the potential to mediate intercellular communication. It has been suggested that certain miRNAs may be selectively exported, although the mechanism has yet to be identified. Manipulation of the miRNA content of EVs will be important for future therapeutic applications. We therefore wished to assess which endogenous miRNAs are enriched in EVs and how effectively an overexpressed miRNA would be exported. Small RNA libraries from HEK293T cells and vesicles before or after transfection with a vector for miR-146 overexpression were analysed by deep sequencing. A subset of miRNAs was found to be enriched in EVs. The global expression data provided by deep sequencing confirms that specific miRNAs are enriched in EVs released by HEK293T cells. Overall design: Cells were transfected with a plasmid to direct overexpression of miR-146a. Extracellular vesicles were isolated by ultracentrifugation from untreated and transfected cells. RNA was isolated from one sample each of untreated and transfected cells and vesicles.Small RNA libraries were prepared for sequencing.","project":"SRP013912"}
{"number_samples":3,"species":"human","abstract":"In this study we investigated the gene expression profiling in three HCC cell lines with different organ-tropism.The parent cell line has low metastasis ability in nude mice model. Subclone 1 has higher metastasis ability specific to lung, and subclone 2 has dual metastasis ability to lung and celiac lymph node. We aimed to explore differentially expressed genes involved in process of HCC metastasis and organ-specific metastasis, and identify their biological functions. Overall design: Examination of different gene expression among the 3 cell types.","project":"SRP013935"}
{"number_samples":3,"species":"human","abstract":"In this study we investigated the small RNAs expression profiling of three HCC cell lines with different organ-tropism.The parent cell line has low mestastasis ability in nude mice model. Subclone 1 has higher metastasis ability specific to lung, and subclone 2 has dual metastasis ability to lung and celiac lymph node. We aimed to explore differentially expressed small RNAs involved in process of HCC metastasis and organ-specific metastasis, and identify their biological functions. Overall design: Examination of  different small RNAs expression among the 3 cell types.","project":"SRP013981"}
{"number_samples":2,"species":"human","abstract":"The CD40 gene, an important immune regulatory gene, is also expressed and functional on non-myeloid derived cells, many of which are targets for tissue specific autoimmune diseases, including d thyroid follicular cells in Graves’ disease (GD). Whether target tissue CD40 expression plays a role in autoimmune disease etiology has yet to be determined. Here we show for the first time, that target-tissue over-expression of CD40 plays a key role in the etiology of autoimmunity. Using a murine model of GD, we demonstrated that thyroidal CD40 over-expression augmented the production of thyroid specific antibodies, resulting in more severe experimental autoimmune Graves’ disease (EAGD), whereas deletion of thyroidal CD40 suppressed disease. Using transcriptome and immune-pathway analyses we showed that in both EAGD mouse thyroids and human primary thyrocytes, CD40 mediates this effect by activating downstream cytokines and chemokines, most notably IL-6. To translate these findings into therapy, we blocked IL-6 during EAGD induction in the setting of thyroidal CD40 over-expression, and showed decreased levels of TSHR stimulating antibodies and frequency of disease. We conclude that target tissue over-expression of CD40 plays a key role in the etiology of organ specific autoimmune disease. Overall design: CD40 in Thyroid Autoimmunity: 1) Incubation of human thyroid cells with G28.5, a CD40 stimulating antibody, and purification of RNA, conversion to cDNA, measurement of mRNA expression using RNAseq. 2) Removal of thyroid tissues from CD40 over-expressing transgenic mice and wild type mice, purification of RNA, conversion to cDNA measurement of mRNA expression using RNAseq.","project":"SRP013984"}
{"number_samples":2,"species":"human","abstract":"In our study we applied a genome-wide DNA methylation analysis approach, MethylCap-seq, to map the differentially methylated regions in 24 tumor and matched normal colon samples. In total, 2687 frequently hypermethylated and 468 frequently hypomethylated regions were identified, which include potential biomarkers for CRC diagnosis. Hypermethylation in the tumor samples was enriched at CpG islands and gene promoters, while hypomethylation was distributed throughout the genome. Using epigenetic data from human embryonic stem cells, we show that frequent differentially methylated regions (DMRs) coincide with bivalent loci in human embryonic stem cells. DNA methylation is commonly thought to lead to cancer gene related silencing, however integration of publically available expression analysis shows that 75% of the frequently hypermethylation genes were most likely already lowly or not expressed in normal tissue. Collectively, our study provides genome-wide DNA methylation maps of colon cancer, comprehensive lists of DMRs, and gives further clues on the role of aberrant DNA methylation in CRC formation. Overall design: To investigate DNA methylation in CRC in a genome-wide unbiased fashion, we applied MethylCap-seq. This method involves capture of methylated DNA using the MBD domain of MeCP2, and subsequent next-generation Illumina sequencing of eluted DNA. In addition, we compared MethylCap with RNA-seq and ChIP-seq profiles of H3K4me3 and H3K27me3 for the colon cancer tumor cell line HCT116 (HCT116 WT) and the cell line of HCT116 with DNMT1 and DNMT3b knockout (HCT116 DKO).","project":"SRP013985"}
{"number_samples":4,"species":"human","abstract":"Numerous studies over the past decade have elucidated a substantial set of long intergenic noncoding RNAs (lincRNAs). It has since become clear that lincRNAs constitute an important layer of genome regulation across a wide spectrum of species. Yet, the factors governing their evolution and origins remain relatively unexplored. One possible factor that may have shaped lincRNA biology are transposable elements (TEs). Here we set out to comprehensively characterize the TE content of lincRNAs relative to genomic averages and protein coding transcripts. Our analysis of the TE composition across 9241 human lincRNAs revealed that, in sharp contrast to protein coding genes, a striking majority (83%) of lincRNAs contain a TE, and TEs comprise 42% of lincRNA transcript sequences. LincRNA TE composition varies significantly from genomic averages, being depleted of LI and Alu elements and enriched for a broad class of endogenous retroviruses (ERVs). Furthermore, specific TE families occur in biased positions and orientations within lincRNAs, particularly at their transcription start sites, suggesting a role in the origin of those lincRNAs. Finally, we find that TEs can drive gene expression regulation of lincRNAs—we observed a dramatic correlation between lincRNAs containing HERVH elements and almost exclusive expression in pluripotent cells. Conversely, those lincRNAs that are devoid of TEs are more highly expressed in testis. Collectively, TEs pervade lincRNAs and have shaped lincRNA evolution and function via bestowing tissue-specific expression from donated transcriptional regulatory signals. Overall design: We extracted profiled the transcriptome expression polyadenylated mRNA-Seq.  We then used these to reconstruct the transcriptome using de-novo assemblers and identify long non coding RNAs and their expression.","project":"SRP013999"}
{"number_samples":4,"species":"human","abstract":"The Drosha-DGCR8 complex (Microprocessor) is required for microRNA (miRNA) biogenesis. DGCR8 contains two double-stranded RNA binding motifs that recognize the RNA substrate, whereas Drosha functions as the endonuclease. We have used high-throughput sequencing of RNAs isolated by crosslinking immunoprecipitation (HITS-CLIP) to identify endogenous RNA targets of DGCR8 in mammalian cells. Unexpectedly, miRNAs were not the most abundant targets. DGCR8-bound RNAs comprised several hundred mRNAs as well as snoRNAs and long non-coding RNAs. We found that DGCR8 together with Drosha controls the abundance of several mRNAs, as well as long non-coding RNAs, such as MALAT-1. By contrast, the DGCR8-mediated cleavage of snoRNAs is independent of Drosha, suggesting the involvement of DGCR8 in cellular complexes with other endonucleases. Interestingly, binding of DGCR8 to cassette exons, acts as a novel mechanism to regulate the relative abundance of alternatively spliced isoforms.  Collectively, these data provide new insights in the complex role of DGCR8 in controlling the fate of several classes of RNAs. Overall design: Comparison of RNAs associated to both endogenous (D8) and overexpressed (T7) DGCR8 in HEK293T cells","project":"SRP014009"}
{"number_samples":20,"species":"human","abstract":"We report data obtaibed from high-throughput sequencing of small RNAs in 20 samples of follicular thyroid tumors. We analyzed a total of 4.7±1.5million reads per sample with 3 different pipelines. The main goal was to evaluate the usefulness of next generation sequencing in small RNA profiling and the concordance of its results with microarrays and qPCR. Additionally we verified published follicular thyroid tumor biomarkers in the set of our samples. Overall design: Small RNA expression profiling  with High Throughput Sequencing of 20 thyroid tumor samples, performed on an Illumina HiScan-SQ.","project":"SRP014020"}
{"number_samples":6,"species":"human","abstract":"Molecular programs that mediate normal cell differentiation are required for oncogenesis and tumor cell survival in certain cancers. How cell-lineage-restricted genes specifically influence metastasis is poorly defined. In lung cancers, we uncovered a transcriptional program that is preferentially associated with distal airway epithelial differentiation and lung adenocarcinoma (ADC) progression. This program is regulated in part by the lineage transcription factors GATA6 and HOPX. These factors can cooperatively limit the metastatic competence of ADC cells, by modulating overlapping alveolar differentiation and invasogenic target genes. Thus, GATA6 and HOPX are critical nodes in a lineage-selective pathway that directly links effectors of airway epithelial specification to the inhibition of metastasis in the lung ADC subtype. Overall design: mRNA profiles of human lung Adenocarcinoma PC9 cell lines infected with lentivirus harboring shRNA of control and shRNA of both GATA6 and HOPX were generated by deep sequencing, in triplicate, using Illumina HiSeq2000.","project":"SRP014027"}
{"number_samples":1,"species":"human","abstract":"Small RNAs regulate the genetic networks through a ribonucleoprotein complex called the RNA induced silencing complexes (RISC), which in mammals contains at its center one of four Argonaute proteins (Ago1-4). A key regulatory event in the RNAi and miRNA pathways is Ago loading, where double stranded small RNA duplexes are incorporated into RISC (pre-RISC) and then become single stranded (mature-RISC), a process that is not well understood. The Agos contain an evolutionary conserved PAZ (Piwi/Argonaute/Zwille) domain whose primary function is to bind the 3’-end of small RNAs. We created multiple Paz domain disrupted Ago mutant proteins and studied their biochemical properties and biological functionality in cells. We found that the Paz domain is dispensable for Ago loading of slicing-competent RISC. In contrast, in the absence of slicer activity or slicer substrate duplex RNAs, Paz-disrupted Agos bound duplex siRNAs but were unable to unwind/eject the passenger strand and form functional RISC complexes. We have discovered that the highly conserved Paz domain plays an important role in RISC activation, providing new mechanistic insights into how miRNAs regulate genes, as well as new insights for future design of miRNA and RNAi-based therapeutics. Overall design: Various Argonautes associated small RNA profiles were generated by deep sequencing the Agos-IP samples in HEK293 Cells transfected with corresponding Argonaute.","project":"SRP014133"}
{"number_samples":15,"species":"human","abstract":"To comprehensively characterize microRNA (miRNA) expression in breast cancer, we performed the first extensive next-generation sequencing expression analysis of this disease. We sequenced small RNA from tumors with paired samples of normal and tumor-adjacent breast tissue. Our results indicate that tumor identity is achieved mainly by variation in the expression levels of a common set of miRNAs rather than by tissue-specific expression. We also report 361 new, well-supported miRNA precursors. Nearly two-thirds of these new genes were detected in other human tissues and 49% of the miRNAs were found associated with Ago2 in MCF7 cells. Ten percent of the new miRNAs are located in regions with high-level genomic amplifications in breast cancer. A new miRNA is encoded within the ERBB2/Her2 gene and amplification of this gene leads to overexpression of the new miRNA, indicating that this potent oncogene and important clinical marker may have two different biological functions. In summary, our work substantially expands the number of known miRNAs and highlights the complexity of small RNA expression in breast cancer. Overall design: Sequencing of approximately 18-35 nt small RNAs from paired samples of normal, tumor and tumor-adjacent tissue for five breast cancer patients","project":"SRP014142"}
{"number_samples":15,"species":"human","abstract":"Studies in model organisms suggest that aged cells can be functionally rejuvenated, but whether this concept applies to human skin is unclear. Here we apply deep sequencing of RNA 3' ends (\"3-seq\") to discover the gene expression program associated with human photoaging and intrinsic skin aging (collectively termed \"skin aging\") and the impact of broadband light (BBL) treatment. We find that skin aging was associated with the significantly altered expression level of 2,265 coding and noncoding RNAs, of which 1,293 became \"rejuvenated\" after BBL treatment, i.e. more similar in expression level of youthful skin. Rejuvenated genes (RGs) included several known key regulators of organismal longevity and their proximal long non-coding RNAs. Skin aging is not associated with systematic changes in 3' end mRNA processing. Hence, BBL treatment can restore the gene expression pattern of photoaged and intrinsically aged human skin to resemble young skin. In addition, our data reveals a novel set of targets that may lead to new insights into the human skin aging process. Overall design: Examination of broadband light treated and untreated human skin transcriptomes of 5 women aged 50 years or more. They were compared to the skin transcriptomes of 5 young women aged 30 years or less.","project":"SRP014146"}
{"number_samples":2,"species":"human","abstract":"Purpose: Identical predicted small interfering RNA (siRNA) sequences targeting Apolipoprotein B100 (siApoB) were embedded in shRNA (shApoB) or miRNA (miApoB) scaffolds and a direct compariso  of the possible aspecific off-target effects in vivo was performed. Next generation sequencing (NGS) of small RNAs originating from shApoB- or miApoB-transfected cells revealed substantial differences in processing, resulting in different siApoB length, 5’ and 3’ cleavage sites and abundance of the guide or passenger strands. Methods [1]: Total liver RNA sequencing libraries for the Illumina sequencing platform were generated using high-quality total RNA as input and the Illumina TrueSeq RNA v2 Sample preparation kit according to the manufacturer’s protocol. Each read file (sample), in the FASTQ format, was individually aligned against the mouse reference genome (15 May 2012 NCBI build 38.1) using CLC Bio-Genomic Workbench and the expression abundance for each gene (RPKM) was calculated according to Montazavi et al. Result [1]s: Based on our previous observations that shApoB and miApoB are differentially processed and that miApoB has a different passenger, we checked for possible aspecific off-target effects in vivo. NGS liver transcriptome analysis was performed 8 weeks p.i. for animals injected with 1x1011 gc AAV encoding shScr, shApoB, miScr and miApoB. We investigated whether shApoB and miApoB processing differences translate into differences in gene expression in injected animals. A total number of 266 genes were significantly changing (p <0,05) in miApoB-injected mice compared to miScr. Additionally 106 genes were found to be significantly up or down-regulated in the shApoB mice compared to shScr. Off-target predictions using Smith-Waterman algorithm for the most abundant guide and passenger strand variants were performed to investigate if any of the observed changes results from aspecific interactions. None of the changing genes had predicted targets for the guide or passenger strands of shApoB or miApoB. Conclusions [1]: An important observation is that none of the changes in gene expression in AAV-miApoB and AAV-shApoB can be explained by possible aspecific down-regulation of non-target transcripts. Methods [2]: For NGS analysis cells were transfected with 4 µg shApoB- or miApoB-expression plasmids using Lipofectamine 2000 reagent and total RNA was isolated from cells 48 hr post-transfection using Trizol (Invitrogen, Carlsbad, CA) according to the manufacturer’s protocol. Total RNA sequencing libraries for the Illumina sequencing platform were generated using high-quality total RNA as input and the Illumina TrueSeq RNA v2 Sample preparation kit according to the manufacturer’s protocol. The NGS small RNA raw data set was analyzed using the CLC_bio genomic workbench (CLC Bio, Aarhus, Denmark). The obtained reads were adaptor-trimmed, which decreased the average read size from ~36bp to ~25bp. The custom adapter sequenced used for trimming all the bases extending 5’ was: GTGACTGGAGTTCC-TTGGCACCCGAGAATTCCA. All reads containing ambiguity N symbols, reads shorter than 15 nt, longer than 55 nt in length and reads represented less than 10 times were discarded. Next, both data sets from shApoB and miApoB samples were grouped based on the match to the reference sequence and the obtained unique small RNAs were aligned to the sequence of pre-miApoB: GATCCTGGAGGCTTGC-TGAAGGCTGTATGCTGATGGACAGGTCAATCAATCTTGTTTTGGCCACTGACTGACAAGATTGAGACCTGTCCATCAGGACACAAGGCCTGTTACTAGCACTCACATGGAACAAATGGCCCAGATCTGGCCGCAG or shApoB: GATCCCCGATTGATTGACCTGTCCATTTCAAGAGAATGGACAGGTCAATCAATC-TTTTTCAGCTT sequence, respectively. To relatively represent the expression counts for the small RNAs obtained in the experiment, reads per million (rpm) or percentage of reads based on the total number of reads matching the reference shApoB or miApoB sequence were calculated Results [2]: The small RNAs were aligned against their reference sequence resulting in 541.939 reads matching shApoB and 1.525.211 reads matching miApoB (Fig S1 and S2). Analysis of the length distribution of the reads indicated that siApoB guide strand originating from shApoB ranged between 19 and 23 nt, with the most abundant one being 21 nt-long. Surprisingly, siApoB guide from miApoB scaffold ranged from 23 to 25 nt with the 24 nt-long strand being the predominant variant. This finding was rather unexpected considering that the predicted guide strand of siApoB was 21 nt long for both shApoB and miApoB scaffolds. Analysis of the processed 5’ ends of the siApoB guide strand indicated that most of the reads matched position +1 relative to the predicted cleavage site for shApoB, while all the reads matched position 0 for miApoB. The 3’ ends of the siApoB guide strand had a more heterogeneous pattern and ranged from -1 to +3 for shApoB and +1 to +4 for miApoB.   Next, we looked at the sequence distribution and percentage of reads for both the guide and passenger siApoB strands originating from the shApoB and miApoB scaffolds. A substantial difference between the two is that the guide from shApoB is in the 3’ arm and hence Dicer or other endonuclease  defines the cleavage position while the guide is present in the 5’ arm of miApoB, where Drosha defines the cleavage. Thus, defining the length and exact cleavage position for the guide and passenger strands is very important since even single nucleotide differences may result in significant changes in the predicted targets of the siRNAs. Moreover, the passenger siRNA* strand, if not degraded efficiently, may bind to unanticipated targets and cause off-target effects. As expected, 44.4% of the reads originating from shApoB matched the siApoB guide strand but processing was shifted at position +1 (Fig. 2d, upper panel). Surprisingly only 12.1% of the reads matched perfectly the predicted siApoB guide strand of 21 nt and starting at position 0. The reads matching the passenger siApoB* strand were represented in much lower percentage with the predominant one being only 5.3%.   Analysis of the guide strand from the siApoB reads originating from the miApoB scaffold indicated that they all started at the predicted cleavage site. Surprisingly, the predominant, 22.3% of reads were 24 nt-long. Furthermore 5.1% reads were 23 nt- and 1.9% were 25 nt-long. A substantial number of 62% of the reads was found matching the passenger siApoB* strand and ranged between 20 and 22 nt in length. In conclusion, both shApoB and miApoB scaffolds did not yield the predicted siApoB guide or siApoB* passenger sequences after processing from the cellular RNAi machinery. The guide from miApoB was cleaved much more precisely by Drosha at its 5’ end compared to shApoB that gave more heterogeneous pools of sequences following processing. Conclusions[2]: An unexpected discovery in the current study was that siRNA processing by the cellular RNAi machinery did not follow the generally accepted and described cleavage sites for both molecules shApoB and miApoB siApoB guide strand originating from the shApoB scaffold was more heterogeneous in cleavage sites and length compared to the product originating from the miApoB scaffold. Most likely, differential cleavage mechanism defined the heterogeneity in the guide and passenger strands. Additionally, shRNAs with 19 bp stem or less are not necessarily recognized by Dicer and maybe be processed differently. The heterogeneity seen with the shApoB can also be explained by the potential for 2 or 3 uridines to be added following termination of pol III transcription. The main pool of guide sequences originating from miApoB was 24 nt-long although we used the scaffold of cellular pri-mir-155 that produces a 23 nt mature miRNA. However, a 24 nt-long siApoB sequence did not compromise efficacy because when placed in the miApoB scaffold, the ApoB target sequence was extended at the 3’ end with 4 nt until the loop.   Another important observation is that the siApoB* Overall design: Liver mRNA profiles of C57BL/6 9 weeks after intravenous AAV injection of 1x1011 gc per animal (~4x1012 gc/kg) of AAV-shRNA, AAV-miRNA or PBS via the tail vein. Next Generation Sequencing on Illumina platform","project":"SRP014157"}
{"number_samples":4,"species":"human","abstract":"In this study we investigated the genome wide DNA binding profile of ZNF143 and ICN1 in human and murine cells. We also analyzed the expression profile in human cells after overexpression or knockdown of ZNF143. Overall design: To identify ZNF143 targets we performed ChIP-seq on 4 human cells lines and 3 murine cell types. We also identified, by ChIP-seq, ICN1 binding sites in HPB-ALL cells. To identify directly regulated genes by ZNF143, we analyzed, by RNA-seq, the expression profile after knockdown and overexpression of the transcription factor.","project":"SRP014190"}
{"number_samples":21,"species":"human","abstract":"We have previously shown that some gefitinib insensitive head and neck squamous cell carcinoma (HNSCC) cell lines exhibit dominant autocrine fibroblast growth factor receptor (FGFR) signaling. Herein, we deployed a whole genome loss-of-function screen to identify genes whose knockdown potentiated the inhibitory effect of the FGFR inhibitor, AZ12908010, in HNSCC cell lines. Three HNSCC cell lines expressing a genome-wide shRNA library were treated with AZ8010 and the abundance of shRNA sequences was assessed by deep sequencing. Synthetic lethal hits were validated through use of specific inhibitors and independent shRNAs. We found that multiple alternate receptors provided protection from FGFR inhibition, including the receptor tyrosine kinases (RTKs), epidermal growth factor receptor 2 (ERBB2) and hepatocyte growth factor receptor (MET).  We showed that specific knockdown of either ERBB2 or MET in combination with FGFR inhibition led to increased inhibition of growth relative to FGFR tyrosine kinase inhibitor (TKI) treatment alone. These results were confirmed using specific small molecule inhibitors of either ERBB family members or MET. Moreover, the combination of FGFR, MET and ERBB family inhibitors showed the largest inhibition of growth as compared to the double combinations. These results reveal a role for alternate RTKs in maintaining pro-growth and survival signaling in HNSCC cells in the setting of FGFR inhibition. Thus, improved therapies for HNSCC patients could involve rationally designed combinations of TKIs targeting FGFR, ERBB family members and MET. Overall design: Using a genome-wide shRNA library in combination with deep sequencing, we screened for gene targets that were synthetic lethal with the FGFR inhibitor, AZ12908010 in HNSCC cells. Three HNSCC cell lines were screened in triplicate and the abundance of shRNA sequences in drug treated cells was compared to control treated cells.","project":"SRP014213"}
{"number_samples":104,"species":"human","project":"SRP014320"}
{"number_samples":88,"species":"human","abstract":"We assessed Smart-Seq, a new single-cell RNA-Seq library preparation method, on a variety of mouse and human RNA samples or cells. Overall design: We generated RNA-Seq libraries for dilution series of MAQC reference RNA and mouse brain RNA to assess technical reproducibility, and for a variety of individual cells including putative circulating tumour cells.","project":"SRP014428"}
{"number_samples":2,"species":"human","abstract":"To investigate the expression characteristic of miRNAs during the development of Down syndrome (DS) fetuses and to identify whether another miRNA gene resides in the Hsa21, we employed high-throughput Solexa sequencing technology to comprehensively characterize the miRNA expression profiles of both DS and normal fetal cord blood mononuclear cells (CBMCs). In total, 200 of 395 identified miRNAs were significantly differentially expressed (fold change > 2.0 and P-value < 0.001) and 2 of 181 candidate novel miRNAs were identified as residing within the \"Down syndrome critical region\" of human chromosome 21 (chr21q22.2-22.3). Additionally, 7 of 14 Hsa21-derived miRNAs genes were detected that three miRNAs (hsa-miR-802, miR-3648, miR-3687) were up-regulated more than 50% and four miRNAs (hsa-miR-99a, let-7c, miR-125b-2, miR-155) were down-regulated in the DS fetal CBMCs compared with the control. Bioinformatics analyses revealed that abnormally expressed miRNAs were major associated with the regulation of transcription, gene expression, cellular biosynthetic process, macromolecule biosynthetic process and nucleic acid metabolic process. The data obtained in our study provides a considerable insight into understanding the expression characteristic of miRNAs in the DS fetal hemopoietic system and the differentially expressed miRNAs may be involved in the hemopoietic abnormalities and the immune defects of DS fetus and newborns. Overall design: A total of 6 DS and 6 matched control fetal cord blood samples (18-22 weeks of gestation) were collected. Three DS and 3 control cord blood samples were combined to form pooled DS and control cord blood samples, respectively, for small RNA library construction and Solexa sequencing. The remaining samples were used as the validation set to confirm the miRNA differential expression patterns by qRT-PCR.","project":"SRP014443"}
{"number_samples":3,"species":"human","abstract":"The correlation of the RNA profiles obtained by microarray analysis was compared with that obtained from RNA-Seq by using reduced complexity sperm datasets. This resolved as a series of discordant probes. The extent of discordancy among other datasets was then determined. Overall design: A correlative study between probe’s signal intensity from Illumina bead arrays with its transcript level detected by next generation sequencing technique was performed. RNAs from sperm and testis samples were applied","project":"SRP014487"}
{"number_samples":1,"species":"human","abstract":"Formation of the blood from self-renewing hematopoietic stem cells to terminal lineages necessarily involves epigenomic modifications of the genome to control regulator and signature gene expression. By analysing the global expression profiles of hematopoietic stem cells (HSCs), in vivo differentiated CD4+ T cells and CD19+ B cells as well as in vitro differentiated erythrocyte precursor cells, we identified hundreds of transcripts showing type-specific expression in these cell types.  To understand the epigenomic changes related to tissue-specific expression during HSC differentiation, we examined the genome-wide distribution of H3K4me1, H3K4me3, H3K27me1, H3K27me3, histone variant H2A.Z, chromatin remodeler BRG1, and RNA Polymerase II in the same four cell types, as well as embryonic stem cells.  Analysis of these datasets revealed that numerous key differentiation genes are primed for expression by Brg1 and Pol II binding, as well as bivalent modifications in the HSCs prior to their expression in downstream differentiated cell types. Much of this bivalency in HSC is retained from embryonic stem cells.  After differentiation, these modified regions resolve to active chromatin modification configuration in the specific lineage, while in parallel differentiated lineages the bivalent modification remains; Pol II and Brg1 are lost in closer lineages but bivalency resolves to silent monovalency in more distant lineages. Correlation of tissue-specific gene expression with the epigenomic changes predicts tens of thousands of potential common enhancers and tissue-specific enhancers, which may critically contribute to the expression patterns. We provide a valuable dataset for further understanding the regulatory mechanisms of differentiation and function of blood lineages. Overall design: RNA-Seq: This submission comprises RNA-Seq profiling of in vivo differentiated human B cells and hematopoietic stem cells. Re-analyzed data for three cell types: The HSCs were previously uploaded as GSM651554 (SRX037948), but processed differently for this upload. The erythrocyte precursors and T cells have also been previously uploaded as GSM651555 (SRX037949) and GSM406414 (SRX005317), respectively.  They were treated as in GSM651554, but processed as here.  The processed files generated by our re-analysis are linked below as supplementary files.","project":"SRP014540"}
{"number_samples":6,"species":"human","abstract":"So far, the annotation of translation initiation sites (TISs) has been based mostly upon bioinformatics rather than experimental evidence. We adapted ribosomal footprinting to puromycin-treated cells to generate a transcriptome-wide map of TISs in a human monocytic cell line. A neural network was trained on the ribosomal footprints at previously annotated AUG translation initiation codons (TICs), and used for the ab initio prediction of TISs in 5062 transcripts with sufficient sequence coverage. Functional interpretation suggested 2994 novel upstream open reading frames (uORFs) in the 5´ UTR (924 AUG, 2070 near-cognate codons), 1406 uORFs overlapping with the coding sequence (116 AUG, 1290 near-cognate) and 546 N-terminal protein extensions (6 AUG, 540 near-cognate). The TIS detection method was validated on the basis of previously published alternative TISs and uORFs. On average, TICs in newly annotated TISs were significantly more conserved among primates than control codons, both for AUGs (p<10-10) and near-cognate codons (p=3.8×10-3). The derived transcriptome-wide map of novel candidate TISs will help to explain how human proteome diversity is influenced by alternative translation initiation and regulation. Overall design: Examination of translational initiation in human cell lines using ribosomal footprinting","project":"SRP014542"}
{"number_samples":1,"species":"human","abstract":"Site identification in high-throughput RNA-protein interaction data","project":"SRP014565"}
{"number_samples":10,"species":"human","abstract":"Site identification in high-throughput RNA-protein interaction data (Ago2_HITS-CLIP)","project":"SRP014566"}
{"number_samples":53,"species":"human","abstract":"Gastric Cancer Whole Genome Sequencing.  Abstract : Whole genome and transcriptome sequencing of gastric cancer tissues and cell lines","project":"SRP014574"}
{"number_samples":6,"species":"human","abstract":"Fragile-X Syndrome (FXS) is a multi-organ disease leading to mental retardation, macro-orchidism in males, and premature ovarian insufficiency in female carriers. FXS is also a prominent monogenic disease associated with autism spectrum disorders (ASD). FXS is typically caused by the loss of FRAGILE X-MENTAL RETARDATION 1 (FMR1) expression, which encodes for the RNA-binding protein (RBP), FMR1 (or FMRP). We report the discovery of the RNA recognition elements (RREs), binding sites, and mRNA targets for wild-type and I304N mutant FMRP isoforms as well as its paralogs, FXR1 and FXR2. RRE frequency, ratio, and distribution determine target mRNA association with FMRP. Among highly-enriched targets, we identified many genes involved in ASD and demonstrate that FMRP can affect their protein levels in cell culture, mice, and human brain. Unexpectedly, we discovered that these targets are also dysregulated in Fmr1-/- mouse ovaries, showing signs of premature follicular overdevelopment. These results indicate that FMRP targets shared signaling pathways across different cellular contexts. As it is become increasingly appreciated that signaling pathways are important to FXS and ASD, our results here provide an invaluable molecular guide towards the pursuit of novel therapeutic targets for these devastating neurological disorders. Overall design: PAR-CLIP profiling for wild-type and I304N mutant FMRP isoforms as well as paralogs, FXR1 and FXR2.","project":"SRP014591"}
{"number_samples":17,"species":"human","abstract":"Hemogenic endothelium (HE) is the source of HSCs in the developing embryo. In this study we have identified the hemogenic endothelial progenitors and their precursors originating from differentiated H1 cells on OP9 stromal cells. Overall design: RNA-seq of  hemogenic endothelial progenitors and their precursors originating from differentiated H1 cells on OP9 stromal cells.","project":"SRP014620"}
{"number_samples":6,"species":"human","abstract":"While Argonaute (AGO) proteins play a major role in transcriptional gene silencing (TGS) in many organisms, their role in the nucleus of somatic mammalian cells remains elusive. Here, we have purified AGO1 and AGO2 chromatin-embedded complexes, and found these proteins associated with previously described partners, but also with chromatin modifiers and, rather unexpectedly, with different splicing factors. Using the CD44 gene as a model for alternative splicing, we show that both AGO1 and AGO2 are required for Protein Kinase C (PKC)-dependent variant exon inclusion. AGO proteins facilitate the spliceosome recruitment and modulate the elongation rate of RNA polymerase II (RNAPII). The recruitment of AGO proteins to CD44 transcribed region is dependent on both the endonuclease Dicer and the chromodomain-containing protein HP1g, and results in locally increased levels of histone H3 lysine 9 (H3K9) methylation on variant exons. Genome wide analysis of splicing in either AGO2 or Dicer null cells showed that the two proteins have similar effects on many splicing events. Finally, sRNAs associated with nuclear AGO2 are mostly in sense orientation relative to protein-coding genes, supporting a role for intragenic antisense non-coding RNAs in the recruitment AGO and splicing factors. Together, our data demonstrate for the first time that the endogenous RNAi pathway is involved in alternative splicing decisions, unravelling a new model in which AGO proteins couple RNAPII elongation and chromatin modification. Overall design: Deep sequencing of small RNAs (approx. 15-80 nucleotides) bound to either cytoplasmic or chromatin-associated AGO2 complex.","project":"SRP014624"}
{"number_samples":2,"species":"human","abstract":"Purpose: Compare the transcriptome of homogeneous XIST+ and XIST- hES cell populations. Methods: We isolated homogeneous XIST+ and XIST- cell populations. The XIST+ cells correspond to cells with a XIST cloud and one ATRX pinpoint. The XIST- cells correspond to cells with no XIST cloud and one ATRX pinpoint. Results: We took advantage of the clonal pattern of X-chromosome inactivation in H9 cells and analyzed the data in an allelic manner.  By comparing the RNA-Seq data with known H9 SNPs, we identified genomic positions which were relaxed from XCI in XIST- cells compared to XIST+ cells. Conclusions: Genic as well as unannotated transcripts are massively relaxed from XCI in H9 cells when XIST expression is lost, however, this reactivation is only partial and a large region around the centromere is protected from relaxation of silencing. Overall design: Total RNA (rRNA depleted) profiles of XIST+ and XIST- human embryonic stem cells","project":"SRP014626"}
{"number_samples":5,"species":"human","abstract":"Understanding translational control in gene expression relies on precise and comprehensive determination of translation initiation sites (TIS) across the entire transcriptome. The recently developed ribosome profiling technique enables global translation analysis, providing a wealth of information about both the position and density of ribosomes on mRNAs. Here we present an approach (global translation initiation sequencing, GTI-seq), by applying in parallel ribosome E site translation inhibitors lactimidomycin (LTM) and cycloheximide (CHX), to achieve simultaneous detection of both initiation and elongation events on a genome-wide scale.","project":"SRP014629"}
{"number_samples":2,"species":"human","abstract":"As genome-scale DNA methylation sequencing technologies have improved it has become apparent that tissue-specific methylation can occur not only at promoters, enhancers, and CpG islands but also over larger genomic regions. In most human tissues, the vast majority of the genome is highly methylated (>70%). However, genomic sequencing of bisulfite-treated DNA (MethylC-seq) has revealed large partially methylated domains (PMDs) in some human cell lines. However, to date only cultured cells and some cancers have shown evidence for PMDs, suggesting that PMDs may not be observed in normal human tissues.  Here we performed MethylC-seq in a set of human tissues and found that full-term human placenta shows clear evidence of PMDs. Overall design: Examination of gene expression in human placenta using RNA-seq, with one biological replicate (taken from same placenta)","project":"SRP014635"}
{"number_samples":7,"species":"human","abstract":"Generation of research quality, clinically relevant cell types in vitro from human pluripotent stem cells (hPSCs) requires detailed understanding of the equivalent cell types in humans. Here we analyzed 130 human fetal samples at 6-20 weeks of development and identified the stages in which human cKIT+ primordial germ cells (PGCs), the precursors of gametes, undergo whole genome epigenetic reprogramming and ultimately initiation of imprint erasure with loss of both 5mC and 5-hydroxy-mC at differentially methylated regions. Using five alternate in vitro differentiation strategies combined with a single-cell microfluidic analysis, high throughput RNA sequencing and a bona fide human cKIT+ PGC signature, we show that hPSC differentiation generates a rare cKIT+ PGC subtype found in both the human fetal gonad and mouse embryo. Taken together, our study creates a resource of human germ line ontogeny that is absolutely essential for future studies aimed at interpreting in vitro differentiation of the human germ line. Overall design: cKIT+ cells analyzed from 2 biological samples for testes and 2 samples for ovaries at 16 and 16.5 weeks. 3 biological replicates of TRA-1-60+ cells sorted from H1 hESCs","project":"SRP014670"}
{"number_samples":9,"species":"human","abstract":"LIN28 is a conserved RNA binding protein implicated in pluripotency, reprogramming and oncogenesis. Previously shown to act primarily by blocking let-7 microRNA (miRNA) biogenesis, here we elucidate distinct roles of LIN28 regulation via its direct messenger RNA (mRNA) targets. Through cross-linking and immunoprecipitation coupled with high-throughput sequencing (CLIP-seq) in human embryonic stem cells and somatic cells expressing exogenous LIN28, we have defined discrete LIN28 binding sites in a quarter of human transcripts. These sites revealed that LIN28 binds to GGAGA sequences enriched within loop structures in mRNAs, reminiscent of its interaction with let-7 miRNA precursors. Among LIN28 mRNA targets, we found evidence for LIN28 autoregulation and also direct but differing effects on the protein abundance of splicing regulators in somatic and pluripotent stem cells. Splicing-sensitive microarrays demonstrated that exogenous LIN28 expression causes widespread downstream alternative splicing changes. These findings identify important regulatory functions of LIN28 via direct mRNA interactions. Overall design: CLIP-seq for LIN28-V5 in stable human Flp-In-293 cells, and LIN28 in hES cells; strand-specific mRNA-seq for uninfected, control KD, and LIN28 KD human H9 ES cells; and strand-specific smallRNA-seq for uninfected, control KD, and LIN28 KD human H9 ES cells.","project":"SRP014671"}
{"number_samples":34,"species":"human","abstract":"MicroRNAs (miRNAs) are small RNAs of ~22 nucleotides in length that are involved in the regulation of a variety of physiological and pathological processes. Advances in high-throughput small RNA sequencing (smRNA-seq), one of the next generation sequencing applications, have reshaped the miRNA research landscape. In this study, we established an integrative database containing analysis pipelines and analysis results of 609 human and mice smRNA-seq results, including public data from GEO and private ones. YM500 collects analysis results for miRNA quantification, for isomiR identification (incl. RNA editing), for arm switching discovery, and, more importantly, for novel miRNA predictions. Wetlab validation on >100 miRNAs confirmed high correlation between miRNA profiling and RT-qPCR results (R=0.84). This database allows researchers to search these 4 different types of analysis results via our interactive web interface. YM500 allows researchers to define the criteria of isomiRs, as well as integrates the information of dbSNP to help researchers to distinguish isomiRs from SNPs. A user-friendly interface is provided to integrate miRNA-related information and existing evidences from hundreds of sequencing datasets. The identified novel miRNAs and isomiRs hold the potentials for both basic research and biotech applications. YM500 is now available on http://ngs.ym.edu.tw/ym500/. Overall design: There are 34 in-house datasets in YM500 . They are 9 human embryonal tumors, 9 human germ cell tumors, and 16 human gliomas.","project":"SRP014675"}
{"number_samples":5,"species":"human","abstract":"RNA sequences are expected to be identical to their corresponding DNA sequences.  Advances in technologies have enabled deep sequencing of nucleic acids that uncovered exceptions to the one-to-one relationship between DNA and RNA sequences.  Previously in human cells, post-transcriptional RNA editing was the only known mechanism that changes RNA sequences from the underlying DNA sequences.  Here, we sequenced nascent RNA  and found all 12 types of RNA-DNA differences.  Using various experimental analyses, we validated this finding.  Our results showed that sequences of nascent RNAs within 40 nucleotides of the exit channel of RNA polymerase II already differ from the corresponding DNA sequences.  These RNA-DNA differences are mediated by RNA processing steps closely coupled with transcription and not by known deaminase-mediated RNA editing mechanisms nor during NTP incorporation by Pol II.  This finding identifies sequence substitution as part of co-transcriptional RNA processing. Overall design: We sequenced nascent RNA using global run-on sequencing, GRO-seq from human B-cells from two individuals and a variant of the GRO-seq procedure, known as precision run-on sequencing, PRO-seq.  The RNAs are prepared after a short run-on assay performed with isolated nuclei in the presence of Br-UTP.  The isolated RNAs are base hydrolyzed to ~100 nucleotides and affinity purified with anti-BrU beads three times at each successive step of preparing the RNAs for orientation-specific sequencing using Illumina technology.  The 5’ ~half of each sequence represents nascent RNA made in the cell and the 3’ ~half  represents sequences made in vitro during the run-on reaction.  The precision variation, PRO-seq, incorporates one or at most a few biotin-labeled nucleoside triphosphates during the run-on, and sequencing from the 3’ end of this affinity purified, nascent RNA maps the cellular location of engaged polymerases with near single nucleotide precision.  We obtained ~ 100 million 100-nucleotide uniquely mapped GRO-seq reads from B-cells of two individuals.  For one subject, we also carried out pGRO-seq and obtained 60 million uniquely mapped reads.  In addition, we sequenced ~135 million uniquely mapped RNA-seq reads, and the corresponding DNA of the two individuals to 30X and 60X coverage. Additionally, we isolated and sequenced nascent RNA with an alternate method described by Wuarin and Schibler (1994) in order to compare chromatin-bound RNA to the very nascent RNA from PRO-seq.  We obtained ~190 million uniquely mapped reads from chormatin-bound RNA-seq.","project":"SRP014688"}
{"number_samples":4,"species":"human","abstract":"Background: Alternative polyadenylation (APA) is emerging as a widespread mechanism of gene regulation. The usage of APA sites allows a single gene to encode multiple mRNA transcripts with different 3'-untranslated region (3'UTR) lengths. Many disease processes reflect the importance of the regulation of APA site switching. The objective of this study was to explore the profiling of tandem APA sites in nasal polyps compared with nasal uncinate process mucosa. Methods: Sequencing of APA sites (SAPAS) based on second-generation sequencing technology was undertaken to investigate the use of tandem APA sites and identify gene expression patterns in samples from the nasal polyps and nasal uncinate process mucosa of two patients with chronic rhinosinusitis with nasal polyps. The findings of the SAPAS analysis were validated via quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Results: First, the results showed a switching of 3'UTR lengths in nasal polyps compared with nasal uncinate process mucosa. From the two patients, 105 overlapping genes in the nasal polyps were switched to distal poly(A) sites, and 90 such genes were switched to proximal poly(A) sites. Several Gene Ontology terms were enriched in the list of genes with switched APA sites, including transcription regulation, cell cycle, apoptosis, and metabolism. Second, we detected genes that showed differential expression with at least a 3-fold difference between nasal polyp tissue and nasal uncinate process mucosa. Between the two sample types, 627 genes exhibited differential expression. The qRT-PCR results confirmed our SAPAS results. Conclusion: APA site-switching events of 3'UTRs are prevalent in nasal polyp tissue, and the regulation of gene expression mediated by APA may play an important role in the formation and persistence of nasal polyps. Our results may provide new insights into the possible pathophysiologic processes involved in nasal polyps. Overall design: Investigate the use of tandem APA sites of 3'ends of mRNAs using second-generation sequencing technology.","project":"SRP014739"}
{"number_samples":5,"species":"human","abstract":"Oncogenic mutations in BRAF and NRAS occur in 70% of melanomas. Here we identify a microRNA, miR-146a, that is highly upregulated by oncogenic BRAF and NRAS. Expression of miR-146a increases the ability of human melanoma cells to proliferate in culture and form tumors in mice, whereas knockdown of miR-146a has the opposite effects. We show these oncogenic activities are due to miR-146a targeting the NUMB mRNA, a repressor of Notch signaling. Previous studies have shown that pre-miR-146a contains a single nucleotide polymorphism (C>G rs2910164). We find that the ability of pre-miR-146a/G to activate Notch signaling and promote oncogenesis is substantially higher than that of pre-miR-146a/C. Analysis of melanoma cell lines and matched patient samples indicates that during melanoma progression pre-miR-146a/G is enriched relative to pre-miR-146a/C, resulting from a C-to-G somatic mutation in pre-miR-146a/C. Collectively, our results reveal a central role for miR-146a in the initiation and progression of melanoma. Overall design: WI-38 cells were either infected with BRAFV600E or Empty retroviral vectors and small RNA were prepared from these cells. As an additional control, WI-38 cells were serum starved and used to generate quiscent cells, which were also used to prepase small RNA.  The small RNA were then used to generate small RNA library and were used on Illumina genome analyzer.","project":"SRP014754"}
{"number_samples":6,"species":"human","abstract":"Flexible detection of differential alternative splicing from replicate RNA-Seq data.","project":"SRP014759"}
{"number_samples":8,"species":"human","abstract":"Hepcidin, a peptide hormone that decreases intestinal iron absorption and macrophage iron release, is a potential drug target for patients with iron overload syndromes because its levels are inappropriately low in these individuals.  Endogenous stimulants of Hepcidin transcription include bone morphogenic protein 6 (BMP) and interleukin-6 (IL-6) via effects on Smad4 or Stat3, respectively.  We conducted a small-scale chemical screen in zebrafish embryos to identify small molecules that modulate hepcidin expression.  We found that treatment with the isoflavone genistein from 28-52 hours post-fertilization in zebrafish embryos enhanced Hepcidin transcript levels as assessed by whole mount in situ hybridization and quantitative realtime RT-PCR.  Genistein’s stimulatory effect was conserved in human hepatocytes: genistein treatment of HepG2 cells increased both Hepcidin transcript levels and Hepcidin promoter activity.  We found that genistein’s effect on Hepcidin expression did not depend on estrogen receptor signaling or increased cellular iron uptake, but was impaired by mutation of either the BMP response elements or the Stat3 binding site in the Hepcidin promoter.  RNA-sequencing of transcripts from genistein-treated hepatocytes indicated that genistein upregulated 68% of the transcripts that were upregulated by BMP6, however genistein raised the levels of several transcripts involved in Stat3 signaling that were not upregulated by BMP6.  Chromatin-immunoprecipitation and ELISA experiments revealed that genistein enhanced Stat3 binding to the Hepcidin promoter and increased phosphorylation of Stat3 in HepG2 cells.  CONCLUSION:  Genistein is the first small molecule experimental drug that stimulates Hepcidin expression in vivo and in vitro.  These experiments demonstrate the feasibility of identifying and characterizing small molecules that increase Hepcidin expression.  Genistein and other candidate molecules may subsequently be developed into new therapies for iron overload syndromes. Overall design: RNA-seq of HepG2 cells treated with DMSO 1%, BMP6 50 ng/ml, or genistein 10 micromolar.  The numbers of biological replicates were 3, 2, and 3.","project":"SRP014790"}
{"number_samples":12,"species":"human","abstract":"The androgen receptor (AR) is a ligand-inducible transcription factor that mediates androgen action in target tissues. Upon ligand binding, the AR binds to thousands of genomic loci and activates a cell-type specific gene program. Prostate cancer growth and progression depend on androgen-induced AR signalling. Treatment of advanced prostate cancer through medical or surgical castration leads to initial response and durable remission, but resistance inevitably develops. In castration-resistant prostate cancer (CRPC), AR activity remains critical for tumor growth despite androgen deprivation. While previous studies have focused on ligand-dependent AR signalling, in this study we explore AR function under the androgen-deprived conditions characteristic of CRPC. Our data demonstrate that the AR persistently occupies a distinct set of genomic loci after androgen deprivation in CRPC. These androgen-independent AR occupied regions have constitutively open chromatin structures that lack the canonical androgen response element and are independent of FoxA1, a transcription factor involved in ligand-dependent AR targeting. Many AR binding events occur at proximal promoters, which can act as enhancers to augment transcriptional activities of other promoters through DNA looping. We further show that androgen-independent AR binding directs a distinct gene expression program in CRPC, which is necessary for the growth of CRPC after androgen withdrawal. Overall design: LNCaP, C4-2B, or 22RV1 cells were cultured in hormone-free media for 3 days and then treated with ethanol vehicle or DHT (10nM) for 4h or 16h prior to ChIP-seq or RNA-seq assays. For siRNA transfection, cells were transfected with AR siRNA or control siRNA for 3 days prior to RNA-seq assays.","project":"SRP014809"}
{"number_samples":59,"species":"human","abstract":"Microarray based analysis of transcriptional profiles and genomic sequencing of tumor samples have greatly improved the biological understanding and molecular classification of breast cancer. However, certain aspects of the transcriptome, including characterization of differential expression of splice variants, are not well captured by these methods. We report results from single-end Illumina-based RNA-sequencing of 53 primary breast cancers and 6 \"normal\" breast samples from mammoplasty patients, using both 29 bp reads (on 29 tumor samples) and 100 bp reads (on 24 tumor samples and 6 normals). Novel as well as widely used computational approaches were used to identify differentially expressed splice variants of known RefSeq genes. This analysis identified a number of variants differentially spliced in subsets of breast tumor samples. Some of these splice variants were specific to either ER+ or ER- breast cancers, including the prostate cancer oncogene TPD52/PrLZ and intronic-start variants of genes IQCG and ACOX2, both of which have been implicated as potentially oncogenic in other cancers. We also found differential splicing in subsets of breast cancer samples which did not associate with ER or HER2 status, including a novel short-form Lactoferrin variant similar to dLTF, and the secreted protein NESP55 at the biallelically imprinted GNAS locus. The expression of several variants were verified in both tumor samples and breast cancer derived cell lines using RT-PCR. These data demonstrate that differential use of transcript variants may play an important role in the phenotype of breast cancer subtypes, and can be robustly identified by RNA sequencing analysis.","project":"SRP014830"}
{"number_samples":12,"species":"human","abstract":"The nuclear lamina (NL) interacts with hundreds of large genomic regions termed lamina-associated domains (LADs). The dynamics of these interactions and the relation to epigenetic modifications are poorly understood. We visualized the fate of LADs in single cells using a novel 'molecular contact memory' approach. In each interphase nucleus, only ~30% of LADs are positioned at the periphery; these LADs are in intermittent molecular contact with the NL but remain constrained to the periphery. Upon mitosis, LAD positioning is not detectably inherited but instead is stochastically reshuffled. Contact of individual LADs with the NL correlates with their degree of H3K9 dimethylation in single cells, and inactivation of the H3K9 methyltransferase G9a reduces the NL contact frequencies. These results indicate that nuclear positioning and histone modification of LADs are both stochastic yet linked in single cells. Collectively, these results highlight principles of the dynamic spatial architecture of chromosomes. Overall design: 12 RNA-seq experiments for 6 samples, each with a biological replicate: m6ATracer-VP16+/DamLaminB1+ m6ATracer-VP16+/DamLaminB1- m6ATracer-VP16-/DamLaminB1- m6ATracer-GFP+/DamLaminB1+ m6ATracer-GFP+/DamLaminB1- m6ATracer-GFP-/DamLaminB1-","project":"SRP014842"}
{"number_samples":2,"species":"human","abstract":"Differences in global levels of histone acetylation occur in normal and cancer cells, although the reason cells regulate these levels has remained unclear. Here we demonstrate a role for histone acetylation in regulating intracellular pH (pHi). As pHi decreases, histones are globally deacetylated by histone deacetylases (HDACs) and the released acetate anions are co-exported with protons out of the cell by monocarboxylate transporters (MCTs), preventing further reductions in pHi. Conversely, global histone acetylation increases at more alkaline pHi, such as when resting cells are induced to proliferate. Inhibition of HDACs or MCTs decreases acetate export and lowers pHi, particularly compromising pHi maintenance in acidic environments. Global deacetylation at low pH is reflected at a genomic level by decreased abundance and extensive redistribution of acetylation at promoters and intergenic regions. Thus acetylation of chromatin functions as a rheostat to regulate pHi with important implications for therapeutic use of HDAC inhibitors. Overall design: To investigate the redistribution of H4K16ac throughout the genome upon treatment at pH 6.5","project":"SRP014844"}
{"number_samples":6,"species":"human","abstract":"Synthetic, innate defense regulators (IDR) peptides, designed based on natural host defenses peptides, have enhanced immunomodulatory activities and reduced toxicity leading to protection in infection and inflammation models that is dependent on macrophages/monocytes. Here we measured the effect of IDR-1018 on macrophage gene expression during differentiation. Differentiation in the presence of IDR-1018 induced a unique signature of immune responses suggesting that IDR-1018 drives macrophage differentiation towards an intermediate M1-M2 state, enhancing anti-inflammatory functions while maintaining certain pro-inflammatory activities important to the resolution of infection. Overall design: RNA-seq was performed using the Illumina Genome Analyzer IIx platform. Monocytes were isolated from 3 healthy donors, and left unstimulated  or stimulated for 4 hours with 20 µg/ml IDR-1018. For library preparation, 500 ng of total RNA was processed according to the Illumina TruSeq RNA sample preparation guide (Illumina catalogue number FC-122-1002). Briefly, mRNA was purified using poly-dT beads, followed by synthesis of the first and second cDNA strands, end repair addition of an poly-A overhang, and ligation of adapters and unique barcodes, as per the manufacturer’s instructions. DNA enrichment was carried out via a 15-cycle PCR. Following quantification, 8 pM of dsDNA was used for cluster generation on a CBOT instrument (Illumina, San Diego, CA). RNA sequencing was done on a GAIIx instrument (Illumina), performed as a single read run with 51 amplification cycles. Data processing was carried out in house, using CASAVA to convert raw data and demultiplex to FASTQ sequence files. Reads were aligned to the reference genome using TOPHAT, and then mapped to genes using the Bioconductor package GenomeRanges.","project":"SRP014856"}
{"number_samples":2,"species":"human","abstract":"N6-methyladenosine (m6A) is the most prevalent internal modification of messenger RNA (mRNA) in higher eukaryotes. Here we report ALKBH5 as a new mammalian demethylase that oxidatively removes the m6A modification in mRNA in vitro and inside cells. This demethylation activity of ALKBH5 significantly affects mRNA export and RNA metabolism as well as the assembly of mRNA processing factors in nuclear speckles. Alkbh5-deficient male mice are characterized by impaired fertility resulting from apoptosis that affects meiotic metaphase-stage spermatocytes. In accordance with this defect, we have identified in mouse testes 1552 differentially expressed genes which cover broad functional categories and include spermatogenesis-related mRNAs involved in the p53 functional interaction network. We show that Alkbh5-deficiency impacts the expression levels of some of these mRNAs, supporting the observed phenotype. The discovery of this new RNA demethylase strongly suggests that the reversible m6A modification plays fundamental and broad functions in mammalian cells. Overall design: RNA-seq in two cell types","project":"SRP014857"}
{"number_samples":6,"species":"human","abstract":"In eukaryotes, the 3' ends of RNA polymerase II-generated transcripts are made in the majority of cases by site-specific endonucleolytic cleavage, followed by the addition of a poly(A) tail. By alternative polyadenylation, a gene can give rise to multiple mRNA isoforms that differ in the length of their 3' UTRs and hence in their susceptibility to post-transcriptional regulatory factors such as microRNAs. A series of recently conducted high-throughput studies of poly(A) site usage revealed an extensive tissue-specific control of 3’ UTR length and drastic changes in 3’ UTR length of mRNAs upon induction of proliferation in resting cells. To understand the dynamics of polyadenylation site usage, we recently identified binding sites of the major pre-mRNA 3’ end processing factors - cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), and cleavage factor Im (CF Im) - and mapped cleaved polyadenylation sites in HEK293 cells. Our present study extends previous findings on the role of CF Im in alternative polyadenylation and reveals that subunits of the CF Im complex generally control 3’ UTR length. More specifically, we demonstrate that the  loss-of-function of CF Im68 and CF Im25 but not of CF Im59 leads to a transcriptome-wide increase of the use of proximal polyadenylation sites. Overall design: 3' ends of transcripts were profiled by high-throughput sequencing in HEK 293 cells under normal conditions, and in HEK 293 cells depleted of 3' end processing factors CF Im25, CF Im59, and CF Im68.","project":"SRP014867"}
{"number_samples":11,"species":"human","abstract":"Normal Karyotype acute myeloid leukemia (NK-AML) represents approximately 50% of all cases of AML which patients develop.  Most AML cell lines are highly abnormal and therefore not good models for investigating NK-AML biology a novel AML cell line, CG-SH, was recently estabished and here we characterize the gene expression and mutations present through high-throughput sequencing of RNA and genomic DNA using a HiSeq 2000 Overall design: The overall design of the experiment was to characterize, at single base pair resolution, all of the genetic defects present in a novel normal karyotype cell line, CG-SH","project":"SRP014906"}
{"number_samples":1,"species":"human","abstract":"Gene regulation by DNA binding small molecules could have important therapeutic applications. This study reports the investigation of a DNA-binding pyrrole-imidazole polyamide targeted to bind the DNA sequence 5’-WGGWWW-3’ with reference to its potency in a subcutaneous xenograft tumor model. The molecule is capable of trafficking to the tumor site following subcutaneous injection and modulates transcription of select genes in vivo. A FITC-labeled analogue of this polyamide can be detected in tumor-derived cells by confocal microscopy. RNA deep sequencing (RNA-seq) of tumor tissue allowed the identification of further affected genes, a representative panel of which were interrogated by qRT-PCR and correlated with cell culture expression levels. Overall design: Xenografts. Grafting with A549-luc-C8. Experiments were performed in female SCID-beige mice (Charles River) between 8 and 12 weeks of age. Cells were injected into the left flank area of the animals as suspensions of 25 x 106 mL-1 in RPMI, 200 µL per injection. Treatment and tumor proliferation monitoring. Mice were treated following the schedule delineated in treatment protocol. Tumor proliferation was monitored using the XENOGEN imaging device. The animals were anesthetized with 2 5 % isoflurane and subsequently transferred to the imaging chamber, whereupon the isoflurane levels were reduced to 1-2.5 %. The floor of the imager was heated to +37 ºC to avoid hypothermia. Breathing frequency was monitored and not allowed to drop below 1 s-1, adjusting the isoflurane levels accordingly at all times. Endpoint criteria and euthanasia. Animal endpoint criteria encompassed weight loss of over 15 %, restriction of motor function by the engrafted tumor, dehydration of over 10 % and moribund behavior. Where appropriate, the animals were euthanized by asphyxiation in a CO2 chamber. Tumor tissue harvest. Animals were resected and tumors excised using standard forceps, scissors and surgical blades. The tumors were combined into one sample per condition and mechanically sheared in TRIZOL, employing a specialized device (tissue tearer, model 985370). Total RNA workup was performed following the standard TRIZOL procedure, followed by a DNAse digest.","project":"SRP014920"}
{"number_samples":2,"species":"human","abstract":"The precise control of microRNA (miRNA) biogenesis is important for various cellular functions, and its dysregulation is often associated with human diseases.  We previously reported that Terminal   uridylyl transferase 4 (TUT4) down-regulates let-7 miRNA biogenesis by oligo-uridylating let-7 precursor (pre-let-7) in mouse embryonic stem cells and that a pluripotency marker Lin28 promotes   a processivity of TUT4.  Here we find that TUT4 positively controls let-7 biogenesis by adding a uridine residue to the 3’ end of pre-let-7 in the absence of Lin28. Such mono-uridylation enhances Dicer   processing by generating an optimal end structure of pre-let-7 for Dicer recognition and may protect pre-miRNA from trimming. Moreover, TUT7, TUT4 and TUT2 redundantly regulate pre-let-7   processing and simultaneous knock down of these TUTs leads to the decrease of mature let-7 and the accumulation of pre-let-7 in cells. This study provides a novel regulation mechanism of miRNA   biogenesis, which may function in development and tumorigenesis. Overall design: HeLa cells were transfected with siRNA two times over a 4~5 day period.","project":"SRP014925"}
{"number_samples":10,"species":"human","abstract":"Papillary type 2B RNA sequencing.","project":"SRP015003"}
{"number_samples":8,"species":"human","abstract":"The purpose of this study was to develop a quantification method that can be used to assess the ability of tag-seq to detect malignant B-cell transcripts. The data support that tumour cell concentration is an important variable with fundamental impact on gene expression pattern. Overall design: We analysed eight serial dilutions of the malignant B-cell line, OCI-Ly8, into the embryonic kidney cell line, HEK293, by tag-sequencing. No technical replicates were performed.","project":"SRP015013"}
{"number_samples":4,"species":"human","abstract":"CD34 positive hematopoietic stem cells were differentiated into erythroid lineage. Next generation sequencing (NGS) of 5hmC affinity pulldown and RNAseq were performed in four time point of different stages of erythroid differentiation. Overall design: 4 RNA-Seq Samples (d0, d3, d7 and d10); 4 affinity-pulldown  (d0, d3, d7 and d10), and 4 input samples  (d0, d3, d7 and d10).","project":"SRP015138"}
{"number_samples":3,"species":"human","abstract":"The mRNA repertoire is specific to a given tissue type.  This specificity, whether it is in the levels of gene expression or the splicing pattern of the transcripts, is important to study to increase our general knowledge of transcription and alternative splicing, as well as to identify novel coding sequence that can be important for elucidating the mechanisms of disease.  We have performed RNA-Seq on three normal human retinal samples and have found a profound level of novel alternative splicing including novel exons, exon skipping, and 3’ and 5’ alternate splice sites.  Additionally, we have identified hundreds of novel genes.  With so many novel splicing events identified in the retinal transcriptome, we set out to determine if these novel features were indeed real.  We developed a high throughput capture set to target 14,696 of the novel features.  We found that 99% of the novel features do validate, including those initially detected at a depth of coverage as low as one.  These results confirm that the transcripts expressed in the retina are much more diverse than previously identified.  The complete human retinal transcriptome described here will be useful for investigators studying multiple aspects of retinal biology and disease. Overall design: Characterization of alternative splicing in the normal human retinal transcriptome.","project":"SRP015336"}
{"number_samples":6,"species":"human","abstract":"We report cytokine specific changes in gene expression in the human neutrophil transcriptome using TNF-alpha and GM-CSF stimulation of healthy neutrophils Overall design: Healthy human neutrophils were stimulated with TNF-alpha or GM-CSF for 1h in vitro. RNA was analysed by SOLiD and Illumina sequencing.  RNA from one biological donor was sequenced on both platforms, and two different biological donors were sequenced by Illumina.","project":"SRP015360"}
{"number_samples":8,"species":"human","abstract":"Breast and ovarian cancer susceptibility genes, BRCA1 and PALB2 have enigmatic roles in cellular growth and mammalian development.  While these genes are essential for growth during early developmental programs, inactivation later in adulthood leads to increased growth and formation of tumors, leading to their designation as tumor suppressors.  We performed genome-wide analysis assessing their chromatin residence and gene expression responsiveness using high throughput sequencing in breast epithelial cells.  These experiments revealed a critical role for BRCA1 and PALB2 in transcriptional responsiveness to NF-kB, a crucial mediator of growth and inflammatory response during development and cancer.  Importantly, we also uncovered a vital role for these proteins in response to retinoic acid (RA), a growth inhibitory signal in breast cancer cells, which may constitute the basis for their tumor suppressor activity. Overall design: Comparison of the genome wide profiles of the BRCA protein complex (BRCA1 and PALB2) and phosphorylated RNAPII (P-Ser2) in MCF10A cells by ChIP-seq. Effect of BRCA1 and PALB2 knockdown (shRNAs)  on transcription was  assessed by RNA-seq.","project":"SRP015361"}
{"number_samples":5,"species":"human","abstract":"MicroRNAs (miRNAs) are major post-transcriptional regulators of gene expression, yet their origins and functional evolution in mammals remain little understood due to the lack of appropriate comparative data. Using RNA sequencing, we have generated extensive and comparable miRNA data for five organs in six species that represent all main mammalian lineages and birds (the evolutionary outgroup), with the aim to unravel the evolution of mammalian miRNAs. Our analyses reveal an overall expansion of miRNA repertoires in mammals, with three-fold accelerated birth rates of miRNA families in placentals and marsupials, facilitated by the de novo emergence of miRNAs in host gene introns. Generally, our analyses suggest a high rate of miRNA family turnover in mammals, with many newly emerged miRNA families being lost soon after their formation. Selectively preserved mammalian miRNA families gradually evolved higher expression levels as well as altered mature sequences and target gene repertoires, and were apparently mainly recruited to exert regulatory functions in nervous tissues. However, miRNAs that originated on the X chromosome evolved high expression levels and potentially diverse functions during spermatogenesis, including meiosis, through selectively driven duplication-divergence processes. Overall, our study thus provides detailed insights into the birth and evolution of mammalian miRNA genes and the associated selective forces. Overall design: 30 main samples from five adult tissues (brain, cerebellum, heart, kidney and testis) collected in six species (human, macaque, mouse, opossum, platypus and chicken) + 5 biological replicates + 3 samples from spermatogenic cells in adult mouse testis  (Sertoli cells, spermatocytes and spermatids)","project":"SRP015370"}
{"number_samples":6,"species":"human","abstract":"We report a novel modular pipeline (iMir) for comprehensive analysis of miRNA-Seq data, from linker removal and sequence quality check to differential expression and biological target prediction, integrating multiple open source modules and resources linker together in an automated flow. Overall design: Development of an integrated pipeline (iMir) for comprehensive analysis of miRNA-Seq experiment.","project":"SRP015409"}
{"number_samples":9,"species":"human","abstract":"We investigated the genomic occupancy of INTS11, in normal condition and after stimulation of EGF. Total RNAPII was profiled in the presence or absence of INTS11, along with the Super Elongation Complex proteins AFF4 and ELL2. Additionally, we extensively examined the transcriptional response to EGF, before and after depletion of INTS11, using RNA-seq on ribosome-depleted total RNA and Global Run-on sequencing (GRO-seq). Overall design: Comparison of the genome-wide profiles of INTS11 and RNAPII in HeLa cells.","project":"SRP015419"}
{"number_samples":1,"species":"human","abstract":"FUS/TLS and TDP-43 are RNA/DNA-binding proteins integrally involved in amyotrophic lateral sclerosis (ALS) and frontal temporal dementia. FUS/TLS is shown to bind RNAs from >5,500 genes in mouse and human brain, primarily through a GUGGU binding motif. A characteristic sawtooth-like binding pattern is identified, supporting co-transcriptional deposition of FUS/TLS. Depletion of FUS/TLS from the adult nervous system is shown to alter levels or splicing of >970 mRNAs, most of which are distinct from the RNAs whose maturation is dependent on TDP-43. Nonetheless, only 55 RNAs are reduced upon depletion of either TDP-43 or FUS/TLS from mouse brain and human neurons differentiated from pluripotent stem cells, including mRNAs transcribed from genes with exceptionally long introns and that encode proteins essential for neuronal integrity. A subset of these is significantly lowered in FUS/TLSR521G and TDP-43G298S mutant fibroblasts and in TDP-43 aggregate-containing motor neurons in sporadic ALS, evidence pointing to a common loss-of-function pathway as one component underlying motor neuron death from misregulation of TDP-43 or FUS/TLS. Overall design: CLIP of Fus/Tls in 8 week mouse brain and adult human brain","project":"SRP015435"}
{"number_samples":11,"species":"human","abstract":"ETS transcription factors ETV2, FLI1 and ERG1 specify pluripotent stem cells into endothelial cells (PSC-ECs). However, these PSC-ECs are unstable and often drift towards non-vascular cell fates. We show that human mid-gestation c-Kit- lineage-committed amniotic cells (ACs) can be reprogrammed into induced vascular endothelial cells (rAC-VECs). Transient ETV2 expression in ACs generated immature iVECs, while co-expression with FLI1/ERG1 endowed rAC-VECs with a vascular repertoire and morphology matching mature ECs. Brief TGFb-inhibition functionalizes VEGFR2 signaling, augmenting specification of ACs into rAC-VECs. Genome-wide transcriptional analyses showed that rAC-VECs are similar to adult ECs in which vascular-specific genes are expressed and non-vascular genes are silenced. Functionally, rAC-VECs form stable vasculature in Matrigel plugs and regenerating livers. Thus, short-term ETV2 expression and TGFb-inhibition along with constitutive ERG1/FLI1 co-expression reprogram mature ACs into generic rAC-VECs with clinical-scale expansion potential. Public banking of HLA-typed rAC-VECs would establish a vascular inventory for treatment of genetically diverse disorders. Overall design: Transcriptome sequencing of clonal and non-clonal rAC-VECs, HUVECs, LSECs, CD34+/Lin-, BMS","project":"SRP015439"}
{"number_samples":5,"species":"human","abstract":"Purpose: Development of resistance to tamoxifen is an important clinical issue in the treatment of patients with breast cancer. Tamoxifen resistance may be the result of the acquisition of epigenetic regulation such as DNA methylation within breast cancer cells resulting in changed mRNA expression of genes being pivotal for estrogen dependent growth. Alternatively, tamoxifen resistance may be due to selection of preexisting resistant cells, which may exhibit cancer stem-like characteristics or a combination of the two mechanisms. Methods: To evaluate the contribution of these possible mechanisms to tamoxifen resistance, we applied modified DNA methylation-specific digital karyotyping (MMSDK) and digital gene expression (DGE) in combination with massively parallel sequencing to analyze a well-established tamoxifen resistant cell line model: MCF-7/S0.5 (tamoxifen sensitive parental cell line) and 4 high-dosage tamoxifen selected resistant offspring sublines (MCF-7/TAMR-1, MCF-7/TAMR-4, MCF-7/TAMR-7 and MCF-7/TAMR-8). MMSDK uses BssHII as mapping enzyme (DNA methylation sensitive enzyme). Both MMSDK and DGE use NlaIII and MmeI to produce 20-21 bp tag. The indexed single-end sequencing was performed by Illumina HiSeq 2000 in BGI-Shenzhen. A dynamic programming algorithm-FASTX-Toolkit implemented in Perl was used to trim the adaptor sequence. The trimmed tags were subjected to quality filtering, so that only tags with sequencing quality higher than 30 for more than 80% of the nucleotides were used for subsequent analysis. For MMSDK tag mapping, we generated a simulated reference library, i.e., BssHII reference library, by in silico enzyme digestion of the human genome (hg19, UCSC) regardless of the methylation state. This library was used as reference for subsequent mapping of the tags in the MMSDK analysis. In the DGE analysis, refMrna (hg19, UCSC) was used as reference for mapping cDNA tags. Subsequently, the Burrows–Wheeler Aligner (BWA) procedure for aligning the MMSDK and DGE tags to the simulated BssHII reference library and refMrna reference library, respectively, was applied. Results: MMSDK libraries using BssHII/NlaIII were generated from the parental tamoxifen sensitive subline MCF-7/S0.5 and the 4 TAMR cell lines: TAMR-1, TAMR-4, TAMR-7 and TAMR-8. The 5 indexed MMSDK libraries were sequenced in one lane and 1.38 Gb clean tag data for all 5 cell lines were obtained, with an average sequencing amount of ~270 Mb per library. On average, 59.5 % of the tags with mapping quality = 20 were mapped back to the simulated BssHII/NlaIII reference library. DGE libraries were also generated from MCF-7/S0.5 and the 4 TAMR cell lines. The 5 indexed DGE libraries were sequenced in one lane and obtained 1.71 Gb clean tag data for all 5 cell lines with an average sequencing amount of ~340 Mb per library. On average, 40.8 % with mapping quality = 20 were mapped back to the simulated NlaIII human transcriptome (refMrna reference library). Our present study demonstrates large differences in global gene expression and DNA methylation profiles between parental tamoxifen-sensitive cell line and 4 high-dosage tamoxifen treatment selected resistant sublines. The tamoxifen resistant cell lines exhibited globally higher methylation level than the parental cell line and an inverse relationship between gene expression and DNA methylation in the promoter regions were noticed. High expression of SOX2 and alterations of other SOX gene family members, E2F gene family members and RB-related pocket protein genes as well as highlighted stem cell pathways imply that cancer initiating cells/stem cells are  involved in the resistance to tamoxifen. Overall design: DNA methylation and mRNA expression profiles from tamoxifen sensitive parental cell line MCF-7/S0.5 and 4 high dosage of tamoxifen selected resistant offspring sublines (MCF-7/TAMR-1, MCF-7/TAMR-4, MCF-7/TAMR-7 and MCF-7/TAMR-8) were analyzed by MMSDK and DGE methods, respectively, in combination of massively parallel sequencing, using Illumina HiSeq 2000","project":"SRP015449"}
{"number_samples":68,"species":"human","abstract":"RNA-Seq is an effective method to study the transcriptome, but can be difficult to apply to scarce or degraded RNA from fixed clinical samples, rare cell populations, or cadavers. Recent studies have proposed several methods for RNA-Seq of low quality and/or low quantity samples, but their relative merits have not been systematically analyzed. Here, we compare five such methods using a comprehensive set of metrics, relevant to applications such as transcriptome annotation, transcript discovery, and gene expression. Using a single human RNA sample, we constructed and deeply sequenced 10 libraries with these methods and two control libraries. We find that the RNase H method performed best for low quality RNA, and can even effectively replace oligo (dT) based methods for standard RNA-Seq. SMART and NuGEN had distinct strengths for low quantity RNA. Our analysis allows biologists to select the most suitable methods and provides a benchmark for future method development. Overall design: Examination of 9 different RNA-Seq libraries starting from total RNA from 5 distinct methods; also 3 control RNA-Seq libraries","project":"SRP015640"}
{"number_samples":33,"species":"human","abstract":"We sought to more precisely characterize the different alpha-synuclein (aSyn) 3’UTR mRNA species in normal and PD human brain.  High-throughput, whole-transcriptome sequencing of the 3’UTR ends of polyadenylated mRNA transcripts (termed pA-RNAseq; see Methods) was performed on a cohort of 17 unaffected and 17 PD cerebral cortical tissue samples.  This revealed 5 aSyn 3’UTR isoforms, with lengths of 290, 480, 560, 1070 and 2520 nt.  Of these, the 560 nt and 2520 nt forms were predominant. The existence and relative preponderance of these species was further confirmed by Northern Blot. We next hypothesized, that aSyn 3’UTR selection might be altered in PD.  Comparison of pA-RNAseq profiles from PD and unaffected cerebral cortex samples revealed an increase in the preponderance of the long 3’UTR species (>560 nt) relative to shorter species (<560 nt).  Such a relative increase in aSynL was confirmed by Quantitative real-time RT-PCR (rt-qPCR) and appeared specific for PD, as the increase was also observed by comparison to RNA from amyotrophic lateral sclerosis patient samples.  We note that the modified aSyn 3’UTR selection associated with PD patient tissue was detected in cerebral cortex tissue, which typically harbors pathological evidence of the disease process without frank cell loss; thus, this phenotype is unlikely to be a secondary consequence of neurodegeneration. Overall design: Comparison of 3'UTR ends of alpha-synuclein in PD and unaffected brain cortex","project":"SRP015668"}
{"number_samples":40,"species":"human","abstract":"Background: West Nile virus is an emerging infection of biodefense concern and there are no available treatments or vaccines. Here we used a high-throughput method based on a novel gene expression analysis, RNA-Seq, to give a global picture of differential gene expression by primary human macrophages of 10 healthy donors infected in vitro with WNV. Results: From a total of 50 million reads per sample, we employed a Bayesian hierarchical mixture model to identify 4,026 transcripts that were differentially expressed after infection. Both predicted and novel gene changes were detected, as were gene isoforms, and while many of the genes were expressed by all donors, some were unique. Knock-down of genes not previously known to be associated with WNV resistance identified their critical role in control of viral infection. Conclusions: Our study distinguishes both common gene pathways as well as novel cellular responses. Such analysis will be valuable for translational studies of susceptible and resistant individuals -- and for targeting therapeutics -- in multiple biological settings. Overall design: Differential gene expression by primary human macrophages of 10 healthy donors infected in vitro with WNV were generated by RNA-Seq.","project":"SRP015670"}
{"number_samples":4,"species":"human","abstract":"We have identfied antibodies that support chromatin immunoprecipitation (ChIP) of 11 components of Polycomb repressive complex 1 (PRC1) and shown that multiple variants of PRC1 associate with the same DNA.  By performing ChIP-seq with antibodies against CBX6, CBX7, CBX8, RING1 and RING2, in two strains of human fibroblasts, we find that  all five proteins co-localize at multiple sites in the genome. The binding profiles have inherent architecture that is conserved between the two strains, but there are also a substantial number of differences between the strains, possibly reflecting their anatomical position. The presence of PcG proteins does not equate with gene silencing and their genomic position does not change at senescence. Overall design: Chromatin immunoprecipitation and sequencing (ChIP-seq) in two strains of  human fibroblasts using antibodies against three Pc orthologs (CBX6, CBX7 and CBX8) and both Psc orthologs (RING1 and 2). Validation by ChIP-PCR and re-ChIP.  Strand specific RNA-seq and validation by qRT-PCR.","project":"SRP015678"}
{"number_samples":1,"species":"human","abstract":"The exon junction complex (EJC) is a central effector of mRNAs fate, linking nuclear processing to mRNA transport, translation and surveillance. Little is known about its transcriptome-wide targets. We used high-throughput sequencing after crosslinking and immunoprecipitation (HITS-CLIP) in human cells to identify the binding sites of the DEAD-box helicase eIF4AIII, an EJC core component. CLIP reads form peaks mainly located in spliced mRNAs. Most expressed exons harbour peaks equally distributed between the canonical EJC region ~24 nucleotides upstream of exonic junctions and other non-canonical regions. Unexpectedly, both are preferentially associated to unstructured and purine-rich sequences containing the motif GAAGA, a potential binding site of EJC-associated factors. Therefore, EJC positions vary spatially and quantitatively between exons. This transcriptome-wide mapping of human eIF4AIII reveals unanticipated aspects of the EJC and broadens its potential impact on post-transcriptional regulation. Overall design: To identify direct RNA binding sites of the EJC core component eIF4AIII, two biological CLIP-seq replicates were performed in HeLa cells. Additionally, mRNA-seq of the HeLa transcriptome was performed to normalize for the mRNA expression levels.","project":"SRP015711"}
{"number_samples":2,"species":"human","abstract":"Gene expression analysis is a widely used and powerful method for investigating the transcriptional behavior of biological systems, for classifying cell states in disease and for many other purposes.  Recent studies indicate that common assumptions currently embedded in experimental and analytical practices can lead to misinterpretation of global gene expression data.  We discuss these assumptions and describe solutions that should minimize erroneous interpretation of gene expression data from multiple analysis platforms. Overall design: Polyadenylated RNA depleted of ribosomal content was used for preparation of two independent sequencing libraries (low-Myc & high-Myc). A panel of synthetic RNA's was added to these populations, based on cell number.","project":"SRP015715"}
{"number_samples":14,"species":"human","abstract":"We undertook an integrative technological approach to compare miRNA detection capability of three high-throughput commercial platforms. Overall design: We artificially introduced human precursor, 2’-O-methyl modified and mature spiked-in miRNAs in a controlled fashion into native human placenta total RNA.","project":"SRP015725"}
{"number_samples":4,"species":"human","abstract":"Identification of EWS regulated genes in Ewing sarcoma.","project":"SRP015733"}
{"number_samples":3,"species":"human","abstract":"We mapped CstF64-RNA interactions at the transcriptome level and studied CstF64-mediated regulation of global mRNA alternative polyadenylation by using iCLIP-seq and direct RNA sequencing analyses. Overall design: CstF64 iCLIP-seq in HeLa cells;  direct RNA sequencing (DRS) of control HeLa cells, CstF64-RNAi cells and CstF64&CstF64tau-RNAi cells","project":"SRP015741"}
{"number_samples":16,"species":"human","abstract":"In order to study molecular changes in the stroma from tissue samples it is recommended to separate tumor tissue from stromal tissue. This is particularly relevant to mouse tumor xenograft models where tumor, particularly metastatic tumors, can be small and difficult to separate from the host tissue. In our research we compared qualitatively the ability of high-throughput mRNA sequencing, RNA-Seq, and microarrays to detect tumor (human) and stromal (mouse) expression from mixed tumor-stromal samples in terms of the genes and pathways that are involved in cross-alignment (RNA-Seq) and cross-hybridization (microarrays). Overall design: Human samples consisted of total RNA obtained from MDA-MB-231 human breast carcinoma cell line and isolated from three independent cultures of sub-confluent MDA-MB-231 cell lines in exponential phase of growth.  Mouse samples were obtained from NOD scid gamma mice, and normal lung tissue was harvested from three independent age-matched mice.","project":"SRP015764"}
{"number_samples":120,"species":"human","abstract":"In this study, we screened human placental samples for allele-specific methylation and subsequently novel imprinted genes associated with these regions. We used reduced representation bisulfite sequencing to identify partially methylated CpG islands (CGIs) in the human placental genome. We were able to delineate potential candidates for allele-specific methylation based on the calculation of a concordance statistic. Amongst the 28 regions chosen for validation based on high levels of expression, two regions were shown to exhibit allele-specific expression. Overall design: Single base-resolution methylation analysis in the placental genome and RNA-Seq","project":"SRP015789"}
{"number_samples":6,"species":"human","abstract":"Gene regulation by BCL11B in Ewing sarcoma.","project":"SRP015799"}
{"number_samples":4,"species":"human","abstract":"Many long non-coding RNA (lncRNA) species have been identified in mammalian cells, but the genomic origin, regulation and function of these molecules in individual cell types is poorly understood.  We have generated comprehensive catalogs of lncRNA species expressed in human and murine embryonic stem cells (ESCs) and mapped their genomic origin.  A surprisingly large fraction of these transcripts (>60%) originate from divergent transcription at promoters of active protein-coding genes.  The divergently transcribed lncRNA/mRNA gene pairs exhibit coordinated changes in transcription when ESCs are differentiated into endoderm.  A significant number of the divergently transcribed lncRNAs/mRNA pairs are conserved between human and mouse ESCs.  Disruption of promoter-associated lncRNA orthologs in a zebrafish model of early development causes gross developmental defects.  Our results reveal that transcription of most lncRNA genes is coordinated with transcription of protein-coding genes and that these lncRNAs have roles in early development. Overall design: Analysis of genome-wide lncRNA transcription in human embryonic stem cells and early differentiation using RNA-seq, GRO-seq and ChIP-seq","project":"SRP015819"}
{"number_samples":6,"species":"human","abstract":"To explore functionally crucial tumor-suppressive (TS)-miRNAs in hepatocellular carcinoma (HCC), we performed integrative function- and expression-based screenings of TS-miRNAs in six HCC cell lines.  The screenings identified seven miRNAs, which showed growth-suppressive activities through the overexpression of each miRNA and were endogenously downregulated in HCC cell lines.  Further expression analyses using a large panel of HCC cell lines and primary tumors demonstrated four miRNAs, miR-101, -195, -378 and -497, as candidate TS-miRNAs frequently silenced in HCCs.  Among them, two clustered miRNAs miR-195 and miR-497 showed significant growth-suppressive activity with induction of G1 arrest.  Comprehensive exploration of their targets using Argonute2-immunoprecipitation-deep-sequencing (Ago2-IP-seq) and genome-wide expression profiling after their overexpression, successfully identified a set of cell-cycle regulators, including CCNE1, CDC25A, CCND3, CDK4, and BTRC.  Our results suggest the molecular pathway regulating cell cycle progression to be integrally altered by downregulation of miR-195 and miR-497 expression, leading to aberrant cell proliferation in hepatocarcinogenesis. Identification of miR-195 and miR-497 target genes by sequencing Ago2-binding mRNAs and total mRNAs of miR-195 or miR-497 overexpressed, or non-treated Hep G2 cell. Overall design: Deep sequencing of RNAs in Ago2-IP fraction and mRNAs extracted from miR-195 or miR-497 overexpressed, or non-treated Hep G2 cell.","project":"SRP015845"}
{"number_samples":2,"species":"human","abstract":"5-methylcytosine (5-mC) can be oxidized to 5-hydroxymethylcytosine (5-hmC). Genome-wide profiling of 5-hmC thus far indicated 5-hmC may not only be an intermediate form of DNA demethylation but could also constitute an epigenetic mark per se. We describe a cost-effective and selective method to detect both the hydroxymethylation and methylation status of cytosines in more than 1.8 million MspI sites in the human genome. This method involves the selective glucosylation of 5-hmC residues, short-sequence tag generation and high-throughput sequencing. We tested this method by screening H9 human embryonic stem cells and their differentiated embroid body cells, and found that differential hydroxymethylation preferentially occur in bivalent genes during cellular differentiation. Especially, our results support hydroxymethylation can regulate key transcription regulators with bivalent marks through demethylation and affect cellular decision on choosing active or inactive state of these genes upon cellular differentiation. Overall design: In order to explore the role of methylation and hyroxymethylation in regulating gene expression upon cellular differentiation to EBs, we examined  the gene expression level in  H9 human embryonic stem cells and their differentiated embroid body cells by Digital gene expression (DGE), respectively.","project":"SRP015853"}
{"number_samples":9,"species":"human","abstract":"In the present work we report the molecular effects of two fullerene derivatives (1-C60 and 2-C60) on MCF-7 cell line by RNA-seq analysis.","project":"SRP015868"}
{"number_samples":4,"species":"human","abstract":"Histone modifications are now well-established regulators of transcriptional programs that distinguish distinct cell states. However, the kinetics of histone modification and their role in mediating rapid, signal-responsive changes in gene expression have been little studied on a genome-wide scale. Vascular endothelial growth factor A (VEGF), a major regulator of angiogenesis, rapidly triggers changes in transcriptional activity of human umbilical vein endothelial cells (HUVECs). Here we used chromatin immunoprecipitation and high throughput sequencing (ChIP-seq) to measure genome-wide changes in histone H3 acetylation at lysine 27 (H3K27ac), a marker of active enhancers {Kharchenko et al., 2011, Nature, 471, 480-5;Zentner et al., 2011, Genome Res, 21, 1273-83;Rada-Iglesias et al., 2011, Nature, 470, 279-83; Creyghton et al., 2010, Proc Natl Acad Sci U S A, 107, 21931-6 }, after 0, 1, 4, and 12 hours of VEGF stimulation. We show that sites with greatest H3K27ac changes were associated tightly with p300, a histone acetyltransferase. This dynamic H3K27ac signature defined transcriptional elements that are functionally linked to angiogenesis, participate in rapid VEGF-stimulated changes in chromatin conformation, and mediate VEGF-induced transcriptional responses. Dynamic H3K27ac deposition required p300 activity and did not involve altered nucleosome occupancy. Our results demonstrate that capture of dynamic changes in H3K27ac provides a new approach to define the activity of functional genomic elements and implicate epigenetic modifications in rapid signal-responsive transcriptional regulation. Overall design: ChiP-seq timecourse of H3K27ac, ETS1, p300 chromatin occupancy, mRNA expression and DNA hypersensitivity of HUVEC cells stimulated with VEGF for 0, 1, 4, and 12 hours","project":"SRP015904"}
{"number_samples":3,"species":"human","abstract":"To examine the role of WTAP in splicing regulation, we performed high-throughput mRNA sequencing (RNA-seq) on RNA isolated from control, WTAP or Virilizer siRNA-treated HUVECs, yielding 12 million uniquely mapped 75nt pair-end tags from each sample. MapSplice software was used for differential expression and differences in transcript splice junctions . Overall design: mRNA profiles of control, WTAP or Virilizer siRNA-treated HUVECs were generated by deep sequencing using Illumina GAII.","project":"SRP015909"}
{"number_samples":2,"species":"human","abstract":"The poly(A)-binding protein nuclear 1 (PABPN1) is a ubiquitously expressed protein that is thought to function during mRNA poly(A) tail synthesis in the nucleus. Despite the predicted role of PABPN1 in mRNA polyadenylation, little is known about the impact of PABPN1 deficiency on human gene expression. This experiment address by high-throughput transcriptome sequencing the global impact of a PABPN1 deficiency on human gene expression.","project":"SRP015926"}
{"number_samples":3,"species":"human","abstract":"We aimed to identify microRNAs that are regulated by YAP in human mammary epithelial cells. Overall design: We utilized deep sequencing technology to identify microRNAs that are induced by YAP overexpression and repressed by YAP knockdown.","project":"SRP015943"}
{"number_samples":6,"species":"human","abstract":"We determined the occupancy of SUMO-1 on chromatin in HeLa cells by use of chromatin affinity purification coupled with next generation sequencing RNA-Seq: The goal is to compare differential gene expression in SUMO-1 depleted cells and control cells (GL2 depleted) to verify the observation in the paper, and to cross-check with RT-PCR results. Overall design: 18 samples examined in syncrhonized HeLa cells: Cells in G1, early S (S0)/mid (S3)/ late S phase (S6), and mitosis (M), triplicates for each sample. 1 ChIP-SUMO1 sample in S0, and 1 ChIP-IgG control RNA-Seq: Six cDNA samples containing three pairs of biological replicates (three SUMO-1 depleted samples and three GL2 control samples) were barcoded, pooled together in equal concentration and subjected to sequencing in one lane of Illumina GAII","project":"SRP015955"}
{"number_samples":2,"species":"human","abstract":"Identification of the all RNA species coding and non-coding in total RNA Overall design: Relationship between DNMT1-RNA interactions, DNA methylation and gene expression","project":"SRP015964"}
{"number_samples":35,"species":"human","abstract":"Short-hairpin RNA (shRNA)-induced RNAi is used for biological discovery and therapeutics. Dicer, whose normal role is to liberate endogenous miRNAs from their precursors, processes shRNAs into different biologically active siRNAs, affecting their efficacy and potential for off-targeting. We found that in cells, Dicer induced imprecise cleavage events around the expected sites based on the previously described 5'/3'-counting rules. These promiscuous non-canonical cleavages were abrogated when the cleavage site was positioned 2 nt from a bulge or loop. Interestingly, we observed that the ~1/3 of mammalian endogenous pre-miRNAs that contained such structures were more precisely processed by Dicer. Implementing a new \"loop-counting rule\", we designed potent anti-HCV shRNAs with substantially reduced off-target effects. Our results suggest that Dicer recognizes the loop/bulge structure in addition to the ends of shRNAs/pre-miRNAs for accurate processing. This has important implications for both miRNA processing and future design of shRNAs for RNAi-based genetic screens and therapies. Overall design: Various shRNAs were expressed in Cell and processed by the RNase III enzyme Dicer.  The profiles of the siRNA products were generated by deep sequencing with or without the Ago2-IP.","project":"SRP015976"}
{"number_samples":7,"species":"human","abstract":"Gene regulation by EWS/FLI1 fusion in Ewing sarcoma.","project":"SRP015989"}
{"number_samples":12,"species":"human","abstract":"We performed Ago HITS-CLIP to identify targets of viral and human miRNAs in latently KSHV-infected PEL cells Overall design: Ago HITS-CLIP was performed in two latently infected PEL cell lines, BCBL-1 and BC-3; Argonaute-immunoprecipitation of UV cross-linked Ago-miRNA-mRNA complexes, followed by RNA isolation, library construction, and high-throughput sequencing (Illumina GAxII); we performed 3 biological replicates for each cell line, two technical (sequencing) replicates of BCBL-1 biological replicate 1","project":"SRP016003"}
{"number_samples":6,"species":"human","abstract":"We have used an integrative high content analysis approach to identify the specific miRNAs implicated in EGF signaling in HeLa cells as potential mediators of cancer mediated functions. We have used microarray and deep-sequencing technologies in order to obtain a global view of the EGF miRNA transcriptome with a robust experimental cross-validation. By applying a procedure based on Rankprod tests, we have delimited a solid set of EGF-regulated miRNAs. After validating regulated miRNAs by RT-qPCR, we have derived protein networks and biological functions from the predicted targets of the regulated miRNAs to gain insight into the potential role of miRNAs in EGF-treated cells. In addition, we have analyzed sequence heterogeneity due to editing relative to the reference sequence (isomirs) among regulated miRNAs. Overall design: Time course experiment comparing HeLa gene expression in response to EGF analyzed by small RNA-seq using Illumina 36-bp read massively parallel sequencing. Three independent experiments were performed where HeLa cells were serum deprived for 24 hours and were either left untreated or treated with EGF for 6h and harvested for RNA extraction.  Thus, a total of 6 samples were analyzed, 3 controls and the 3 respective treated counterparts. These same samples were also analyzed in parallel on two different microarray platforms.","project":"SRP016004"}
{"number_samples":8,"species":"human","abstract":"We performed RNA-seq experiments to identify differentially expressed intergenic transcripts between gastric cancer and normal tissues/cells. Overall design: Three primary cell culture samples from gastric cancer tissues, three gastric cancer cell lines and two normal tissue samples were used for the experiments.","project":"SRP016059"}
{"number_samples":9,"species":"human","abstract":"The RNA samples from HT-29 (ATCC) colon cancer cell line were reverse transcribed to build cDNA libraries and categorized into 3 groups with different concentrations of 5-aza-deoxy-cytidine (5-Aza); in each group three replicative 150 mm cultures were treated with: 1) dimethyl sulfoxide (vehicle alone, 0 µM 5-Aza); 2) 5µM 5-Aza and 3) 10 µM 5-Aza; for five days. This experiment was also performed parallel on a commercial Affymetrix microarray [GSE41364] and the aim of the study was to compare the two platforms on gene expression measurements and differentially expressed gene (DEG) detection. The results showed a strong correlation between the two platforms, yet it also confirmed the existence of fixed and proportional biases on the gene expression measurements between microarray and RNA-Seq. The DEG analysis indicated the relative superiority of DESeq method in terms of its performance; high consistency was confirmed between DESeq, baySeq methods from RNA-Seq and SAM/eBayes from microarray data. Overall design: Experiment on human colon cancer HT29 cell line treated with 3 concentrations of 5-aza-deoxy-cytidine","project":"SRP016118"}
{"number_samples":6,"species":"human","abstract":"Epstein-Barr virus (EBV) is a human herpesvirus linked to a number of B cell cancers and lymphoproliferative disorders. During latent infection, EBV expresses 25 viral pre-microRNAs (miRNAs) and induces the expression of specific host miRNAs, such as miR-155 and miR-21, which potentially play a role in viral oncogenesis. To date, a limited number of EBV miRNA targets have been identified; thus, the role of EBV miRNAs in viral pathogenesis is not well defined. Here, we used photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) combined with deep sequencing and computational analysis to comprehensively examine the viral and cellular miRNA targetome in EBV B95-8-infected lymphoblastoid cell lines (LCLs). We identified 7,827 miRNA-interaction sites in 3,492 cellular 3'UTRs. 531 of these sites contained seed-matches to viral miRNAs. 24 PAR-CLIP-identified miRNA:3'UTR interactions were confirmed by reporter assays. Our results reveal that EBV miRNAs predominantly target cellular transcripts during latent infection, thereby manipulating the host environment. Furthermore, targets of EBV miRNAs are involved in multiple cellular processes that are directly relevant to viral infection, including innate immunity, cell survival, and cell proliferation. Finally, we present evidence that myc-regulated host miRNAs from the miR-17/92 cluster can regulate latent viral gene expression. This comprehensive survey of the miRNA targetome in EBV-infected B cells represents a key step towards defining the functions of EBV-encoded miRNAs, and potentially, identifying novel therapeutic targets for EBVassociated malignancies. Overall design: Ago2 (Argonaute 2) PAR-CLIP and small RNA deep sequencing of Epstein-Barr virus B95.8-infected lymphoblastoid cell lines (LCLs).","project":"SRP016130"}
{"number_samples":5,"species":"human","abstract":"Corneal endothelium is composed of a monolayer of corneal endothelial cells (CECs) in the inner layer of cornea, which is essential for maintaining corneal transparency. In order to better characterize CECs in different developmental stages, we profiled mRNA transcriptomes in human fetal and adult corneal endothelium with the goal to identify novel molecular markers in these cells. By comparing CECs with 12 other types of tissues, we identified 245 and 284 signature genes that are highly expressed in fetal and adult CECs, respectively. Functionally, these genes are characteristic of CECs, involving in cell adhesion, proteoglycan and sulfur metabolic process. Importantly, several of these genes are disease target genes in hereditary corneal dystrophies, consistent with their functional significance in CEC physiology. By comparing fetal and adult CECs, we also identified stage-specific markers associated with CEC maturation, such as the activation of the Wnt pathway genes in fetal, but not in adult CECs. Lastly, by immunohistochemistry of ocular tissues, we further confirmed the unique protein expression patterns for Wnt5a, S100A4, S100A6, and IER3, the four novel markers for either fetal or adult CECs.  The identification of a new panel of molecular markers for fetal and mature CECs would be very useful for characterizing and quality controlling CECs through ex vivo expansion or stem cell differentiation for cell replacement therapy. Overall design: mRNA profile between adult and fetal CECs by high-throughput sequencing","project":"SRP016140"}
{"number_samples":29,"species":"human","abstract":"Reprogramming human somatic cells into induced pluripotent stem cells (iPSC) has been suspected of causing de novo copy number variations (CNVs). To explore this issue, we performed a whole-genome and transcriptome analysis of 20 human iPSC lines derived from primary skin fibroblasts of 7 individuals using next-generation sequencing. We find that, on average, an iPSC line manifests two CNVs not apparent in the fibroblasts from which the iPSC was derived. Using qPCR, PCR, and digital droplet PCR (ddPCR) to amplify across the CNVs' breakpoints, we show that at least 50% of those CNVs are present as low frequency somatic genomic variants in parental fibroblasts and are manifested in iPSC colonies due to their clonal origin. Hence, reprogramming does not necessarily lead to de novo CNVs in iPSC, since most of line-manifested CNVs reflect somatic mosaicism in the human skin. Moreover, our findings demonstrate that clonal expansion, and iPSC lines in particular, can be used as a discovery tool to reliably detect low frequency CNVs in the tissue of origin. Overall, we estimate that approximately 30% of the fibroblast cells have somatic CNVs, suggesting widespread somatic mosaicism in the human body. Our study paves the way to understanding the fundamental question of the extent to which cells of the human body normally acquire structural alterations in their DNA post-zygotically. Overall design: We have generated and characterized hiPSC lines derived from skin fibroblasts collected from seven members of two families, which were competent to be differentiated into neuronal progenitors and neurons","project":"SRP016568"}
{"number_samples":6,"species":"human","abstract":"The transcriptomic profiling of psoriasis has led to an increased understanding of disease pathogenesis. Although microarray technologies have been instrumental in this regard, it is clear that these tools detect an incomplete set of DEGs. RNA-seq can be used to supplement these prior technologies. Here, the use of RNAseq methods substantially increased the number of psoriasis-related DEGs. Furthermore, DEGs that were uniquely identified by RNA-seq, but not in other published microarray studies, further supported the role of IL-17 and tumor necrosis factor-a synergy in psoriasis. Examination of one of these factors at the protein level confirmed that RNA-seq is a powerful tool that can be used to identify molecular factors present in psoriasis lesions, and may be useful in the identification of therapeutic targets that to our knowledge have not been reported previously.  Further studies are in progress to determine the biological significance of DEGs uniquely discovered by RNA-seq. Overall design: To define the transcriptomic profile of psoriatic skin, three pairs of lesional and nonlesional skin biopsy specimens were taken from patients with untreated  moderate-to-severe plaque psoriasis.","project":"SRP016583"}
{"number_samples":1,"species":"human","abstract":"The prediction of genomic piRNA clusters relies on bioinformatics. We developed a new software for prediction of piRNA clusters that reduces false positive/negative annotations without leading to an artificial underestimation of transposon-related piRNA loci.","project":"SRP016589"}
{"number_samples":2,"species":"human","abstract":"miRNAs are small non-coding RNAs that inhibit translation and promote mRNA decay.  The levels of mature miRNAs are the result of different rates of transcription, processing, and turnover. The non-canonical polymerase Gld2 has been implicated in the stabilization of miR-122 possibly by catalyzing 3’ monoadenylation, however, there is little evidence that this relationship is one of cause and effect.  Here, we biochemically characterize Gld2 involvement in miRNA monoadenylation and its effect on miRNA stability.  We find that Gld2 directly monoadenylates and stabilizes specific miRNA populations in human fibroblasts and that sensitivity to monoadenylation-induced stability depends on nucleotides in the miRNA 3‘ end.  These results establish a novel mechanism of miRNA stability and resulting post-transcriptional gene regulation. Overall design: Sequencing of miRNAs to assess amount and 3' end monoadenylation state upon Gld2 knock-down.","project":"SRP016626"}
{"number_samples":3,"species":"human","abstract":"Human pluripotent stem cells are a promising source of diverse cells for developmental studies, cell transplantation, disease modeling, and drug testing.  However, their widespread use even for intensely studied cell types like spinal motor neurons, is hindered by the long duration and low yields of existing protocols for in vitro differentiation and by the molecular heterogeneity of the populations generated. We report a combination of small molecules that induce up to 50% motor neurons within 3 weeks from human pluripotent stem cells with defined subtype identities that are relevant to neurodegenerative diseases.  Despite their accelerated differentiation, motor neurons expressed combinations of HB9, ISL1 and column-specific markers that mirror those observed in vivo in human fetal spinal cord. They also exhibited spontaneous and induced activity, and projected axons towards muscles when grafted into developing chick spinal cord.  Strikingly, this novel protocol preferentially generates motor neurons expressing markers of limb-innervating lateral motor column motor neurons (FOXP1+/LHX3-). Access to high-yield cultures of human limb-innervating motor neuron subtypes will facilitate in-depth study of motor neuron subtype-specific properties, disease modeling, and development of large-scale cell-based screening assays. Overall design: We analyzed 3 samples including 2 positive samples and 1 negative sample. Descriptions are as follows: a) Positive Sample 1: SHH-derived, day 21 GFP-high FACS-purified motor neurons. b) Positive Sample 2: S+P-derived, day 21 GFP-high FACS-purified motor neurons. c) Negative: S+P condition, day 21 GFP-off FACS-purified non-motor neurons. Initial analysis of data was performed on ~40% of fastq reads (Amoroso et al., J Neurosci 2013 Jan 9;33(2):574-86. PMID: 23303937).  Further processing of the full dataset has since been carried out and the updated rpkm file and expression analysis reflecting all aligned reads can be accessed at: http://scholar.harvard.edu/amorosornaseq/","project":"SRP016629"}
{"number_samples":2,"species":"human","abstract":"Linker histones are essential components of chromatin but the distributions and functions of many during cellular differentiation is not well understood. Here, we show that H1.5 binds to genic and intergenic regions, forming blocks of enrichment, in differentiated human cells from all three embryonic germ layers but not in embryonic stem cells. In differentiated cells, H1.5, but not H1.3, binds preferentially to genes that encode membrane and membrane-related proteins. Strikingly, 37% of H1.5 target genes belong to gene family clusters, groups of homologous genes that are located in proximity to each other on chromosomes. H1.5 binding is associated with gene repression and is required for SIRT1 binding, H3K9me2 enrichment and chromatin compaction. Depletion of H1.5 results in loss of SIRT1 and H3K9me2, increased chromatin accessibility, deregulation of gene expression and decreased cell growth. Our data reveal for the first time a specific and novel function for linker histone subtype H1.5 in maintenance of condensed chromatin at defined gene families in differentiated human cells. Overall design: Examine mRNA expression in control and H1.5 knockdown IMR90 cells","project":"SRP016790"}
{"number_samples":6,"species":"human","abstract":"We performed translatome and transcriptome sequencing of A549, H1299 and HBE cells and analyzed the translation ratio (TR) of all genes Overall design: Total RNA and ribosome-bound RNA were extracted from A549, H1299 and HBE cells, and the polyA+ mRNA was sequenced using Illumina GAIIx and HiSeq-2000. The reads were mapped to RefSeq RNA reference sequences and the expression level were quantified by rpkM calculation.","project":"SRP017019"}
{"number_samples":2,"species":"human","abstract":"High-throughput RNA sequencing of a formalin-fixed, paraffin-embedded (FFPE) autopsy lung tissue samples from the 1918 and 2009 influenza pandemics. The goal of this project is to identify the influenza viruses in these preserved FFPE samples.","project":"SRP017105"}
{"number_samples":9,"species":"human","abstract":"We have performed genome-wide DNA methylation analysis at single base resolution and gene expression analysis, resulted in hypermethylation in all autosomes in DS samples, mediated by down-regulation of all three TET family genes, and down-regulation of REST/NRSF. Genes located on chr21 were up-regulated by a median of 45% in DS compared to normal villi, while genes with promoter hypermethylation were down regulated. Overall design: Comparison of RNA expression in human placenta between 5 normal and 4 Trisomy 21 samples.","project":"SRP017123"}
{"number_samples":16,"species":"human","abstract":"RNA-Seq analysis of atypical chronic myeloid leukemia samples Overall design: We sequenced leukemic mRNA from 13 Atypical Cronic Mieloid Leukemia (aCML) samples  by Illumina GAIIx. Transcriptomic profiles, differentially expressed genes and pathway enrichment analysis were obtained comparing 7 SETBP1-mutated samples and 6 non-mutated (WT) samples by using TopHat aligner and SAMMate gene expression quantifier. We focused on the gene expression profile of known coding transcripts. A dataset of 20,907 protein-coding Ensembl Genes was obtained from the RNA-Seq by using the Human Ensembl GTF annotation file vs54 dowloaded from ftp://ftp.ensembl.org/pub/release-54/gtf/homo_sapiens/.","project":"SRP017138"}
{"number_samples":6,"species":"human","abstract":"The goals of this study is to analyse transcriptionnal changes  in senescent cells and to compare them with ChIPseq data for several histones marks and proteins of the SUMO machinery Overall design: mRNA profiling of proliferative versus Ras-induced senescent humain primary fibroblasts 5 days post-infection","project":"SRP017142"}
{"number_samples":4,"species":"human","abstract":"The progression of cancer to metastatic disease is a major cause of death. We identified miR-708 being transcriptionally repressed by polycomb repressor complex (PRC2)-induced H3-K27 trimethylation in metastatic breast cancer.  miR-708 targets the endoplasmic reticulum protein neuronatin (Nnat) to decrease intracellular calcium (Ca2+) level, resulting in reduction of activation of ERK and FAK, decreased cell migration, and impaired metastases. Functional complementation experiments with Nnat-3’UTR mutant, which is refractory to suppression by miR-708, rescued cell migration and metastasis defects. In breast cancer patients, miR-708 expression was decreased in lymph node and distal metastases, suggesting a metastasis-suppressive role. Our findings uncover a mechanistic role for miR-708 in metastasis and provide a rationale for developing miR-708 as a therapeutic agent against metastatic breast cancer. Overall design: Sequencing miRNAs from Human breast cancer cells: MCF10A, MCF7, MDA-MB-231, MDA-MB-LM2","project":"SRP017190"}
{"number_samples":5,"species":"human","abstract":"We report on abundance and transcript profile characteristics of sperm RNAs. Overall design: Examination of RNA population and distribution in spermatozoa","project":"SRP017199"}
{"number_samples":1,"species":"human","abstract":"Homo sapiens HKCI5a Transcriptome.","project":"SRP017253"}
{"number_samples":43,"species":"human","abstract":"Therapy-related and de novo acute myeloid leukemia transcriptome sequencing","project":"SRP017262"}
{"number_samples":19,"species":"human","abstract":"Translational control permits cells to respond swiftly to changing environment. Rapid attenuation of global protein synthesis under stress conditions has been largely ascribed to the inhibition of translation initiation. To monitor the co-translational response to proteotoxic stress especially during elongation stage, we performed ribosome profiling (deep sequencing of ribosome-protected mRNA fragments) and investigated the roles of chaperone molecules in this process.","project":"SRP017263"}
{"number_samples":4,"species":"human","abstract":"To investigate whether U1C plays a role in splicing regulation in human system, we performed siRNA-mediated knockdown of U1C in HeLa cells and analyzed alternative splicing patterns by high-throughput RNA sequencing (RNAseq) Overall design: RNAseq performed with poly(A)+ selected total RNA from U1C-knockdown and control-treated HeLa cells","project":"SRP017294"}
{"number_samples":4,"species":"human","abstract":"Purpose: To identify all of the APA targets of CFIm25 on a global scale and develop an algorithm that can idenitify APA events from standard RNA-seq data Methods: RNA from HeLa cells treated with control siRNA and CFIm25 siRNA were subject to RNA-Seq. Using a custom-designed algorithm to mine RNA-seq data for novel APA events regulated by CFIm25. Results: We identified over 1,400 genes with shortened 3’UTRs after CFIm25 knockdown.  Importantly, we show that as a consequence of APA, many of these mRNAs have greatly enhanced protein expression due to the loss of destabilizing features within the 3’UTR. Conclusions: Our study underscored the critical function of the CFIm complex members in governing APA  and establish a previously unknown link between APA and metabolic pathways important for tumor progression. Overall design: Hela cell line mRNA profiles of control treated and CFIm25 Knockdown were generated by RNA-Seq using Illumina GAIIx.","project":"SRP017305"}
{"number_samples":2,"species":"human","abstract":"Although liganded nuclear receptors have been established to regulate RNA polymerase II (Pol II)-dependent transcription units, their role in regulating Pol III-transcribed DNA repeats remains largely unknown. Here we report that ~2-3% of the ~100,000-200,000 total human DR2 Alu repeats located in proximity to activated Pol II transcription units are activated by the retinoic acid receptor (RAR) in human embryonic stem cells to generate Pol III-dependent RNAs. These transcripts are processed, initially in a DICER-dependent fashion, into small RNAs (~28-65 nt) referred to as repeat-induced RNAs that cause the degradation of a subset of crucial stem-cell mRNAs, including Nanog mRNA, which modulate exit from the proliferative stem-cell state. This regulation requires AGO3-dependent accumulation of processed DR2 Alu transcripts and the subsequent recruitment of AGO3-associated decapping complexes to the target mRNA. In this way, the RAR-dependent and Pol III-dependent DR2 Alu transcriptional events in stem cells functionally complement the Pol II-dependent neuronal transcriptional program. Overall design: RNA-sequencing of polyA selected RNA molecules in NTera2/D1 cells and Global Run On (GRO) assay followed by high throughput sequencing (GRO-seq).","project":"SRP017330"}
{"number_samples":13,"species":"human","abstract":"Having developed a noninvasive method to sequence epithelial cells shed from the intestinal lining, this allowed testing of infants. We compared pooled RNA isolated from full term infants and pretern infants. This allowed for comparison of the differences in the healthy full term infant transcriptome and the extremely preterm infant transcriptome.","project":"SRP017352"}
{"number_samples":9,"species":"human","abstract":"We applied in parallel RNA-Seq and Ribosome-profiling analyses to immortalized human primary BJ fibroblast cells under the following conditions: normal proliferation, quiescence (induced by serum depletion), senescence (induced by activation of the oncogenic RASG12V gene, and examined at early (5 days; pre-senescent state) and late (14 days; fully senescent state) time points), and neoplastic transformation (induced by RASG12V in the background of stable p53 and p16INK4A knockdowns and SV40 small-T expression. Overall design: RNA-seq, using Illumina HiSeq 2000, was applied to BJ cells under 5 conditions: proliferation, quiescence, pre-senescence, full-senescence, and transfomed. Ribosome profiling, using Illumina HiSeq 2000, was applied to BJ cells under 5 conditions: proliferation, quiescence, pre-senescence, full-senescence, and transfomed.","project":"SRP017378"}
{"number_samples":13,"species":"human","abstract":"Submit for publication for Ting Ni","project":"SRP017395"}
{"number_samples":15,"species":"human","abstract":"The induction of pluripotency or trans-differentiation of one cell type to another  can be accomplished with cell lineage-specific transcription factors. Here we report that  repression of a single RNA binding protein PTB, which occurs during normal brain  development via the action of miR-124, is sufficient to induce trans-differentiation of  fibroblasts into functional neurons. Besides its traditional role in regulated splicing, we  show that PTB has a previously undocumented function in the regulation of microRNA  functions, suppressing or enhancing microRNA targeting by competitive binding on  target mRNA or altering local RNA secondary structure. A key event during neuronal  induction is the relief of PTB-mediated blockage of microRNA action on multiple  components of the REST complex, thereby de-repressing a large array of neuronal genes,  including miR-124 and multiple neuronal-specific transcription factors, in non-neuronal  cells. This converts a negative feedback loop to a positive one to elicit cellular  reprogramming to the neuronal lineage. Overall design: Examination of PTB regulated  AGO2/microRNA targeting in Hela cells by CLIP-seq (two biological replicates) , paired-end RNA-seq (control and PTB knockdown) and 3’end stability RNA-seq  (control and PTB knockdown)","project":"SRP017411"}
{"number_samples":8,"species":"human","abstract":"Uveal melanoma is the most common primary malignancy of the eye. By studying the expression of genes in primary uveal melanoma tumors we hope to gain an understanding of the underlying molecular biology. Expression analysis between uveal melanoma classes may make particularly important contributions to diagnosis and treatment decisions.","project":"SRP017413"}
{"number_samples":18,"species":"human","abstract":"Purpose: Asess the transcritpional changes induced upon RAB7 knock-down in melanoma (SK-Mel-28 and UACC-62) and in colon cancer (HCT-116) cell lines. Methods: mRNA profiles of tumor cell lines (SK-Mel-28, UACC-62, HCT-116) stably expressing scrambled shRNA or RAB7 shRNA (harvested at day 3 after lentiviral infection) were generated by deep sequencing, using three biological replicates per condition. The sequence reads that passed quality filters were analyzed with TopHat and Cufflinks. Validation of induced / silenced genes was performed by western blot. Results show a differential impact of RAB7 expression in the transcriptomic profile of melanoma vs non-melanoma cell lines, and support a lineage-specific role of this small GTPase in melanoma. Overall design: Examination of the mRNA profiles RAB7-depleted vs wild type cells, performed in parallel in 3 different tumor cell lines (Melanomas: SK-Mel-28 and UACC-62, Non-melanoma: HCT-116) harvested at day 3 after lentiviral infection.","project":"SRP017435"}
{"number_samples":16,"species":"human","abstract":"Assessing the transcriptional response of human white blood cells elicited by treating samples with inflammation inducing LPS.","project":"SRP017464"}
{"number_samples":21,"species":"human","abstract":"RNA sequencing data for human cell lines.","project":"SRP017465"}
{"number_samples":53,"species":"human","abstract":"Purpose:  Multiple studies from last decades have shown that the microenvironment of carcinomas plays an important role in the initiation, progression and metastasis of cancer.  Our group has previously identified novel cancer stroma gene expression signatures associated with outcome differences in breast cancer by gene expression profiling of two tumors of fibroblasts as surrogates for physiologic stromal expression patterns.  The aim of this study is to find additional new types of tumor stroma gene expression patterns. Results:  53 tumors were sequenced by 3SEQ with an average of 29 million reads per sample.  Both the elastofibroma (EF) and fibroma of tendon sheath (FOTS) gene signatures demonstrated robust outcome results for survival in the four breast cancer datasets.  The EF signature positive breast cancers (20-33% of the cohort) demonstrated significantly better outcome for survival.  In contrast, the FOTS signature positive breast cancers (11-35% of the cohort) had a worse outcome.  The combined stromal signatures of EF, FOTS, and our previously identified DTF, and CSF1 signatures characterize, in part, the stromal expression profile for the tumor microenvironment for between 74%-90% of all breast cancers. Conclusions:  We defined and validated two new stromal signatures in breast cancer (EF and FOTS), which are significantly associated with prognosis. Overall design: Gene expression profiling by 3SEQ was performed on 8 additional types of fibrous tumors, to identify different fibrous tumor specific gene expression signatures.  We then determined the significance of the fibrous tumor gene signatures in four publically available breast cancer datasets (GSE1456, GSE4922, GSE3494, NKI Dataset).","project":"SRP017575"}
{"number_samples":6,"species":"human","abstract":"Centromeres are essential to ensure proper chromosome segregation in eukaryotes. Their definition relies on the presence of a centromere-specific H3 histone variant CenH3, known as CENP-A in mammals. Its overexpression in aggressive cancers raises questions concerning its effect on chromatin dynamics and contribution to tumorigenesis. We find that CenH3 overexpression in human cells leads to ectopic enrichment at sites of active histone turnover. Furthermore, in over-expressing conditions, we see the formation of a novel heterotypic particle (CenH3-H4/H3.3-H4) that occludes CTCF binding, but this occlusion has only a minor effect on gene expression. Ectopic localization and CTCF occlusion depends on the H3.3 chaperone DAXX, rather than its dedicated chaperone HJURP. This DAXX-dependent occlusion also occurs in naturally overexpressing cancer cells. Furthermore, our cellular model reveals a DAXX-dependent survival advantage when challenged by DNA damage. Our findings illustrate how changes in histone variant levels can disrupt chromatin dynamics in disease and provides a possible mechanism for cell resistance to anti-cancer treatments. Overall design: Examination of CenH3 ChIP-seq and RNA-seq transcriptional programs in two conditions - WT and CenH3 over-expressing.","project":"SRP017577"}
{"number_samples":4,"species":"human","abstract":"To investigate the efficacy of nicotinamide treatment using our ex-vivo primary lymphocyte model, we performed high-throughput RNA sequencing on libraries generated from untreated and nicotinamide treated samples. Overall design: PBMC isolated from FRDA affected individuals were cultured to prepare the primary lymphocyte cell lines. The primary cultured cells were either treated with 10mM nicotinamide or without the addition of drug during the 3-days treatment. RNA was extracted after the treatment and then RNA-seq libraries were generated by standard protocols.","project":"SRP017580"}
{"number_samples":6,"species":"human","abstract":"The placenta is a key component in understanding the physiological processes involved in pregnancy. Characterizing genes critical for placental function can serve as a basis for identifying mechanisms underlying both normal and pathologic pregnancies. Detailing the placental tissue transcriptome could provide a valuable resource for genomic studies related to placental disease. We have conducted a deep RNA sequencing (RNA-Seq) study on three tissue components (amnion, chorion, and decidua) of 5 human placentas from normal term pregnancies. Our data demonstrate unique aspects of gene expression and splicing in placental tissues that provide a basis for disease investigation related to disruption of these mechanisms.","project":"SRP017583"}
{"number_samples":14,"species":"human","abstract":"Millions of cis-regulatory sequences have recently been found in the human genome, but the function of most cis-elements are not yet clear, in part due to the difficulty in determining their regulatory targets, which are often located millions of base pairs away and separated by one or more unrelated genes. To address this problem, the Hi-C method has been developed to identify long-range looping interactions in a genome-wide, unbiased fashion.  However, current data analysis of Hi-C datasets cannot fully resolve regulatory interactions between enhancers and promoters due to the low resolution.  Here, we generated a high-depth Hi-C dataset and applied a new analysis method that offers improved resolution permitting genome-wide identification of nearly one million chromatin interactions.  We demonstrated the use of Hi-C to identify target promoters of enhancers regulated by NF-?B signaling and signal-dependent dynamic chromatin interaction at these enhancers in human cells. Surprisingly, our results showed that most NF-?B binding sites are looped to their regulatory targets prior to activation of the signaling pathway, and appear to undergo little change during signaling. This observation suggests that the chromatin organization landscape, once established in a cell type, is rather stable and may influence the selection and activation of target genes by a transcription factor. Overall design: We performed Hi-C analysis using a human fibroblast cell line IMR90 before and after NF-?B activation. In the meantime, we also performed ChIP-seq experiments to map the location of NF-?B p65 subunit, RNA polymerase II, p300, and several histone modifications (including H3K4me1, H3K4me3, H3K27ac and H3K36me3) in IMR90 cells before and after transient TNF-a stimulation. Additionally, to monitor the dynamic transcription profiles, we also performed Global Run-On sequencing (GRO-seq).","project":"SRP017631"}
{"number_samples":3,"species":"human","abstract":"Aberrant DNA hypermethylation of CpG island (CGI) promoters are associated with transcriptional repression of many tumor suppressor genes and lead to tumor progression in many cancers. Most recently, one research group observed that aberrantly hypermethylated genes in multiple cancers are already repressed, but their promoters are maintained in a hypomethylated state in pre-cancerous tissues.Their studies didn't provide a clue to explain by what mechanisms those genes were repressed in pre-cancerous tissues. Another research group found that many genes with de novo promoter hypermethylation in colon cancer were among the subset of genes \"bivalently\" marked in embryonic stem cells and adult stem/progenitor cells by repressive Polycomb group proteins (PcG), which are known for maintaining low, but poised, transcription.These observations provide a clue that CGI promoter hypermethylation in cancers is associated with PcG target genes in pre-cancerous tissues.we took advantage of ChIP-BS-seq technology and applied it to examine H3K27me3 and H3K4me3 profiles for one normal lymphoblastoid cell line (YH) and three cancer cell lines including one cervical cancer cell line (Hela) and two gastric cancer (GC) cell lines (BGC-823 and AGS). Overall design: We aplied ChIP-BS technology to examine H3K27me3 marks, which are catalyzed by the SET domain histone methyltransferase EZH2 and have a repressive function with 50bp pair-end sequencing. found H3K27me3 marks were enriched preferentially at CpG islands, (+/-500) transcription start sites (TSSs) and exons in two GC cell lines (BGC-823 and AGS). In YH cells, H3K27me3 marks were only preferentially enriched at CpG islands. In contrast, Hela cells presented a reverse pattern with highest H3K27me3 enrichment in intergenic regions. To confirm this result in Hela cells, we performed two independent replicates of ChIP-Seq and ChIP-BS-seq. Cause of useful was relative small. we still sequenced one 100bp pe reads replicate for H3K4me3 and two replicate for H3K27me3 ChIP-BS-seq.","project":"SRP017644"}
{"number_samples":4,"species":"human","abstract":"By using  NGS-derived retinal transcriptome profiling (RNA-seq) to compare the gene expression profiling between 4 differently treated NPC cells Overall design: Examination of different gene expression in EBV-miRNA-BART1/3/7 lentivirus and their control infected nasopharyngeal carcinoma cells.","project":"SRP017670"}
{"number_samples":6,"species":"human","abstract":"In this study, we investigated its suitability for disease modeling by carrying out gene expression profiling, using RNA-seq, on neurons derived from iPSCs made from dental pulp extracted from deciduous teeth (T-iPSCs) and fibroblasts (F-iPSCs). Overall design: Comparison of expression profiles of iPSC derived from dental pulp and skin-fibroblast","project":"SRP017684"}
{"number_samples":2,"species":"human","abstract":"Small RNA expression was analysed in total RNA of HeLa cells treated with siRNA toward Luciferase (negative cotrol) or ADAR1. Overall design: Small RNA: 2 samples examined: HeLa cell with siLuc (negative cotrol), with siADAR1","project":"SRP017699"}
{"number_samples":2,"species":"human","abstract":"FUS binds the CTD of RNA polymerase II and regulates its phosphorylation at Ser2.","project":"SRP017717"}
{"number_samples":5,"species":"human","abstract":"TAF15, an RNA binding protein was recently implicated in Amyotrophic Lateral Sclerosis (ALS). ALS is a fatal neurodegenerative disease. We report the identification of  the conserved neuronal RNA targets of TAF15 and the assessment of the impact of TAF15 depletion on the neuronal transcriptome. Our study uncovers regulation of splicing of sets of neuronal RNAs encoding proteins with essential roles in synaptic activities including glutamergic receptors such as zeta-1 subunit of the glutamate N-methyl-D-aspartate (NMDA) receptor (Grin1). Overall design: Identification of TAF15 neuronal targets using normal human brain samples and mouse neurons.  Mouse background: E14Tg2a.4 wildtype cells derived from 129P2/OlaHsd.","project":"SRP017777"}
{"number_samples":6,"species":"human","abstract":"6 DGE sequencing human","project":"SRP017786"}
{"number_samples":6,"species":"human","abstract":"CXCL12 and IGF1 are key secreting molecules produced by cancer-associated fibroblasts in breast cancer. These factors promote the survival of disseminated cancer cells in the bone marrow. To assess the combined responses elicited by CXCL12 and IGF1, we examined the translating transcriptome of cancer cells in response to these two factors by Translating Ribosome Affinity Purification (TRAP)-RNAseq. Overall design: MDA-MB-231 cells were engineered to express an EGFP-tagged version of ribosomal protein L10a. This allows the retrieval of polysome-associated mRNA by anti-GFP pull down (TRAP) and profiling the translating transcriptome by RNAseq. EGFP-L10a+ cancer cells were serum starved (0.2% serum) for 24 hours, and then treated with CXCL12 (30ng/mL) + IGF1 (10ng/mL) or CXCL12 (300ng/mL) + IGF1 (100ng/mL) for 6hrs. Two biological replicates were profiled for each condition.","project":"SRP017788"}
{"number_samples":9,"species":"human","abstract":"A mesenchymal rich stroma such as cancer-associated fibroblasts (CAFs) in breast tumors favors the selection of cancer clones with enhanced bone metastatic ability. To determine the cancer cell transcriptomic response to the mesenchymal stroma, we supplemented experimental mammary tumours with or without exogenous mesenchymal cells. We used bone marrow-derived human mesenchymal stem cells (MSCs) as a source of mesenchymal stroma, as MSCs have been shown to undergo CAF-like differentiation. We engineered the cancer cells to express an EGFP-tagged version of ribosomal protein L10a (EGFP-L10a). This allows the retrieval of cancer cell specific transcripts rapidly from whole tumor lysates by translating ribosome affinity purification (TRAP) and direct profiling of cancer cell gene expression patterns when they are in situ. Overall design: EGFP-10a+ MDA-MB-231 cells were orthotopically injected into the mammary fat pad with or without 1:1 ratio of MSCs. The mammary tumors were retrieved for TRAP-RNAseq profiling after 3 weeks.","project":"SRP017789"}
{"number_samples":4,"species":"human","abstract":"The surprising observation that virtually the entire human genome is transcribed means we know very little about the function of many emerging classes of RNAs, except their astounding diversity. Traditional RNA function prediction methods rely on sequence or alignment information, which are limited in their ability to classify classes of non-coding RNAs (ncRNAs). To address this, we developed CoRAL, a machine learning-based approach for classification of RNA molecules. CoRAL uses biologically interpretable features including fragment length, cleavage specificity, and antisense transcription to distinguish between different ncRNA classes. We evaluated CoRAL using genome-wide small RNA sequencing (smRNA-seq) datasets from two human tissue types (brain and skin [GSE31037]), and were able to classify six different types of RNA transcripts with 79~80% accuracy in cross-validation experiments, and with 71~73% accuracy when CoRAL uses one tissue type for training and the other as validation. Analysis by CoRAL revealed that long intergenic ncRNAs, small cytoplasmic RNAs, and small nuclear RNAs show more tissue specificity, while microRNAs, small nucleolar, and transposon-derived RNAs are highly discernible and consistent across the two tissue types. The ability to consistently annotate loci across tissue types demonstrates the potential of CoRAL to characterize ncRNAs using smRNA-seq data in less characterized organisms. Overall design: Four samples were sequenced, each one coming from frozen brain tissue (frontal cortex) of a deceased female human patient with no remarkable pathology.","project":"SRP017809"}
{"number_samples":6,"species":"human","abstract":"RIPK4 but not the related kinases RIPK1, RIPK2, and RIPK3 caused similar transcriptional changes to Wnt3a. Overall design: PA1 cells were transfected by 8ug RIPK1, RIPK2, RIPK3, or RIPK4 for 48h, RNA were extracted and sequenced.","project":"SRP017843"}
{"number_samples":19,"species":"human","project":"SRP017875"}
{"number_samples":25,"species":"human","abstract":"Cap analysis of gene expression (CAGE) and massive parallel sequencing were used to profile the promoterome of aged human brains from five regions, namely: caudate, frontal cortex, hippocampus, putamen and temporal cortex. Overall design: 25 RNA  libraries from post-mortem brain tissue (five caudate, five frontal, 5 hippocampus, 5 putamen, five temporal RNA libraries from seven individuals) were processed using CAGE protocol and CAGE tags derived from the 25 libraries were sequenced with Illumina.","project":"SRP017933"}
{"number_samples":4,"species":"human","abstract":"RNA-seq and ribosome footprinting libraries of mouse 3T3 and human 293T cellsrelated to Shalgi et al.  2013 Overall design: 36bases paired-end RNA-seq, and ribosome footprinting libraries for: 3T3 cells - Control, 8 hours of mild heat shock (42) and 2 hours of severe heat shock (44) - in replicates, as well as 3T3 cells treated by mild followed by severe heat shock. In addition, 3T3 cells treated with Hsp70 inhibitor VER-155008 (Massey et al. 2010), and 293T cells transfected with Hspa1a or GFP, before and after 2 hours of severe heat shock.","project":"SRP017942"}
{"number_samples":12,"species":"human","abstract":"Only a minuscule fraction of long non-coding RNAs (lncRNAs) are well characterized. The evolutionary history of lncRNAs can provide insights into their functionality, but comparative analyses have been precluded by our ignorance of lncRNAs in non-model organisms. Here, we use RNA sequencing to identify lncRNAs in eleven tetrapod species and we present the first large-scale evolutionary study of lncRNA repertoires and expression patterns. We identify ~11,000 primate- specific lncRNA families, which show evidence for selective constraint during recent evolution, and ~2,400 highly conserved lncRNAs (including ~400 genes that likely originated more than 300 million years ago). We find that lncRNAs, in particular ancient ones, are generally actively regulated and may predominantly function in embryonic development. lncRNA X-inactivation patterns reveal an extremely female-biased monotreme-specific lncRNA, which may partially compensate X-dosage in this lineage. Most lncRNAs evolve rapidly in terms of sequence and expression levels, but global patterns like tissue specificities are often conserved. We compared expression patterns of homologous lncRNA and protein-coding families across tetrapods to reconstruct an evolutionarily conserved co-expression network. This network, which surprisingly contains many lncRNA hubs, suggests potential functions for lncRNAs in fundamental processes like spermatogenesis or synaptic transmission, but also in more specific mechanisms such as placenta growth suppression through miRNA production. Overall design: [Batch 1 and 2] To broaden our understanding of lncRNA evolution, we used an extensive RNA-seq dataset to establish lncRNA repertoires and homologous gene families in 11 tetrapod species. We analyzed the poly- adenylated transcriptomes of 8 organs (cortex/whole brain without cerebellum, cerebellum, heart, kidney, liver, placenta, ovary and testis) and 11 species (human, chimpanzee, bonobo, gorilla, orangutan, macaque, mouse, opossum, platypus, chicken and the frog Xenopus tropicalis), which shared a common ancestor ~370 millions of years (MY) ago. Our dataset included 47 strand-specific samples, which allowed us to confirm the orientation of gene predictions and to address the evolution of sense-antisense transcripts. See also GSE43721 (Soumillon et al, Cell Reports, 2013) for three strand-specific samples for mouse brain, liver and testis.","project":"SRP017959"}
{"number_samples":2,"species":"human","abstract":"Topoisomerases are necessary for the expression of neurodevelopmental genes, and are mutated in some patients with autism spectrum disorder (ASD). We have studied the effects of inhibitors of Topoisomerase 1 (Top1) and Topoisomerase 2 (Top2) enzymes on mouse cortical neurons. We find that topoisomerases selectively inhibit long genes (>100kb), with little effect on all other gene expression. Using ChIPseq against RNA Polymerase II (Pol2) we show that the Top1 inhibitor topotecan blocks transcriptional elongation of long genes specifically. Many of the genes inhibited by topotecan are candidate ASD genes, leading us to propose that topoisomerase inhibition might contribute to ASD pathology. Overall design: [Mouse] 5 biological replicates of transcriptome sequencing (RNAseq) from topotecan treated neurons and vehicle treated controls; Pol2 ChIPseq of topotecan and vehicle treated neurons [Human] Transcriptome sequencing (RNAseq) from topotecan treated neurons and vehicle treated control.","project":"SRP017966"}
{"number_samples":12,"species":"human","abstract":"We report the application of bi-species RNAseq for investigating mechanisms of reprogramming towards pluripotency using heterokaryons (mouse embryonic stem cell X human fibroblast cell fusions). The use of mixed species allows one to monitor reprogramming of the human somatic nuclei independently of contributions from the mouse nuclei using nucleotide differences. We used RNAseq to monitor heterokaryon reprogramming over a 3-day timecourse, generating transcriptome-wide data for cell fusion based reprogramming of human fibroblasts towards pluripotency. Overall design: Examination of cellular reprogramming over a heterokaryon timecourse (mouse embryonic stem cells X human fibroblasts)","project":"SRP017972"}
{"number_samples":4,"species":"human","abstract":"A pair of stage III colorectal cancer (CRC) tumor and adjacent normal tissue were collected from a patient without Transcatheter Arterial Infusion chemotherapy (TAI) during surgical resection of CRC tumors. Another pair of stage III CRC tumor and adjancent normal tissue were collected from another patient who had been treated with TAI one week before surgical resection. Total RNA of the collected tissues were extracted using TRIzol reagent (Invitrogen) according to the manufacturer's instructions. The integrity of the RNA was checked with an ultraviolet spectrophotometry and 2100 BioAnalyzer (Agilent Technologies, Santa Clara, CA, USA). Construction of small RNA libraries from size fractionated RNA was carried out by following Solexa protocol and the obtained libraries were sequenced by Illumina HiSeq 2000 sequencer. We identified some de-regulated miRNAs in CRC tumor tissues. The expression levels of miR-142-5p were significantly reduced in tumor tissues of stage III CRC, then were increased significantly in tumor tissues receiving TAI and higher than tumor tissues without TAI. miR-142-5p is a potential tumor suppressor in CRC and is upregulated in tumor tissues after TAI, suggesting its potential clinical values for testing the functionality of TAI and predicting progress of CRC. Overall design: Examination of small RNA transcriptomes in colorectal tumor and adjancent normal tissues without and with Transcatheter Arterial Infusion chemotherapy using Illumina HiSeq2000 sequencer.","project":"SRP017979"}
{"number_samples":94,"species":"human","abstract":"We sequenced the RNA of 42 bladder cancer patients.","project":"SRP018008"}
{"number_samples":6,"species":"human","abstract":"DNA Topoisomerase I (Top1) relaxes DNA supercoiling and is inhibited with high specificity by camptothecin, a natural product of chinese tree Camptotheca acuminata with anticancer activity. Topoisomerase activity is required at transcribing regions to modulate DNA supercoils generated by RNA polymerases. However, Top1 functions at promoters and molecular responses to CPT are not fully understood. We found that camptothecin increases antisense RNA polymerase II transcripts at active divergent CpG-island promoters in a replication-independent manner. Kinetics investigations of the formation of Top1-DNA cleavage complexes and non-B DNA structures showed that CPT interferes with Top1 modulation of negative DNA supercoiling at promoters. The present findings will be a resource to establish the role of such antisense RNAs in transcription regulation and to discover additional components of the response pathway. Moreover, the transcriptional camptothecin effects can be the molecular basis of the therapeutic activity in cancer as well as neurological syndromes.","project":"SRP018010"}
{"number_samples":1,"species":"human","abstract":"Circular RNAs (circRNAs) in animals are an enigmatic class of RNAs with unknown function. To systematically explore circRNAs, we sequenced and computationally analyzed human, mouse and nematode RNA. We detected thousands of well-expressed, stable circRNAs, with oftentimes tissue/developmental stage specific expression. Sequence analysis suggested important regulatory functions for circRNAs. Indeed, we discovered that human circRNA CDR1as is densely bound by miRNA effector complexes and harbors 63 conserved binding sites for the ancient miRNA miR-7. Further analyses indicated that CDR1as functions to bind miR-7 in neuronal tissues. Human CDR1as expression in zebra fish impaired midbrain development similar to knocking down miR-7, suggesting that CDR1as is a miRNA antagonist with a miRNA binding capacity ten times higher than any other known transcript. Together, our data provide evidence that circRNAs form a large class of post-transcriptional regulators. Numerous circRNAs form by head-to-tail splicing of exons, indicating previously unrecognized regulatory potential of coding sequences. Overall design: 1 Sample","project":"SRP018014"}
{"number_samples":16,"species":"human","abstract":"RNA-Seq technique was applied to investigate the effects of two semen collection methods (Pelleted vs Liquefied) and two sperm purification methods (SCLB vs PS) to the integrity of isolated RNAs at different perspectives. Overall design: The same set of semen samples were applied to investigate the qualitative and quantitative effect of semen collection methods and sperm cell purification methods on sperm transcript profiling.","project":"SRP018020"}
{"number_samples":7,"species":"human","abstract":"Gene regulation by GLI1 in Ewing sarcoma.","project":"SRP018022"}
{"number_samples":3,"species":"human","abstract":"SF3B1, splicing factor 3b subunit 1 is a component of the RNA splicing machinery and it has been found to be highly mutated in refractory anemia with ring sideroblasts (RARS) and RARS associated with thrombocytosis (RARS-T). The aim of this study is to investigate the RNA splicing patterns in bone marrow from patients with SF3B1 somatic mutations compared to normal physiologic splicing. We compared the RNA splicing pattern as exon usage, differential gene expression, and pathway analysis with normal splicing Overall design: mRNA profiles, as measured by RNA-Seq, in bone marrow derived mononuclear cells from 2 RARS patients with SF3B1 somatic mutations were compared to the mRNA profile of a normal donor.","project":"SRP018028"}
{"number_samples":10,"species":"human","abstract":"Background: In recent years, a variety of small RNAs derived from other RNAs with well-known functions such as tRNAs and snoRNAs, have been identified. The functional relevance of these RNAs is largely unknown. To gain insight into the complexity of snoRNA processing and the functional relevance of snoRNA-derived small RNAs, we sequenced long and short RNAs, small RNAs that co-precipitate with the Argonaute 2 protein and RNA fragments obtained in photoreactive nucleotide-enhanced crosslinking and immunoprecipitation (PAR-CLIP) of core snoRNA-associated proteins. Results: Analysis of these data sets revealed that many loci in the human genome reproducibly give rise to C/D box-like snoRNAs, whose expression and evolutionary conservation are typically less pronounced relative to the snoRNAs that are currently catalogued. We further found that virtually all C/D box snoRNAs are specifically processed inside the regions of terminal complementarity, retaining in the mature form only 4-5 nucleotides upstream of the C box and 2-5 nucleotides downstream of the D box. Sequencing of the total and Argonaute 2-associated populations of small RNAs revealed that despite their cellular abundance, C/D box-derived small RNAs are not efficiently incorporated into the Ago2 protein. Conclusions: We conclude that the human genome encodes a large number of snoRNAs that are processed along the canonical pathway and expressed at relatively low levels. Generation of snoRNA-derived processing products with alternative, particularly miRNA-like, functions appears to be uncommon. Overall design: PAR-CLIP profiling for snoRNP core proteins NOP56, NOP58, Fibrillarin, and Dyskerin in HEK293 cells. Small RNA profiling using RNA-seq in HEK293 and HeLa cells, small RNA profiling using IP-seq of Ago2 associated small RNAs.","project":"SRP018104"}
{"number_samples":26,"species":"human","abstract":"The poor clinical outcome in pancreatic ductal adenocarcinoma (PDA) has been attributed to intrinsic resistance to chemotherapy and a growth-permissive tumor microenvironment.  Quiescent pancreatic stellate cells (PSCs) are neuroendocrine, nestin-positive, lipid-accumulating cells whose homologues in the liver are the principal repository of Vitamin A esters.  Upon activation, lipid droplets are lost and via transdifferentiation they become the key cell type responsible for driving the severe desmoplasia that characterizes PDA.  Despite their critical role in PDA progression and chemoresistance, therapeutic strategies targeting PSCs are lacking.  Here we identified the vitamin D receptor (VDR) as a master genomic regulator of PSC activation and function.  In vitro we demonstrate that VDR activation reduces expression in PSCs of genes implicated in activation, inflammation, and extracellular matrix production, as well as restoring lipid droplet integrity. In vivo, the VDR ligand calcipotriol enhances the anti-tumor effects of gemcitabine by increasing intratumoral concentration 5-fold, reducing tumor volume to near baseline and lowering metastases by more than 65%.  These findings implicate VDR as a master regulator of PSC activation and identify a novel therapeutic approach for the treatment of pancreatic cancer. Overall design: RNA-Seq analyses was used to characterize cancer-associated changes between pre-activated (3-day culture) and activated (7-day culture) primary mouse PSCs, as well as control and PDA human PSCs.  RNA-Seq was also used to assess the impact of VDR activation (DMSO vs calcipotriol) in a human PSC line (MiaPaCa-2), the mouse primary PSCs","project":"SRP018218"}
{"number_samples":2,"species":"human","abstract":"Vascular permeability is frequently associated with inflammation and it is triggered by chemokines and by a cohort of secreted permeability factors, such as VEGF.  In contrast, here we showed that the physiological vascular permeability that precedes implantation is directly controlled by progesterone receptor (PR) and it is independent of VEGF. Both global and endothelial-specific deletion of PR block physiological vascular permeability in the uterus while misexpression of PR in the endothelium of other organs results in ectopic vascular leakage. Integration of genome-wide transcriptional profile of endothelium and ChIP-sequencing revealed that PR induces a NR4A1 (Nur77/TR3) specific transcriptional program that broadly regulates vascular permeability in response to progesterone.  This program triggers concurrent suppression of several junctional proteins and leads to an effective, timely and venule-specific regulation of vascular barrier function. Silencing NR4A1 blocks PR-mediated permeability responses indicating a direct link between PR and NR4A1. These results reveal a previously unknown function for progesterone receptor on endothelial cell biology with consequences to physiological vascular permeability and implications to the clinical use of progestins and anti-progestins on blood vessel integrity. Overall design: Examination of PR target genes in human umbilical vein endothelial cells (HUVECs) using RNA-seq (PR infected only -PR only and PR infected followed by ligand treatment-PR+P)","project":"SRP018234"}
{"number_samples":4,"species":"human","abstract":"The androgen receptor (AR) mediates the action of androgens by binding to androgen-responsive elements (AREs) and subsequently regulating target genes involved in prostate carcinogenesis. The precise locations, true nature, and functional roles of AREs in human prostate cancer are still unknown. Here we redefine AREs by motif-resolution AR chromatin immunoprecipitation-exonuclease (ChIP-exo) assay in human prostate cancer cells and tumors. Surprisingly, we find that, in addition to canonical full-length AREs and half-site-like AREs, a significant portion of the four redefined ARE categories comprises non-canonical full-length AREs. The redefined AREs in enhanced AR binding regions in prostate tumors versus paired non-malignant adjacent tissues regulate a prostate cancer-relevant gene network not only centered on AR, but more interestingly, on novel AR target genes mTOR, BIRC5 and BCL2L1 involved in prostate cancer cell growth and survival. The precise redefinition of AREs has important implications for understanding how AR contributes to prostate carcinogenesis. Overall design: To accurately define ARE in human genome and identify cancer related AR binding site, AR ChIP-exo is performed in LNCaP cells before/after androgen stimulation, malignant prostate tumors and non-malignant adjacent tissues. Each experiment includes two replicates.","project":"SRP018241"}
{"number_samples":5,"species":"human","abstract":"We explored the RNA binding properties of LINE-1 ORF1p, both free and in the L1 RNP, using a recently developed photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation technique (PAR-CLIP) to comprehensively identify ORF1p binding sites in the transcriptome of human cells (HEK293T). Our results show that ORF1p binds to a wide range of cellular mRNAs, with an enrichment for binding at the 3’ UTR. Our data also show that ORF1p binds very strongly with retrotransposable RNA, i.e., L1, Alu and SVA. Overall design: PAR-CLIP analysis of L1 RNPs and free ORF1p RNA binding profiles, comparison to HuR RNA binding profile","project":"SRP018242"}
{"number_samples":10,"species":"human","abstract":"AML1-ETO, a fusion protein generated by the t(8;21) translocation in acute myeloid leukemia, is a transcription factor implicated in both gene repression and activation. We now show that, in leukemic cells, AML1-ETO resides in and functions through a stable protein complex (AETFC) that contains several hematopoietic transcription factors and cofactors. In conjunction with biochemical and leukemia pathological studies, the ChIP-seq and RNA-seq analyses of the AETFC components in leukemic cells reveal that these components stabilize the complex through multivalent interactions, provide multiple DNA-binding domains for diverse target genes, colocalize genome-wide, cooperatively regulate gene expression, and contribute to leukemogenesis. Overall design: RNA-seq analyses gene expression upon knockdown of each AETFC component, including AML1-ETO, HEB, E2A, LYL1, LDB1 and LMO2, and double-knockdown of HEB and E2A, in Kasumi-1 cells. ChIP-seq analyses of four AETFC components, namely AML1-ETO, HEB, E2A and LMO2, in Kasumi-1 cells.","project":"SRP018254"}
{"number_samples":6,"species":"human","abstract":"In this study, we used Global Run-On sequencing (GRO-seq), a method that assays the genome-wide location and orientation of all active RNA polymerases. We generated a global profile of active transcription at ERa binding sites in MCF-7 human breast cancer cells in response to short time course of E2 treatment. This method enabled us to detect active transcription at enhancers and define a class of primary transcripts transcribed uni- or bidirectionally from the ERa binding sites. The raw data used in this study is from GSE27463 but sequenced to a greater depth. Overall design: Using GRO-seq over a time course (0, 10, 40 min) of estrogen signaling in ER-alpha positive MCF-7 human breast cancer cells.","project":"SRP018256"}
{"number_samples":4,"species":"human","abstract":"Colorectal carcinoma (CRC) is one of the most common cancers worldwide. Re-evaluating our current knowledge on CRC and developing novel therapeutic strategies is still crucial. Accumulating evidence suggests that cancer cells possess characters reminiscent of those of normal stem cells. Unveiling small RNAs responsible for cell stemness and chemoradioresistance should eventually lead to the development of novel therapeutic approaches. Overall design: Expression profiles of parental CRC cells and cancer spheres expanded under stem cell medium cultivation were generated for identifying key regulators.","project":"SRP018284"}
{"number_samples":1,"species":"human","abstract":"Access RNA-seq data as a source of SNP calls.","project":"SRP018304"}
{"number_samples":2,"species":"human","abstract":"We report high throughput transcriptomic profiling with RNA-Sequencing (RNA-Seq) to uncover network responses in human THP-1 monocytes treated with high glucose (HG). Overall design: Examination of differential expression between normal and high glucose condition in THP1 cells.","project":"SRP018312"}
{"number_samples":4,"species":"human","abstract":"AGO-PAR-CLIP was employed to identify microRNA binding sites in BCBL-1, a Kaposi's sarcoma-associated herpesvirus (KSHV) infected B-cell line and DG75, a KSHV negative B-cell line as a control. By using our novel computational method (PARma) and differential analysis of PAR-CLIP data, highly accurate target sites of KSHV microRNAs can be defined. Overall design: Examination of microRNA target sites in two different cell lines using replicate PAR-CLIP experiments","project":"SRP018317"}
{"number_samples":3,"species":"human","abstract":"We report the transcriptome human primary hepatocytes and liver sinusoidal endothelial cells. Hepatocytes were obtained from commercial sources.  LSECs were isolated based on the coexpression of Tie2 and CD32b, te strategy of purification controlled by RNA-Seq. Overall design: Comparison of the expression of the Tie-2, CD32b, SELP, FVIII, VWF, Alb, Fg, F7genes","project":"SRP018359"}
{"number_samples":2,"species":"human","abstract":"Prostate glands predominantly exhibit androgen dependence, but increasing evidence suggests that estrogen receptor signaling is involved in its development and pathogenesis. By integrating ChIP sequencing for estrogen receptor alpha (ERa) with transcriptome sequencing data from prostate cancer samples, we found ERa to significantly influence the noncoding transcriptome in prostate cancer. We identified one such long noncoding RNA, NEAT1, to play an important role in prostate cancer progression through direct regulation of transcription of its target genes. NEAT1, in an ERa dependent manner, promotes prostate tumorigenesis by interacting with and modulating chromatin state at promoters of prostate cancer specific signature genes. NEAT1 expression is positively correlated with PSMA in prostate adenocarcinoma and with B3GAT1 in neuroendocrine prostate cancer. This study identifies NEAT1 as a novel biomarker or therapeutic target in prostate cancer and also suggests that co-targeting ERa and androgen receptor (AR) may be effective for a subset of patients with advanced prostate cancer and with NEAT1 overexpression. mRNA profiles of MEF cell lines prepared from E13.5 embryos of wild-type (WT) and NEAT1 knockout (KO; NEAT1-/-) mice  were generated by deep sequencing, using Illumina HiSeq 2000. Strand specific mRNA profiles of VCaP and VCaP ERa cell lines were generated by deep sequencing, using Illumina GA IIx. Overall design: RNA sequencing of MEF (WT vs NEAT1 KO) and VCaP (control vs ERa)","project":"SRP018377"}
{"number_samples":4,"species":"human","abstract":"A multilayered transcription regulatory system is unveiled, where protein- and RNA-based repressors are super-imposed in combinatorial fashion to govern the timely triggering of an essential step of erythropoiesis Overall design: Analyses of transcriptional profiles and chromatin state in KAP1 WT and KO (or KD)  cells","project":"SRP018403"}
{"number_samples":29,"species":"human","abstract":"Mammalian preimplantation development is a complex process involving dramatic changes in the transcriptional architecture. Through single-cell RNA-sequencing (RNA-seq), we report here a comprehensive analysis of transcriptome dynamics from oocyte to morula in both human and mouse embryos. Based on single nucleotide variants (SNVs) in blastomere mRNAs and paternal-specific SNPs, we identify novel stage-specific monoallelic expression patterns for a significant portion of polymorphic gene transcripts (25-53%). By weighted gene co-expression network analysis (WGCNA), we find that each developmental stage can be concisely delineated by a small number of functional modules of co-expressed genes. This result indicates a sequential order of transcriptional changes in pathways of cell cycle, gene regulation, translation, and metabolism in a step-wise fashion from cleavage to morula.  Cross-species comparisons reveal that the majority of human stage-specific modules (7 out of 9) are remarkably preserved, only to diverge in developmental specificity and timing in mice. We further identify conserved key members (or hub genes) of the human and mouse networks. These genes represent novel candidates that are likely key players in driving mammalian preimplantation development. Collectively, we demonstrate that mammalian preimplantation development is orchestrated by evolutionarily conserved genetic programs that diverge in developmental timing.  Our results provide a valuable resource to dissect gene regulatory mechanism underlying progressive development of early mammalian embryos. Overall design: single-cell RNA-seq of human and mouse blastomeres","project":"SRP018525"}
{"number_samples":8,"species":"human","abstract":"We have develped a novel method of making siRNAs (named pro-siRNA for prokaryotic siRNA). To evaluate off-targeting of pro-siRNA, we compared the mRNA expression profiles of HeLa-d1EGFP cells transfected with 4 nM EGFP siRNAs and pro-siRNAs by microarray. Overall design: We used microarray to study the off-target effect of siRNAs in the HeLa-d1EGFP cell line. After transfection of siRNAs for 24 hrs, RNA were extracted using Trizol. Deep sequencing libraries were generated using the NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #E7530). HeLa-d1EGFP cells are HeLa cells stably expressing d1EGFP gene. EGFP siRNA is a siRNA made by chemical synthesis. EGFP100 and EGFPFL are pro-siRNAs made from either a 100 bp hairpin or a full length hairpin targeting EGFP coding sequence.","project":"SRP018552"}
{"number_samples":12,"species":"human","abstract":"Recent studies of genome-wide chromatin interactions have revealed that the human genome is partitioned into many self-associating topological domains. The boundary sequences are enriched for binding sites of CTCF and the cohesin complex, implicating these two factors in the establishment or maintenance of topological domains.  To determine the role of cohesin and CTCF in higher order chromatin architecture in human cells, we proteolytically cleaved the cohesin complex from interphase chromatin and examined changes in chromosomal organization as well as transcriptome. We observed a general loss of local chromosomal interactions upon disruption of cohesin complex, but the topological domains remain intact. However, we found that depletion of CTCF by RNA interference in these cells not only reduced intra-domain interactions but also increased inter-domain interactions. Further more, distinct groups of genes become mis-regulated upon depletion of cohesin and CTCF. Taken together, these observations suggest that CTCF and cohesin contribute in different ways to chromatin organization and gene regulation. Overall design: Hi-C and mRNA-seq experiments in Cohesin and CTCF depleted HEK293 cells","project":"SRP018571"}
{"number_samples":24,"species":"human","abstract":"When adapting to environmental stress, cells attenuate and reprogram their translational output.  In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs.  The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking.  Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq.  Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction.  Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction.  To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq.  Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated.  Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. Overall design: In this sub-series, CLIP and RNAseq data from the arsenite treatment experiment are presented.  Experiments on 293S cells +/- arsenite treatment, in 4 biological replicates.  On each treatment/replicate biological sample, both a CLIP-seq protocol and an RNA-seq protocol were performed, so these datasets are “paired” in addition to the treatment pairing.","project":"SRP018716"}
{"number_samples":14,"species":"human","abstract":"When adapting to environmental stress, cells attenuate and reprogram their translational output.  In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs.  The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking.  Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq.  Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction.  Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction.  To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq.  Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated.  Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. Overall design: In this sub-series, CLIP and RNAseq data from the emetine treatment experiment are presented.  Experiments on 293S cells +/- emetine treatment, in 1 biological replicate.  Here, for each treatment/replicate sample, an aliquot was used for RNA-seq, and the rest was split into three aliquots to perform 3 parallel CLIP-seq protocols with different antibodies.  So, each RNA-seq dataset here corresponds to 3 CLIP-seq datasets.","project":"SRP018717"}
{"number_samples":16,"species":"human","abstract":"When adapting to environmental stress, cells attenuate and reprogram their translational output.  In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs.  The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking.  Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq.  Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction.  Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction.  To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq.  Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated.  Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. Overall design: In this sub-series, CLIP and RNAseq data from the hippuristanol treatment experiment are presented.  Experiments on 293S cells +/- hippuristanol treatment, in 2 biological replicates.  Here, for each treatment/replicate sample, an aliquot was used for RNA-seq, and the rest was split into three aliquots to perform 3 parallel CLIP-seq protocols with different antibodies.  So, each RNA-seq dataset here corresponds to 3 CLIP-seq datasets.","project":"SRP018718"}
{"number_samples":16,"species":"human","abstract":"When adapting to environmental stress, cells attenuate and reprogram their translational output.  In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs.  The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking.  Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq.  Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction.  Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction.  To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq.  Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated.  Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. Overall design: In this sub-series, RNAseq data from sucrose gradient fractions with arsenite treatment are presented.","project":"SRP018719"}
{"number_samples":6,"species":"human","abstract":"Autosomal-recessive loss of the NSUN2 gene has been recently identified as a causative link to intellectual disability disorders in humans. NSun2 is an RNA methyltransferase modifying cytosine-5 in transfer RNAs (tRNA). Whether NSun2 methylates additional RNA species is currently debated. Here, we adapted the individual-nucleotide resolution UV cross-linking and immunoprecipitation method (iCLIP) to identify NSun2-mediated methylation in RNA transcriptome. We confirm site-specific methylation in tRNA and identify messenger and non-coding RNAs as potential methylation targets for NSun2. Using RNA bisulfite sequencing we establish Vault non-coding RNAs as novel substrates for NSun2 and identified six cytosine-5 methylated sites. Furthermore, we show that loss of cytosine-5 methylation in Vault RNAs causes aberrant processing into argonaute-associating small RNA fragments (svRNA). Thus, impaired Vault non-coding RNA processing may be an important contributor to the etiology of NSUN2-deficieny human disorders. Overall design: Identification of Nsun2 targets by miCLIP in Embryonic kidney (HEK293) cells","project":"SRP018723"}
{"number_samples":1,"species":"human","abstract":"We have used stranded RNA-seq to explore RNA editing in H9 cells Overall design: Examination RNA editing with stranded RNA-seq","project":"SRP018777"}
{"number_samples":2,"species":"human","abstract":"We proposed that besides TET family dioxygenase oxidizing 5mC to 5hmC, there is a non-enzyme pathway which is due to hydroxyl radica l(OH) or OH-like species also involvement in demethylation of 5mC forming 5hmC. This pathway includes classical fenton reagents such as H2O2 and Fe2+, and more important redox-activity quinoid compounds, especially, tetrachloro-1,4-benzoquinone (TCBQ), which was reported producing hydroxyl radicals independent of transition metal Overall design: Examination of 5hmC and transcriptome levels with TCBQ and DMSO in human MRC-5 cell lines","project":"SRP018778"}
{"number_samples":6,"species":"human","abstract":"Recent genome wide association studies have identified a number of genes that contribute to the risk for coronary heart disease. One such gene, TCF21, encodes a basic-helix-loop-helix transcription factor believed to serve a critical role in the development of epicardial progenitor cells that give rise to coronary artery smooth muscle cells (SMC) and cardiac fibroblasts. Using reporter gene and immunolocalization studies with mouse and human tissues we have found that vascular TCF21 expression in the adult is restricted primarily to adventitial cells associated with coronary arteries and also medial SMC in the proximal aorta of mouse. Genome wide RNA-Seq studies in human coronary artery SMC (HCASMC) with siRNA knockdown found a number of putative TCF21 downstream pathways identified by enrichment of terms related to CAD, including “vascular disease,” “disorder of artery,” and “occlusion of artery” as well as disease-related cellular functions including “cellular movement,” and “cellular growth and proliferation.” In vitro studies in HCASMC demonstrated that TCF21 expression promotes proliferation and migration and inhibits SMC lineage marker expression. Detailed in situ expression studies with reporter gene and lineage tracing revealed that vascular wall cells expressing Tcf21 before disease initiation migrate into vascular lesions of ApoE-/- and Ldlr-/- mice. While Tcf21 lineage traced cells are distributed throughout the early lesions, in mature lesions they contribute to the formation of a subcapsular layer of cells, and others become associated with the fibrous cap. The lineage traced fibrous cap cells activate expression of SMC markers and growth factor receptor genes. Taken together, these data suggest that TCF21 may have a role regulating the differentiation state of SMC precursor cells that migrate into vascular lesions and contribute to the fibrous cap and more broadly, in view of the association of this gene with human CAD, provide evidence that these processes may be a mechanism for CAD risk attributable to the vascular wall. Overall design: RNA-Seq: 3 versus 3 technical replicates, siControl versus siTCF21","project":"SRP018779"}
{"number_samples":8,"species":"human","abstract":"Autosomal-recessive loss of the NSUN2 gene has been recently identified as a causative link to intellectual disability disorders in humans. NSun2 is an RNA methyltransferase modifying cytosine-5 in transfer RNAs (tRNA). Whether NSun2 methylates additional RNA species is currently debated. Here, we adapted the individual-nucleotide resolution UV cross-linking and immunoprecipitation method (iCLIP) to identify NSun2-mediated methylation in RNA transcriptome. We confirm site-specific methylation in tRNA and identify messenger and non-coding RNAs as potential methylation targets for NSun2. Using RNA bisulfite sequencing we establish Vault non-coding RNAs as novel substrates for NSun2 and identified six cytosine-5 methylated sites. Furthermore, we show that loss of cytosine-5 methylation in Vault RNAs causes aberrant processing into argonaute-associating small RNA fragments (svRNA). Thus, impaired Vault non-coding RNA processing may be an important contributor to the etiology of NSUN2-deficieny human disorders. Overall design: mRNA-seq in Embryonic kidney (HEK293) cells transfected with siRNA against Nsun2 vs control","project":"SRP018786"}
{"number_samples":12,"species":"human","abstract":"During development neuronal progenitors compete for growth factors such as nerve growth factor NGF and require the prolyl hydroxylase EglN3 and the kinesin KIF1Bß for developmental apoptosis. Inherited KIF1Bß loss-of-function mutations in neuroblastomas and pheochromocytomas implicate KIF1Bß as a 1p36.2 tumor suppressor, however the mechanism of tumor suppression is unknown. We found that KIF1Bß interacts with the RNA helicase A (DHX9) resulting in DHX9 nuclear accumulation to regulate apoptosis. KIF1Bß-dependent DHX9 nuclear localization leads to transcription of the apoptotic target XIAP-associated factor 1. DHX9 is induced when NGF is limiting and required for apoptosis in cells deprived of NGF. Overall design: NB1 cells were transduced to incorporate shRNA against DHX9 or a scrambled control, and transfected with a KIF1Bß expression vector or control, then transfected cells were isolated and lysed after 48h.","project":"SRP018815"}
{"number_samples":6,"species":"human","abstract":"Human LIN28A and B are RNA-binding proteins (RBPs) conserved in animals with important roles during development and stem cell reprogramming. We used Photoactivatable-Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation (PAR-CLIP) in HEK293 cells and identified a largely overlapping set of ~3,000 mRNAs at ~9,500 sites located in the 3’UTR and CDS. In vitro and in vivo, LIN28 preferentially bound single-stranded RNA containing a uridine-rich element and one or more flanking guanosines, and appeared to be able to disrupt base-pairing to access these elements when embedded in predicted secondary structure. In HEK293 cells, LIN28 protein binding mildly stabilized target mRNAs and increased protein abundance. The top targets were its own mRNAs and those of other RBPs and cell-cycle regulators. Alteration of LIN28 protein levels also negatively regulated the abundance of some, but not all let-7 miRNA family members, indicating sequence-specific binding of let-7 precursors to LIN28 proteins and competition with cytoplasmic miRNA biogenesis factors. Overall design: To assess whether miRNAs are regulated by LIN28B we analyzed the miRNA levels of LIN28B overexpressing and LIN28B-depleted cells using small RNA cDNA library sequencing. The RBP LIN28B was depleted by siRNAs and the expression levels was compared to mock-transfected HEK293 cells.","project":"SRP018836"}
{"number_samples":4,"species":"human","abstract":"Human LIN28A and B are RNA-binding proteins (RBPs) conserved in animals with important roles during development and stem cell reprogramming. We used Photoactivatable-Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation (PAR-CLIP) in HEK293 cells and identified a largely overlapping set of ~3,000 mRNAs at ~9,500 sites located in the 3’UTR and CDS. In vitro and in vivo, LIN28 preferentially bound single-stranded RNA containing a uridine-rich element and one or more flanking guanosines, and appeared to be able to disrupt base-pairing to access these elements when embedded in predicted secondary structure. In HEK293 cells, LIN28 protein binding mildly stabilized target mRNAs and increased protein abundance. The top targets were its own mRNAs and those of other RBPs and cell-cycle regulators. Alteration of LIN28 protein levels also negatively regulated the abundance of some, but not all let-7 miRNA family members, indicating sequence-specific binding of let-7 precursors to LIN28 proteins and competition with cytoplasmic miRNA biogenesis factors. Overall design: LIN28 protein PAR-CLIP","project":"SRP018837"}
{"number_samples":62,"species":"human","abstract":"RNA-seq transcriptome measurements are typically performed by isolating RNA from large numbers of cells in culture or tissues. While highly informative, such experiments mask the variability in gene expression patterns that exists between individual cells. To gain insight into the dynamics of gene expression on the level of single-cells, we have carried out the transcriptomes of single-cells from the GM12878 cell line using RNA-seq. Overall design: Single GM12878 cells were picked and RNA-seq libraries were generated using the SMART-seq protocol. We also carried out RNA-seq experiments on pools of 10, 30 and 100 cells, on 100pg and 10ng of total RNA, and on pools of 10 cells that were subsequently split into 10 separate sample and processed as if they were single cells in order to assess technical variation in our experiments.","project":"SRP018838"}
{"number_samples":3,"species":"human","abstract":"We surveyed RNA-Seq data to identify those TEs that are transcriptionally active uniquely in human pluripotent cells. We identified one endogenous retrovirus (HERV-H) family, uniquely found in primates as being unusually abundant in the transcriptome. Overall design: Poly(A)+ RNA extraction and further sequencing library construction was done for two types of cells (iPS, HFF-1) and hiPSC-derived embryoid bodies following Illumina TruSeq RNA Sample Preparation Kit protocol, which was further sequenced on Illumina HiSeq machine with single-end 101 cycles.","project":"SRP018848"}
{"number_samples":80,"species":"human","abstract":"Type 1 diabetes (T1D) is an autoimmune disease characterized by the destruction of pancreatic insulin-producing ß cells. CD4+ T cells are integral to the pathogenesis of T1D, but biomarkers that define their pathogenic status in T1D are lacking. miRNAs have essential functions in a wide range of tissues/organs, including the immune system. We reasoned that CD4+ T cells from individuals at high risk for T1D (pre-T1D) might be distinguished by an miRNA signature. We sorted CD4+ T cells from 9 healthy and 7 pre-T1D individuals into 6 subsets, namely naïve, resting regulatory (rTreg), activated regulatory (aTreg), transitional memory (Ttm), central memory (Tcm) and effector memory (Tem) cells, and then compared miRNA profiles between these subsets and between pre-T1D and healthy individuals by deep sequencing. Differential expression of miRNAs was detected in each of the CD4+ T cell subsets. For example, expression of miRNAs that induce apoptosis (miR-15a) or FOXP3 instability (miR-31) was increased in rTreg and aTreg cells, respectively, in pre-T1D individuals, whereas miR-150 was increased in Tem cells of pre-T1D individuals. Importantly, increased miR-150 expression could be detected by qRT-PCR in total CD4+ T and PBMCs of pre-T1D individuals. Consistent with it being a marker of pathogenic CD4+ T cells, we showed that miR-150 regulates IFN-? production in mouse CD4+ T cells. Thus, comprehensive profiling identifies miRNA profiles that not only distinguish CD4+ T cell subsets but also discriminate individuals with preclinical T1D. The ability to detect differentially expressed miRNAs in total CD4+ T cells or PBMCs should facilitate clinical application of miRNAs as biomarkers. Overall design: CD4+T cells from healthy and individuals at high risk for autoimmune type 1 diabetes were sorted into 6 subsets, which resulted in 80 samples, 38 for healthy and 42 for high risk individuals. Each sample was barcoded and miRNA libraries were constructed and subsequently subjected to deep-sequencing on the Illumina GAII or HiSeq platform. The Fastq files are have deconvoluted and stripped of the barcode adaptor sequences.","project":"SRP018853"}
{"number_samples":21,"species":"human","abstract":"In mammalian cells, the Myc oncoprotein binds to thousands of promoters. During mitogenic stimulation of primary lymphocytes, Myc promotes an increase in expression of virtually all genes. In contrast, Myc-driven tumour cells differ from normal cells in expression of specific sets of up- and downregulated genes that have significant prognostic value. To understand this discrepancy, we studied the consequences of inducible expression and depletion of Myc in human cells and murine tumour models. Changes in Myc levels activate and repress specific sets of direct target genes that are characteristic of Myc-transformed tumour cells. Three factors account for this specificity: First, the magnitude of response parallels the change in occupancy by Myc at each promoter. Functionally distinct classes of target genes differ in the E-box sequence bound by Myc, arguing that different cellular responses to physiological and oncogenic Myc levels are controlled by promoter affinity. Secondly, Myc both positively and negatively affects transcription initiation independent of its effect on transcriptional elongation. Third, complex formation with Miz1 mediates repression of multiple target genes by Myc and the ratio of Myc and Miz1 bound to each promoter correlates with the direction of response. Overall design: Myc, Miz1 and RNA polymerase II ChIPseq as well as RNAseq experiments in two human cancer cell lines and murine carcinoma cells as well as fibroblasts from Miz1?POZ mice. All sequencing experiment were performed on an Illumina Genome Analyzer IIx.","project":"SRP018861"}
{"number_samples":8,"species":"human","abstract":"The airways of the human lung are lined by an epithelium made up of ciliated and secretory luminal cells and undifferentiated p63+ Krt5+ basal cells. The integrity of this epithelium and its ability to act as a selective barrier are critical for normal lung function. In other epithelia there is evidence that transcription factors of the evolutionarily conserved grainyheadlike (GRHL) family play key roles in co-ordinating the expression of numerous proteins required for epithelial morphogenesis, differentiation, remodeling and repair. However, little is known about their function in the adult lung. Here, we focus on the role of GRHL2 in primary human bronchial epithelial (HBE) cells, using either shRNA or a dominant negative protein (DN-GRHL2) to inhibit its function. We follow changes in epithelial phenotype, and in gene transcription using RNA-seq or microarray analysis, both in undifferentiated basal cells and in cells differentiating in air-liquid interface culture into a mucociliary epithelium with transepithelial electrical resistance. We identify several hundreds of genes that are directly or indirectly regulated by GRHL2. Using ChIP-seq to map sites of GRHL2 binding in the basal cells we identify 7,687 potential primary targets, and confirm that GRHL2 binding is strongly enriched near GRHL-regulated genes. Different subsets of the large cohort of potential GRHL2 targets appear to be active in basal and differentiated cells. Taken together, the results strongly support the hypothesis that GRHL2 plays a key role in regulating many physiological functions of human airway epithelium, including those involving cell adhesion, polarity and morphogenesis. Overall design: Frozen primary human bronchial epithelial (HBE) cells were obtained from three donors. Passage 2 cells at 40% confluence were infected with H2B-GFP or DN-GRHL2 lentivirus and 1 mg/ml puromycin added 48 h later. At confluence, Doxycycline 0.5 mg/ml was added for 24 h. RNA-seq was performed on all six samples, as well as samples from two donors that were not infected.","project":"SRP018883"}
{"number_samples":2,"species":"human","abstract":"Cumulus cells are biologically distinct from other follicular cells and perform specialized roles, transmitting signals within the ovary and supporting oocyte maturation during follicular development. The bi-directional communication between the oocyte and the surrounding cumulus cells is crucial for the acquisition of oocyte competence. Using Illumina/deep-sequencing technology, we dissected the small RNAome of pooled human mature MII oocytes and cumulus cells. Overall design: Cumulus cells and MII mature oocytes small RNA profiles were generated by deep-sequencing, using Illumina 1G sequencer","project":"SRP018933"}
{"number_samples":2,"species":"human","abstract":"The gastric cancer cell line AGS was purchased from the American Type Culture Collection (ATCC). And the AGS-EBV is the AGS cell line was infected by EBV","project":"SRP018972"}
{"number_samples":7,"species":"human","abstract":"We report the design and implementation of a \"breakpoint analysis\" pipeline to discover novel gene fusions by tell-tale transcript level or genomic DNA copy number transitions occurring within genes.  We use this method to prioritize candidate rearrangements from high density array CGH datasets as well as exon-resolution expression microarrays.  We mine both publicly available data as well as datasets generated in our laboratory.  Several gene fusion candidates were chosen for further characterization, and corresponding samples were profiled using paired end RNA sequencing to discover the identity of the gene fusion.  Using this approach, we report the discovery and characterization of novel gene fusions spanning multiple cancer subtypes including angiosarcoma, pancreatic cancer, anaplastic astrocytoma, melanoma, breast cancer, and T-cell acute lymphoblastic leukemia.  Taken together, this study provides a robust approach for gene fusion discovery, and our results highlight a more widespread role of fusion genes in cancer pathogenesis. Overall design: Breakpoint analysis for the discovery of novel gene fusions across human cancers","project":"SRP019207"}
{"number_samples":4,"species":"human","abstract":"Epstein-Barr virus (EBV) has evolved exquisite controls over its host cells, human B lymphocytes, not only directing these cells during latency to proliferate and thereby expand the pool of infected cells, but also to survive and thereby persist for the lifetime of the infected individual.  Although these activities ensure the virus is successful, they also make the virus oncogenic, particularly when infected people are immunosuppressed. Here we show, strikingly, that one set of EBV’s miRNAs both sustain BL (Burkitt’s lymphoma) cells in the absence of other viral oncogenes and promote the transformation of primary B lymphocytes.  Burkitt’s Lymphoma cells were engineered to lose EBV and found to die by apoptosis and could be rescued by constitutively expressing viral miRNAs in them.  Two of these EBV miRNAs were found to target Caspase 3 to inhibit apoptosis at physiological concentrations. Overall design: Examination of RISC associated transcripts under 4 conditions in Sav S1-1 cells","project":"SRP019222"}
{"number_samples":2,"species":"human","abstract":"DNA methylation plays a significant role in assuring cell identity, thus potentiating its application in molecular classification of cancers in respect of tissue origins or clinically and aetiologically distinct subtypes. In this study, we adapted our liquid hybridization capture-based bisulfite sequencing approach on the targeted sequencing of promoter methylomes. We detected ten cell lines originated from different tissue origins and demonstrated a similar potentiality of promoter methylomes as classifiers for cancer cell lines from different tissue origins in comparison with gene expression profiles. Furthermore, promoter methylome can sensitively differentiate two different cell lines from the same tissue origin in respect of the CpG island methylator phenotype (CIMP), as in the case of AGS and BGC-823 gastric cancer cell lines. These results potentiate the targeted sequencing of promoter methylomes as a means for comprehensive screening and classifying cancer cells with respect to tissue-origins and CIMP subtypes in the future studies. Overall design: We proved the potentiality of promoter-targeted LHC-BS that requires reduced experimental cost and less amount of initial DNA samples in comparison with a previous design [23] using YH cell line. In addition, we generated single-base promoter methylomes for ten cell lines, including eight cancer cell lines generated from four types of tissues and one pair of model cell lines for ovarian cancer (T29 and T29H). We proved the potentiality of promoter-targeted LHC-BS that requires reduced experimental cost and less amount of initial DNA samples in comparison with a previous design using YH cell line. In addition, we generated single-base promoter methylomes for ten cell lines, including eight cancer cell lines generated from four types of tissues and one pair of model cell lines for ovarian cancer (T29 and T29H).","project":"SRP019240"}
{"number_samples":17,"species":"human","abstract":"Long non-coding RNAs (lncRNAs) regulate diverse biological processes including cell lineage specification. Here we report transcriptome profiling of human endoderm and pancreatic cell lineages using purified cell populations. Analysis of the data sets allowed the identification of hundreds of lncRNAs exhibiting differentiation stage-specific expression patterns. As a first step in characterizing these lncRNAs, we focus on an endoderm-specific lncRNA DEANR1 (Definitive Endoderm Associated long Non-coding RNA1) and demonstrate that it plays an important role in human endoderm differentiation. DEANR1 contributes to endoderm differentiation by positively regulating endoderm factor FOXA2 expression. Importantly, overexpression of FOXA2 is able to rescue endoderm differentiation defects caused by DEANR1 depletion. Mechanistically, DEANR1 facilitates FOXA2 activation by helping SMAD2/3 recruitment to the FOXA2 promoter. Thus, our study not only reveals a large set of differentiation stage-specific lncRNAs, but also characterizes a novel lncRNA important for endoderm differentiation. Overall design: Here we perform RNA-seq based transcriptome profiling using undifferentiated human ESCs (one sample), definitive endoderm (two replicates), pancreatic progenitors(three replicates) as well as sorted human primary alpha cells (two replicates), beta cells (two replicates) and exocrine cells (one sample). Moreover, we perform RNA-seq based transcriptome profiling for WT, FOXA2 knockdown, and DEANR1 knockdown of differentiated definitive endoderm samples (two replicates for each).","project":"SRP019241"}
{"number_samples":4,"species":"human","abstract":"In this study, we hypothesized that IL-27 could induce the expression of novel miRNAs in macrophages which may have functional relevance in terms of anti-viral activity. In this study, primary monocytes were differentiated into macrophages using M-CSF (M-Mac) or with a combination of M-CSF and IL-27 (I-Mac) for seven days. Following this, total RNA was extracted from these cells and deep sequencing was performed, in parallel with gene expression microarrays. Using the novel miRNA discovery software, miRDeep, seven novel miRNAs were discovered in the macrophages, four of which were expressed higher in I-Mac (miRNAs 2.1, 8.1, 9.1 and 14.2) whilst three were detected in both M-Mac and I-Mac (miRNAs 9.3, 13.6 and 15.8). The expression of six of the seven novel miRNAs was highly correlated with qRT-PCR using specific primer/probes designed for the novel miRNAs. Gene expression microarray further demonstrated that a number of genes were potentially targeted by these differentially expressed novel miRNAs. Overall design: screening novel and known miRNAs which may have antiviral properties in 2 different treatments in 2 donors.","project":"SRP019248"}
{"number_samples":15,"species":"human","abstract":"RBM10 is an RNA binding protein that was identified as a component of spliceosome complex, suggesting its potential role in splicing regulation. However, the direct experimental evidence for this function has been lacking. Here we characterized in vivo RBM10-RNA interactions and investigated the role of RBM10 in splicing regulation at the global level. We observed significant RBM10-RNA interactions in the vicinity of splice sites and identified hundreds of splicing changes following perturbation of cellular RBM10 abundance. A RNA splicing map integrating the binding pattern and splicing profiles revealed a significant correlation between RBM10-enhanced exon skipping events and its binding close to the splicing sites of both upstream and downstream introns. Furthermore, we demonstrated the splicing defects in a patient carrying a RBM10 mutation. Overall, our data provided insights into the mechanistic model of RBM10-mediated splicing regulation and established genomic resources for future studies on its function  in different pathophysiological contexts. Overall design: We sequenced the mRNA of HEK293 cells and LCL cells, and we determined the RBM10 binding sites using PARCLIP in HEK293 cells. In total we sequenced four mRNA-Seq libraries for KD and two for OE in HEK293 cells; for each of these libraries, we also sequenced one control library. We also sequenced the mRNA of one patient LCL and two normal LCL libraries. Two replicates of PARCLIP sequencing were perfomed.","project":"SRP019250"}
{"number_samples":2,"species":"human","abstract":"Combinatorial transcription factor (TF) interactions regulate hematopoietic stem cell formation, maintenance and differentiation, and are increasingly recognised as drivers of stem cell signatures in cancer.  However, genome-wide combinatorial binding patterns for key regulators do not exist in primary human hematopoietic stem/progenitor cells (HSPCs) and have constrained analysis of the global architecture of the molecular circuits controlling these cells.  Here we provide new high-resolution genome-wide binding maps of seven key TFs (FLI1, ERG, GATA2, RUNX1, SCL, LYL1 and LMO2) in human CD34+ HSPCs together with quantitative RNA and microRNA expression profiles.  We catalogue binding of TFs at coding genes and microRNA promoters and report that combinatorial binding of all seven TFs is favoured and is associated with differential expression of genes and microRNA in HSPCs.   We also uncover a hitherto unrecognized association between FLI1 and RUNX1 pairing in HSPCs, establish a correlation between the density of histone modifications, which mark active enhancers and the number of overlapping TFs at a peak and identify complex relationships between specific miRNAs and coding genes regulated by the heptad.  Taken together, this study demonstrates that a heptad of TFs forms a dense auto-regulatory core in human HSPCs with binding of all seven TFs at tissue specific regulatory elements of heptad genes and collectively regulates miRNAs that in turn target components of the heptad and genes regulated by the heptad. Overall design: Examination of cominatorial binding by 7 transcription factors, 1 IgG control along with mRNA and small RNA sequencing in human CD34+ cells","project":"SRP019263"}
{"number_samples":10,"species":"human","abstract":"Long noncoding RNAs (lncRNAs) have emerged as an important layer of genome regulation with common mechanistic themes including the formation of ribonucleoprotein complexes. Here, we present a novel X-linked lncRNA termed linc-Firre that escapes X-chromosome inactivation and forms trans-chromosomal interactions required for adipogenesis. Linc-Firre is exclusively nuclear and forms punctate expression foci on chromatin near its site of transcription on both X-chromosomes in human and mouse. Both the localization of linc-Firre and the association with the nuclear matrix protein hnRNPU require a conserved repeating RNA domain, R2D2. Collectively, these results reveal a lincRNA that escapes X-chromosome inactivation with a critical role in driving cell fate decisions by trans-chromosomal interactions. Overall design: Replicate RNA-Seq analyses of oligo-mediated knockdowns of linc-FIRRE and hnRNPU in two different cellular contexts; HeLa cells and mouse embryonic stem cells.  Also included are Firre knockout and wild-type mouse embryonic stem cells.","project":"SRP019270"}
{"number_samples":362,"species":"human","abstract":"The genetics of messenger RNA expression has been extensively studied in humans and other organisms, but little is known about genetic factors contributing to microRNA (miRNA) expression. We examined natural variation of miRNA expression in adipose tissue in a population of 200 men who have been carefully characterized for metabolic syndrome phenotypes as part of the METSIM study. We genotyped the subjects using high-density SNP microarrays and quantified the mRNA abundance using genome-wide expression arrays and miRNA abundance using next generation sequencing. We reliably quantified 356 miRNA species that were expressed in human adipose tissue, a limited number of which made up most of the expressed miRNAs. We mapped the miRNA abundance as an expression quantitative trait and determined cis regulation of expression for 9 of the miRNAs and of the processing of one miRNA (miR-28). The degree of genetic variation of miRNA expression was substantially less than that of mRNAs. For the majority of the miRNAs, genetic regulation of expression was independent of the host mRNA transcript expression. We also showed that for 108 miRNAs, mapped reads displayed widespread variation from the canonical sequence. We found a total of 24 miRNAs to be significantly associated with metabolic syndrome traits. We suggest a regulatory role for miR-204-5p which was predicted to inhibit ACACB, a key fatty acid oxidation enzyme that has been shown to play a role in regulating body fat and insulin resistance in adipose tissue. Overall design: miRNA expression profiling of adipose tissue isolated from 200 humans","project":"SRP019272"}
{"number_samples":8,"species":"human","abstract":"Next-generation sequencing has revolutionized cancer biology by accelerating the unbiased discovery of novel mutations across human cancers. Transforming such discoveries into a conceptual framework of cancer progression requires narrowing the vast number of mutations down to the driver elements, and further reducing these to mutations that govern cancer progression as opposed to tumor initiation. By integrating next-generation RNA-sequencing (RNA-seq) with in vivo selection, we devise an approach that identifies a series of novel recurrent non-synonymous amino acid mutations that are enriched in metastatic breast cancer cells and predicted to significantly alter protein function. These mutations, found in PANX1, RBFA, REST, KRIT1 and ZSWIM6, are detected at higher frequencies in the transcriptomes of two patients’ highly metastatic sub-lines relative to their poorly metastatic parental lines. We functionally characterize the cellular and molecular roles of one of these mutations—a nonsense alteration that yields a truncated pannexin-1 (PANX11-89) plasma membrane megachannel subunit—in metastatic progression. PANX11-89 interacts with full-length PANX1 and augments PANX1 channel activity to promote the survival of cancer cells as they are mechanically deformed. Protection from deformation-induced cell death requires PANX1 channels to release ATP, which acts as a cell autonomous survival signal during mechanical stress. Functional characterization of additional nonsense and missense PANX1 mutations detected in epithelial cancers of the colon, lung, and prostate reveals that these mutants also enhance PANX1-mediated ATP release. In vivo testing of one such truncating mutation detected in a metastatic colorectal tumor also enhances early survival, dissemination and liver metastatic colonization by human colon cancer cells. Finally, pharmacological inhibition of PANX1 inhibits breast cancer metastasis, implicating PANX1 as a novel therapeutic target in cancer. Our findings reveal that ATP release through mechanosensitive PANX1 channels enables cancer cells to overcome a major metastasis suppressive barrier—deformation-induced death in the microvasculature. Overall design: To systematically identify base-pair mutations present in metastatic cells that may drive cancer progression, we performed whole-transcriptome RNA-sequencing (RNA-seq) of in vivo-selected, highly metastatic human breast cancer cell sub-lines, CN-LM1A and MDA-LM2, as well as the CN34 and MDA-MB-231 parental lines from which they were derived. To minimize the false positive rate and allow for subsequent statistical analysis, we sequenced biological replicates of each cell line. Mutations conferring enhanced metastatic capacity should be enriched in the transcriptomes of highly metastatic cells relative to their less metastatic parental populations.","project":"SRP019275"}
{"number_samples":4,"species":"human","abstract":"The goal of this study was to determine the similarity between human dermal microvascular endothelial cells, induced endothelial cells from fibroblasts, and fibroblasts through RNA-seq expression analysis.  RNA samples from independently induced cultures, plus fibroblast and human dermal microvascular endothelial cultures were converted into individual cDNA libraries using Illumina TruSeq methods and subjected to single-end 50 base-sequence analysis at 20-30 million read depths. Overall design: Examination of one fibroblast culture, one human dermal mibrovascular endothelial cell culture, and two induced endothelial cell cultures.","project":"SRP019374"}
{"number_samples":3,"species":"human","abstract":"Androgen-stimulated growth of the molecular apocrine breast cancer is mediated by an androgen receptor (AR)-regulated transcriptional program.  Through profiling the genomic licalizations of AR and its co-regulators FOXA1 and TCF7L2 in MDA-MB-453 breast cancer cells, we revealed the molecular details of the AR-centered regulatory network. We further identified that  c-MYC is a key downstream target co-regulated by AR, FOXA1 and TCF7L2, and reinforces the transctiopnal activation of androgen-responsive genes in this subtype of breast cancers. Overall design: MDA-MB-453 breast cancer cells were transfected with control of MYC siRNA for 48 h, followed by treatment with 10nM DHT or vehicle for 6 h. The cells were subjected to mRNA purification and library praparation for RNA-seq on Illumina HiSeq2000 platform.","project":"SRP019498"}
{"number_samples":8,"species":"human","abstract":"VCaP Xenografts in mice.","project":"SRP019503"}
{"number_samples":8,"species":"human","abstract":"RNA-seq data from HT-29 cells treated with IFN-? for 24 hr, MCF10A cells, and MDA-MB-436 cells. Overall design: mRNA profiles of HT-29, MCF10A, and MDA-MB-436 were generated by deep sequencing using Illumina HiSeq 2000. All RNA sequencing data was generated by the Genomics Services Lab at the HudsonAlpha Institute for Biotechnology (Huntsville, AL).","project":"SRP019758"}
{"number_samples":14,"species":"human","abstract":"While Neanderthals are extinct, fragments of their genome still persist in the genomes of contemporary humans. Here, we show that such Neanderthal-like sequences are not distributed randomly in contemporary human genomes. Specifically, while genome-wide frequency of Neanderthal-like sites is close to 6% in all out-of-Africa populations, genes involved in lipid catabolism contain large excess Neanderthal-like sequences in Europeans (24.3%), but not in Asians (12.4%). While lipid catabolism cannot be assayed in Neanderthals, we took advantage of genetic divergence between human populations, chimpanzees and Neanderthals to predict metabolic divergence expected from the observed excess of Neanderthal gene flow into Europeans. We confirmed predicted changes in lipid catabolism using hydrophobic metabolome measurements in the brain tissue and further linked these metabolic changes to gene expression divergence. Overall design: 14 human and 6 chimpanzee samples were sequenced.","project":"SRP019762"}
{"number_samples":12,"species":"human","abstract":"Methods: RNA profiles of 12 tissues were generated by deep sequencing using Illumina HiSeq 2000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with TopHat followed by Cufflinks. Results: This work has focused on identification of non-coding RNAs expressed in human tissues. We have identified a set of non-coding RNAs that are both expressed and conserved, as described in the manuscript accompagnying this work. Overall design: total RNA profiles of 12 normal human tissues, no replicates.","project":"SRP019807"}
{"number_samples":17,"species":"human","abstract":"Genome-wide view of the interplay between methylation and RNA expression in this colorectal cancer model was obtained. Overall design: RNA-sequencing of HCT116 cell line, the DNMT1 and DNMT3B double knock-out HCT116 cell line (DKO), and 7 additional colorectal cell lines.","project":"SRP019810"}
{"number_samples":4,"species":"human","abstract":"To improve our understanding of the relationships between methylation and expression we profiled mRNA expression and single-base resolution methylation levels for two breast cancer cell lines, MCF7 and T47D. Expression was profiled using RNA-seq.  Methylation was assayed using Methyl-MAPS, which uses methylation-sensitive and -dependent restriction enzyme digests followed by high-throughput sequencing to identify methylation levels at individual CpGs (Edwards et al. 2010, Genome Research). Overall design: RNA-Seq was used to generate mRNA expression profiles of MCF7 and T47D cells under standard growth conditions.","project":"SRP019817"}
{"number_samples":32,"species":"human","abstract":"The goal of our study is to build an integrated transcriptome landscape model for HER2 positive breast tumors and identify the crucial signaling pathways associated with HER2 tumors. Genomic features include, 685 genes that were differentially expressed only in HER2-positive tumors, 102 genes that were alternatively spliced in a pattern that is unique to HER2-positive tumors, and 303 genes that expressed single nucleotide sequence variants (eSNVs) that were unique to HER2-positive tumors. Network analysis was performed to integrate the genomic features into a transcriptome landscape model that identified 12 highly interconnected cellular processes that appear to be critical to the establishment and maintenance of HER2-positive tumors. We observed that integrin signaling was linked to lapatinib sensitivity in vitro and strongly associated with risk of relapse in the NCCTG N9831 adjuvant trastuzumab clinical trial dataset. Overall design: We analyzed RNA-seq data from a survey panel consisting of 8 benign breast lesions, 8 ER+, 8 triple negative, and 8 HER2-positive primary breast tumors to identify genomic features that were uniquely associated with HER2-positive tumors","project":"SRP019936"}
{"number_samples":9,"species":"human","abstract":"Many new alternative splice forms have been detected at the transcript level using next generation sequencing (NGS) methods, especially RNA-Seq, but it is not known how many of these transcripts are being translated.  Leveraging the unprecedented capabilities of NGS, we collected RNA-Seq and proteomics data from the same cell population (Jurkat cells) and created a bioinformatics pipeline that builds customized databases for the discovery of novel splice-junction peptides. Results: Eighty million paired-end Illumina reads and ~500,000 tandem mass spectra were used to identify 12,873 transcripts (19,320 including isoforms) and 6,810 proteins. We developed a bioinformatics workflow to retrieve high-confidence, novel splice junction sequences from the RNA data, translate these sequences into the analogous polypeptide sequence, and create a customized splice junction database for MS searching. Overall design: Jurkat T-cell mRNA was analyzed on an Illumina HiSeq2000.  ~80 million paired end reads (2x200bp, ~350bp lengths) were collected.","project":"SRP019939"}
{"number_samples":2,"species":"human","abstract":"SFMBT1 is a poorly characterized mammalian MBT domain-containing protein homologous to Drosophila SFMBT, a Polycomb group protein involved in epigenetic regulation of gene expression. Here, we show that SFMBT1 regulates transcription in somatic cells and during spermatogenesis through the formation of a stable complex with LSD1 and CoREST.  When bound to its gene targets, SFMBT1 recruits its associated proteins and causes chromatin compaction and transcriptional repression.  SFMBT1, LSD1, and CoREST share a large fraction of target genes including those encoding replication-dependent histones.  Simultaneous occupancy of histone genes by SFMBT1, LSD1, and CoREST is regulated during the cell cycle and correlates with the loss of RNA polymerase II at these promoters during G2, M, and G1. The interplay between the repressive SFMBT1–LSD1–CoREST complex and RNA polymerase II contributes to the timely transcriptional regulation of histone genes in human cells. SFMBT1, LSD1, and CoREST also form a stable complex in germ cells and their chromatin binding activity is regulated during spermatogenesis. Overall design: RNA-seq in HeLaS3 cells ctrl compared to triple knockdown for SFMBT1, CoREST, and LSD1","project":"SRP019946"}
{"number_samples":2,"species":"human","abstract":"Our goal of this study was to perform quantitative and global assessment of EBV gene expression in gastric carcinomas and assess EBV associated cellular pathway alterations. Overall design: Examination of a gastric carcinoma cell line naturally infected with EBV, SNU-719 using poly-A and ribodepletion RNA-seq data sets","project":"SRP019961"}
{"number_samples":8,"species":"human","abstract":"RNA sequencing data for four cell lines representing different stages during malignant transformation.","project":"SRP019968"}
{"number_samples":4,"species":"human","abstract":"Previous investigations of the core gene regulatory circuitry that controls embryonic stem cell (ESC) pluripotency have largely focused on the roles of transcription, chromatin and non- coding RNA regulators. Alternative splicing (AS) represents a widely acting mode of gene regulation, yet its role in the regulation of ESC pluripotency and differentiation is poorly understood. Here, we identify the Muscleblind-like RNA binding proteins, MBNL1 and MBNL2, as conserved and direct negative regulators of a large program of AS events that are differentially regulated between ESCs and other cell types. Knockdown of MBNL proteins in differentiated cells causes switching to an ESC-like AS pattern for at least half of these AS events. Among the events is an ESC-specific AS switch in the forkhead family transcription factor FOXP1 that controls pluripotency. Consistent with a central and negative regulatory role for MBNL proteins in pluripotency, their knockdown significantly enhances the expression of key pluripotency genes and the formation of induced pluripotent stem cells (iPSCs) during somatic cell reprogramming. Overall design: mRNA profiles of various embryonic stem cells, tissues and cell lines from human and mouse using high-throughput sequencing data and the role of MBNL proteins in regulation of ESC-differential alternative splicing","project":"SRP019989"}
{"number_samples":6,"species":"human","abstract":"microRNAs (miRNAs) are small non-coding RNAs that function in literally all cellular processes. miRNAs interact with Argonaute (Ago) proteins and guide them to specific target sites located in the 3’ untranslated region (UTR) of target mRNAs leading to translational repression and deadenylation-induced mRNA degradation. Most miRNAs are processed from hairpin-structured precursors by the consecutive action of the RNase III enzymes Drosha and Dicer. However, processing of miR-451 is Dicer-independent and cleavage is mediated by the endonuclease Ago2. Here we have characterized miR-451 sequence and structure requirements for processing as well as sorting of miRNAs into different Ago proteins. Pre-miR-451 appears to be optimized for Ago2 cleavage and changes result in reduced processing. In addition, we show that the mature miR-451 only associates with Ago2 suggesting that mature miRNAs are not exchanged between different members of the Ago protein family. Based on cloning and deep sequencing of endogenous miRNAs associated with Ago1-3, we do not find evidence for miRNA sorting in human cells. However, Ago identity appears to influence the length of some miRNAs, while others remain unaffected. Overall design: Examination of miRNAs associated with endogenous human Ago1-4 in HeLa cells","project":"SRP019990"}
{"number_samples":99,"species":"human","abstract":"Leiomyosarcoma (LMS) is a malignant neoplasm with smooth muscle differentiation.  Little is known about its molecular heterogeneity and no targeted therapy currently exists for LMS.  We demonstrate the existence of 3 molecular subtypes in a cohort of 99 cases and an independent cohort of 82 LMS.  Two new FFPE tissue-compatible diagnostic immunohistochemical markers are identified: LMOD1 for subtype I LMS and ARL4C for subtype II LMS. Subtype I and subtype II LMS are associated with good and poor prognosis, respectively.  The LMS subtypes show significant differences in expression levels for genes for which novel targeted therapies are being developed. Overall design: Gene expression profiling was performed by 3' End RNA Sequencing (3SEQ), a next generation sequencing approach that does not rely on frozen tissue but can be performed on archival FFPE tissue. Samples included 99 LMS, 6 Undifferentiated Pleomorphic Sarcomas (UPS), 3 leiomyomas, 4 normal myometrium samples, and 1 case of Lymphangioleiomyomatosis (LAM). This study only includes the 99 LMS Samples. After gene expression levels were quantified by 3SEQ analysis pipeline, Consensus Clustering with bootstrap method was used to determine that the dataset contained three robust subtypes, and Silhouette analysis was performed to validate the subtype assignments. Two class SAM analysis (Significance Analysis of Microarrays) was performed to identify genes expressed differentially between each subtype of LMS with FDR of 0.05. Immunohistochemical staining was used to validate the potential diagnostic and prognostic markers from 3SEQ data on a tissue microarray.","project":"SRP019994"}
{"number_samples":20,"species":"human","abstract":"The Office of Cancer Genomics (OCG) at the National Cancer Institute is sponsoring a series of studies as part of the Cancer Genome Characterization Initiative (CGCI) to assess novel emerging sequencing technologies in cancer. The CGCI program includes comprehensive characterization of the genetic aberrations found in different pediatric and/or adult tumors. CGCI is currently characterizing a number of B-cell non-Hodgkin lymphomas (including diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), and adult and pediatric Burkitt lymphomas).  In combination, the lymphoid cancers (non-Hodgkin lymphoma, Hodgkin lymphoma, myeloma and chronic lymphocytic leukemia), constitute the fourth most common malignancy in both men and women in North America. Lymphomas typically have characteristic abnormal chromosomes, including translocations, indicating the relevance of mutations to how NHLs develop and behave. This project uses detailed analysis on a specific... (for more see dbGaP study page.)","project":"SRP020237"}
{"number_samples":16,"species":"human","abstract":"Human milk fat globules, by enveloping cell contents during their secretion into milk, are a rich source of mammary cell RNA. Here, we pair this non-invasive mRNA source with RNA sequencing technology to probe the milk fat layer transcriptome during three stages of lactation: colostral, transitional, and mature milk production. We find that transcriptional profiles cluster not by postpartum day, but by milk Na:K ratio, indicating that women sampled during the same postpartum time frame could be at markedly different stages of gene expression. Each stage of lactation is characterized by a dynamic range (105-fold) in transcript abundances not previously observed with microarray technology. We discovered that transcripts for isoferritins and cathepsins are strikingly abundant during colostrum production, highlighting the potential importance of these proteins for neonatal health. Two transcripts, encoding ß-casein (CSN2) and a-lactalbumin (LALBA), make up 45% of the total pool of mRNA in mature lactation. Genes significantly expressed across all stages of lactation are associated with making, modifying, transporting, and packaging milk proteins. Stage-specific transcripts are associated with immune defense during the colostral stage, up-regulation of the machinery needed for milk protein synthesis during the transitional stage, and the production of lipids during mature lactation. We observed strong modulation of key genes involved in lactose synthesis and insulin signaling. In particular, PTPRF may serve as a biomarker linking insulin resistance with insufficient milk supply. This study provides the methodology and reference data set to enable future targeted research on the physiological contributors to sub-optimal lactation in humans. Overall design: Milk fat mRNA profiles were generated from Day 2 and mature milk samples obtained from lactating mothers","project":"SRP020470"}
{"number_samples":33,"species":"human","abstract":"Exosomes, endosome-derived membrane microvesicles, contain a specific set of RNA transcripts that are involved in cell-cell communication and hold a great potential as disease biomarkers. To systemically characterize exosomal RNA profiles, we performed RNA sequencing analysis using three human plasma samples and evaluated efficacies of small RNA library preparation protocols from 3 manufacturers. Overall design: We tested the six samples (A1 and A2, B1 and B2, C1 and C2) using two small RNA library preparation kits: NEBNext Multiplex Small RNA library Prep Set from New England Biolab (NEB) and NEXTflex Small RNA Sequencing Kit from Bioo Scientific (BS). We also tested Illumina’s TrueSeq Small RNA Sample Preparation Kit (ILMN) in sample A1 and A2. Together, we tested these plasma samples by sequencing 14 indexed libraries. This study allowed direct comparison of current small RNA library preparation protocols and identified the most suitable strategy for future exosomal RNA sequencing analysis.","project":"SRP020486"}
{"number_samples":55,"species":"human","abstract":"Daily sampling of peripheral blood from human subjects vaccinated for influenza was done immediately before vaccination and for 10 days after vaccination. In B cells, 90% of transcriptomic variation in subjects who received influenza vaccine within the previous three years was explained by a single temporal pattern unique to the individual. A common set of 742 genes was strongly correlated with the migration of differentiating plasma cell subtypes. Overall design: Five subjects, 11 time points per subject (pre-vaccination and daily for 10 days post-vaccination)","project":"SRP020491"}
{"number_samples":55,"species":"human","abstract":"Daily sampling of peripheral blood from human subjects vaccinated for influenza was done immediately before vaccination and for 10 days after vaccination. Temporal patterns of gene expression, determined by RNA-seq, in unfractionated PBMC suggested migration of myeloid/dendritic cell lineage cells one day after vaccination. Overall design: Five subjects, 11 time points per subject (pre-vaccination and daily for 10 days post-vaccination)","project":"SRP020492"}
{"number_samples":14,"species":"human","abstract":"Recurrent mutations in histone modifying enzymes in multiple cancer types imply key roles in tumorigenesis. However, the functional relevance of these mutations remains unknown. Here we show that the JARID1B histone H3 lysine 4 demethylase is frequently amplified and overexpressed in luminal breast tumors and a somatic point mutation of JARID1B leads to the gain of luminal-specific gene expression programs. Downregulation of JARID1B in luminal breast cancer cells induces the expression of basal cell-specific genes and growth arrest, which is partially rescued by the inhibition of TGFBR thereby indicating a key role for TGFb signaling. Integrated genome-wide analysis of JARID1B chromatin binding, histone H3 lysine trimethyl (H3K4me3) and dimethyl (H3K4me2) patterns, and gene expression profiles in luminal and basal-like breast cancer cells suggest a key role for JARID1B in luminal cell-specific gene expression programs. A significant fraction of JARID1B binding-sites overlaps with CTCF in both luminal and basal-like breast cancer cells. CTCF also co-immunoprecipitates with JARID1B and it may influence its histone demethylase (HDM) activity as the H3K4me3/me2 ratio is lower at the CTCF-overlapping compared to JARID1B-unique sites. Additionally, a heterozygous JARID1B missense mutation (K1435R) in the HCC2157 basal-like breast cancer cell line is associated with unique JARID1B chromatin-binding and gene expression patterns implying gain of luminal features. In line with this, exogenous expression of this mutant in basal-like breast cancer cells leads to a gain of JARID1B binding at many luminal-specific genes. A PARADIGM score reflecting JARID1B activity in luminal breast cancer cells is associated with poor clinical outcome in patients with luminal breast tumors. Together, our data imply that JARID1B is a luminal lineage-driving oncogene and that its therapeutic targeting may represent a novel therapeutic strategy in treatment-resistant luminal breast tumors. Overall design: RNA-Seq in breast cancer cell-lines transfected with JARID1B/CTCF/control siRNA. 50 cycles of sequencing on Illumina platform.","project":"SRP020493"}
{"number_samples":1,"species":"human","abstract":"Pancreatic ductal adenocarcinoma cell line Pt45P1.","project":"SRP020496"}
{"number_samples":5,"species":"human","abstract":"RNA-Seq of TNFa time course in human A549 cancer cell line.","project":"SRP020499"}
{"number_samples":5,"species":"human","abstract":"We applied in parallel RNA-Seq and Ribosome-profiling analyses to immortalized human primary BJ fibroblast cells in which p53 was induced by Nutlin-3a Overall design: RNA-seq, using Illumina HiSeq 2000, was applied to BJ cells treated with Nutlin-3a, at 5 timepoints: 0, 2, 4, 6, 19 hrs Ribosome profiling was applied to BJ cells treated with Nutlin-3a, at 5 timepoints: 0, 2, 4, 6, 19 hrs","project":"SRP020544"}
{"number_samples":11,"species":"human","abstract":"We present an approach for globally monitoring RNA structure in native conditions in vivo with single nucleotide precision. This method is based on in vivo modification with dimethyl sulfate (DMS), which reacts with unpaired adenine and cytosine residues9, followed by deep sequencing to monitor modifications. Our data from yeast and mammalian cells are in excellent agreement with known mRNA structures and with the high-resolution crystal structure of the Saccharomyces cerevisiae ribosome10. Comparison between in vivo and in vitro data reveals that in rapidly dividing cells there are vastly fewer structured mRNA regions in vivo than in vitro. Even thermostable RNA structures are often denatured in cells, highlighting the importance of cellular processes in regulating RNA structure. Indeed, analysis of mRNA structure under ATP-depleted conditions in yeast reveals that energy-dependent processes strongly contribute to the predominantly unfolded state of mRNAs inside cells. Our studies broadly enable the functional analysis of physiological RNA structures and reveal that, in contrast to the Anfinsen view of protein folding, thermodynamics play an incomplete role in determining mRNA structure in vivo. We use Dimethyl Sulfate to probe the structure of rRNA and mRNA in yeast in vivo, in vitro, and at different temperatures in vitro. We obtain a great agreement between in vivo data and known mRNA structures as well as the ribosome crystal structure. We find that in contrast  to ribosomal rna, mRNAs are less structured in vivo than in vitro, and the structures present in vivo can only partially be explained by thermodynamic stability. In addition, we identify new regulatory structures present in vivo. Overall design: Examination of RNA structure in yeast under different conditions - in vivo and in vitro at five different temperatures (30,45,60,75,95) We adapt our DMS-seq assay for use in mammalian cells and probe RNA structure genome-wide in K562 cells. We probe the RNA structure of primary fibroblast using DMS on a genome-wide scale to confirm the presence of more structures in vitro. In addition we probe the RNA structure in yeast upon ATP depleted conditions to investigate whether active (ATP-dependent) processed are modulating RNA structure in vivo.","project":"SRP020556"}
{"number_samples":6,"species":"human","abstract":"The functional importance of gene enhancers in regulated gene expression is well established. In addition to widespread transcription of long non-coding RNA (ncRNA) transcripts in mammalian cells, bidirectional ncRNAs referred to as eRNAs are present on enhancers. However, it has remained unclear whether these eRNAs are functional, or merely a reflection of enhancer activation. Here, we report that 17 ß-estradiol (E2)-bound estrogen receptor alpha (ERa) on enhancers causes a global increase in eRNA transcription on enhancers adjacent to E2 upregulated coding genes. These induced eRNAs, as functional transcripts, appear to exert important roles for the observed ligand-dependent induction of target coding genes, causing an increased strength of specific enhancer:promoter looping initiated by ERa binding. Cohesin, present on many ERa-regulated enhancers even prior to ligand treatment, apparently contributes to E2-dependent gene activation by stabilizing E2/ERa/eRNA-induced enhancer:promoter looping. Our data indicate that eRNAs are likely to exert important functions in many regulated programs of gene transcription. Overall design: The ChIP-seqs in this study measure the binding landscape of master transcription regulator of estrogen signaling - ERa, together with common histone marks including H3K27ac and H3K4me1 in MCF7 cells. These data serve as the basis to understand the enhancer map and subsequent analysis of eRNA expression using GRO-seq. The GRO-seq measures the trancription of nascent RNAs in the genome. From MCF7 cells treated with veichle or estrodial, we could identify estrogen-regulated eRNAs and subsequently could study their functions.","project":"SRP020561"}
{"number_samples":4,"species":"human","abstract":"The human double-homeodomain retrogene DUX4 is normally expressed at high levels in germ cells of the testis. When aberrantly expressed in muscle its protein product causes facioscapulohumeral muscular dystrophy (FSHD), perhaps partly by inducing inappropriate expression of germline genes. DUX4 can bind >60,000 locations in the human genome that contain a strongly enriched sequence motif. Numerous long terminal repeat (LTR) class repetitive elements are enriched among DUX4 binding sites, including many from the mammalian apparent LTR-retrotransposon (MaLR) family as well as some ERVL and ERVK types, with MaLRs comprising ~1/3 of DUX4’s binding sites. We performed RNA-seq on myoblasts over-expressing DUX4 and find that DUX4 binding activates transcription of some but not all bound repeat types. Some of these activated repetitive elements comprise novel promoters for genes, long non-coding RNAs and antisense transcripts. We show that some of these chimeric repeat-initiated transcripts are expressed in testis and FSHD patient myotubes. The acquisition of MaLR-LTR elements during mammalian evolution may therefore have allowed rewiring of the transcriptional network. We also find that the pericentromeric satellite HSATII can be bound by DUX4 and that its transcription is massively induced by DUX4 over-expression. Our findings suggest a role for repetitive element transcripts in muscle disease and in the biology of normal testis. Overall design: RNA-seq of two myoblast cell lines transduced with lentivirus carrying DUX4, and two control myoblast lines","project":"SRP020646"}
{"number_samples":4,"species":"human","abstract":"Transcribed regions in adult temporal lobe, hippocampus and frontal lobe were assesed by strand specific next generation sequencing of polyA RNA. Overall design: Strand specific mRNA expression profiles of three human adult brain regions were generated by next generation sequencing using Illumina GAIIx","project":"SRP020661"}
{"number_samples":5,"species":"human","abstract":"Immunodeficiency, Centromeric Instability, and Facial Anomalies Type I (ICF1) Syndrome is a rare genetic disease caused by mutations in DNMT3B, a de novo DNA methyltransferase. However, the molecular basis of how DNMT3B-deficiency leads to ICF1 pathogenesis is unclear. Induced pluripotent stem cell (iPSC) technology facilitates the study of early human developmental diseases via facile in vitro paradigms. Here, we generate iPSCs from ICF Type 1 Syndrome patient fibroblasts followed by directed differentiation of ICF1-iPSCs to mesenchymal stem cells (MSCs). By performing genome-scale bisulfite sequencing, we find that DNMT3B-deficient iPSCs exhibit global loss of non-CG methylation and select CG hypomethylation at gene promoters and enhancers. Further unbiased scanning of ICF1 iPSC methylomes also identifies large megabase regions of CG hypomethylation typically localized in centromeric and subtelomeric regions.  RNA sequencing of ICF1 and control iPSCs reveals abnormal gene expression in ICF1 iPSCs relevant to ICF Syndrome phenotypes, some directly associated with promoter or enhancer hypomethylation. Upon differentiation of ICF1 iPSCs to mesenchymal stem cells (MSCs), we find virtually all CG hypomethylated regions remained hypomethylated when compared to either wild-type iPSC-derived MSCs or primary bone-marrow MSCs. Collectively, our results show specific methylome and transcriptome defects in both ICF1-iPSCs and differentiated somatic cell lineages, providing a valuable stem cell system for further in vitro study of the molecular pathogenesis of ICF1 Syndrome. Overall design: MethylC-seq and RNA-Seq in ICF Syndrome patient fibroblast derived induced pluripotent stem cells.","project":"SRP021039"}
{"number_samples":6,"species":"human","abstract":"Two islet preparations (from different donors) were subjected to RNA-sequencing. Within each preparation, subsets of islets were treated with 5 or 8 mM glucose, or 8 mM glucose in a specialized, conditioned medium containing Wnt and R-spondin ligands with Rho and ROCK inhibitors (L-WRN+) for 48 h immediately after initial receipt and hand-selection. L-WRN+ promotes proliferation with undue de-differentiation. Polyadenylated mRNA was obtained from 1 mcg of human islet total RNA and 50 nt, single-end sequencing reads were obtained from an Illumina HiSeq 2500.","project":"SRP021048"}
{"number_samples":5,"species":"human","abstract":"Different mechanisms for CBF-MYH11 function in acute myeloid Leukemia (AML) with inv(16) have been proposed such as tethering of RUNX1 outside the nucleus, interference with transcription factor complex assembly and recruitment of histone deacetylases, all resulting in transcriptional repression of RUNX1 target genes. Here, through genome-wide CBF-MYH11 binding site analysis and quantitative interaction proteomics we found that CBF-MYH11 localizes to RUNX1 occupied promoters where it interacts with TAL1, FLI1 and TBP associated factors (TAFs) in the context of the hematopoietic transcription factors ERG, GATA2 and PU.1/SPI1 and the co regulators EP300 and HDAC1. Transcriptional analysis revealed that upon fusion protein knock down a subset of the CBF-MYH11 target genes show increased expression, confirming a role in transcriptional repression. However, the majority of CBF-MYH11 target genes, including genes implicated in hematopoietic stem cell (HSC) self-renewal such as ID1, LMO1 and JAG1, are actively transcribed and upon fusion protein knock down repressed. Together these results suggest an essential role for CBF-MYH11 in regulating expression of genes involved in maintaining a stem cell phenotype. Overall design: 17 ChIP-seq samples using antibodies recognizing the indicated proteins and one RNA-seq file from ME-1 cells were analyzed. In addition 2 ChIP-seq profiles were generated using patient AML cells. A CBFß-MYH11 inducible U937 system (U937CM) was used to examine binding patterns before (1 profile) and after (2 profiles) induction of CBFb-MYH11. In addition, expression was measured through RNA-seq analysis of the two states. The U937CM cells were maintained in the presence of tetracyclin (Tet, 1 uM) and grown in the absence of tetracycline for 3 days to induce expression of CBFß-MYH11. Finally, a CBFb-MYH11 knock down system was developed in ME-1 cells. Two ME-1 cell lines were created, one with a stably integrated shRNA construct that targets CBFb-MYH11 (ME-1_knockdown) and one with a scrambled shRNA construct (ME-1_SCR). Expression of the shRNA constructs  was induced  using doxycyclin (dox; 1 mM) treatment for 3 days.","project":"SRP021072"}
{"number_samples":30,"species":"human","abstract":"Defining the role of epigenetic regulators in normal hematopoiesis has become critically important, as recurrent mutations or aberrant expression of these genes has been identified in both myeloid and lymphoid hematological malignancies. We have found that PRMT4, a type I arginine methyltransferase, whose function in normal and malignant hematopoiesis is unknown, is overexpressed in AML patient samples. In support of an oncogenic role for PRMT4, we find that its overexpression blocks the myeloid differentiation of human stem/progenitor cells (HSPCs) while its knockdown (KD) is sufficient to induce myeloid differentiation of HSPCs and multiple AML cell lines. Although classically thought of as a co-activator, we found that PRMT4 functions to repress the expression of miR-223 in HSPCs via the methylation of RUNX1, which triggers the assembly of a multi-protein repressor complex that includes DPF2. As part of a feedback loop, PRMT4 expression is repressed post-transcriptionally by miR-223 during the normal differentiation process. These data reveal an unidentified role of PRMT4 in myeloid differentiation and its unexpected repressive role in transcriptional regulation. Furthermore, depletion of PRMT4 results in the differentiation of myeloid leukemia cells in vitro and their decrease proliferation in vivo. Thus, targeting PRMT4 holds potential as a novel therapy for acute myelogenous leukemia. Overall design: Purified human primary CD34+ cells were transduced with lentiviruses carrying PRMT4KD or scramble control shRNAs. Total RNA was extrated. RNAseq was performed to identify target genes that are regulated by PRMT4. Experiments were performed in triplicate.","project":"SRP021085"}
{"number_samples":20,"species":"human","abstract":"MicroRNAs (miRNAs) are small (20-22 nucleotides) regulatory non-coding RNAs that strongly influence gene expression. Most prior studies addressing the role of miRNAs in neurodegenerative diseases (NDs) have focused on individual controls (n = 2), AD (n = 5), dementia with Lewy bodies (n = 4), hippocampal sclerosis of aging (n = 4), and frontotemporal lobar dementia (FTLD) (n = 5) cases, together accounting for the most prevalent ND subtypes. All cases had short postmortem intervals, relatively high-quality RNA, and state-of-the-art neuropathological diagnoses. The resulting data (over 113 million reads in total, averaging 5.6 million reads per sample) and secondary expression analyses constitute an unprecedented look into the human cerebral cortical miRNome at single nucleotide resolution. While we find no apparent changes in isomiR or miRNA editing patterns in correlation with ND pathology, our results validate and extend previous miRNA profiling studies with regard to quantitative changes in NDs. In agreement with this idea, we provide independent cohort validation for changes in miR-132 expression levels in AD (n = 8) and FTLD (n = 14) cases when compared to controls (n = 8). The identification of common and ND-specific putative novel brain miRNAs and/or short-hairpin molecules is also presented. The challenge now is to better understand the impact of these and other alterations on neuronal gene expression networks and neuropathologies. Overall design: Using RNA deep sequencing, we sought to analyze in detail the small RNAs (including miRNAs) in the temporal neocortex gray matter from non-demented controls (n = 2), AD (n = 5), dementia with Lewy bodies (n = 4), hippocampal sclerosis of aging (n = 4), and frontotemporal lobar dementia (FTLD) (n = 5) cases, together accounting for the most prevalent ND subtypes.","project":"SRP021130"}
{"number_samples":8,"species":"human","abstract":"Differentiation of human embryonic stem cells (hESCs) provides a unique opportunity to study the epigenetic mechanisms that facilitate cellular transitions in a human context. To that end, we performed comprehensive transcriptional and epigenetic profiling of early populations derived through directed differentiation of hESCs representing each of the three embryonic germ layers. Integration of whole genome bisulfite sequencing, chromatin immunoprecipitation-sequencing and RNA-Sequencing reveals unique events associated with specification towards each lineage. While we observe expected dynamics such as loss of DNA methylation and gain of H3K4me1 at distal putative regulatory elements, we frequently found a germ layer specific switch to H3K27me3 at sites of high DNA methylation in the undifferentiated state. By carefully dissecting these initial events, we may be able to devise more faithful differentiation strategies and gain novel insights in to the robust rewiring of regulatory programs during cellular transitions. Overall design: 10 Samples in total, 5 in replicate. To better understand the interplay of epigenetic dynamics and transcription factor binding upon in vitro specification of human embryonic stem cells we profiled OCT4, SOX2 and NANOG in hESC and the endoderm master regulatory factor FOXA2 in in vitro derived endoderm cells (dEN). To gain further insights into the relation of DNA methylation and TF binding, we carried out ChIP-bisulfite sequencing for FOXA2 in dEN. Lastly, we were interested in the fate of genes bound by FOXA2 in dEN upon further differentiation and therefore differentiated dEN further towards a hepatocyte like state and performed RNA-Seq.","project":"SRP021134"}
{"number_samples":10,"species":"human","abstract":"We first demonstrate that non-genetically determined inter-individual differentially methylated regions (iiDMRs) can be temporally stable for at least two years. Then, we show that iiDMRS are associated with concomitant changes in chromatin state as measured by inter-individual differences in the levels of the histone variant H2A.Z. However, the correlation of promoter iiDMRs with gene expression is negligible and this correlation is not improved even by integrating H2A.Z information. We find that most promoter epialleles, whether genetically or non-genetically determined, are associated with low levels of transcriptional activity, depleted for house keeping genes, and either depleted for H3K4me3/enriched for H3K27me3, or lacking both these marks in human embryonic stem cells. These findings validate in an independent cohort. Interestingly, the key features of iiDMRs are reminiscent of those previously observed for promoters that undergo hyper-methylation in various cancers, in vitro cell culture, and human chronological ageing. Overall design: H2A.z ChIP-seq, RNA-seq, and DNA methylation data (submitted separately) were collected for five normal individuals.   T21/T22 and T31/T32 are monozygotic twins.","project":"SRP021191"}
{"number_samples":80,"species":"human","abstract":"Complete transcriptome profiling in human failing and non-failing control hearts using next-gen sequencing Overall design: Poly-A selected RNA and small RNA sequencing carried out in 5 groups of samples: NF, ICM, NICM, ICM+LVAD, NICM+LVAD","project":"SRP021193"}
{"number_samples":8,"species":"human","abstract":"Through RNA-seq analyses of nascent transcripts, we found large numbers of RNA transcripts that extend beyond the 3’ ends of protein-coding genes; we refer to these extended transcripts as geRNAs. These findings demonstrate that transcription of most human protein-coding genes does not terminate within close proximity to poly(A) signals; rather, it terminates at sites far beyond these signals (up to 50 kb). Overall design: Examination of different RNA species in HelaS3 cells by strand-specific RNA-seq. The 4sU-labeled RNA was isolated by using organomercurial affinity matrix.","project":"SRP021214"}
{"number_samples":2,"species":"human","project":"SRP021221"}
{"number_samples":3,"species":"human","abstract":"A time course of infection of the alphavirus Sindbis virus (SINV) was used to investigate the presence of viral specific vsRNA and the changes in miRNAs profiles in human embryonic kidney 293 cells (HEK293) by high throughput DNA sequencing. Deep sequencing of small RNAs early in SINV infection (4 and 6 hpi) showed low abundance (0.8%) of viral specific RNAs (vsRNAs) , with a random uniform distribution not typical of Dicer products, suggesting they arise from non-specific degradation. Sequencing showed little variation of cellular microRNAs (miRNAs) at 4 and 6 hpi compared to uninfected cells. Twelve miRNAs exhibiting some minor differential expression by sequencing, showed insignificant modulation by Northern blot analysis. Overall design: RNA was isolated from mock infected and SINV inoculated HEK 293 cells at 4hpi and 6hpi cDNA libraries were generated for the small RNA (sRNA) content of the cells and sequenced using Illumina GA II, which yielded between 29.1M and 30.5M reads per sample","project":"SRP021459"}
{"number_samples":8,"species":"human","abstract":"Transcript and miRNA profiling using a novel reverse transcriptase.","project":"SRP021468"}
{"number_samples":16,"species":"human","abstract":"Recent advances in RNA sequencing (RNA-Seq) have enabled the discovery of novel transcriptomic variations that are not possible with traditional microarray-based methods. Tissue and cell specific transcriptome changes during pathophysiological stress, in disease cases versus controls and in response to therapies are of particular interest to investigators studying cardiometabolic diseases. Thus, knowledge on the relationships between sequencing depth and detection of transcriptomic variation is needed for designing RNA-Seq experiments and for interpreting results of analyses. Using deeply sequenced RNA-Seq data derived from adipose of a healthy individual before and after systemic administration of endotoxin (LPS), we investigated the sequencing depths needed for studies of gene expression and alternative splicing (AS). We found that to detect expressed genes and AS events, ~100 million (M) filtered reads were needed. However, the requirement on sequencing depth for the detection of LPS modulated differential expression (DE) and differential alternative splicing (DAS) was much higher. To detect 80% of events, ~300M filtered reads were needed for DE analysis whereas at least 400M filtered reads were necessary for detecting DAS. Although the majority of expressed genes and AS events can be detected with modest sequencing depths (~100M filtered reads), the estimated gene expression levels and exon/intron inclusion levels were less accurate. We report the first study that evaluates the relationship between RNA-Seq depth and the ability to detect DE and DAS in human adipose. Our results suggest that a much higher sequencing depth is needed to reliably identify DAS events than for DE genes. Overall design: Random sampling the RNA-seq data in different depth for gene and alternative-splicing analysis","project":"SRP021478"}
{"number_samples":43,"species":"human","abstract":"The EZH2 histone methyltransferase is highly expressed in germinal center (GC) B-cells and targeted by somatic mutations in B-cell lymphomas. Here we find that EZH2 deletion or pharmacologic inhibition suppresses GC formation and functions in mice. EZH2 represses proliferation checkpoint genes and helps establish bivalent chromatin domains at key regulatory loci to transiently suppress GC B-cell differentiation. Somatic mutations reinforce these physiological effects through enhanced silencing of EZH2 targets in B-cells, and in human B-cell lymphomas. Conditional expression of mutant EZH2 in mice induces GC hyperplasia and accelerated lymphomagenesis in cooperation with BCL2. GCB-type DLBCLs are mostly addicted to EZH2, regardless of mutation status, but not the more differentiated ABC-type DLBCLs, thus clarifying the therapeutic scope of EZH2 targeting. Overall design: RNA sequencing and H3K27me3 ChIP sequencing of human DLBCL cell lines and murine BCL1 cell line. RNA sequencing, H3K27me3 and H3K4me3 ChIP sequencing of B cells from de-identified human tonsills.","project":"SRP021509"}
{"number_samples":4,"species":"human","abstract":"We describe the discovery of sno-lncRNAs, a class of nuclear-enriched intron-derived long noncoding RNAs (lncRNAs) that are processed on both ends by the snoRNA machinery. During exonucleolytic trimming, the sequences between the snoRNAs are not degraded, leading to the accumulation of lncRNAs flanked by snoRNA sequences but lacking 5' caps and 3' poly(A) tails. Such RNAs are widely expressed in cells and tissues and can be produced by either box C/D or box H/ACA snoRNAs. Importantly, the genomic region encoding one abundant class of sno-lncRNAs (15q11-q13) is specifically deleted in Prader-Willi Syndrome (PWS). The PWS region sno-lncRNAs do not colocalize with nucleoli or Cajal bodies, but rather accumulate near their sites of synthesis. These sno-lncRNAs associate strongly with Fox family splicing regulators and alter patterns of splicing. These results thus implicate a previously unannotated class of lncRNAs in the molecular pathogenesis of PWS. Overall design: We have used deep sequencing to explore the gene expression from poly(A)+ RNAs in embryonal carcinoma (EC) line PA-1 cells treated with scrambled or specific antisense oligodeoxynucleotides (ASOs).","project":"SRP021524"}
{"number_samples":2,"species":"human","abstract":"Altered DNA methylation patterns represent an attractive mechanism for the phenotypic changes associated with human aging. Several studies have described age-related methylation changes to various extents, but their functional significance has remained largely unclear. We have now used an integrated methylome and transcriptome sequencing approach to characterize age-related methylation changes in the human epidermis and to analyze their impact on gene expression. Our results show limited and localized methylation differences between young and old methylomes at single-base resolution. Similarly, the comparison of transcriptomes from young and old samples revealed a highly defined set of differentially expressed genes with functional annotations in skin homeostasis. Further data analysis showed a robust correlation between age-related promoter hypermethylation and gene silencing, particularly at promoters that were pre-marked with stem cell-specific chromatin features. In addition, we also observed age-related methylation changes at transcription factor binding sites, with a significant enrichment of stem cell regulatory networks. Our results provide a high-resolution analysis of age-related methylation changes and suggest that they result in highly defined alterations in the transcriptional programme of the human epidermis. Interestingly, several of our findings can be interpreted to reflect epigenetic changes in aging stem cells, thus supporting a critical role of stem cells in human aging. Overall design: Whole transcriptome analysis of H. sapiens. Two samples were analyzed, one sample containing RNA from young, one sample containing RNA from old human skin.","project":"SRP021891"}
{"number_samples":6,"species":"human","abstract":"Steroid hormones are well-recognized suppressors of the inflammatory response, however, their cell- and tissue-specific effects in the regulation of inflammation are far less understood, particularly for the sex-related steroids. To determine the contribution of progesterone in the endothelium, we have characterized and validated an in vitro culture system in which human umbilical vein endothelial cells constitutively express human progesterone receptor (PR). Using next generation RNA-sequencing, we identified a selective group of cytokines that are suppressed by progesterone both under physiological conditions and during pathological activation by lipopolysaccharide. In particular, IL-6, IL-8, CXCL2/3, and CXCL1 were found to be direct targets of PR, as determined by ChIP-sequencing. Regulation of these cytokines by progesterone was also confirmed by bead-based multiplex cytokine assays and quantitative PCR. These findings provide a novel role for PR in the direct regulation of cytokine levels secreted by the endothelium. They also suggest that progesterone-PR signaling in the endothelium directly impacts leukocyte trafficking in PR-expressing tissues Overall design: Examination of PR target genes in human umbilical vein endothelial cells (HUVECs) using RNA-seq PR infected only (PR); PR infected followed by ligand treatment (PR+P); PR infected followed by 4h LPS treatment (PR+LPs_4h); PR infected followed by 8h LPS treatment (PR+LPs_8h); PR infected followed by 4h LPS and progesterone treatment (PR+LPS+P_4h); PR infected followed by 8h LPS and progesterone treatment (PR+LPS+P_8h)","project":"SRP021908"}
{"number_samples":12,"species":"human","abstract":"The granulosa cells in the mammalian ovarian follicle respond to gonadotropin signalling and are involved in the processes of folliculogenesis and oocyte maturation. Studies on gene expression and regulation in human granulosa cells are of interest due to their potential for estimating the oocyte viability and IVF success. However, the post-transcriptional gene expression studies on miRNA level in the human ovary have been scarce. The current study determined the miRNA profile by deep sequencing of the two intrafollicular somatic cell types: mural and cumulus granulosa cells isolated from women undergoing controlled ovarian stimulation and in vitro fertilization. Overall design: Paired cumulus and mural granulosa samples were analysed from 3 women participating in IVF procedure. Libraries of all 6 samples were sequenced twice, generating 2 technical replicates for each sample. Differential gene expression study was performed on the pooled results of technical replicates.","project":"SRP021911"}
{"number_samples":6,"species":"human","abstract":"The granulosa cells in the mammalian ovarian follicle respond to gonadotropin signalling and are involved in the processes of folliculogenesis and oocyte maturation. Studies on gene expression and regulation in human granulosa cells are of interest due to their potential for estimating the oocyte viability and IVF success. The current study determined the mRNA profile by deep sequencing of the two intrafollicular somatic cell types: mural and cumulus granulosa cells isolated from women undergoing controlled ovarian stimulation and in vitro fertilization. Overall design: Paired cumulus and mural granulosa samples were analysed from 3 women participating in IVF procedure. Differential gene expression study was performed. The identified gene expression profile was also used for predicting targets for miRNAs that were also identified from the same samples (GSE46489).","project":"SRP021912"}
{"number_samples":2,"species":"human","abstract":"Epstein-Barr virus is a gamma-herpes virus that is causally associated with several lymphomas and carcinomas. This virus encodes at least 25 pre-miRNAs, which are expressed in infected cells to yield more than 50 detected mature miRNAs. miRNAs are small, non-coding RNAs that inhibit gene expression by promoting the inhibition of translation or of degradation of mRNAs. Currently, the function of these viral miRNAs and the contribution they provide to EBV's life-cycle remain largely unknown, due to difficulties in identifying cellular and viral genes regulated by these miRNAs. We have compared and contrasted two methods to identify targets of viral miRNAs in order to identify the advantages and limitations of each method to aid in uncovering the functions of EBV's miRNAs. Overall design: Examination of RISC (RNA Induced Silencing Complexes) associated transcripts under 2 conditions in BJAB cells","project":"SRP021916"}
{"number_samples":15,"species":"human","abstract":"RNA-sequencing of SSP RNA from patients with serrated polyposis syndrome identifies VSIG1 and MUC17 as potential diagnostic markers for SSPs Overall design: 5' capped RNA from seven ascending SSPs, six patient matched uninvolved right colon and two normal right colon samples was used for RNA sequencing (15 samples total)","project":"SRP021917"}
{"number_samples":3,"species":"human","abstract":"This study provides, for the first time, TIA1 and TIAR linked-transcriptomic analysis by using RNA-Seq next-generation sequencing technology. Illumina RNA-Seq was used to survey transcriptome profiles from permanent TIA1- and TIAR-(shRNA-mediated) deficient HeLa cells. Analysis of the transcriptomes with the Cufflinks tool revealed that differentially expressed genes, isoforms produced by alternative splicing and/or promoter usage as well as microRNAs generated a great transcriptomic heterogeneity which might reflect the complexity linked to these cell phenotypes. The data of differential expression were validated by using genome-wide microarrays and QPCR analysis. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes term enrichment analysis revealed over-representation of genes associated with cell differentiation, multicellular organismal development, signal transduction, axon guidance and cell adhesion and under-representation of genes associated with positive regulation of migration, cell adhesion, response to organic substance, prostaglandin metabolic process and blood coagulation. Taken together, these results indicate that differential gene expression, alternative pre-mRNA isoforms, promoter usage and microRNA profiling contribute to define the molecular expression phenotypes implied in the progression of proliferative phenotypes associated to the absence of TIA proteins and prioritize candidates for future study. Overall design: Each library was run on one RNASeq Multiplex of 76 bp using sequencing from Illumina Genome Analyzer (GAIIx). Three samples were analyzed in this manner, taken from control, TIA1 and TIAR shRNA-depleted HeLa cells.","project":"SRP021918"}
{"number_samples":5,"species":"human","abstract":"We report the application of high-throughput RNA sequencing to the human prefrontal cortex. The brain dataset was obtained by sequencing total RNAs extracted from the dorsolateral prefrontal cortex of five deceased human patients with no apparent pathology, followed by depletion of ribosomal RNA to obtain all non-rRNA coding and non-coding RNAs in the human brain transcriptome. Overall design: Five samples were sequenced, four coming from frozen brain tissue (frontal cortex) of deceased female human patients with no remarkable pathology, and one from a male patient with no remarkable pathology.","project":"SRP021924"}
{"number_samples":4,"species":"human","abstract":"Transcription start sites are the focal points of transcriptional regulation, where information from regulatory elements is integrated to stabilize initiation of transcription. In humans, most genes have more than one transcription start site, and these often exhibit different tissue specificity, serving as distinct regulatory frameworks for the same gene.  Usage of such promoters can also result in differential gene function manifested on the protein level. Alternative promoter usage has been shown to be increased in several disease states, especially cancer.   In this study, we have applied the nanoCAGE method to create a genome-wide map of TSS usage in sorted leukemic blasts from acute myeloid leukemia patients, and corresponding normal controls.   We show that the nanoCAGE method can replace similar experiments made with microarrays in terms of expression, but also that it uniquely, allow for the identification of alternative promoter usage in cancer cells. We identify 2,162 putative promoters that are significantly differentially regulated between APL and controls. Interestingly, promoters whose usage is upregulated in APL have an increased propensity to be downstream alternative promoters, and conversely, the promoters producing the annotated longest gene variants are commonly downregulated in cancer. We show examples of genes with upregulated downstream promoters, and demonstrate protein domain loss that could contribute to leukemic induction and maintenance. In conclusion, we present the first genome wide promoterome study from rare purified human leukemic cells. Overall design: nanoCAGE-seq of human acute myeloid leukemia samples vs. normal hematopoietic counterparts, both in replicates","project":"SRP022025"}
{"number_samples":8,"species":"human","abstract":"Individuals with 22q11.2 Deletion Syndrome (22q11.2 DS) are a specific high-risk group for developing schizophrenia (SZ), schizoaffective disorder (SAD) and autism spectrum disorders (ASD). Several genes in the deleted region have been implicated in the development of SZ, e.g., PRODH and DGCR8. However, the mechanistic connection between these genes and the neuropsychiatric phenotype remains unclear. To elucidate the molecular consequences of 22q11.2 deletion in early neural development, we carried out RNA-seq analysis to investigate gene expression in differentiating human neurons derived from induced pluripotent stem cells (iPSCs) of 22q11.2 DS SZ and SAD patients. Eight cases (ten iPSC-neuron samples in total including duplicate clones) and seven controls (nine in total including duplicate clones) were subject to RNA sequencing. Using a systems level analysis, differentially expressed genes/gene-modules and pathway of interests were identified. We observed ~2-fold reduction in expression of almost all genes in the 22q11.2 region in SZ (37 genes reached p-value < 0.05, 36 of which reached a false discovery rate < 0.05). Outside of the deleted region, 745 genes showed significant differences in expression between SZ and control neurons (p<0.05). Function enrichment and network analysis of the differentially expressed genes uncovered converging evidence on abnormal expression in key functional pathways, such as apoptosis, cell cycle and survival, and MAPK signaling in the SZ and SAD samples. Overall design: Eight cases (ten iPSC-neuron samples in total including duplicate clones) and seven controls (nine in total including duplicate clones)","project":"SRP022028"}
{"number_samples":2,"species":"human","abstract":"We cultured a parental cell line (M14) in the presence of increasing concentrations of BRAFi to select for BRAFi resistant mutants. We then performed RNAseq on the parental cell line and the derived BRAFi resistant mutant cell line to elucidate the mechanisms of acquired BRAFi resistance.","project":"SRP022029"}
{"number_samples":70,"species":"human","abstract":"We applied Next-Generation Sequencing (NGS) to miRNAs from blood samples of 48  AD (Alzheimer's Disease) patients and 22 unaffected controls, yielding a total of 140 unique mature miRNAs with significantly changed expression level. Of these, 82 were higher and 58 lower abundant in samples from AD patients. We selected a panel of 12 miRNAs for a qRT-PCR analysis on a larger cohort of 202 samples including not only AD patients and healthy controls but also patients with other CNS illnesses: Multiple Sclerosis, Parkinson's Disease, Major Depression, Bipolar Disorder, Schizophrenia, and Mild Cognitive Impairment, which is assumed to represent a transitional period before the development of AD. MiRNA target enrichment analysis of the selected 12 miRNAs indicated an involvement of miRNAs in nervous system development, neuron projection, neuron projection development, and neuron projection morphogenesis, respectively. Using this 12-miRNA signature we were able to differentiate between AD and controls with an accuracy of 93.3%, a specificity of 95.1%, and a sensitivity of 91.5%. The differentiation of AD from other neurological diseases was possible with accuracies between 73.8% and 77.8%. The differentiation of the other CNS disorders from controls yielded even higher accuracies. Overall design: Examination of the miRNA profile in blood samples of 48 AD patients and 22 controls","project":"SRP022043"}
{"number_samples":5,"species":"human","abstract":"High-throughput sequencing has opened numerous possibilities for the identification of regulatory RNA-binding events. Cross-linking and immunoprecipitation of Argonaute protein members can pinpoint microRNA target sites within tens of bases, but leaves the identity of the microRNA unresolved. A flexible computational framework that integrates sequence with cross-linking features reliably identifies the microRNA family involved in each binding event, considerably outperforms sequence-only approaches, and quantifies the prevalence of noncanonical binding modes. Overall design: Ago2 (Argonaute 2) PAR-CLIP and RNA deep sequencing of Epstein-Barr virus B95.8-infected Lymphoblastoid Cell Lines (LCLs)","project":"SRP022052"}
{"number_samples":38,"species":"human","abstract":"MiRNAs are discussed as diagnostic and therapeutic molecules. However, effective miRNA drug treatments with miRNAs are so far hampered by the complexity of the miRNA networks. To identify potential miRNA drugs in colorectal cancer, we profiled miRNA and mRNA expression in matching normal, tumor and metastasis tissues of eight patients by Illumina sequencing. We identified miRNA-1 as top candidate differentially expressed in tumor and metastasis. Furthermore, miRNA-1 was de-regulated in 16 additional tumor entities underscoring its central role in tumor pathogenesis. Functional analyses showed an additive effect of miRNA-1 with camptothecin treatment. We used a systems-biology simulation of cellular cancer models implemented in PyBios to investigate miRNA-1 function and assessed the effects of depletion as well as overexpression in terms of miRNA-1 as a potential treatment option. In this system miRNA-1 treatment reverted the disease phenotype with different effectiveness among the patients. Scoring the gene expression changes obtained through mRNA-Seq from the same patients we show that the combination of deep sequencing and systems biological modeling can help to identify patient-specific responses to miRNA treatments. We present this data as guideline for future pre-clinical assessments of new and personalized therapeutic options. Overall design: Examination of miRNA expression values by Illumina sequencing of matched benign, tumor and metastasis tissues of 8 colorectal cancer patients. For 4 of these patients all tissues have been resequenced to obtain mRNA expression values.","project":"SRP022054"}
{"number_samples":25,"species":"human","abstract":"A specific set of genes involved in regulating cellular immune response, antigen presentation, and T cell activation and survival were down-regulated 7 days after LVAD placement. 6 months following LVAD placement, the expression levels of these genes were significantly increased; yet importantly, remained significantly lower than age and sex-matched samples from healthy controls. Overall design: Examination of the effect of LVAD implant on peripheral blood transcriptome.  Blood was drawn before LVAD placement, 7 days post implant, and 180 days post implant.  RNA sequencing was performaed on all samples.","project":"SRP022133"}
{"number_samples":3,"species":"human","abstract":"This project contains whole genome, exome and transcriptome sequence from the HCC1395 (ATCC CRL-2324) breast carcinoma and corresponding HCC1395BL (ATCC CRL-2325) B lymphoblastoid cell lines of a 43 year old white female with a TNM stage I, grade 3 primary ductal carcinoma and a prior history of cancer.","project":"SRP022140"}
{"number_samples":4,"species":"human","abstract":"Acute myeloid leukemia (AML) continues to have the lowest survival rates of all leukemias. Therefore, new therapeutic strategies are urgently needed to improve clinical outcomes for AML patients. Here, we report a novel role for Wilms’ tumor 1-associated protein (WTAP) in pathogenesis of AML. We have performed RNA-Seq in K562 cells with knockdown of WTAP to ascertain which genes it regulates. Overall design: We have 2 replicates of total RNA for K562 cells and 2 replicates with WTAP knocked down","project":"SRP022166"}
{"number_samples":2,"species":"human","abstract":"Melanocytes within benign human nevi are the paradigm for tumor suppressive senescent cells in a pre-malignant neoplasm. These cells typically contain mutations in either the BRAF or N-RAS oncogene and express markers of senescence, including p16. However, a nevus can contain 10s to 100s of thousands of clonal melanocytes and approximately 20-30% of melanoma are thought to arise in association with a pre-existing nevus. Neither observation is indicative of fail-safe senescence-associated proliferation arrest and tumor suppression. We set out to better understand the status of nevus melanocytes. Proliferation-promoting Wnt target genes, such as cyclin D1 and c-myc, were repressed in oncogene-induced senescent melanocytes in vitro, and repression of Wnt signaling in these cells induced a senescent-like state. In contrast, cyclin D1 and c-myc were expressed in many melanocytes of human benign nevi. Specifically, activated Wnt signalling in nevi correlated inversely with nevus maturation, an established dermatopathological correlate of clinical benignancy. Single cell analyses of lone epidermal melanocytes and nevus melanocytes showed that expression of proliferation-promoting Wnt targets correlates with prior proliferative expansion of p16-expressing nevus melanocytes. In a mouse model, activation of Wnt signaling delayed, but did not bypass, senescence of oncogene-expressing melanocytes, leading to massive accumulation of proliferation-arrested, p16-positive non-malignant melanocytes. We conclude that clonal hyperproliferation of oncogene-expressing melanocytes to form a nevus is facilitated by transient delay of senescence due to activated Wnt signaling. The observation that activation of Wnt signaling correlates inversely with nevus maturation, an indicator of clinical benignancy, supports the notion that persistent destabilization of senescence by Wnt signaling contributes to the malignant potential of nevi. Overall design: We used RNA-Seq to detail the global programme of gene expression in primary human melanocytes which were Uninfected and BRAF600V induced cells","project":"SRP022259"}
{"number_samples":7,"species":"human","abstract":"Melanocytes within benign human nevi are the paradigm for tumor suppressive senescent cells in a pre-malignant neoplasm. These cells typically contain mutations in either the BRAF or N-RAS oncogene and express markers of senescence, including p16. However, a nevus can contain 10s to 100s of thousands of clonal melanocytes and approximately 20-30% of melanoma are thought to arise in association with a pre-existing nevus. Neither observation is indicative of fail-safe senescence-associated proliferation arrest and tumor suppression. We set out to better understand the status of nevus melanocytes. Proliferation-promoting Wnt target genes, such as cyclin D1 and c-myc, were repressed in oncogene-induced senescent melanocytes in vitro, and repression of Wnt signaling in these cells induced a senescent-like state. In contrast, cyclin D1 and c-myc were expressed in many melanocytes of human benign nevi. Specifically, activated Wnt signalling in nevi correlated inversely with nevus maturation, an established dermatopathological correlate of clinical benignancy. Single cell analyses of lone epidermal melanocytes and nevus melanocytes showed that expression of proliferation-promoting Wnt targets correlates with prior proliferative expansion of p16-expressing nevus melanocytes. In a mouse model, activation of Wnt signaling delayed, but did not bypass, senescence of oncogene-expressing melanocytes, leading to massive accumulation of proliferation-arrested, p16-positive non-malignant melanocytes. We conclude that clonal hyperproliferation of oncogene-expressing melanocytes to form a nevus is facilitated by transient delay of senescence due to activated Wnt signaling. The observation that activation of Wnt signaling correlates inversely with nevus maturation, an indicator of clinical benignancy, supports the notion that persistent destabilization of senescence by Wnt signaling contributes to the malignant potential of nevi. Overall design: We used RNA-Seq to detail the global programme of gene expression in human melanoma cell lines","project":"SRP022260"}
{"number_samples":6,"species":"human","abstract":"A673 Ewing sarcoma cells were transfected with siRNAs targeting ZEB2 (siZEB2-5 and siZEB2-6) or a control siRNA (siControl) and RNAseq was performed to identify ZEB2 regulated genes in Ewing sarcoma.","project":"SRP022361"}
{"number_samples":6,"species":"human","abstract":"The conserved human LIN28 RNA-binding proteins function in development, maintenance of pluripotency and oncogenesis. We used PAR-CLIP and a newly developed variant of this method, iDo-PAR-CLIP, to identify LIN28B targets as well as sites bound by the individual RNA binding domains of LIN28B in the human transcriptome at nucleotide resolution. The position of target binding sites reflected the known structural relative orientation of individual LIN28B binding domains, validating iDo-PAR-CLIP. Our data suggest that LIN28B directly interacts with most expressed mRNAs and members of the let-7 microRNA family. The Lin28 binding motif detected in pre-let-7 was enriched in mRNA sequences bound by LIN28B. Upon LIN28B knock down, cell proliferation and the cell cycle were strongly impaired. Quantitative shotgun proteomics of LIN28B depleted cells revealed significant reduction of protein synthesis from its RNA targets that function in translation, mRNA splicing and cell cycle control. Computational analyses provided evidence that the strength of protein synthesis reduction correlated with the location of LIN28B binding sites within target transcripts. Overall design: We used PAR-CLIP and a newly developed variant of this method, iDo-PAR-CLIP, to identify LIN28B targets as well as sites bound by the individual RNA binding domains of LIN28B in the human transcriptome at nucleotide resolution.","project":"SRP022591"}
{"number_samples":4,"species":"human","abstract":"Administration of mesenchymal stem cells (MSCs) has the potential to ameliorate degenerative disorders and to repair damaged tissues. The homing of transplanted MSCs to injured sites is a critical property of engraftment. Our aim was to identify microRNAs involved in controlling MSC proliferation and migration. MSCs can be isolated from bone marrow and umbilical cord Wharton's jelly (BM-MSCs and WJ-MSCs, respectively), and WJ-MSCs show poorer motility yet have a better amplification rate compared to BM-MSCs. Overall design: One human BM-MSC (pooled sample of 4 independent donors), one human WJ-MSC (pooled sample of 3 independent donors), and differentiated osteocytes and adipocytes derived from BM-MSCs after 2 weeks induction.","project":"SRP022772"}
{"number_samples":4,"species":"human","abstract":"RNA-seq and ChIP-seq on MCF-7 breast cancer cell line upon activation of p53 by the non-genotoxic small molecule Nutlin-3a Overall design: RNA-seq on MCF7 without (NS) or with Nutlin-3a stimulation (S), in duplicate, using illumina HiSeq 2000","project":"SRP022871"}
{"number_samples":6,"species":"human","abstract":"SOX2 is a transcription factor essential for pluripotent stem cells, and development and maintenance of squamous epithelium. We previously reported SOX2 an oncogene subject to highly recurrent genomic amplification in squamous cell carcinomas (SCCs)1.  Here we demonstrate in SCCs that SOX2 interacts with another master squamous transcription factor p63, and through ChIP-seq show that genomic occupancy of SOX2 overlaps with that of p63 at a large number of loci and that they cooperatively regulate gene expression including ETV4, which we find essential for SOX2-amplified SCC cell survival. Furthermore, SOX2 binds to distinct genomic loci in SCCs than in embryonic stem cells and the SOX2-p63 coordinate binding is unique to SCC.  In addition, a subset of SOX2 genomic binding sites in SCC that lack p63 co-occupancy are co-occupied by the AP-1 transcriptional complex.  These demonstrate that SOX2’s actions in SCC differ substantially from its role in pluripotency and identify novel SOX2 interactions that will enable deeper characterization of SOX2’s function in SCC. Overall design: KYSE70 cells with stable expression of either pLKO-Tet-Op-shSOX2 or pLKO-Tet-Op-shTp63 were treated with 50ng/ml of doxycyline for 4 days. Total RNA was extracted, polyA+ selected, reverse transcribed, library constructed and sequencing was performed with Illumina HiSeq 2000. Differencial gene expression between the stable cell lines with Dox-induced and non-Dox treated was analyzed to determine the effects by suppression of either SOX2 or TP63 in KYSE70 cells.","project":"SRP022876"}
{"number_samples":3,"species":"human","abstract":"The RNA-binding protein RC3H1 (also known as ROQUIN) promotes TNFalpha mRNA decay via a 3'UTR constitutive decay element (CDE). Here, we applied PAR-CLIP to human RC3H1 to identify about 3800 mRNA targets with more than 16000 binding sites. A large number of sites are distinct from the consensus CDE and revealed a structure-sequence motif with U-rich sequences embedded in hairpins. RC3H1 binds preferentially short-lived and DNA damage induced mRNAs, indicating a role of this RNA-binding protein in the posttranscriptional regulation of the DNA damage response. Intriguingly, RC3H1 affects expression of NF-kB pathway regulators such as IkBalpha and A20. RC3H1 uses roquin and Zn-finger domains to contact a binding site in the A20 3'UTR, demonstrating a not yet recognized mode of RC3H1 binding. Knockdown of RC3H1 resulted in increased A20 protein expression, thereby interfering with IkB kinase and NF-kB activities, demonstrating that RC3H1 can modulate the activity of the IKK/NF-kB pathway. Overall design: One PAR-CLIP Sample with human HEK293 cells","project":"SRP022892"}
{"number_samples":2,"species":"human","abstract":"We sequenced mRNA from SA cells with SFRS9 knocking down or    non-silencing control. Overall design: Examination of alternative splicing between SFRS9 KD    and NS.","project":"SRP022905"}
{"number_samples":26,"species":"human","abstract":"Synthetic transcription factors can be applied in many areas of biotechnology, medicine, and basic research.  In contrast to current methods based on engineering new DNA-binding proteins, we show that Cas9 fused to a transcriptional activation domain can be targeted by combinations of guide RNA molecules to induce the expression of endogenous human genes. This simple approach for targeted gene activation circumvents the need for engineering new proteins and will enable widespread synthetic gene regulation. Overall design: HEK293T cells were transfected with plasmid expressing Cas9-VP64 fusion protein and a guide RNA. As a control, empty guide RNA was transfected. Gene expression was then measured using mRNA-seq, and differential expression calculated using DESeq. All experiments were performed in biological duplicates or triplicates.","project":"SRP022913"}
{"number_samples":6,"species":"human","abstract":"In this report, we examine transcripts on a genome-wide level between the synchronized cell cycles of hESCs and hESC-derived endoderm cells. We found 10347, 10299 and 10362 genes expressed in the G1, G1/S, and S phase of hESCs, respectively; 10333, 10227 and 10215 genes expressed at the G1, G1/S and S phase of derived endoderm. We compared the transcriptome between these data sets and identified genes with differentiated phase expression between hESCs and hESC-derived endoderm cells. Overall design: Examination of the transcriptome at the G1-S transition in hESCs compared to hESC-derived endoderm cells.","project":"SRP022920"}
{"number_samples":6,"species":"human","abstract":"The advent of high-throughput sequencing has led to an explosion of studies into the diversity, expression, processing, and lifespan of RNAs.  Recently, three different high-throughput sequencing-based methods have been developed to specifically study RNAs that are in the process of being degraded.  All three methods—genome-wide mapping of uncapped and cleaved transcripts (GMUCT), parallel analysis of RNA ends (PARE), and degradome sequencing—take advantage of the fact that Illumina sequencing libraries use T4 RNA ligase 1 to ligate an adapter to the 5’ end of RNAs that have a free 5’-monophosphate.  This condition for T4 RNA ligase 1 substrates means that mature mRNAs are not substrates of the enzyme because they have a 5’-cap.  As a result, these sequencing libraries are specifically made up of clones of decapped or degrading mRNAs and the 3’ fragment of cleaved miRNA and siRNA targets.  In this paper, we present a massively streamlined protocol for GMUCT that takes 2-3 days and can be initiated with as little as 5 µg of starting total RNA and involves only one gel size-selection step.  We show that the results are similar to libraries made using the previous GMUCT protocol and PARE. Overall design: GMUCT libraries were made from flower buds of the Arabidopsis thaliana accession Columbia (Col-0) and three human cell lines—the cervical cancer cell line HeLa, human embryonic kidney 293 T (HEK293T) cell line, and the human chronic myelogenous leukaemia cell line K562—two replicates of each.  Matched mRNA-seq libraries were made for each Col-0 GMUCT replicate. These libraries were sequenced on the Illumina HiSeq 2000 and the reads were trimmed to remove the adapter sequences (TGGAATTCTCGGGTGCCAAGGAACTCCAGTCA). The abundance of each unique read was determined, and unique reads were mapped to the Arabidopsis and human genomes, respectively.","project":"SRP022925"}
{"number_samples":3,"species":"human","abstract":"HCC827 sensitive (parental).","project":"SRP022942"}
{"number_samples":3,"species":"human","abstract":"Cellular mRNAs are permanently associated to proteins in the form of ribonucleoprotein particles. In addition to the hnRNP and SR protein families, the double-stranded RNA-binding (DRB) proteins play important roles in mRNA synthesis, modification, activity and decay (Dreyfuss et al, 2002; Stutz & Izaurralde, 2003). Staufen is a DRB protein first described as a maternal factor involved in the localised translation of specific mRNAs during early development in the fly (Riechmann & Ephrussi, 2001). One of the human homologues, human Staufen1 (hStau1) forms ribonucleoprotein complexes known as RNA granules that contain proteins involved in translation regulation as well as cytoskeleton and motor proteins to allow the movement of the granule on the microtubules. Although many cellular mRNAs have been found associated to these RNA granules, the specificity of such binding and the mechanims of hStau1-RNA interaction are still unclear. Here we identified a protected sequence signature with high homology to human Alu family of short interspersed elements at the 3’-untranslated region of mRNAs specifically associated to hStau1 protein. Using a combination of affinity chromatography, RNAse-protection, deep-sequencing and bioinformatic analyses to compare the mRNAs differentially associated to hStau1 or a mutant protein unable to bind RNA we defined a collection of mRNAs specifically associated to wt hStau1. A common sequence signature consisting of two opposite-polarity Alu motifs was present in the hStau1-associated mRNAs and was shown to be sufficient for binding to hStau1 and hStau1-dependent stimulation of protein expression. Our results unravel how hStau1 identifies a wide spectrum of cellular target mRNAs to control their localisation, expression and fate. We suggest that mammalian Staufen1 protein has adapted to use the evolutionary recent Alu elements as recognition signals thereby increasing its possibilities for rapid identification to new mRNA targets. Overall design: Staufen (wild type or mutant) associated RNAs were extracted from HEK293T cells and sequenced according the illumina mRNA-seq potocol (HiSeq2000 sequencer). A third sample, of RNA, associated to a wild type Staufen, was treated with RNase T1 so only RNA fragments directly attached to Staufen were purified and sequenced.","project":"SRP023111"}
{"number_samples":6,"species":"human","abstract":"We sequenced mRNAs from HeLa cells transduced with either scrambled shRNA or shRNA targeting ASCC3. The results have shown differential gene expression in ASCC3 knockdown cells, suggesting a regulatory role for ASCC3 in certain cellular pathway. Overall design: mRNA from HeLa cells transduced with either scrambled shRNA (ni) or shRNA targeting ASCC3 (ai) were harvested and used for deep sequencing by Illumina HiSeq 2000 sequencing instruments.","project":"SRP023199"}
{"number_samples":3,"species":"human","abstract":"Recently, gene-trap mutagenesis in near haploid human cells has been used to disrupt genes and identify novel host-pathogen interactions and elucidate mechanisms of drug action. Building on this technology, we here report the generation of a human gene-trap mutant collection of individual clones, which currently covers over a quarter of the expressed genome. Overall design: Strand specific RNA-Seq of ribosomal RNA depleted total cellular RNA to measure transcript abundance and to detect fusion transcripts in KBM7 cells at standard culture conditions.","project":"SRP023233"}
{"number_samples":80,"species":"human","abstract":"The earliest recognizable stages of breast neoplasia are lesions that represent a heterogeneous collection of epithelial proliferations currently classified based on morphology. Their role in the development of breast cancer is not well understood but insight into the critical events at this early stage will improve efforts in breast cancer detection and prevention. These microscopic lesions are technically difficult to study so very little is known about their molecular alterations.  To characterize the transcriptional changes of early breast neoplasia, we sequenced 3'- end enriched RNAseq libraries from formalin-fixed paraffin-embedded tissue of early neoplasia samples and matched normal breast and carcinoma samples from 25 patients. We find that gene  expression patterns within early neoplasias are distinct from both normal and breast cancer patterns and identify a pattern of pro-oncogenic changes, including elevated transcription of ERBB2, FOXA1, and GATA3 at this early stage. We validate these findings on a second independent gene expression profile data set generated by whole transcriptome sequencing. Measurements of protein expression by immunohistochemistry on an independent set of early neoplasias confirms that ER pathway regulators FOXA1 and GATA3, as well as ER itself, are consistently upregulated at this early stage. The early neoplasia samples also demonstrate coordinated changes in long non-coding RNA expression and microenvironment stromal gene expression patterns.  This study is the first examination of global gene expression in early breast neoplasia, and the genes identified here represent candidate participants in the earliest molecular events in the development of breast cancer. Overall design: 3SEQ was performed on 72 FFPE human breast samples from 25 patients:  24 normal, 25 early neoplasia, 9 carcinoma in situ, and 14 invasive cancer","project":"SRP023262"}
{"number_samples":12,"species":"human","abstract":"Although the function of DNA methylation in gene promoter regions is well established in transcriptional repression, the function of the evolutionarily conserved widespread distribution of DNA methylation in gene body regions remains incompletely understood. Here, we show that DNA methylation is enriched in included alternatively spliced exons (ASEs) and inhibiting DNA methylation results in aberrant splicing of ASEs. The methyl-CpG binding protein MeCP2 is enriched in included ASEs, particularly those that are also highly DNA methylated, and inhibition of DNA methylation disrupts specific targeting of MeCP2 to exons. Interestingly, ablation of MeCP2 results in increased nucleosome acetylation and aberrant skipping events of ASEs. We further show that inhibition of histone deacetylases leads to a highly significant overlap of exon skipping events caused by knocking-down MeCP2. Together, our data indicate that intragenic DNA methylation operates in exon definition to modulate alternative splicing and can enhance exon recognition via recruitment of the multifunctional protein MeCP2, which thereby maintains local histone hypoacetylation through its established interaction with HDACs. Overall design: RNA-Seq in IMR90 and HCT116","project":"SRP023270"}
{"number_samples":8,"species":"human","abstract":"Purpose: We aimed to investigate in depth the regulation of microRNA expression by hypoxia in the breast cancer cell line MCF-7, establish the relationship between microRNA expression and HIF binding sites, pri-miRNA transcription and microRNA processing gene expression. Methods: microRNA sequencing data and gene expression microarray data were generated from MCF-7 cells submitted to an hypoxia timecourse (16h, 32h and 48h at 1% Oxygen). Data was integrated to 500 published high-stringency HIF binding sites identified in MCF-7 cells. Results: We identified 41 microRNAs significantly up- and 28 down- regulated, of which 38 mature and 20 star forms are reported in conjunction with hypoxia for the first time. HIF-1a and HIF-2a binding sites within 50kb distance of microRNA loci were found by integration of HIF ChIP-seq data, showing overall association between binding sites and up-regulation. Gene expression profiling analysis showed no full coordination between pri-miRNA and microRNA expression, pointing towards additional levels of regulation. Several transcripts playing a role in microRNA processing were found regulated by hypoxia, of which two were HIF dependent. Conclusions: The data support the hypothesis that microRNA expression under hypoxia is regulated at transcriptional and post-transcriptional level. HIF is involved at both levels, regulating the transcription of certain microRNAs and also the expression of key elements of the microRNA processing pathway. Overall design: microRNA-seq profiles of MCF-7 exposed to hypoxia (1% Oxygen) for 16h (2 replicates), 32h (2 replicates) and 48h (2 replicates) and to normoxia (2 replicates) were generated using Illumina sequencing platform.","project":"SRP023533"}
{"number_samples":1,"species":"human","abstract":"LX22 SCLC primay xenograft line. Host species: mouse. Graft specied: human. The experiment goal was to separate and identify species-specific NGS reads.","project":"SRP023538"}
{"number_samples":6,"species":"human","abstract":"Transcriptome sequencing of HBV+ HKCI cell-lines.","project":"SRP023539"}
{"number_samples":1,"species":"human","abstract":"LX33 SCLC primary xenograft line. Host species: mouse. Graft Species: human. The experiment goal was to separate and identify species-specific NGS reads.","project":"SRP023540"}
{"number_samples":16,"species":"human","abstract":"Degraded RNA - RNA with low RIN.  U-251 MG brain gliablastoma cell line.","project":"SRP023548"}
{"number_samples":10,"species":"human","abstract":"Understanding cellular and molecular differences between human and non-human primates (NHPs) is essential to the basic comprehension of the evolution and diversity of our own species. Until now, preserved tissues have been the main source of most comparative studies between humans, chimpanzees (Pan troglodytes) and bonobos (Pan paniscus). However, these tissue samples do not fairly represent the distinctive traits of live cell behavior, are not amenable to genetic manipulation and do not allow translation of observed differences into phenotypical divergence. We hypothesized that induced pluripotent stem cells (iPSCs) could provide a unique biological resource to elucidate relevant phenotypical differences between human and the great apes and that those differences could have potential adaptation and speciation value. Here, we describe the generation and initial characterization of iPSCs from chimpanzees and bonobos as novel tools to explore our most recent evolution. Comparative gene expression analysis of human and NHP iPSCs revealed differences in regulation of Long Interspersed Nuclear Element (LINE-1 or L1) transposons. A force of change in mammalian evolution, L1 elements are retrotransposons that have remained active during primate evolution. We observed decreased levels of L1 restricting factors APOBEC3B (A3B)7 and PIWIL28 in NHP iPSCs which was correlated with increased human and chimpanzee L1 mobility and endogenous L1 mRNA levels. Moreover, results from manipulation of A3B and PIWIL2 levels in iPSCs suggested a causal inverse relationship between levels of these proteins and L1 activity. Finally, we found increased copy numbers of species-specific L1 elements in the genome of chimpanzees compared to humans, supporting the idea that increased L1 mobility in NHPs is not limited to iPSCs in culture and may have also occurred in the germline during primate evolution. We propose that differences in L1 mobility may have differentially shaped the genomes of humans and NHPs and could have had an adaptive significance. Overall design: polyA RNA-Seq profiling of iPS cells from human, chimpanzee, and bonobo, and small RNA-Seq profiling of human iPS cells.","project":"SRP023550"}
{"number_samples":1,"species":"human","abstract":"LX36 SCLC primary xenograft line. Host species: mouse. Graft species: human. The experiment goal was to separate and identify species-specific NGS reads.","project":"SRP024236"}
{"number_samples":2,"species":"human","abstract":"Natural CD4+FOXP3+ regulatory T (Treg) cells constitute a unique T-cell lineage that plays a pivotal role in maintaining immune homeostasis and immune tolerance. Recent studies provide evidence for the heterogeneity and plasticity of the Treg cell lineage. However, the fate of human Treg cells after loss of FOXP3 expression and the underlying epigenetic mechanisms remain to be fully elucidated. Here, we compared gene expression profiles and histone methylation status on two histone H3 lysine residues (H3K4me3 and H3K27me3) of expanded FOXP3+ and corresponding FOXP3-losing Treg cells. DGE assay showed that human Treg cells down-regulated Treg signature genes, whereas up-regulated a set of Th lineages-associated genes, especially for Th2, such as GATA3, GFI1 and IL13, after in vitro expansion. Furthermore, we found that reprogramming of Treg cells was associated with histone modifications, as shown by decreased abundance of permissive H3K4me3 within down-regulated Treg signature genes, such as FOXP3, CTLA4 and LRRC32 loci, although with no significant changes in H3K27me3 modification. Thus, our results indicate that human Treg cells could convert into a Th-like cells upon in vitro expansion, displaying a gene expression signature dominated by Th2 lineage associated genes, and the histone methylation might contribute to such conversion. Overall design: mRNA profiles of in-vitro-expanded FOXP3+ Treg and FOXP3-losing Treg cells generated by deep sequencing.","project":"SRP024244"}
{"number_samples":20,"species":"human","abstract":"Purpose: upper tract urothelial carcinoma (UTUC) is the predominant subtype of the renal pelvis carcinoma but current knowledge about the molecular properties and prognostic markers is sparse. In this study, we examined the genome-wide mRNA expression spectrum of UTUC aiming to characterize the molecular basis of this cancer, and identify potential prognostic markers and thus facilitate the clinical practices. Experimental Design: we compared the whole mRNA expression spectrum of cancer and matched normal tissues in 10 patients with UTUC using massively parallel sequencing, thereafter the protein levels and prognostic roles of ALDH2, CCNE1 and SMAD3 were evaluated under an independent validation set comprising of 104 patients. Results: mRNA down-regulation of ALDH2 and up-regulation of SMAD3 and CCNE1 in UTUC were revealed by expression profiling. And low protein expression of ALDH2 was associated with an adverse outcome for patients (p < 0.0001). Whereas high CCNE1 and SMAD3 were associated with adverse clinical outcome (p < 0.0001). And multivariate analysis revealed that all these three molecular markers were independent prognostic predictors. Besides, compared to the pathological TNM classification, All ALDH2, CCNE1 and SMAD3 were more competent in identifying patient subgroup with high mortality risk, and the molecular markers were able to predict the survival difference in the patients of T2 and T3 subgroup (p < 0.001), which could not be achieved by TNM staging. Conclusions: This is the ?rst prospective study that characterizes genome-wide mRNA expression profile of UTUC. We revealed the prognostic significance of ALDH2, CCNE1 and SMAD3, and these molecular marker were more robust than TNM system in clinical outcome prediction. Overall design: Whole mRNA expression spectrum of cancer and matched normal tissues in 10 patients with UTUC was obtained by using massively parallel sequencing","project":"SRP024268"}
{"number_samples":52,"species":"human","abstract":"Rationale: Genome-wide association studies (GWAS) and candidate gene studies have identified a number of loci linked to susceptibility of chronic obstructive pulmonary disease (COPD), a smoking-related disorder that originates in the airway epithelium. Objectives: Since airway basal cell (BC) stem/progenitor cells exhibit the earliest abnormalities associated with smoking (hyperplasia, squamous metaplasia), we hypothesized that smoker BC have a dysregulated transcriptome linked, in part, to known GWAS/candidate gene loci. Methods: Massive parallel RNA sequencing was used to compare the transcriptome of BC purified from the airway epithelium of healthy nonsmokers (n=10) and smokers (n=7). The chromosomal location of the differentially expressed genes was compared to loci identified by GWAS and candidate gene studies to confer risk for COPD. Measurements and Main Results: Smoker BC have 676 known genes differentially expressed compared to nonsmoker BC, dominated by smoking up-regulation. Strikingly, 166 (25%) of these genes are located on chromosome 19, with 13 localized to 19q13.2 (p<10-4 compared to chance), including TGFB1, LTBP4, EGLN2 and NFKBIB, genes associated with risk for COPD. Conclusions: These observations provide the first direct link of known genetic risks for smoking-related lung disease with the specific population of lung cells that undergoes the earliest changes associated with smoking. Overall design: The human airway basal cell transcriptome of 7 smokers versus 10 nonsmokers was compared using massive parallel RNA sequencing (Illumina HiSeq 2000).","project":"SRP024274"}
{"number_samples":4,"species":"human","abstract":"While thousands of long non-coding RNAs (lncRNAs) are expressed in higher eukaryotes, the potential regulatory roles of lncRNAs in regulated gene transcription programs remain rather poorly understood. Here, we report that two lncRNAs highly overexpressed in aggressive prostate cancer, PRNCR1 and PCGEM1, bind successively to the androgen receptor (AR) and strongly enhance both ligand-dependent and ligand-independent AR-mediated gene activation programs and proliferation in prostate cancer cells. Binding of PRNCR1 to the C-terminally acetylated AR on enhancers and its association with DOT1L appear to be required for recruitment of the second lncRNA, PCGEM, to the N-terminally methylated AR. Unexpectedly, recognition of the H3K4me3 promoter  mark by the PHD finger-domain of Pygopus2, recruited by PCGEM1, proves to enhance selective looping of AR-bound enhancers to target gene promoters in these cells, revealing a novel aspect of ligand-induced enhancer-promoter interactions. In “resistant” prostate cancer cells, these overexpressed lncRNAs can interact with, and are required for the robust activation of both truncated and full length AR, causing DHT-independent activation of the AR transcriptional program and cell proliferation. Conditionally-expressed short hairpin RNA (shRNA)-mediated targeting of these lncRNAs in these resistant cancer cell lines strongly suppressed xenograft growth in vivo. Together, these results suggest that these overexpressed lncRNAs can potentially serve as a required component of castration-resistance in prostatic tumors. Overall design: Global Run On (GRO) assay followed by high throughput sequencing (GRO-seq); after knocking down PYGO2 LNCaP cells were grown to 30-50% confluence and siRNA/ASO transfections were carried out using Lipofectamine 2000 (Invitrogen) according to the  manufacturer’s instructions. Control samples were transfected with scramble  ASO and control siRNA, respectively. On the following day of transfection,  the cells were cultured in UltraCULTURE (Phenol red free) + 5% Charcoal  Dextran Stripped (CDS) serum for 48 hours. For androgen induction, we treat  cells with DHT from a 100 uM stock in 70% ethanol to a final concentration of 100 nM for 1 hour Control siRNA, -DHT Control siRNA, +DHT Pygo2 siRNA, -DHT Pygo2 siRNA, +DHT","project":"SRP024669"}
{"number_samples":6,"species":"human","abstract":"While thousands of long non-coding RNAs (lncRNAs) are expressed in higher eukaryotes, the potential regulatory roles of lncRNAs in regulated gene transcription programs remain rather poorly understood. Here, we report that two lncRNAs highly overexpressed in aggressive prostate cancer, PRNCR1 and PCGEM1, bind successively to the androgen receptor (AR) and strongly enhance both ligand-dependent and ligand-independent AR-mediated gene activation programs and proliferation in prostate cancer cells. Binding of PRNCR1 to the C-terminally acetylated AR on enhancers and its association with DOT1L appear to be required for recruitment of the second lncRNA, PCGEM, to the N-terminally methylated AR. Unexpectedly, recognition of the H3K4me3 promoter  mark by the PHD finger-domain of Pygopus2, recruited by PCGEM1, proves to enhance selective looping of AR-bound enhancers to target gene promoters in these cells, revealing a novel aspect of ligand-induced enhancer-promoter interactions. In “resistant” prostate cancer cells, these overexpressed lncRNAs can interact with, and are required for the robust activation of both truncated and full length AR, causing DHT-independent activation of the AR transcriptional program and cell proliferation. Conditionally-expressed short hairpin RNA (shRNA)-mediated targeting of these lncRNAs in these resistant cancer cell lines strongly suppressed xenograft growth in vivo. Together, these results suggest that these overexpressed lncRNAs can potentially serve as a required component of castration-resistance in prostatic tumors. Overall design: Global Run On (GRO) assay followed by high throughput sequencing (GRO-seq); after knocking-down lincRNAs PCGEM1 and PRNCR1. LNCaP cells were grown to 30-50% confluence and siRNA/ASO transfections were carried out using Lipofectamine 2000 (Invitrogen) according to the  manufacturer’s instructions. Control samples were transfected with scramble  ASO and control siRNA, respectively. On the following day of transfection,  the cells were cultured in UltraCULTURE (Phenol red free) + 5% Charcoal  Dextran Stripped (CDS) serum for 48 hours. For androgen induction, we treat  cells with DHT from a 100 uM stock in 70% ethanol to a final concentration of 100 nM for 1 hour Scramble ASO, -DHT Scramble ASO, +DHT PRNCR1 ASO, -DHT PRNCR1 ASO, +DHT PCGEM1 ASO, -DHT PCGEM1 ASO, +DHT","project":"SRP024674"}
{"number_samples":4,"species":"human","abstract":"The condensin complex is required for chromosome condensation during mitosis. However its role in interphase is not clear. Neuroblastoma is the most common extracranial childhood tumor of sympathetic neuron. In human neuroblastoma, MYCN amplification correlates with poor prognosis. Results identify the genes that increased/decreased when Smc2 knockdowned in MYCN-overexperssing or control SH-EP neuroblastoma cell line.","project":"SRP025977"}
{"number_samples":1720,"species":"human","abstract":"We present primary results from the Sequencing Quality Control (SEQC) project, coordinated by the United States Food and Drug Administration. Examining Illumina HiSeq, Life Technologies SOLiD and Roche 454 platforms at multiple laboratory sites using reference RNA samples with built-in controls, we assess RNA sequencing (RNA-seq) performance for sequence discovery and differential expression profiling and compare it to microarray and quantitative PCR (qPCR) data using complementary metrics. At all sequencing depths, we discover unannotated exon-exon junctions, with >80% validated by qPCR. We find that measurements of relative expression are accurate and reproducible across sites and platforms if specific filters are used. In contrast, RNA-seq and microarrays do not provide accurate absolute measurements, and gene-specific biases are observed, for these and qPCR. Measurement performance depends on the platform and data analysis pipeline, and variation is large for transcriptlevel profiling. The complete SEQC data sets, comprising >100 billion reads (10Tb), provide unique resources for evaluating RNA-seq analyses for clinical and regulatory settings. Overall design: The well-characterized reference RNA samples A (pooled cell lines) and B (human brain) from the MAQC consortium, adding spike-ins of synthetic RNA from the External RNA Control Consortium (ERCC).  Samples C and D were then constructed by combining A and B in known mixing ratios, 3:1 and 1:3, respectively.  All samples were distributed to several independent sites for RNA-Seq library construction and profiling by Illumina HiSeq 2000 and LifeTech SOLiD 5500 platforms.  Also, vendors created their own cDNA libraries that were then distributed to each test site, in order to examine the degree of a ?site effect? that was independent of the library preparation process.  To support an assessment of gene models, samples A and B were also sequenced at independent sites by the Roche 454 platform, providing longer reads.  For comparison to other technologies, these data were also compared to the MAQC-I Affymetrix U133 Plus2 microarray, several current microarray platforms, and also assessed by 20,801 PrimePCR reactions. Sample A: Universal Human Reference RNA (UHRR) from Stratagene and ERCC Spike-In controls Sample B: Human Brain Reference RNA (HBRR) from Ambion and ERCC Spike-In controls Sample C: Mix of A and B (3:1) Sample D: Mix of A and B (1:3) Sample E: Ambion ERCC Spike-In Control Mix 1 Sample F: Ambion ERCC Spike-In Control Mix 2","project":"SRP025982"}
{"number_samples":1,"species":"human","abstract":"MiRNA-mediated regulation depends on the stoichiometry between miRNAs and their mRNA targets. To decipher dynamic function of this complex layer, it is critical to characterize individual miRNA species within a specific cellular context. Small RNA cloning followed by deep sequencing is uniquely positioned as a genome-wide profiling method to quantify miRNA expression with potentially unlimited dynamic range and provide single-nucleotide resolution for precise miRNA classification and de novo discovery. However, significant biases introduced by RNA ligation steps in the current RNA cloning protocol often lead to inaccurate miRNA quantification by >1000-fold deviation. As a result, it has greatly hindered the broad application of this method. Here we report a highly efficient RNA cloning method that achieves over 90% efficiency for both 5’ and 3’ ligations with diverse small RNA substrates. When applied to a pool of either equimolar or differentially mixed synthetic miRNAs, the deviation of the cloning frequency for each miRNA is minimized to less than 2-fold of the anticipated value. By using samples obtained from multiple tissues and cells, we further demonstrate the accurate quantification of miRNA expression over a dynamic range of four orders of magnitude. Our results also reveal that most cistronic miRNAs are expressed at similar levels and, in each cell population, miRNAs repress their cognate targets in a dosage dependent manner. Collectively, our high-efficiency RNA cloning method combining with deep sequencing establishes a cost-effective approach for accurate genome-wide miRNA profiling. Overall design: We designed an artificial system composed of synthetic miRNAs for benchmarking biases in small RNA cDNA cloning for NGS.","project":"SRP025989"}
{"number_samples":16,"species":"human","abstract":"Objectives: The sequencing by the PolyA selection is the most common approach for library preparation. With limited amount or degraded RNA, alternative protocols such as the NuGEN have been developed. However, it is not yet clear how the different library preparations affect the downstream analyses of the broad applications of RNA sequencing. Methods and Materials: Eight human mammary epithelial cell (HMEC) lines with high quality RNA were sequenced by Illumina’s mRNA-Seq PolyA selection and NuGEN ENCORE library preparation. The following analyses and comparisons were conducted: 1) the numbers of genes captured by each protocol; 2) the impact of protocols on differentially expressed gene detection between biological replicates; 3) expressed single nucleotide variant (SNV) detection; 4) non-coding RNAs, particularly lincRNA detection; and 5) intragenic gene expression. Results: Sequences from the NuGEN protocol had lower (75%) alignment rate than the PolyA (over 90%). The NuGEN protocol detected fewer genes (12-20% less) with a significant portion of reads mapped to non-coding regions. A large number of genes were differentially detected between the two protocols. About 17-20% of the differentially expressed genes between biological replicates were commonly detected between the two protocols. Significantly higher numbers of SNVs (5-6 times) were detected in the NuGEN samples, which were largely from intragenic and intergenic regions. The NuGEN captured fewer exons (25% less) and had higher base level coverage variance. While 6.3% of reads were mapped to intragenic regions in the PolyA samples, the percentages were much higher (20-25%) for the NuGEN samples. The NuGEN protocol did not detect more known non-coding RNAs such as lincRNAs, but targeted small and “novel” lincRNAs. Conclusion: Different library preparations can have significant impacts on downstream analysis and interpretation of RNA-seq data. The NuGEN provides an alternative for limited or degraded RNA but it has limitations for some RNA-seq applications. Overall design: Eight human mammary epithelial cell (HMEC) lines with high quality RNA were sequenced by Illumina’s mRNA-Seq PolyA selection and NuGEN ENCORE library preparation. The goal was to look the impacts of protocols on results and intepretation.","project":"SRP026013"}
{"number_samples":3,"species":"human","abstract":"hESC have morphologic, genetic and genomic alternatiions when cells cultured in different passaging condition. Overall design: Cells had initially mechanic passaging. From the initiating passage p13, cell were switched to enzymatic passaging after 15 passage (p13+EP15). Another sample was from cells after 15 mechanic passages (p13+MP15) which was designed as control. The transcripome of p13, p13+EP15 and p13+MP15 was analyzed with RNA sequencing by Illumina HiSeq2000 next-generation sequencer.","project":"SRP026033"}
{"number_samples":84,"species":"human","abstract":"Environmental stimuli are known to contribute to psoriasis pathogenesis and that of other autoimmune diseases, but the mechanism is unknown. Here we show that the aryl hydrocarbon receptor (AhR), a transcription factor that senses environmental stimuli, modulates pathology in psoriasis. AhR-activating ligands reduced inflammation in the lesional skin of psoriasis patients, whereas AhR antagonists upregulated inflammation. Similarly, AhR signaling via the endogenous FICZ ligand reduced the inflammatory response in the imiquimod-induced model of psoriasis and AhR deficient mice exhibited a substantial exacerbation of the disease, compared to AhR sufficient controls. Non-haematopoietic cells, in particular keratinocytes, were responsible for this hyper-inflammatory response, which involved increased reactivity to IL-1beta and upregulation of AP-1 family members of transcription factors. Thus, our data suggest a critical role for AhR in the regulation of inflammatory responses and open the possibility for novel therapeutic strategies in chronic inflammatory disorders. Overall design: Total RNA obtained from skin explants taken from psoriatic patients or healthy donors cultured in the presence of AhR agonist or antagonist","project":"SRP026042"}
{"number_samples":3,"species":"human","abstract":"The small nuclear RNA (snRNA) genes have been widely used as a model system for understanding transcriptional regulation due to unique aspects of their promoter structure, selectivity for either RNA Polymerase (Pol) II or III and a unique mechanism of termination that is tightly linked with the promoter.  Recently, we identified the Little Elongation Complex (LEC) in Drosophila that is required for the expression of Pol II-transcribed snRNA genes. Here, we identify the molecular mechanism by which LEC specifically regulates Pol II-dependent snRNA gene transcription. We present genetic and molecular evidence from both Drosophila and mammals that LEC regulates both initiation and elongation stages of transcription of Pol II-transcribed snRNA genes. Overall design: In human HCT116 cells we performed: ChIP-seq of ICE1, ICE2, ZC3H8, ELL, and AFF4; total RNA-seq following ICE1 knock-down and non-targeting (GFP) knock-down; ChIP-seq of ICE1 and Pol II following non-targetting (shGFP) and ICE1 knock-down (shICE1).  In fly S2 cells we performed: Ice1 ChIP-seq following small hairpin knock-down of GFP (shGFP/non-targeting control) and Ice1 (knock-down of Ice1); ChIP-seq of Pol II following small hairpin knock-down of GFP (shGFP/non-targeting control), Ice1 (knock-down of Ice1), and Ell (knock-down of Ell).","project":"SRP026044"}
{"number_samples":16,"species":"human","abstract":"Whole-genome single-base resolution methylcytosine and hydroxymethylcytosine maps reveal profound changes that occur during frontal cortex development in humans and mice. Overall design: MethylC-Seq, TAB-Seq, RNA-Seq and hmC-IP (CMS-IP and biotin-glucosyl tagging) from Homo sapiens and Mus musculus frontal cortex tissue and neural sorted cell populations. Additionally, MethylC-Seq was performed on the HUES6 embryonic stem cell line (Homo sapiens).","project":"SRP026048"}
{"number_samples":5,"species":"human","abstract":"The RNA helicase UPF1 is best known for its key function in mRNA nonsense-mediated mRNA decay (NMD), but has also been implicated in additional mRNA turnover mechanisms, telomere homeostasis, and DNA replication. In NMD, UPF1 recruitment to target mRNAs is thought to occur through interaction with release factors at terminating ribosomes, but evidence for translation-independent interaction of UPF1 with the 3’ untranslated region (UTR) of mRNAs has also been reported. To map UPF1 binding sites transcriptome-wide, we performed individual-nucleotide resolution UV crosslinking and immunoprecipitation (iCLIP) in human cells, untreated or after inhibiting translation by puromycin. We found a strong association of UPF1 with 3’ UTRs in undisturbed, translationally active cells and a significant increase in UPF1 binding to coding sequence (CDS) after translation inhibition. These results indicate that UPF1 binds RNA before translation and gets displaced from the CDS by translating ribosomes. This evidence for translation-independent UPF1-RNA interaction, which is corroborated by RNA immunoprecipitations experiments and by our observation that UPF1 also crosslinks to long non-coding RNAs, suggests that the decision to trigger NMD occurs after association of UPF1 with the mRNA, presumably through activation of RNA-bound UPF1 by aberrant translation termination. Overall design: Examination of Upf1 binding preferences via iCLIP in untreated HeLa cells and HeLa cells, where translation is blocked by puromycin treatment in vivo crosslinking and immunoprecipitation strategy (iCLIP)","project":"SRP026052"}
{"number_samples":8,"species":"human","abstract":"Alternative mRNA splicing is a major mechanism for gene regulation and transcriptome diversity. Despite the extent of the phenomenon, the regulation and specificity of the splicing machinery are only partially understood. Adenosine-to-inosine (A-to-I) RNA editing of pre-mRNA by ADAR enzymes has been linked to splicing regulation in several cases. Here we used bioinformatics approaches, RNA-seq and exon-specific microarray of ADAR knockdown cells to globally examine how ADAR and its A-to-I RNA editing activity influence alternative mRNA splicing. Although A-to-I RNA editing only rarely targets canonical splicing acceptor, donor, and branch sites, it was found to affect splicing regulatory elements (SREs) within exons. Cassette exons were found to be significantly enriched with A-to-I RNA editing sites compared with constitutive exons. RNA-seq and exon-specific microarray revealed that ADAR knockdown in hepatocarcinoma and myelogenous leukemia cell lines leads to global changes in gene expression, with hundreds of genes changing their splicing patterns in both cell lines. This global change in splicing pattern cannot be explained by putative editing sites alone. Genes showing significant changes in their splicing pattern are frequently involved in RNA processing and splicing activity. Analysis of recently published RNA-seq data from glioblastoma cell lines showed similar results. Our global analysis reveals that ADAR plays a major role in splicing regulation. Although direct editing of the splicing motifs does occur, we suggest it is not likely to be the primary mechanism for ADAR-mediated regulation of alternative splicing. Rather, this regulation is achieved by modulating trans-acting factors involved in the splicing machinery. Overall design: HepG2 and K562 cell lines were stably transfected with plasmids containing siRNA designed to specifically knock down ADAR expression (ADAR KD). This in order to examine how ADAR affects alternative splicing globally.","project":"SRP026084"}
{"number_samples":1,"species":"human","abstract":"We have used RNA-seq to examine circular RNAs from RNase R treated poly(A)-/ribo- RNAs in human embryonic stem cells Overall design: Examine circular RNAs in human embryonic stem cells","project":"SRP026089"}
{"number_samples":422,"species":"human","abstract":"Next-generation sequencing (NGS) technology applications like RNA-sequencing (RNA-seq) have dramatically expanded the potential for novel genomics discoveries, but the proliferation of various platforms and protocols for RNA-seq has created a need for reference data sets to help gauge the performance characteristics of these disparate methods.  Here we describe the results of the ABRF-NGS Study on RNA-seq, which leverages replicate experiments across multiple sites using two reference RNA standards tested with four protocols (polyA selected, ribo-depleted, size selected, and degraded RNA), and examined across five NGS platforms (Illumina’s HiSeqs, Life Technologies’ Personal Genome Machine and Proton, Roche 454 GS FLX, and Pacific Biosciences RS).  These results show high (R2 >0.9) intra-platform consistency across test sites, high inter-platform concordance (R2 >0.8) for transcriptome profiling, and a large set of novel splice junctions observed across all platforms.  Also, we observe that protocols using ribosomal RNA depletion can both salvage degraded RNA samples and also be readily compared to polyA-enriched fractions.  These data provide a broad foundation for standardization, evaluation and improvement of RNA-seq methods. Overall design: Two reference RNA standards tested with four protocols (polyA selected, ribo-depleted, size selected, and degraded RNA), and examined across five NGS platforms (Illumina’s HiSeqs, Life Technologies’ Personal Genome Machine and Proton, Roche 454 GS FLX, and Pacific Biosciences RS). Please note that the samples were named following the ABRF-Platform-Site-Sample-Replicate# format. For example, ABRF-454-CNL-A-1 means Sample A was run on 454 platform at Cornell and this is the first replicate, and  ABRF-454-CNL-A-2 means the same exact sample was ran with same machine at same location and is 2nd replicate.","project":"SRP026126"}
{"number_samples":3,"species":"human","abstract":"An in-depth analysis of miRNomes in 3 human myeloid leukemia cell lines was carried out to comprehensively identify miRNAs that distinguish acute and chronic myeloid leukemias and relate to myeloid cell differentiation. Overall design: Characterization the miRNomes in 3 myeloid leukemia cell lines.","project":"SRP026144"}
{"number_samples":4,"species":"human","abstract":"The C4-12/Flag.ERß cell line which stably expressed Flag.ERß is used to study ERß genomic functions without ERa interference. Mapping ERß binding sites in these cells reveals ERß unique distribution and motif enrichment patterns. Accompanying our mapping results, nascent RNA profiling is performed on cells at the same treatment time. The combined results allow the identification of ERß target genes. Gene ontology analysis reveals that ERß targets are enriched in differentiation, development and apoptosis. Concurrently, E2 treatment suppresses proliferation in these cells. Within ERß binding sites, while the most prevalent binding motif is the canonical ERE, motifs of known ER interactors are also enriched in ERß binding sites. Moreover, among enriched binding motifs are those of GFI, REST and EBF1, which are unique to ERß binding sites in these cells. Further characterization confirms the association between EBF1 and the estrogen receptors, which favors the N-terminal region of the receptor. Furthermore, EBF1 negatively regulates ERs at the protein level. In summary, by studying ERß genomic functions in our cell model, we confirm the anti-proliferative role of ERß and discover the novel cross talk of ERß with EBF1 which has various implications in normal physiology. Overall design: C4-12/Flag.ERß cells were treated with 10nM E2 (or ethanol as vehicle control) for 1 hour. Nuclei were extracted and processed with run-on assay. The resultant run-on RNA was reverse-transcribed to generate cDNA library which was subsequently sequenced by Illumina Genome Analyzer II or HiSeq2000. Two samples for each treatment were included in this experiment.","project":"SRP026204"}
{"number_samples":30,"species":"human","abstract":"Changes in gene expression contribute to the pathogenesis of heart failure. The sequence, expression level, and structure of the human cardiac transcriptome are incompletely described, as are their changes in heart disease. High throughput transcriptome sequencing (RNA-seq) is a quantitative and unbiased approach to measure transcript level and to identify novel transcribed elements or transcript splicing. Here we acquired 975.2 x 106 mapped RNA-seq reads in 15 control and 15 ischemic cardiomyopathy (ICM) hearts, obtained at the time of heart transplantation. We identified over 1000 differentially expressed transcripts, and thousands of novel transcribed elements, some of which were differentially expressed in between control and ICM groups. We found that transcript processing of several cardiac genes was deranged in ICM. For instance, the ratio between specific MYH6 exons was significantly changed in ICM compared to controls, while this type of inter-exon variation was not observed for the adjoining gene MYH7. This RNA-seq study of the human heart failure transcriptome revealed the diversity of transcripts expressed in the human heart and their complex patterns of expression in the diseased heart. Overall design: Transcriptome profiling (RNA-seq) of 15 control and 15 ischemic cardiomyopathy (ICM) hearts using Illumina GAII and SOLiD","project":"SRP026208"}
{"number_samples":10,"species":"human","abstract":"This paper shows, for the first time, a novel function of the OVO-like proteins (OVOL1and OVOL2) as critical inducers of mesenchymal to epithelial transition (MET) in human cancer. Overall design: Examination of the effects of OVOL1 and OVOL2 overexpression in a prostate cancer model.","project":"SRP026256"}
{"number_samples":5,"species":"human","abstract":"RNA samples were extracted before and after infection at different timepoints (0.5hpi, 2hpi, 6hpi, 18hpi)","project":"SRP026257"}
{"number_samples":2,"species":"human","abstract":"This project looks into experimentally identifying all minor introns by knocking down the minor spliceosome's catalytic snRNP, U6atac. Overall design: Knockdown of U6atac by antisense morpholino followed by examining mRNA splicing by RNA-seq","project":"SRP026297"}
{"number_samples":8,"species":"human","abstract":"Sequencing of 5' and 3'ends and RNA-seq of PROMPT and mRNA molecules from control and exosome-depleted cells. Overall design: CAGE, 3'TAG and RNAseq library construction from RNA extracted from control and exosome-depleted cells.","project":"SRP026315"}
{"number_samples":2,"species":"human","abstract":"Using mRNA-seq, we determined intron retaining genes that were differentially regulated in FACS purified cells at three progressive stages of mouse granulopoiesis; CD34+Kit+Gr-1low promyelocytes, CD34-Kit-Gr-1mid myelocytes and CD34-Kit-Gr-1high granulocytes. We found that IR affects 86 genes, including those specific to granulocyte (Lyz2 and MMP8) and nuclear architecture (Lmnb1 and Lbr). IR was associated with the decrease in protein levels measured by mass spectrometry (P=0.0015, binomial test). There was a significant overlap of IR between human and mouse (P=2.85E-22, hypergeometric test), showing that IR is conserved.Inhibition of NMD in granulocytes resulted in marked accumulation of 39/86 intron retaining mRNAs (P<0.05, RUV procedure with Holm-Bonferroni correction), indicating that IR triggers NMD to downregulate mRNA and protein expression. Overall design: Sequencing of polyadenylated RNA from three types of myeloid cells (promyelocytes, myelocytes and granulocytes) using Illumina GAIIx","project":"SRP026331"}
{"number_samples":3,"species":"human","abstract":"Objectives: Long non-coding RNAs (lncRNAs) have been shown to play important roles in the development and progression of cancer. However, functional lncRNAs and their downstream mechanisms are largely unknown in the molecular pathogenesis of esophageal adenocarcinoma (EAC) and its progression. Design: lncRNAs that are abnormally upregulated in EACs were identified by RNA-seq analysis, followed by quantitative RT-PCR (qRTPCR) validation using tissues from 31 EAC patients. Cell biological assays in combination with siRNA-mediated knockdown were performed in order to probe the functional relevance of these lncRNAs. Results: We discovered that a lncRNA, HNF1A-AS1, is markedly upregulated in human primary EACs relative to their corresponding normal esophageal tissues (mean fold change 7.2, p<0.01). We further discovered that HNF1A-AS1 knockdown significantly inhibited cell proliferation and anchorage independent growth, suppressed S-phase entry, and inhibited cell migration and invasion in multiple in vitro EAC models (p<0.05). A gene ontological analysis revealed that HNF1A-AS1 knockdown preferentially affected genes that are linked to assembly of chromatin and the nucleosome, a mechanism essential to cell cycle progression. The well-known cancer-related lncRNA, H19, was the gene most markedly inhibited by HNF1A-AS1 knockdown. Consistent to this finding, there was a significant positive correlation between HNF1A-AS1 and H19 expression in primary EACs (p<0.01). Overall design: In order to identify novel oncogenic lncRNAs in esophageal adenocarcinogenesis, we carried out RNA-seq of a matched NE-BE-EAC tissue pair","project":"SRP026333"}
{"number_samples":20,"species":"human","abstract":"Strand specific RNA-Seq of T2D","project":"SRP026359"}
{"number_samples":14,"species":"human","abstract":"Androgen ablation therapy (AAT) is standard treatment for locally-advanced/metastatic prostate cancer (PCa).  Many patients develop castration-resistance (CRPCa) after ~2-3 years, with a poor prognosis.  The molecular mechanisms underlying CRPCa progression are unclear.  mRNA-Seq was performed on tumours from 7 patients with locally-advanced/metastatic PCa before and ~22 weeks after AAT initiation.  Differentially regulated genes were identified in treatment pairs. Overall design: Tumour biopsies from 7 patients were taken before and after AAT treatment","project":"SRP026387"}
{"number_samples":12,"species":"human","abstract":"Transcriptome Profiling of Activated T Cells","project":"SRP026389"}
{"number_samples":3,"species":"human","abstract":"Recognition of modified histones by “reader” proteins plays a critical role in the regulation of transcription1. H3K36 trimethylation (H3K36me3) is deposited onto the nucleosomes in the transcribed regions following RNA polymerase II (Pol II) elongation. In yeast, this mark in turn recruits epigenetic regulators to reset the chromatin at an appropriate state to suppress cryptic transcription2,3. However, much less is known about the role of H3K36me3 in transcription regulation in mammals. This is further complicated by the transcription-coupled incorporation of the histone variant H3.3 in gene bodies4. Here we show that the candidate tumor suppressor ZMYND11 specifically recognizes H3K36me3 on H3.3 (H3.3K36me3) and regulates Pol II elongation. Structural studies reveal that in addition to the trimethyl-lysine binding by an aromatic cage within the PWWP domain, the H3.3-dependent recognition is mediated by the encapsulation of the H3.3-specific “Ser31” residue in a composite pocket formed by the tandem bromo-PWWP domains of ZMYND11. ChIP-sequencing analysis reveal a genome-wide colocalization of ZMYND11 with H3K36me3 and H3.3 in gene bodies, and its occupancy requires the pre-deposition of H3.3K36me3. Although ZMYND11 is associated with highly expressed genes, it functions as an unconventional transcription corepressor via modulating the transition of the promoter-proximal paused Pol II to elongation. ZMYND11 is critical for the repression of a transcriptional program that is essential for tumor cell growth; higher expression of ZMYND11 is observed in triple-negative breast cancer patients with better prognosis. Consistently, overexpression of ZMYND11 suppresses cancer cell growth and tumor formation in mice. Together, this study identifies ZMYND11 as an H3.3-specific reader of H3K36me3 that links the histone variant-mediated transcription elongation control to tumor suppression. Overall design: ChIP-seq analysis of ZMYND11, H3K36me3 in U2OS cells and ZMYND11 knockdown cells; ChIP-seq of H3.3 in Flag-H3.3 stable U2OS cells; RNA-seq of ZNYMD11 depleted U2OS cells.","project":"SRP026454"}
{"number_samples":64,"species":"human","abstract":"56 breast cancer cell lines were profiled to identify patterns of gene expression associated with subtype and response to therapeutic compounds. Overall design: Cell lines were profiled in their baseline, unperturbed state.","project":"SRP026537"}
{"number_samples":7,"species":"human","abstract":"Sequencing from Ovarian Cancer cell lines","project":"SRP026553"}
{"number_samples":12,"species":"human","abstract":"We report the high-throughput profiling of microRNAs (miRNAs) from the prefrontal cortex of controls, early and late-stages Alzheimer's disease subjects. We show miRNA expression changes between the two groups and down-regulation of miR-132-3p in the late-stages group. Overall design: Deep-sequencing of microiRNAs in 6 controls/early-stages and 6 late-stages Alzheimer's disease patients","project":"SRP026562"}
{"number_samples":20,"species":"human","abstract":"ieASV transcriptome project.","project":"SRP026597"}
{"number_samples":28,"species":"human","abstract":"We sequenced the WGS/transcriptome of 2 HCC patients and 2 Cellline.","project":"SRP026600"}
{"number_samples":12,"species":"human","abstract":"RNA-seq analysis was performed on TICs and the parental counterparts of 4 LSCC cell lines. In addition, PRKCI knock down (KD) variants of these cells were sequenced. Subsequent analysis revealed that activation of HH signaling, a known driver of the TIC phenotype, is dependent on PRKCI expression. Overall design: Examination of TICS, parental cells, parental PRKCI KD cells, and TIC PRKCI KD cells","project":"SRP026620"}
{"number_samples":2,"species":"human","abstract":"This experiment performed RNA-seq of transcriptome and translatome (translating mRNA) of Caco-2 cells Overall design: We extracted transcriptome and translatome from Caco-2 cells and deep sequenced them","project":"SRP026621"}
{"number_samples":8,"species":"human","abstract":"Human innate lymphoid cells: insights into function at homeostasis and during filarial infection.","project":"SRP026648"}
{"number_samples":16,"species":"human","abstract":"The anti-cancer drug camptothecin inhibits replication and transcription by trapping DNA topoisomerase I (Top1) covalently to DNA in a “cleavable complex”. To examine the effects of camptothecin on RNA synthesis genome-wide we used Bru-Seq and show that camptothecin treatment affected transcription initiation, elongation, termination, splicing and enhancer activity. Following removal of camptothecin, transcription spread as a wave from the 5’-end of genes with no recovery of transcription apparent from RNA polymerases stalled in the body of genes. As a result, camptothecin preferentially inhibited the expression of large genes such as proto-oncogenes, and anti-apoptotic genes while smaller ribosomal protein genes, pro-apoptotic genes and p53 target genes showed relative higher expression. In addition, a set of mitotic regulator genes and histone genes were inhibited in a size-independent manner. Cockayne syndrome group B fibroblasts showed a very similar RNA synthesis recovery profile to normal fibroblasts suggesting that transcription-coupled repair is not involved in the repair of transcription-blocking TOP1 lesions. These findings of the effects of camptothecin on transcription have important implications for its anti-cancer activities and may aid in the design of improved combinatorial treatments involving Top1 poisons. Overall design: Analysis of the effect of Camptothecin (CPT) on transcription. Normal fibroblasts and Cockayne syndrome group B fibroblasts were exposed to CPT for 60 minutes, after which a washout was performed. Nascent RNA was labeled using bromouridine for 15 minutes starting at time points: 1) 15 minutes before the washout; 2) Immediately after the washout; 3) 15 minutes after the washout. Test samples are compared to control cells that were not exposed to CPT.","project":"SRP026690"}
{"number_samples":12,"species":"human","abstract":"Here we use an integrated systems-level examination of transcription, translation, and proteolysis to explore how cancer cells struggle with a chemotherapeutic drug prior to succumbing to apoptosis. As a model system we study myeloma cells exposed to the proteasome inhibitor bortezomib, a first-line clinical treatment. Despite robust transcriptional changes, unbiased quantitative proteomics detects production of only a few critical anti-apoptotic proteins against a background of general translation inhibition. Ribosome profiling further reveals potential translational regulation of stress response genes following bortezomib treatment. Once the apoptotic machinery is engaged, degradation by caspases is largely independent of changes at the transcriptional level. Moreover, previously uncharacterized non-caspase proteolytic events also participate in cellular deconstruction. As suggested by these data, we find that inhibition of the anti-apoptotic response regulator HSF1 promotes cell death by bortezomib. Thus, monitoring global cellular dynamics after chemotherapy offers in-depth insight into apoptosis and can also guide potential therapeutic combinations. Overall design: We examined MM1.S myeloma cells exposed to 20 nM bortezomib across a time course with independent samples of poly(A) mRNA and ribosome footprints isolated at each of six time points (0h (untreated), 1.5h, 3h, 6h, 9h, 12h). Sequencing was performed on a Illumina HiSeq 2000 with single-end.","project":"SRP027015"}
{"number_samples":36,"species":"human","abstract":"Epidemiological studies provide strong evidence that consumption of cruciferous vegetables, such as broccoli, can significantly reduce the risk of developing cancers. Sulforaphane (SFN), a phytochemical derived from cruciferous vegetables, induces anti-proliferative and pro-apoptotic responses in prostate cancer cells, but not in normal prostate cells. The mechanisms responsible for these specific chemopreventive properties remain unclear. We utilized RNA sequencing to test the hypothesis that SFN modifies the expression of genes that are critical in prostate cancer progression. Normal prostate epithelial cells, and androgen-dependent and androgen-independent prostate cancer cells were treated with 15 µM SFN and the transcriptome was determined at 6 and 24 hour time points. SFN altered the expression of ~3,000 genes in each cell line and the response was highly dynamic over time. SFN influenced the expression of genes in functional groups and pathways that are critical in cancer including cell cycle, apoptosis and angiogenesis, but the specific effects of SFN differed depending on the state of cancer progression. Network analysis suggested that a transcription factor that is overexpressed in many cancers, Specificity protein 1 (Sp1), is a major mediator of SFN-induced changes in gene expression. Nuclear Sp1 protein was significantly decreased by 24 hour SFN treatment in prostate cancer cells, while a related transcription factor, Sp3 protein was only modestly decreased in androgen-independent prostate cancer cells.  Overall, the data show that SFN significantly affects gene expression in normal and cancer cells, with key targets in chemopreventive processes, making it a promising dietary anti-cancer agent. Overall design: Examination of how the transcriptome of normal and prostate cancer cells is altered by treatment with sulforaphane","project":"SRP027258"}
{"number_samples":8,"species":"human","abstract":"Small RNAs play a critical role in host-pathogen interaction. In insects, for instance, small RNA-mediated silencing or RNA interference (RNAi) represents the main antiviral defense system. However, the antiviral role of RNAi has not been clearly proven in higher vertebrates. On the contrary, it is well established that the cell response relies on the recognition of viral RNAs by host pattern recognition receptors (PRR) to trigger the activation of the interferon pathway. Based on this evidence, we wished to contribute to this research field by identifying and characterizing small non-coding RNAs produced in mammalian cells upon RNA virus infection. We focused on Sindbis virus (SINV), the prototypical arbovirus, which by definition, is able to infect both vertebrate hosts and invertebrate vectors and triggers the interferon pathway or RNAi, respectively. Overall design: Taking advantage of large scale sequencing, we cloned and sequenced small RNAs from both mock and SINV-infected mammalian cells (HEK 293 and VERO). We identified a novel population of viral small RNAs (vsRNAs) that accumulate as 20 to 30 nt species during infection. We assessed that this viral small RNA population is modified in 3'end and derived from the activation of the cellular antiviral endoribonuclease RNaseL. Altogether our results indicate a potential role for the SINV-derived small RNAs in the host defense mechanism.","project":"SRP027345"}
{"number_samples":136,"species":"human","abstract":"We analysed the transcriptome of different HSC-enriched subpopulations of cells sorted from human umbilical cord blood and isolated from several individuals with different genetic backgrounds. We aim at identifying new cell surface markers associated with human HSC and downstream mature hematopoietic cell activity. Overall design: RNA-seq of CD34+CD45RA- cord blood cells from 17 non-pooled individuals.","project":"SRP027358"}
{"number_samples":11,"species":"human","abstract":"Exposure to ionizing radiation during childhood markedly increases the risk of developing papillary thyroid cancer. We identified non-overlapping somatic driver mutations in all 26 cases of post-Chernobyl thyroid cancers we studied through candidate gene assays and next generation RNA-sequencing. We found that 22/26 harbored fusion oncogenes arising primarily through intrachromosomal rearrangements. Altogether 23/26 of the oncogenic drivers identified in this cohort aberrantly activate MAPK signaling, including the two novel somatic rearrangements ETV6-NTRK3 and AGK-BRAF. Two other tumors harbored distinct fusions leading to overexpression of the nuclear receptor PPAR?. A lower prevalence of fusion oncogenes was found in a cohort of pediatric thyroid cancers from children from the same geographical regions that were not exposed to radiation. Radiation-induced thyroid cancers are a paradigm of tumorigenesis driven by fusion oncogenes that activate MAPK signaling or, less frequently, a PPAR?-driven transcriptional program. Overall design: Examination of transcriptome profiles and genetic somatic changes in thyroid cancer.","project":"SRP027364"}
{"number_samples":274,"species":"human","abstract":"We detected fusion genes in 274 fresh surgical samples of gliomas using whole transcriptome sequencing. Using this approach we screened a panel of glioma samples and identified a number of activating novel fusion transcripts. Overall design: Fusion detection in 274 glioma patients","project":"SRP027383"}
{"number_samples":2,"species":"human","abstract":"We developed a novel approach combining next generation sequencing, bioinformatics and mass spectrometry to assess the impact of non-MHC polymorphisms on the repertoire of MHC I-associated peptides (MIPs). We compared the genomic landscape of MIPs eluted from B lymphoblasts of two MHC-identical siblings and determined that MIPs mirror the genomic frequency of non-synonymous polymorphisms but they behave as recessive traits at the surface level. Moreover, we showed that 11.7% of the MIP coding exome is polymorphic at the population level. Our method provides fundamental insights into the relation between the genomic self and the immune self and accelerates the discovery of polymorphic MIPs (also known as minor histocompatibility antigens), which play a major role in allo-immune responses. Overall design: RNA-seq of human B lymphoblasts derived from peripheral blood mononuclear cells from 2 HLA-identical female siblings.","project":"SRP027508"}
{"number_samples":4,"species":"human","abstract":"Ribosome profiling and RNAseq data on human BJ fibroblasts and cybrid cells using an adapted ribosome profiling protocol to improve detection of mitochondrial ribosome protected fragments Overall design: 51-base length single read ribosome profiling data on human fibroblasts and cybrid cells using an adapted ribosome profiling protocol; and 51-base length single read RNAseq on polyA enriched RNA","project":"SRP027514"}
{"number_samples":20,"species":"human","abstract":"CONTEXT:  BRAF V600E mutation (BRAF-mut.) confers aggressiveness in papillary thyroid carcinoma, but unidentified genomic abnormalities may be required for full phenotypic expression.  OBJECTIVE:  To perform deep sequencing to identify genes differentially expressed between BRAF-mut. and BRAF-wild-type (BRAF-WT) tumors, and to compare to patient clinical status.  DESIGN:  BRAF-mut. and BRAF-WT tumors were identified in patients with T1N0 and with T23N1 tumors.  Expression levels of genes were determined from RNA sequencing (RNA-Seq) data and fusion transcripts were detected.  NanoString was used to validate the RNA-Seq data for immune genes.  SETTING:  Patients were seen at two sites of a major referral medical center.  PATIENTS:  Twenty patients were studied.  BRAF-mut. patients included 9 women, 3 men; 9 were TNM stage I and 3 were stage III; 3 (25%) had lymphocytic thyroiditis.  BRAF-WT included 5 women; 3 men; all were stage I; 5 (62.5%) had lymphocytic thyroiditis.  RESULTS:  560 of 13,085 genes were differentially expressed by RNA-Seq, and MetaCore analysis identified  55 immune function genes that were differentially expressed as a function of  BRAF mutational status.  Immune function genes were broadly underexpressed in BRAF-mut. tumors, with only 4 genes (HLA-G, CXCL14, TIMP1, IL1RAP) more highly expressed.  NanoString validated the RNA Seq data for immune genes.  Eleven high confidence fusion transcripts were detected, four being inter-chromosomal and seven intra-chromosomal.  CONCLUSION:  BRAF-mut. papillary thyroid cancers have less expression of immune and inflammatory response genes than BRAF-WT tumors.  Thirteen of 20 (65%) tumors had between one and three fusion transcripts.  Functional studies will be required to determine the potential role of the newly identified genomic abnormalities in contributing to the aggressiveness of BRAF-mut. and wild-type tumors. Overall design: RNA-seq was performed on 20 thyroid carcinoma tumors","project":"SRP027530"}
{"number_samples":42,"species":"human","abstract":"MicroRNAs (miRNAs) have been recently detected in the circulation of cancer patients, where they are associated with clinical parameters. Discovery profiling of circulating small RNAs has not been previously reported in breast cancer (BC), and was carried out in this study to identify blood-based small RNA markers of BC clinical outcome. Overall design: The pre-treatment sera of 42 stage II–III locally advanced and inflammatory BC patients who received neoadjuvant chemotherapy (NCT) followed by surgical tumor resection were analyzed for marker identification by deep sequencing all circulating small RNAs.","project":"SRP027589"}
{"number_samples":5,"species":"human","abstract":"Human induced pluripotent stem (hiPS) cells and human embryonic stem (hES) cells differentiate into cells of the endothelial lineage, but derivation of cells with human umbilical cord blood endothelial colony forming cell (ECFC)-like properties has not been reported. Here we describe a novel serum- and stromal cell-free ECFC differentiation protocol for the derivation of clinically relevant numbers of ECFCs (> 108) from hiPS and hES cells. We identified NRP-1+CD31+ selected cells that displayed a stable endothelial phenotype exhibiting high clonal proliferative potential, extensive replicative capacity, formation of human vessels that inosculated with host vasculature upon transplantation, but lacking in teratoma formation in vivo. We also identified NRP-1-VEGF165-KDR-mediated activation of KDR as a critical mechanism for the emergence and derivation of ECFCs from hiPS and hES cells.  This protocol advances the field by generating highly replicative but stable endothelial cells for use as a potential cell therapy for human clinical disorders. Overall design: Transcriptome sequencing of undifferentiated day 0 hiPS cells, Day 3 differentiated hiPS-derived mesoderm proginator cells, Day 12 hiPS-derived NRP-1+CD31+ cells, Day 12 H9-hES-derived NRP-1+CD31+ cells and cord blood-derived Endothelial colony forming cells.","project":"SRP028118"}
{"number_samples":73,"species":"human","abstract":"Determination of the genes regulated by ERRalpha nuclear receptor in MDA-MB231 cells Overall design: MDA-MB231 cells were inactivated for ERRalpha using siRNA. Three different siRNAs were used (siE1, siE2, siE3). Cells treated with a control siRNA (siC samples) were used for comparison. Duplicate samples were analyzed. Transcriptomic analysis was performed by RNA-Seq","project":"SRP028155"}
{"number_samples":48,"species":"human","abstract":"Global Run-On has been performed on WT or KD for RECQL5 cells after release from DRB.  When RECQL5 is knocked-down the transcriptional wave front is more advanced, suggesting that transcription is faster. Overall design: Constitutive knock-down cell lines expressing or not endogenous levels of shRNA resistant RECQL5 under a Doxycycline inducible promoter were treated with high doses of DRB to block transcription. Upon release into fresh medium we were able to follow how much and how fast the RNA Pol II progresses through genes by mapping nascent RNA by Run-On.  The experiment was performed in two cell line clones.","project":"SRP028170"}
{"number_samples":24,"species":"human","abstract":"Lung squamous cell carcinoma (SCC) is thought to arise from premalignant lesions in the airway epithelium, therefore studying these lesions is critical for understanding lung carcinogenesis. We performed RNA sequencing on laser-microdissected representative cell populations along the SCC pathological continuum of patient-matched normal basal cells, premalignant lesions, and tumor cells. We discovered transcriptomic changes and identified genomic pathways altered with initiation and progression of SCC within individual patients. We used immunofluorescent staining to confirm gene expression changes in premalignant lesions and tumor cells, including increased expression of SLC2A1, CEACAM5, and PTBP3 at the protein level and increased activation of MYC via nuclear translocation. Cytoband enrichment analysis revealed coordinated loss and gain of expression in chromosome 3p and 3q regions, respectively, during carcinogenesis. This is the first gene expression profiling of airway premalignant lesions with patient-matched samples that provides insight into the mechanisms of stepwise lung carcinogenesis. Overall design: Profiling of mRNA expression in laser-microdissected normal airway basal cells, premalignant airway lesions, and lung SCC tumor cells by massively parallel RNA sequencing.","project":"SRP028180"}
{"number_samples":12,"species":"human","abstract":"To study the fibroblast to myofibroblast differentiation in a normal or pathological situation, NHDF (Normal human dermal fibroblast) obtained from two different healthy donors (donors A and B) were either left untreated (T-E-) either treated with TGF-beta alone (T+E-), or with exudate from chronic wounds (T-E+) or both (T+E+). For each different treatment we performed mRNA deep sequencing 3 times : twice with cells from donor A and once with cells from donor B.We focused our study on gene expression profile as a representation of cell fate. We performed mRNA deep sequencing analysis of the 4 different conditions. For each condition, mRNA deep sequencing was performed 3 times : twice with cells from donor A and once with cells from donor B.Comparing the mRNA abundance between the different treatments, we identified 3 lists of genes characterizing the different gene expression states: 171 genes in List I representing the genes differentially expressed during normal fibroblast to myofibroblast differentiation, 409 genes in List II representing the genes differentially expressed upon exudate-only treatment and 1006 genes in List III representing the genes differentially expressed upon the combination of exudate and TGF-beta treatments. Overall design: 4 different treatments are compared : (T-E-) NHFD left untreated, (T+E-) NHDF treated with TGFbeta  for 4 days, (T-E+) NHDF treated with exudate for 4 days and (T+E+) NHDF treated with both TGFbeta and exudate. NHDF : Normal Human Dermal Fibroblast","project":"SRP028190"}
{"number_samples":10,"species":"human","abstract":"Androgen receptor (AR) is the major therapeutic target in aggressive prostate cancer. However, targeting AR alone can result in drug resistance and disease recurrence. Therefore, simultaneous targeting of multiple pathways could in principle be an effective new approach to treating prostate cancer. Here we provide proof-of-concept that a small molecule inhibitor of nuclear ß-catenin activity (called C3) can inhibit both the AR and ß-catenin signaling pathways that are often misregulated in prostate cancer. Treatment with C3 ablated prostate cancer cell growth by disruption of both ß-catenin/TCF and ß-catenin/AR protein interaction, reflecting the fact that TCF and AR have overlapping binding sites on ß-catenin. Given that AR interacts with, and is transcriptionally regulated by ß-catenin, C3 treatment also resulted in decreased occupancy of ß-catenin on the AR promoter and diminished AR and AR/ß-catenin target gene expression. Interestingly, C3 treatment resulted in decreased AR binding to target genes accompanied by decreased recruitment of an AR and ß-catenin cofactor, CARM1, providing new insight into the unrecognized function of ß-catenin in prostate cancer. Importantly, C3 inhibited tumor growth in an in vivo xenograft model, and blocked renewal of bicalutamide-resistant sphere forming cells, indicating the therapeutic potential of this approach. Overall design: Compare and contrast the expression profile of prostate cancer cells treated with a Wnt inhibitor (C3) with respect to ß-catenin and AR knockdown (all samples in duplicates).","project":"SRP028282"}
{"number_samples":78,"species":"human","abstract":"Micro-RNA sequencing of adrenocortical tumors and normal adrenal samples. Overall design: miRNA sequencing of  45 adrenocortical carcinomas (ACC), 30 adrenocortical adenomas (ACA) and 3 normal adrenal samples.","project":"SRP028291"}
{"number_samples":63,"species":"human","abstract":"Improved Smart-Seq for sensitive full-length transcriptome profiling in single cells. Overall design: Cells of four different origins were profiled using commercial SMARTer and compared to five variants of an improved protocol (Smart-Seq2).","project":"SRP028301"}
{"number_samples":1,"species":"human","abstract":"Research human liver cancer.","project":"SRP028316"}
{"number_samples":18,"species":"human","abstract":"N6-methyladenosine (m6A) is the most prevalent internal modification present in the mRNA of all higher eukaryotes. Here we present that m6A is selectively recognized by human YTH domain family (YTHDF2) protein to regulate mRNA degradation. By using crosslinking and immunoprecipitation, we have identified over 4000 substrate RNA of YTHDF2 with conserved core motif of G(m6A)C. We further estabilshed the role of YTHDF2 in RNA metabolism by a combination of ribosome profiling, RNA sequencing, m6A level quantification and cell-based imaging: the C-terminal domain of YTHDF2 selectively binds to m6A of mRNA and the N-terminal domain is responsive for localizing mRNA from translatable pool to processing body where mRNA decay occurs. Overall design: PAR-CLIP and RIP was used to identify YTHDF2 binding sites followed by ribosome profling and RNA seq to assess the consequences of YTHDF2 siRNA knock-down","project":"SRP028325"}
{"number_samples":60,"species":"human","abstract":"This dataset was generated with the goal of comparative study of gene expression in three brain regions and two non-neural tissues of humans, chimpanzees, macaque monkeys and mice. Using this dataset, we performed studies of gene expression and gene splicing evolution across species and search of tissue-specific gene expression and splicing patterns. We also used the gene expression information of genes encoding metabolic enzymes in this dataset to support a larger comparative study of metabolome evolution in the same set of tissues and species. Overall design: 120 tissue samples of prefrontal cortex (PFC), primary visual cortex (VC), cerebellar cortex (CBC), kidney and skeletal muscle of humans, chimpanzees, macaques and mice. The data accompanies a large set of metabolite measurements of the same tissue samples. Enzyme expression was used to validate metabolite measurement variation among species.","project":"SRP028336"}
{"number_samples":7,"species":"human","abstract":"EWS/ERG function in Ewing sarcoma","project":"SRP028340"}
{"number_samples":7,"species":"human","abstract":"Gene regulation by EWS/ERG in Ewing sarcoma","project":"SRP028344"}
{"number_samples":7,"species":"human","abstract":"The effect on gene regulation in Ewing sarcoma cells treated with LSD1 inhibitor HCI-2509","project":"SRP028346"}
{"number_samples":12,"species":"human","abstract":"miRNA-Sequencing was performed on human aortic valve interestitial cells (AVICs) exposed to 14% stretch at 1 hz or static conditions for 24h. Overall design: Six static control and six samples exposed to cyclic stretch 14% for 24h","project":"SRP028526"}
{"number_samples":39,"species":"human","abstract":"Ribosome pofile","project":"SRP028530"}
{"number_samples":12,"species":"human","abstract":"RNA sequencing of human leukemia Overall design: The goals of this project are to obtain a comprehensive study of mutations and gene expression in human T ALL (acute lymphoblastic leukemia). Methods: T ALL cells were thawed. DNA and RNA (polyA) was extracted and sequences were obtained with an Illumina HiSeq 2000 sequencer.","project":"SRP028554"}
{"number_samples":32,"species":"human","abstract":"We performed whole-genome stability measurements for MDA-MB-231 and its highly metastatic derivative MDA-LM2. Our goal was to identify post-transcriptonal regulons that are deregulated en route to higher metastatic capacity. Overall design: Cells were pulsed with 4-thiouridine for 2 hours and then RNA was extracted at 0, 2, 4, and 7 hr time-points in quadruplicate from each cell line. 4sU labeling followed by RNA-seq was then used to measure the abundance of transcripts in each population. A decay rate was estimated based on the rate at which transcript abundance was reduced at these time-points.","project":"SRP028570"}
{"number_samples":54,"species":"human","abstract":"RNA sequencing of human leukemia Overall design: The goals of this project are to obtain a comprehensive study of mutations and gene expression in human acute myeloid leukemia (AML). Methods: AML cells were thawed. DNA and RNA (polyA) was extracted and sequences were obtained with an Illumina HiSeq 2000 sequencer.","project":"SRP028594"}
{"number_samples":12,"species":"human","abstract":"Variation in gene regulation is thought to have played an important role in the evolution of primates, and many studies have documented differences in mRNA expression levels across primate species. However, it is not yet known to what extent measurements of divergence in mRNA levels reflect divergence in protein expression levels, which are more directly tied to phenotypic differences. To address this question, we used high-resolution, quantitative mass spectrometry to collect thousands of protein expression measurements from human, chimpanzee, and rhesus macaque lymphoblastoid cell lines (LCLs). We also used RNA sequencing to collect transcript expression data from the same samples. Considering the two datasets jointly we found that there is much more inter-species divergence at the mRNA level than at the protein level. Remarkably, we found dozens of genes with significant expression differences between species at the mRNA level yet little or no difference in protein expression. Overall, our data suggest a much stronger evolutionary constraint on protein expression levels than on mRNA levels. We conclude that inter-species mRNA expression differences may often have limited functional consequences due to either buffering or compensatory changes in post-transcriptional or posttranslational regulation of proteins. Overall design: Gene expression patterns were compared between human, chimpanzee, and rhesus macaque lymphoblastoid cell lines (5 individuals from each species) using RNA-Seq. These gene expression patterns were also compared to protein expression patterns based on mass-spec data, which are available separately (see publication for details).","project":"SRP028612"}
{"number_samples":10,"species":"human","abstract":"A large number of computational methods have been recently developed for analyzing differential gene expression (DE) in RNA-seq data. We report on a comprehensive evaluation of the commonly used DE methods using the SEQC benchmark data set and data from ENCODE project. We evaluated a number of key features including: normalization, accuracy of DE detection and DE analysis when one condition has no detectable expression. We found significant differences among the methods. Furthermore, computational methods designed for DE detection from expression array data perform comparably to methods customized for RNA-seq. Most importantly, our results demonstrate that increasing the number of replicate samples significantly improves detection power over increased sequencing depth. Overall design: The Sequencing Quality Control Consortium generated two datasets from two reference RNA samples in order to evaluate transcriptome profiling by next-generation sequencing technology. Each sample contains one of the reference RNA source and a set of synthetic RNAs from the External RNA Control Consortium (ERCC) at known concentrations. Group A contains 5 replicates of the Strategene Universal Human Reference RNA (UHRR), which is composed of total RNA from 10 human cell lines, with 2% by volume of ERCC mix 1. Group B includes 5 replicate samples of the Ambion Human Brain Reference RNA (HBRR) with 2% by volume of ERCC mix 2. The ERCC spike-in control is a mixture of 92 synthetic polyadenylated oligonucleotides of 250-2000 nucleotides long that are meant to resemble human transcripts.","project":"SRP028705"}
{"number_samples":2,"species":"human","abstract":"The role of long noncoding RNA (lncRNA) in adult hearts is unknown; also unclear is how lncRNA modulates nucleosome remodelling. An estimated 70% of mouse genes undergo antisense transcription1, including myosin heavy chain 7 (Myh7), which encodes molecular motor proteins for heart contraction2. Here we identify a cluster of lncRNA transcripts from Myh7 loci and demonstrate a new lncRNA–chromatin mechanism for heart failure. In mice, these transcripts, which we named myosin heavy-chain-associated RNA transcripts (Mhrt), are cardiac-specific and abundant in adult hearts. Pathological stress activates the Brg1–Hdac–Parp chromatin repressor complex3 to inhibit Mhrt transcription in the heart. Such stress-induced Mhrt repression is essential for cardiomyopathy to develop: restoring Mhrt to the pre-stress level protects the heart from hypertrophy and failure. Mhrt antagonizes the function of Brg1, a chromatin-remodelling factor that is activated by stress to trigger aberrant gene expression and cardiac myopathy3. Mhrt prevents Brg1 from recognizing its genomic DNA targets, thus inhibiting chromatin targeting and gene regulation by Brg1. It does so by binding to the helicase domain of Brg1, a domain that is crucial for tethering Brg1 to chromatinized DNA targets. Brg1 helicase has dual nucleic-acid-binding specificities: it is capable of binding lncRNA (Mhrt) and chromatinized—but not naked—DNA. This dual-binding feature of helicase enables a competitive inhibition mechanism by which Mhrt sequesters Brg1 from its genomic DNA targets to prevent chromatin remodelling. A Mhrt–Brg1 feedback circuit is thus crucial for heart function. Human MHRT also originates from MYH7 loci and is repressed in various types of myopathic hearts, suggesting a conserved lncRNA mechanism in human cardiomyopathy. Our studies identify a cardioprotective lncRNA, define a new targeting mechanism for ATP-dependent chromatin-remodelling factors, and establish a new paradigm for lncRNA–chromatin interaction. Overall design: Overexpression of murine non-coding RNAs in human cell line, comparison of RNA-sequencing with Ribosome Profiling","project":"SRP028720"}
{"number_samples":16,"species":"human","abstract":"MiRNAs are important negative regulators of protein coding gene expression, and have been studied intensively over the last few years. Several measurement platforms, designed to determine their relative RNA abundance levels in biological samples, have been developed. In this study, we systematically compared 12 commercially available miRNA expression platforms by measuring an identical set of 20 standardized positive and negative control samples, including human universal reference RNA, human brain RNA and titrations thereof, human serum samples, and synthetic spikes from miRNA family members with varying homology. We developed novel and robust quality metrics to objectively assess platform performance of very different technologies such as small RNA sequencing, RT-qPCR and (microarray) hybridization. We assessed reproducibility, sensitivity, accuracy, specificity, and concordance of differential expression. The results indicate that each method has its strengths and weaknesses, which helps to guide informed selection of a quantitative miRNA gene expression platform in function of particular study goals. Overall design: Sequencing of 20 miRQC samples on Illumina Genome Analyzer IIx System","project":"SRP028738"}
{"number_samples":3,"species":"human","abstract":"Our data provided a genome-wide DNA methylation landscape of human early development embryos, including human MII oocytes, sperm, zygotes, 2-cell to 8-cell embryos, morula, blastocyst and postimplantation embryos at single base resolution. Overall design: In total, 44 samples including biological and technical replicates, from 12 different human embryo development stages were analyzed, including two metaphase II oocytes, two zygotes, three first polar bodies, two second polar bodies, four sperm samples, two 2-cell-stage embryos, two 4-cell-stage embryos, three 8-cell-stage embryos, three morulae, three inner cell masses (ICMs) and three trophectoderms (TEs) seperated from late blastocysts, and three post-implantation embryos. In addition, 12 different human embryo development stages were analyzed, including metaphase II oocytes, zygotes, first polar bodies, second polar bodies, sperm samples, 2-cell-stage embryos, 4-cell-stage embryos, 8-cell-stage embryos, morulae, inner cell masses (ICMs) and trophectoderms (TEs) seperated from late blastocysts, and post-implantation embryos.","project":"SRP028804"}
{"number_samples":5,"species":"human","abstract":"We report that full length TET1 (TET1-FL) overexpression fails to induce global DNA demethylation in HEK293T cells. The preferential binding of TET1-FL to hypomethylated CpG islands (CGIs) through its CXXC domain leads to its inhibited 5-hydroxymethylcytosine (5hmC) production as methylation level increases. TET1-FL-induced 5hmC accumulates at CGI edges, while TET1 knockdown induces methylation spreading from methylated edges into hypomethylated CGIs. However, TET1 can regulate gene transcription independent of its dioxygenase catalytic function. Thus, our results identify TET1 as a maintenance DNA demethylase that does not purposely decrease methylation levels, but specifically maintains the DNA hypomethylation state of CGIs in adult cells. Overall design: Genome-wdie profiling of gene expression in HEK293T cells following overexpression of wild type or catalytically mutant TET1-FL or TET1-CD","project":"SRP028814"}
{"number_samples":2,"species":"human","abstract":"Sequencing of 5' ends of RNA molecules from control and exosome-depleted HeLa-S3 cells. Overall design: CAGE library construction from RNA extracted from control and exosome-depleted cells.","project":"SRP028815"}
{"number_samples":5,"species":"human","abstract":"The second trimester fetal transcriptome can be assessed based on cell-free RNA found within the amniotic fluid supernatant. The objective of this study was to compare the suitability of two technologies for profiling the human fetal transcriptome: RNA-Seq and expression microarray. Comparisons were based on total numbers of gene detected, rank-order gene expression, and functional genomic analysis. Fewer gene transcripts were observed using RNA-Seq than microarray (4,158 vs 8,842). Correlation of total expression within each sample ranged from R=0.43 to R=0.57. On average, there was 59% concordance in gene identity among the top 10% of genes ranked by expression. The RNA-Seq data yielded more significant pathways enrichment within the ?Physiological Systems Development and Function? categories of IPA. Alternative splicing of many well-known genes, including those previously studied in fetal development, such as H19 and IGF2  is detected by RNA-Seq. Also included in this paper is discussion of the technical challenges inherent to working with cell-free fetal RNA and possible solutions. Overall design: Cell-free fetal RNA from the amniotic fluid supernatant of five second trimester fetuses was divided and prepared in tandem for analysis using either the Illumina HiSeq 2000 or Affymetrix HG-U133 Plus 2.0 GeneChip microarray.","project":"SRP028822"}
{"number_samples":27,"species":"human","abstract":"Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We have developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing (Baltz and Munschauer et al. 2012). Our current work focuses on streamlining and extending protein occupancy profiling on poly(A)-RNA. Our objectives are to identify previously unknown protein-bound transcripts and, more importantly, to assess global and local differences in protein occupancy across different biological conditions. To this end, we have implemented poppi, the first pipeline for differential analysis of protein occupancy profiles. We have applied our analysis pipeline to pinpoint changes in occupancy profiles of MCF7 cells against already published HEK293 cells [GSE38157]. Overall design: We generated protein occupancy cDNA libraries for two biological replicates. Briefly, we crosslinked 4SU-labeled MCF7 cells and purified protein-mRNA complexes using oligo(dT)-beads. The precipitate was treated with RNAse I to reduce the protein-crosslinked RNA fragments to a length of about 30-60 nt. To remove non-crosslinked RNA, protein-RNA complexes were precipitated with ammonium sulfate and blotted onto nitrocellulose. The RNA was recovered by Proteinase K treatment, ligated to cloning adapters, and reverse transcribed. The resulting cDNA libraries were PCR-amplified and next-generation sequenced.","project":"SRP028887"}
{"number_samples":6,"species":"human","abstract":"We sequenced the total mRNA and translating mRNA (RNC-mRNA) of three hepatocellular carcinoma cell lines Hep3B, HCCLM3 and MHCC97H Overall design: For each cell line, samples prepared from three independent and identical cell cultures were pooled with equal amounts. C-HPP China Team","project":"SRP028902"}
{"number_samples":18,"species":"human","abstract":"mRNA synthesis, processing, and destruction involve a complex series of molecular steps that are incompletely understood.  Because the RNA intermediates in each of these steps have finite lifetimes, extensive mechanistic and dynamical information is encoded in total cellular RNA.  Here we report the development of SnapShot-Seq, a set of computational methods that allow the determination of in vivo rates of pre-mRNA synthesis, splicing, intron degradation, and mRNA decay from a single RNA-Seq snapshot of total cellular RNA.  SnapShot-Seq can detect in vivo changes in the rates of specific steps of splicing, and it provides genome-wide estimates of pre-mRNA synthesis rates comparable to those obtained via labeling of newly synthesized RNA.  We used SnapShot-Seq to investigate the origins of the intrinsic bimodality of metazoan gene expression levels, and our results suggest that this bimodality is partly due to spillover of transcriptional activation from highly expressed genes to their poorly expressed neighbors.  SnapShot-Seq dramatically expands the information obtainable from a standard RNA-Seq experiment. Overall design: These data are total RNA-Seq data, from RNA sequencing of rRNA-depleted total cellular RNA -- except the LCL 4SU data, which derive from 4SU-labeled RNA.  Please see the associated paper for more details.","project":"SRP028912"}
{"number_samples":6,"species":"human","abstract":"RNA-seq data from human colon cancer tumors and xenografts in mice","project":"SRP028952"}
{"number_samples":18,"species":"human","abstract":"Small RNA-seq on MCF10A, HCT116 and HCT116p53-/- cell lines after induction of DNA damage (5 Gy Irradiation). Overall design: Small RNA-seq on MCF10A, HCT116 and HCT116p53-/- at 4 and 24 hours after induction of DNA damage (5 Gy Irradiation), done in duplicate with respective control (0 hour) using illumina Genome Analyzer IIx","project":"SRP028963"}
{"number_samples":5,"species":"human","abstract":"Purpose:  The goal of this study was to identify differentially expressed genes and pathways between the cumulus of compact/unstimulated cumulus-oocyte-complex  (COC) and the cumulus of expanded/stimulated COC. Methods: mRNA profiles of  Compact/unstimulated cumulus cells (CCs) from germinal vesicle (GV) COC obtained from two patients undergoing unstimulated IVM procedure and  expanded/stimulated CCs from metaphase 2 (MII) COC obtained from three patients undergoing IVF/ICSI were generated by deep sequencing using Illumina HiSeq 2000. The sequence reads that passed quality filters were mapped to the human genome (hg19) using Tophat software. Differential expression analysis was done using DESeq bioconductor package. qRT–PCR validation was performed using SYBR Green assays. Results: A total of 40-80 million sequence reads per sample were mapped to the human genome (hg19). A total of 1746 differentially expressed genes between compact and expanded CCs with fold change > 2, and adjusted p value < 0.05 were identified. Gene ontology analysis of differentially expressed genes revealed a number of cellular processes regulated during the periovulatory interval including cellular movement, inflammatory response, immune cell trafficking, tissue development, lipid metabolism,  tissue morphology, DNA replication and cell cycle.  A total of 116 of the differentially expressed genes were annotated as long non-coding RNAs, 10 of them coded from introns of genes known to be involved in granulosa cell processes suggesting that unique non coding RNA transcripts may contribute to the regulation of cumulus expansion and oocyte maturation. Results were validated using qRT-PCR. Conclusions: Using global transcriptome sequencing we identified new important genes and non coding RNAs involved in COC maturation and cumulus expansion, which may contribute to improve the process of in vitro maturation of immature oocytes utilized in IVM cycles Overall design: mRNA profiles of  Compact/unstimulated cumulus cells (CCs) from germinal vesicle (GV) COC obtained from two patients undergoing unstimulated IVM procedure and  expanded/stimulated CCs from metaphase 2 (MII) COC obtained from three patients undergoing IVF/ICSI were  generated by deep sequencing using Illumina HiSeq 2000.","project":"SRP029207"}
{"number_samples":4,"species":"human","abstract":"Polycomb repressive complex-2 (PRC2) is a histone methyltransferase required for epigenetic silencing during development and cancer. Long non-coding RNAs (lncRNAs) recruit PRC2 to chromatin, but the general role of RNA in maintaining repressed chromatin is unknown. ChIP-seq, combined with RNA-seq, indicating that PRC2 is also associated with active genes, but most of these are not regulated by PRC2. These results were complemented by in vitro binding assays measuring the binding constant of human PRC2 to various RNAs and find comparable affinity for human lncRNAs targeted by PRC2 and irrelevant transcripts from ciliates and bacteria. PRC2 binding is size-dependent, with lower affinity for shorter RNAs. These findings support a model in which promiscuous binding of PRC2 to RNA transcripts allows it to scan for target genes that have escaped repression, leading to maintenance of the repressed state. Such RNAs may also provide a decoy for PRC2. Overall design: Cell culture: HEK293T/17 cells were cultured in DMEM with 10% FBS for no longer than 15 passages. ON-TARGETplus SMARTpool for human SUZ12 (Thermo Scientific, Dharmacon cat # L-006957-00-0005) was used to knockdown SUZ12 (siSUZ12) and ON-TARGETplus Non-targeting Pool (Dharmacon cat # D-001810-10-05) was used as a negative control (siCtrl). A total of 25 nM siRNA was transfected in 6-well dishes using Lipofectamine™ RNAiMAX Reagent (Life Technologies, Invitrogen) following the manufacturer’s recommendations. RNA-seq: Polyadenylated RNA was purified from 4 ug of RNA. cDNA libraries were prepared and double-stranded cDNA was fragmented using DNase I according to Illumina specifications, prior to adaptor ligation. Sequencing libraries were amplified and sequenced using an Illumina HiSeq 2000 sequencer. ChIP-seq: 30 million cells at 80% to 90% confluent culture were crosslinked in 1% v/v formaldehyde for 10 min, quenched in 150 mM glycine, washed with cold 1xPBS and harvested by scraping. Cells were lysed in Lysis Buffer (1% w/v SDS, 10 mM EDTA, 50 mM Tris-HCl pH 8.1) with 1x Complete® protease inhibitors (Roche). Cells were sonicated for 10 to 15 min using a Bioruptor™ UCD-200 (Diagenode) with 30 sec pulses at maximum power. Lysates were diluted to 10 ml in IP Buffer (0.01% w/v SDS, 1.1% v/v Triton-X, 1.2 mM EDTA, 16.7 mM Tris-HCL pH 8.0, 167 mM NaCl) and 10 to 20 ng of antibodies were added and incubated overnight at 4 °C with rotation. Antibodies were immunoprecipitated with 60 µl protein G Plus/protein A Agarose Suspension (Calbiochem cat # IP05) and washed sequentially with 1 ml Low Salt Wash Buffer (0.1% w/v SDS, 1% v/v Triton X-100, 2 mM EDTA, 20 mM Tris-HCl pH 8.0, 150 mM NaCl), 1 ml High Salt Wash Buffer (0.1% w/v SDS, 1% v/v Triton X-100, 2 mM EDTA, 20 mM Tris-HCl pH 8.0, 500 mM NaCl), 1 ml LiCl Wash Buffer (0.25 M LiCl, 1% v/v Nonidet P-40, 1% w/v deoxycholate, 1 mM EDTA, 10 mM Tris-HCl pH 8.0) and 1 ml TE buffer (Qiagen). DNA was eluted with 0.4 ml of 0.1 M NaHCO3 and 1% w/v SDS. Eluent was transferred into a new tube and NaCl was added to a final concentration of 200 mM. Crosslinks were reversed by incubation at 65 °C for 2 hours. Next, proteins and RNA were digested by adding 33 µl of Digestion Reagent (0.6 M Tris-HCl pH 6.5, 152 mM EDTA, 61 ng/ml RNase A (Invitrogen cat # AM2274) and 0.61 mg/ml Proteinase K (NEB cat # P8102)) following by 1 hour incubation at 37 °C. DNA was extracted by phenol:chloroform and ethanol precipitated. DNA was resuspended in Milli-Q pure water and concentration was measured using Qubit™ (Invitrogen). At least 10 ng of recovered DNA was used to synthesize sequencing libraries using the ChIP-seq Sample Preparation kit (Illumina). Between 6 and 10 pmoles were used for sequencing on the HiSeq2000 sequencer.","project":"SRP029245"}
{"number_samples":89,"species":"human","abstract":"Here we harnessed the potential of RNA sequencing in 89 human pancreatic islet donors to identify genes and exons regulated in this relevant tissue for T2D. Overall design: mRNA profiles of 89 human pancreatic islet donors having different levels of blood glucose (HbA1c) with and without T2D. The data was generated by deep sequencing using Illumina HiSeq 2000.","project":"SRP029262"}
{"number_samples":8,"species":"human","abstract":"Insulin-secreting ß cells and glucagon-secreting a cells maintain physiological blood glucose levels, and their malfunction drives diabetes development. Using ChIP sequencing and RNA sequencing analysis, we determined the epigenetic and transcriptional landscape of human pancreatic a, ß, and exocrine cells. We found that, compared with exocrine and ß cells, differentiated a cells exhibited many more genes bivalently marked by the activating H3K4me3 and repressing H3K27me3 histone modifications. This was particularly true for ß cell signature genes involved in transcriptional regulation. Remarkably, thousands of these genes were in a monovalent state in ß cells, carrying only the activating or repressing mark. Our epigenomic findings suggested that a to ß cell reprogramming could be promoted by manipulating the histone methylation signature of human pancreatic islets. Indeed, we show that treatment of cultured pancreatic islets with a histone methyltransferase inhibitor leads to colocalization of both glucagon and insulin and glucagon and insulin promoter factor 1 (PDX1) in human islets and colocalization of both glucagon and insulin in mouse islets. Thus, mammalian pancreatic islet cells display cell-type–specific epigenomic plasticity, suggesting that epigenomic manipulation could provide a path to cell reprogramming and novel cell replacement-based therapies for diabetes. Overall design: Pancreatic islets were collected post-mortem from 6 human donors and subjected to FACS to separate populations of alpha, beta, and exocrine cells. Depending on the availability of resulting material, sorted islet cell populations were used for H3K4me3, H3K27me3 ChIP-seq, or RNA-seq analysis. All ChIP-seq samples have a corresponding input from the same sample.","project":"SRP029281"}
{"number_samples":5,"species":"human","abstract":"Background: RNA-seq is a powerful technique for identifying and quantifying transcription and splicing events, both known and novel. However, given its recent development and the proliferation of library construction methods, understanding the bias it introduces is incomplete but critical to realizing its value. Results: We present a method, in vitro transcription sequencing (IVT-seq), for identifying and assessing the technical biases in RNA-seq library generation and sequencing at scale. We created a pool of over 1,000 in vitro transcribed RNAs from a full-length human cDNA library and sequenced them with polyA and total RNA-seq, the most common protocols. Because each cDNA is full length, and we show in vitro transcription is incredibly processive, each base in each transcript should be equivalently represented. However, with common RNA-seq applications and platforms, we find 50% of transcripts have more than two-fold and 10% have more than 10-fold differences in within-transcript sequence coverage. We also find greater than 6% of transcripts have regions of dramatically unpredictable sequencing coverage between samples, confounding accurate determination of their expression. We use a combination of experimental and computational approaches to show rRNA depletion is responsible for the most significant variability in coverage, and several sequence determinants also strongly influence representation. Conclusions: These results show the utility of IVT-seq for promoting better understanding of bias introduced by RNA-seq. We find rRNA depletion is responsible for substantial, unappreciated biases in coverage introduced during library preparation. These biases suggest exon-level expression analysis may be inadvisable, and we recommend caution when interpreting RNA-seq results. Overall design: 5 rRNA-depleted samples with duplicates, 1 polyA selected, 1 total RNA, and 1 plasmid library all without replicates.","project":"SRP029334"}
{"number_samples":154,"species":"human","project":"SRP029341"}
{"number_samples":4,"species":"human","abstract":"To identify genes regulated by BRD4 and to provide insight into new mechanisms de-regulated by BRD4, such as the response to oxidative stress, we integrated BRD4-binding regions with BRD4 gene expression data. For this analysis we performed BRD4 chromatin immunoprecipitation  experiments and BRD4 knock down experiments followed by RNA-Seq analyses. By integration of both gene lists we identified top candidate genes regulated by BRD4. Overall design: HEK cells have been investigated for genomewide BRD4 binding sites and expression changes after knock down of BRD4. Illumina sequencing was used to gather data of the type ChIP Seq and mRNA Seq.","project":"SRP029365"}
{"number_samples":16,"species":"human","abstract":"The fragile X mental retardation protein FMRP is an RNA binding protein that regulates translation of its bound mRNAs through incompletely defined mechanisms. FMRP has been linked to the microRNA pathway and we show here that it is associated with MOV10, a putative helicase that is also associated with the microRNA pathway.  We show that FMRP associates with MOV10 in an RNA-dependent manner and facilitates MOV10-association with RNAs in brain. We identified the RNA sequences recognized by MOV10 using iCLIP and found an increased number of G-quadruplexes in the CLIP sites. We provide evidence that MOV10 facilitates microRNA-mediated translation regulation and also has the novel role of increasing the expression of a subset of RNAs by sterically hindering Argonaute2 association. In summary, we have identified a new mechanism for FMRP-mediated translational regulation through its association with MOV10. Overall design: Comparison of MOV10 siRNA knockdown, irrelevant siRNA control and MOV10 overexpression on total RNA levels","project":"SRP029367"}
{"number_samples":4,"species":"human","abstract":"To identify the miRNAs that are differentially expressed and secreted between the MDA-MB-231 metastatic breast cancer cells and the MCF-10A non-cancerous human mammary epithelial cells, we profiled the cellular and exosomal small RNAs (between 17 and 52 nt) isolated from these two cell lines by Solexa deep sequencing.  MiRNAs that are significantly different between the two cell lines are identified. Overall design: RNA was extracted from cultured MDA-MB-231 and MCF-10A cells or purified exosomes secreted by these cells, and subjected to library construction and Solexa deep sequencing.","project":"SRP029401"}
{"number_samples":9,"species":"human","abstract":"Using a chromatin regulator-focused shRNA library, we found that suppression of sex determining region Y-box 10 (SOX10) in melanoma causes  resistance to BRAF and MEK inhibitors. To investigate how SOX10 loss leads to drug resistance, we performed transcriptome sequencing (RNAseq) of both parental A375 (Ctrl. PLKO) and A375-SOX10KD (shSOX10-1, shSOX10-2) cells. To ask directly whether SOX10 is involved indrug resistance in BRAF(V600E) melanoma patients, we isolated RNA from paired biopsies from melanoma patients (pre- and post- treatment) , that had gained  BRAF or MEK inhibitor resistance . We performed RNAseq analysis to determine changes in transcriptome upon drug resistance. Overall design: Investigate genes regulated by SOX10 and differntial gene expression between pre- and post-treatment biopsies. We use short hairpin RNA to suppression SOX10 in A375 cells and cells were harvested with trizol reagent for RNA isolation. For paired biopsies (patient samples) we collected the first biopsy before the initiation of treatment and the second biopsy after drug resistance developed. RNA was isolated from FFPE samples and subjected for RNA sequencing.","project":"SRP029434"}
{"number_samples":8,"species":"human","abstract":"The transcription factor FOXP1 is implicated in the pathogenesis of B-cell lymphomas through immunoglobulin heavy chain (IGH) locus-related chromosomal translocations leading to dysregulated expression of FOXP1. Translocations of FOXP1 with non-IG gene sequences have been also reported, but the molecular consequences of such aberrations remain undetermined. Here, using molecular cytogenetics and molecular biology studies, we comprehensively analyzed four lymphoma cases with non-IG rearrangements of FOXP1 and compared these with cases harboring t(3;14)(p13;q32)/IGH-FOXP1 and FOXP1-expressing lymphomas without underlying t(3p13/FOXP1). We found that non-IG rearrangements are usually acquired during evolution of lymphoma and constantly target the coding region of FOXP1, promiscuously fusing with coding and non-coding gene sequences at various reciprocal breakpoints (2q36, 10q24 and 3q11). Intriguingly, these rearrangements do not generate functional chimeric genes but commonly disrupt the full-length FOXP1 transcript leading to an aberrant expression of N-truncated FOXP1 isoforms, as shown by QRT-PCR and Western blot analysis. In contrast, cases with t(3;14)(p13;q32)/IGH-FOXP1 overexpress the full-length FOXP1. Collectively, our findings point to a dual mechanism through which FOXP1 is implicated in B-cell lymphomagenesis. The primary t(3;14)(p13;q32)/IGH-FOXP1 produces the full-length protein with potent oncogenic activity, whereas the secondary non-IG 17 rearrangements of FOXP1 generate N-truncated FOXP1 isoforms, likely driving progression of disease. Overall design: Using molecular cytogenetics and molecular biology studies (including RNA-seq), we comprehensively analyzed four lymphoma cases with non-IG rearrangements of FOXP1 and compared these with cases harboring t(3;14)(p13;q32)/IGH-FOXP1 and FOXP1-expressing lymphomas without underlying t(3p13/FOXP1).","project":"SRP029452"}
{"number_samples":4,"species":"human","abstract":"label the cells overexpressed Myc tagged METTL3 and Flag tagged WTAP with 4-SU, the RNA bound by METT3,WTAP can be got by Myc or Flag IP followed by RNA isolation by using the TRIzol (Invitrogen) reagent by following the company manual.the RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 (Illumina, San Diego, CA, USA) according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 (Illumina) in single-read mode, creating reads with a length of 50 bp. Sequencing chemistry v2 (Illumina) was used and samples were multiplexed in two samples per lane. Overall design: discovery of the binding motif of METTL3,WTAP in METTL3,WTAP overexpressed Human 293T cells","project":"SRP029513"}
{"number_samples":6,"species":"human","abstract":"RNA was isolated from and METTL3,WTAP deficient Human HeLa cells using the TRIzol (Invitrogen) reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100. All samples showed a RIN (RNA integrity number) of higher than 9. Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 (Illumina, San Diego, CA, USA) according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 (Illumina) in single-read mode, creating reads with a length of 50 bp. Sequencing chemistry v2 (Illumina) was used and samples were multiplexed in two samples per lane. Overall design: Examination of gene expressive levels in normal and METTL3,WTAP deficient Human HeLa cells","project":"SRP029515"}
{"number_samples":12,"species":"human","abstract":"This study sought to characterize changes in mRNA translation during cell cycle progression. Understanding how translation is regulated during each phase of the cell cycle will provide a greater understanding of normal cell growth and division as well as guide the development of potential cancer therapies.","project":"SRP029589"}
{"number_samples":10,"species":"human","abstract":"Angioimmunoblastic T cell lymphoma","project":"SRP029591"}
{"number_samples":10,"species":"human","abstract":"Purpose: To chart the human myometrial transcriptomes before and after the onset of labour. Methods: Tophat splice junction mapping of paired-end reads, HTSeq to generate counts, cufflinks to track transcripts, DESeq, edgeR and baySeq to detect differentially expressed genes and principal component analysis for clustering analyses. Results: We mapped on average 14 million paired-end reads per sample (counting each end individually) to the human genome (build hg19) and covered the expressed transcriptome about 13 times with a TopHat-HTSeq workflow. We performed a comparative analysis with an analogous microarray study (Mittal et al., 2010) and found some overlap between the RNA-seq and the microarray data. Conclusions: Our study is the first RNA-seq study of the human myometrium before and after the onset of labour. We show that while microarray and RNA-seq studies may complement each other, RNA-seq has a much greater resolution. Overall design: At term with and at term without labour human myometrial mRNA profiles were generated by deep sequencing, using Illumina GAIIx (five biological replicates each).","project":"SRP029592"}
{"number_samples":8,"species":"human","abstract":"We report the gene expression profile of 8 metastatic castration resisistant prostate cancer samples analyzed by paired-end RNA-seq. We found evidence of extensive abnormal splicing as well as several novel fusion genes. Finally, we also observed several recurrent high-confidence somatic mutations. Overall design: Paired-end RNA-seq by rRNA depletion","project":"SRP029603"}
{"number_samples":18,"species":"human","abstract":"In parallel to the genetic code for protein synthesis, a second layer of information is embedded in all RNA transcripts in the form of RNA structure. The ability of RNA to base pair with itself and other nucleic acids endow RNA with the capacity to form extensive structures, which are known to influence practically every step in the gene expression program1. Yet the nature of most RNA structures or effects of sequence variation on structure are not known.  Here we report the initial landscape and variation of RNA secondary structures (RSS) in a human family trio, providing a comprehensive RSS map of human coding and noncoding RNAs. We identify unique RSS signatures that demarcate open reading frames, splicing junctions, and define authentic microRNA binding sites.  Comparison of native deproteinized RNA isolated from cells versus refolded purified RNA suggests that the majority of the RSS information is encoded within RNA sequence. Over one thousand transcribed single nucleotide variants (~15% of all transcribed SNVs) alter local RNA structure; these “RiboSNitches”2 occur in disease-associated variants.  We discover simple sequence and spacing rules that determine the ability of point mutations to impact RSS. Selective depletion of RiboSNitches versus structurally synonymous variants at precise locations suggests selection for specific RNA shapes at thousands of sites, including 3’UTRs, binding sites of miRNAs and RNA binding proteins genome-wide. These results highlight the potentially broad contribution of RNA structure and its variation to gene regulation. Overall design: RNA structure probing is performed at 37°C on poly(A)+ selected RNAs from GM12878, GM12891 and GM12892 cell lines, as well as on native proteinized RNAs from GM12878.  The structure probed RNAs is then cloned into a sequencing library using modied Ambion RNA sequencing kit compatible with the Illumina platform.  The samples were deep sequenced using Illumina's Hi-Seq platform. AGO CLIP was performed as reported. Cells were crosslinked with UV and lysed using published protocols. AGO2 was enriched using immunopurification. The RNA-protein complex was digested with ribonuclease and purified by gel electrophoresis. Purified RNA was reverse transcribed and cDNA molecules were amplified and sequenced as described.","project":"SRP029656"}
{"number_samples":18,"species":"human","abstract":"The goal of this study is to identify the transcriptome differences between the two major subtypes of diffuse large B cell lymphoma (DLBCL).  DLBCL is the most common form of non-Hodgkin’s lymphoma and has two major subtypes: germinal center B-cell-like (GCB) and activated B-cell-like (ABC).  When compared to the GCB form, ABC lymphomas respond much more poorly to current therapies. To investigate how gene expression changes might contribute to this aggressive phenotype, we have used RNA-Seq to profile the whole transcriptome in 8 DLBCL cell lines (4 GCB subtype, 4 ABC) that are derived from patient tumors.  1,545 genes are differentially expressed between subtypes (FDR < 0.05), approximately 7% of the transcriptome.  The vast majority of these genes (81%, n = 1251) are more highly expressed in the ABC cell lines.  In contrast, only 294 genes (19%) are more highly expressed in the GCB cell lines.  Half (n = 765) of the genes with greater ABC subtype expression demonstrate very low read counts (< 5) in the GCB cell types.  Conversely, only 21 genes that are more highly expressed in GCB are unique to that subtype.  The prevalence of such “on/off” genes indicates that the major differences between ABC and GCB DLBCL are due almost exclusively to additional gene expression in ABC, rather than the two subtypes having divergent but equally active genetic programs. Overall design: Measurement and comparison of gene expression in 8 cell lines representing the 2 subtypes of DLBCL.  4 cell lines are subtyped as ABC and 4 are subtyped as GCB.  2 replicates are present for each cell line.  (Cell line OCI-Ly19 was not included in the analysis because its gene expression clustered in between the subtypes, probably due to its EBV+ status.  However, its sequencing runs have been included for completeness.)","project":"SRP029739"}
{"number_samples":54,"species":"human","abstract":"The objective of this study is to identify a prognostic signature in colorectal cancer (CRC) patients with diverse progression and heterogeneity of CRCs. We generated RNA-seq data of 54 samples (normal colon, primary CRC, and liver metastasis) from 18 CRC patients and, from the RNA-seq data, identified significant genes associated with aggressiveness of CRC. Through diverse statistical methods including generalized linear model likelihood ratio test, two significantly activated regulators were identified. In the validation cohorts, two activated regulators were independent risk factors and potential chemotherapy-sensitive agenets in colorectal cancers. Overall design: RNA-seq data of 54 samples (normal colon, primary CRC, and liver metastasis) were generated from 18 CRC patients. Total RNA was isolated by RNeasy Mini Kit (Qiagen, CA, USA), according to the manufacturer's protocol. The quality and integrity of the RNA were confirmed by agarose gel electrophoresis and ethidium bromide staining, followed by visual examination under ultraviolet light. Sequencing library was prepared using TruSeq RNA Sample Preparation kit v2 (Illumina, CA, USA) according to the manufacturer’s protocols. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads, fragmented, and converted into cDNAs. Then, adapters were ligated and the fragments were amplified on a PCR. Sequencing was performed in paired end reads (2x100 bp) using Hiseq-2000 (Illumina).","project":"SRP029880"}
{"number_samples":7,"species":"human","abstract":"Induced pluripotent stem cells (iPSCs) are regarded as a central tool to understand human biology in health and disease. Similarly, iPSCs from closely related species should be a central tool to understand human evolution and to identify conserved and variable patterns of iPSC disease models. Here, we have generated human, gorilla, bonobo and cynomolgus monkey iPSCs. We show that these cells are well comparable in their differentiation potential and generally similar to human, cynomolgus and rhesus monkey embryonic stem cells (ESCs). RNA sequencing reveals that expression differences among clones, individuals and stem cell type are all of very similar magnitude within a species. In contrast, expression differences between closely related primate species are three times larger and most genes show significant expression differences among the analysed species. However, pseudogenes differ more than twice as much, suggesting that evolution of expression levels in primate stem cells is rapid, but constrained. These patterns in pluripotent stem cells are comparable to those found in other tissues except testis. Hence, primate iPSCs reveal insights into general primate gene expression evolution and should provide a rich source to identify conserved and species-specific gene expression patterns for cellular phenotypes. Contributors: Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany Max Planck Institute for Evolutionary Anthropology, Deutscher Platz 6, D-04103 Leipzig, Germany Overall design: We used expression profiling to characterize five gorilla, two bonobo and three macaque iPS clones as well as three iPS clones from two human individuals, three human embryonic stem (ES) cell lines and three macaque ES cell lines. We generated tagged RNA-Seq libraries from these 19 samples including four technical replicates (23 samples). Over 100 million single end reads were generated on the Illumina platform.","project":"SRP029888"}
{"number_samples":25,"species":"human","abstract":"We have quantified gene expression in five tissues (brain, heart, kidney, liver and testis) from humans, chimpanzees and rhesus macaques using the Illumina NlaIII Digital Gene Expression (DGE) protocol. This dataset extends a previous microarray study by Khaitovich et al. (Khaitovich et al. 2005) with the rhesus macaque outgroup and complements other previously generated tissue transcriptome profiles from primates (Enard et al. 2002; Khaitovich et al. 2006; Somel et al. 2009; Babbitt et al. 2010; Blekhman et al. 2010; Wetterbom et al. 2010). contributor: Max Planck Institute for Evolutionary Anthropology, Deutscher Platz 6, D-04103 Leipzig, Germany Overall design: Samples were obtained from brain (pre-frontal cortex), heart, kidney, liver, and testis tissues of male humans, chimpanzees and rhesus macaques. Illumina NlaIII DGE libraries for all samples were generated in tissue batches, randomizing species in library preparation and sequencing. Human samples originate from different, probably unrelated, individuals for each tissue. For chimpanzees and rhesus macaques the libraries for all tissues come from the same set of individuals and among these are individuals related at the half- and full-sibling level. Due to limited access to samples, the analysis could not be limited to individuals of similar age. Human individuals vary between 5 and 88 years of age, chimpanzees between 6 years and 35 years of age and rhesus macaques between 3 and 9 years of age.","project":"SRP029889"}
{"number_samples":36,"species":"human","abstract":"Inappropriate or sustained activation of innate immunity is a pathologic feature of several common cardio-metabolic disorders. Little is known, however, about transcriptomic modulation during inflammatory stress in disease-relevant human tissues. We applied deep RNA sequencing (RNA-seq) during low-dose experimental endotoxemia (LPS) in healthy humans to interrogate, in an unbiased manner, inflammatory tissue-level transcriptome responses of relevance to complex cardio-metabolic diseases. We utilized adipose and blood samples from three individuals who underwent a standardized inpatient endotoxemia protocol. Our comprehensive analysis revealed substantial, highly tissue- and subject-specific LPS-modulated changes in the expression of protein-coding genes and linc-RNAs as well as alternative splicing (AS). We also confirmed adipocytes and macrophages as potential cell sources of selective LPS-modulated linc-RNAs and AS events. Finally, we defined disease relevance of a subset of findings in obese adipose tissue and through interrogation of overlap with genome-wide association study loci for cardio-metabolic traits. Our findings provide novel insights into tissue-level genomic regulation, not detectable through analysis of DNA variations alone, of relevance to common cardio-metabolic diseases. Overall design: Using RNA-seq data to study LPS-modulated changes in lincRNA expression for adipose and blood of a healthy individual.","project":"SRP029899"}
{"number_samples":17,"species":"human","abstract":"More than half of human genes use alternative cleavage and polyadenylation (ApA) to generate mRNA transcripts that differ in the length of their 3' untranslated region (3'UTR), thus altering the post-transcriptional fate of the message and hence the protein output. The extent of 3'UTR variation across tissues and the functional role of ApA remain poorly understood. We developed a sequencing method to quantitatively map the 3' ends of the transcriptome of diverse human tissues and isogenic transformation systems. We found that cell type-specific gene expression is accomplished by two complementary programs. Tissue-restricted genes tend to have single 3'UTRs whereas a majority of ubiquitously transcribed genes generate multiple 3'UTRs. During transformation and differentiation, single-UTR genes change their mRNA abundance levels while multi-UTR genes mostly change 3'UTR isoform ratios to achieve tissue-specificity. However, both regulation programs target genes that function in the same pathways and processes that characterize the new cell type. Instead of finding global shifts in 3'UTR length during transformation and differentiation, we identify tissue-specific groups of multi-UTR genes that change their 3'UTR ratios – these changes in 3'UTR length are largely independent from changes in mRNA abundance. Finally, tissue-specific usage of ApA sites appears to be mechanism for changing the landscape targetable by ubiquitously expressed microRNAs.","project":"SRP029953"}
{"number_samples":4,"species":"human","abstract":"RBFOX over-expression in 293T cells","project":"SRP029987"}
{"number_samples":12,"species":"human","abstract":"Sterile immunity to Plasmodium falciparum infection can be induced experimentally in humans after few exposures. An example is the induction of immunity using whole parasites by exposure of malaria-naive volunteers to infectious mosquito bites while using chloroquine prophylaxis (CPS immunization). Chloroquine kills blood-stage parasites but leaves liver-stage parasites unaffected, thereby exposing the liver-stage and early blood-stage antigens to the immune system. Upon subsequent challenge, volunteers are completely protected from infection, but protective efficacy decreases when fewer infectious mosquito bites are used for CPS immunization. Efforts to understand the mechanisms of this immunity, and how it differs from naturally-acquired immunity, may provide critical insights that could aid malaria vaccine development.  In this pilot study, transcriptomic features are derived from blood samples collected before and after challenge with infectious mosquito bites. Overall design: 12 samples; paired pre- and post-challenge for 5 individuals, plus two controls","project":"SRP029990"}
{"number_samples":12,"species":"human","abstract":"Early chemotherapy for advanced/metastatic non-castration resistant prostate cancer (PCa) may improve overall patient survival.  We studied the safety, tolerability and early efficacy of up-front docetaxel chemotherapy and androgen deprivation therapy (ADT) versus ADT alone for patients with newly-diagnosed advanced/metastatic PCa.  As proof of concept, we undertook in vivo gene expression profiling by next generation RNA sequencing (RNA-Seq). Overall design: Tumour biposies from 6 patients were taken before and after treatment with combined ADT and docetaxcel for 6 weeks","project":"SRP030027"}
{"number_samples":28,"species":"human","abstract":"RNAseq human HCC","project":"SRP030040"}
{"number_samples":36,"species":"human","abstract":"The majority of disease-associated variants lie outside protein-coding regions, suggesting a link between variation in regulatory regions and disease predisposition. We studied differences in chromatin states using five histone modifications, cohesin, and CTCF in lymphoblastoid lines from 19 individuals of diverse ancestry. We found extensive signal variation in regulatory regions, which often switch between active and repressed states across individuals. Enhancer activity is particularly diverse among individuals, whereas gene expression remains relatively stable. Chromatin variability shows genetic inheritance in trios, correlates with genetic variation and population divergence, and is associated with disruptions of transcription factor binding motifs. Overall, our results provide insights into chromatin variation among humans. Overall design: To characterize human variation in diverse types of regulatory elements, we studied the chromatin state of lymphoblastoid cell lines (LCLs) derived from 19 individuals: 5 European (CEU), 7 Yoruban (YRI), and 2 Asian individuals from the 1000 Genomes Project (including two mother-father-daughter trios), an additional Caucasian individual), and four individuals from the San population). We used RNA-Seq to measure expression and Chromatin Immunoprecipitation followed by high-throughput sequencing (ChIP-Seq) to map five histone modifications (H3K27ac, H3K4me1, H3K4me3, H3K36me3, and H3K27me3) and two general factors (CTCF, and SA1, a subunit of cohesin). We generated 2 replicates per factor.","project":"SRP030041"}
{"number_samples":27,"species":"human","abstract":"Advantages of RNA-Seq over array based platforms are quantitative gene expression and discovery of expressed single nucleotide variants (eSNVs) and fusion transcripts from a single platform, but the sensitivity for each of these characteristics is unknown. We measured gene expression in a set of manually degraded RNAs, nine pairs of matched fresh-frozen, and FFPE RNA isolated from breast tumor with the hybridization based, NanoString nCounter, (226 gene panel) and with whole transcriptome RNA-Seq using RiboZeroGold ScriptSeq V2 library preparation kits. We performed correlation analyses of gene expression between samples and across platforms. We then specifically assessed whole transcriptome expression of lincRNA and discovery of eSNVs and fusion transcripts in the FFPE RNA-Seq data. For gene expression in the manually degraded samples, we observed Pearson correlation of >0.94 and >0.80 with NanoString and ScriptSeq protocols respectively. Gene expression data for matched fresh-frozen and FFPE samples yielded mean Pearson correlations of 0.874 and 0.783 for NanoString (226 genes) and ScriptSeq whole transcriptome protocols respectively. Specifically for lincRNAs, we observed superb Pearson correlation (0.988) between matched fresh-frozen and FFPE pairs. FFPE samples across NanoString and RNA-Seq platforms gave a mean Pearson correlation of 0.838. In FFPE libraries, we detected 53.4% of high confidence SNVs and 24% of high confidence fusion transcripts. Sensitivity of fusion transcript detection was not overcome by an increase in depth of sequencing up to 3-fold (increase from ~56 to ~159 million reads). Both NanoString and ScriptSeq RNA-Seq technologies yield reliable gene expression data for degraded and FFPE material. The high degree of correlation between NanoString and RNA-Seq platforms suggests discovery based whole transciptome studies from FFPE material will produce reliable expression data. The RiboZeroGold ScriptSeq protocol performed particularly well for lincRNA expression from FFPE libraries but detection of eSNV and fusion transcripts was less sensitive. Overall design: We performed RNASeq on RNA from nine matched pairs of fresh-frozen and FFPE tissues from breast cancer patients. The goal was to test the RiboZeroGold ScriptSeq complete low input library preparation kit for degraded RNA samples.","project":"SRP030401"}
{"number_samples":1,"species":"human","abstract":"We use these data to estimate phylogenetic relationships among Actias luna, Ceratomia undulosa, Enyo lugubris, Hemaris diffinis, Darapsa myron, and Attacus atlas and other Bombycoids moth.","project":"SRP030471"}
{"number_samples":8,"species":"human","abstract":"We used transcription activator-like effector nucleases (TALENs) to generate knockout cells for two related microRNAs (miRNAs), mir-141 and mir-200c, which belong to the deeply conserved mir-200 family. By carrying out deep sequencing, we identified the target genes of each miRNA. Interestingly, miR-141 and miR-200c, despite their overall similarity, suppressed largely non-overlapping groups of targets. Overall design: Analysis of global mRNA level change in mir-141 and mir-200c knockout compared to wild type cells","project":"SRP030475"}
{"number_samples":113,"species":"human","abstract":"We generated single-cell transcriptomes from a large number of single cells using several commercially available platforms, in both microliter and nanoliter volumes, and compared performance between them. We benchmarked each method to conventional RNA-seq of the same sample using bulk total RNA, as well as to multiplexed qPCR, which is the current gold standard for quantitative single-cell gene expression analysis. In doing so, we were able to systematically evaluate the sensitivity, precision, and accuracy of various approaches to single-cell RNA-seq.  Our results show that it is possible to use single-cell RNA-seq to perform quantitative transcriptome measurements of individual cells, that it is possible to obtain quantitative and accurate gene expression measurements with a relatively small number of sequencing reads, and that when such measurements are performed on large numbers of cells, one can recapitulate the bulk transcriptome complexity, and the distributions of gene expression levels found by single-cell qPCR. Overall design: 109 single-cell human transcriptomes were analyzed in total; 96 using nanoliter volume sample processing on a microfluidic platform, Nextera library prep (biological replicates); 3 using the SMARTer cDNA synthesis kit, Nextera library prep (biological replicates); 3 using the Transplex cDNA synthesis kit, Nextera library prep (biological replicates); 7 using the Ovation Nugen cDNA synthesis kit (biological replicates) where 3 used Nextera library prep and 4 used NEBNext library prep. In addition, 4 bulk RNA samples were sequenced: bulk RNA generated using ~1 million pooled cells was used to make bulk libraries, 2 of which were made using SMARTer cDNA synthesis kit (technical replicates) and 2 made using Superscript RT kit with no amplification (technical replicates). All 4 bulk samples were made into libraries using Nextera.","project":"SRP030617"}
{"number_samples":38,"species":"human","abstract":"78 tissue samples from prefrontal cortex (PFC) in human and macaque Overall design: The data from human and macaque PFC samples with different ages were used to estimate gene expression changes, including both protein-coding genes and lincRNAs, in PFC along lifespan in the two species.","project":"SRP030628"}
{"number_samples":9,"species":"human","abstract":"BS69 (aka ZMYND11) was initially discovered as a direct target of adenoviral E1A oncoprotein1. Subsequent studies implicated BS69 as a tumor suppressor and a transcriptional regulator2. But exactly how BS69 regulates gene expression has remained elusive. BS69 contains tandemly arranged PHD, BROMO and PWWP domains, which are known to function as chromatin recognition modalities. Here we show that BS69 selectively recognizes histone variant H3.3 lysine 36 trimethylation (H3.3K36me3) via these chromatin recognition modules. Using a proteomics approach, we further identified association of BS69 with a host of RNA splicing regulators including EFTUD2 and other U5 snRNP components of the spliceosome. Remarkably, RNA-seq analysis shows that BS69 mainly regulates intron retention (IR), which is one of the least well-understood RNA alternative splicing events in mammalian cells. Specifically, loss of BS69 results in a decrease in IR, while in contrast, knockdown of EFTUD2 leads to an increase in IR, consistent with its role as a core member of the splicesome machinery. Importantly, wildtype BS69, but not a BS69 mutant defective in its interaction with EFTUD2, rescues the IR events. We also show that knockdown of SETD2, the main enzyme that mediates H3K36 trimethylation3, similarly results in a decreased retention of the same introns regulated by BS69. Significantly, a BS69 mutant unable to bind H3.3K36me3 in vitro fails to rescue the reduced IR phenotype associated with the loss of BS69. Taken together, our findings identify an antagonistic relationship between BS69 and the core splicing machinery and demonstrate that BS69-mediated RNA splicing regulation is dependent both on its ability to bind chromatin decorated by H3K36me3 and its ability to physically interact with the core spliceosome machinery. Our study reveals a novel and unexpected role of BS69 in connecting histone H3.3K36 trimethylation to regulated RNA splicing, providing significant new insights into chromatin regulation of RNA splicing. Overall design: HeLa cells were infected with lentiviruses carrying control and BS69 shRNA and deep sequencing data were generated, in triplicate, using Illumina HiSeq 2000","project":"SRP030639"}
{"number_samples":23,"species":"human","abstract":"High-throughput sequencing of primary cutaneous follicle center lymphoma (PCFCL), primary cutaneous diffuse large B-cell lymphoma, leg type (PCLBCL-LT) and in vitro activated peripheral blood B-cells. We performed high-throughput sequencing analysis on frozen tumor biopsies from 19 cases of PCFCL and PCLBCL-LT to establish microRNA profiles. Cluster analysis of the complete microRNome could not distinguish between the two subtypes, but 16 single microRNAs were found to be differentially expressed. Overall design: Lymphoma miRNA profiles of were generated by deep sequencing, using Illumina Genome Analyzer II.","project":"SRP031459"}
{"number_samples":38,"species":"human","abstract":"Motivation: RNA-seq is replacing microarrays as the primary tool for gene expression studies. Many RNA-seq studies have used insufficient biological replicates, resulting in low statistical power and inefficient use of sequencing resources. Results: We show the explicit trade-off between more biological replicates and deeper sequencing in increasing power to detect differentially expressed (DE) genes. In the human cell line MCF-7, adding more sequencing depth after 10M reads gives diminishing returns on power to detect DE genes, while adding biological replicates improves power significantly regardless of sequencing depth. We also propose a cost-effectiveness metric for guiding the design of large scale RNA-seq DE studies. Our analysis showed that sequencing less reads and perform more biological replication is an effective strategy to increase power and accuracy in large scale differential expression RNA-seq studies, and provided new insights into efficient experiment design of RNA-seq studies Overall design: Treatment (10nM E2 treatment for 24h) and control MCF7 cells are both replicated 7 times, and collected for mRNA-seq. Reads are then subsampled for statistical analysis.","project":"SRP031476"}
{"number_samples":12,"species":"human","abstract":"To gain insight into the molecular changes during OSCC carcinogenesis, we performed unbiased, whole genome deep sequencing (RNA-seq) using RNA isolated from cultured, human TERT-immortalized, non-tumorigenic OKF6-TERT1R and OSCC SCC-9 cells. Overall design: OKF6-TERT1R cells and SCC-9 cells were  plated in 10 cm2 tissue culture plates at the density of 2 × 106 cells/plate and treated with 1 µM RA or vehicle (0.1% ethanol) for 48 hours. Experiment includes 3 independent biological replicates.","project":"SRP031478"}
{"number_samples":20,"species":"human","abstract":"Macrophage activation is associated with profound transcriptional reprogramming. Although much progress has been made in the understanding of macrophage activation, polarization and function, the transcriptional programs regulating these processes remain poorly characterized. We stimulated human macrophages with diverse activation signals, acquiring a dataset of 299 macrophage transcriptomes. Analysis of this dataset revealed a spectrum of macrophage activation states extending the current M1 versus M2-polarization model. Network analyses identified central transcriptional regulators associated with all macrophage activation complemented by regulators related to stimulus-specific programs. Applying these transcriptional programs to human alveolar macrophages from smokers and patients with chronic obstructive pulmonary disease (COPD) revealed an unexpected loss of inflammatory signatures in COPD patients. Finally, by integrating murine data from the ImmGen project we propose a refined, activation-independent core signature for human and murine macrophages. This resource serves as a framework for future research into regulation of macrophage activation in health and disease. Overall design: Since transcriptional programs are further modulated on several levels including miRNAs we assessed the global spectrum of miRNA expression by miRNA-Seq in macrophages stimulated with IFN?, IL4 or with the combination of TNFa, PGE2 and P3C","project":"SRP031496"}
{"number_samples":5,"species":"human","abstract":"We report cell-type specific ribosome profiling in a mouse glioma model. Overall design: We report a strategy for cell-type specific ribosome profiling in vivo. Our strategy was applied to the characterization of a mouse glioma model.","project":"SRP031501"}
{"number_samples":2,"species":"human","abstract":"We have identified differentially PR-regulated genes in T47D cells and mapped PR-binding sites by using next generation sequencing Overall design: RNA-seq  and ChIP-seq in progestin treated cells","project":"SRP031503"}
{"number_samples":4,"species":"human","abstract":"Using the iCLIP protocol we have identified the cellular RNA entities that are bound by MOV10. We report the location and sequence of the MOV10 binding region on each RNA entity. Overall design: To identify the RNAs that bound MOV10, we UV-cross-linked HEK293F cells and immunoprecipitated with an irrelevant antibody (ir or \"control\") followed by a MOV10-specific antibody (MOV10) to isolate associated RNAs after stringent washing.","project":"SRP031507"}
{"number_samples":15,"species":"human","abstract":"Functional characterization of an identified physical connection between the cap-binding complex and the RNA exosome","project":"SRP031620"}
{"number_samples":20,"species":"human","abstract":"Global miRNA expression profiling of human malignancies is gaining popularity in both basic and clinically driven research. But to date, the majority of such analyses have used microarrays and quantitative real-time PCR.  With the introduction of digital count technologies, such as next-generation sequencing (NGS) and the NanoString nCounter System, we have at our disposal, many more options. To make effective use of these different platforms, the strengths and pitfalls of several miRNA profiling technologies were assessed, including a microarray platform, NGS technologies and the NanoString nCounter System.  These results were compared to gold-standard quantitative real-time PCR. Overall design: Comparison of non-small cell lung cancer cell lines grown in vitro (n = 5) and in vivo (n = 5) as xenograft models.","project":"SRP031698"}
{"number_samples":20,"species":"human","abstract":"Aging signatures developed from a longitudinal study design are dominated by reduced transcription of genes involved in protein synthesis Aging is a multifactorial process where the impact of singular components still remains unclear. Furthermore, previous studies were focused on measuring specific traits such as DNA -methylation and used categorical group-wise designs, unable to capture intra-individual signature changes. Here we have developed a new method for a longitudinal, age-related analysis combining the merits of a pair-wise design with the statistical power of gene set enrichment analysis. We present an integrated analysis, including transcriptional changes and genome-wide epigenetic changes in DNA- methylation, H3K4- and H3K27- histone methylation in promoter regions.  We tested our method on a rare collection of paired skin fibroblast samples from male middle age to old age transitions and obtained functional, age-related clusters. By using a set of only ten individuals, we could demonstrate a high overlap of functional terms to previously established tissue-independent age signatures including extracellular matrix, apoptosis and oxidative stress. Importantly, we identify protein translation-related processes as the main cluster of age-driven, specific down regulation. Overall design: Evaluation of transcriptional changes in matched sample pairs of primary skin fibroblasts from middle and old age.","project":"SRP031776"}
{"number_samples":9,"species":"human","abstract":"The recent reports of two circular RNAs (circRNAs) with strong potential to act as microRNA (miRNA) sponges suggest that circRNAs might play important roles in regulating gene expression. However, the global properties of circRNAs are not well understood. We developed a computational pipeline to identify circRNAs and quantify their relative abundance from RNA-seq data. Applying this pipeline to a large set of non-poly(A)-selected RNA-seq data from the ENCODE project, we annotated 7,112 human circRNAs that were estimated to comprise at least 10% of the transcripts accumulating from their loci. Most circRNAs are expressed in only a few cell types and at low abundance, but they are no more cell-type–specific than are mRNAs with similar overall expression levels. Although most circRNAs overlap protein-coding sequences, ribosome profiling provides no evidence for their translation. We also annotated 635 mouse circRNAs, and although 20% of them are orthologous to human circRNAs, the sequence conservation of these circRNA orthologs is no higher than that of their flanking linear exons. The previously proposed miR-7 sponge, CDR1as, is one of only two circRNAs with more miRNA sites than expected by chance, with the next best miRNA-sponge candidate deriving from a primate-specific zinc-finger gene, ZNF91. These results provide a new framework for future investigation of this intriguing topological isoform while raising doubts regarding a biological function of most circRNAs. Overall design: Examination of 9 samples in 1 cell type Note: The ENCODE data we used are under GEO SuperSeries GSE26284 (all samples labeled \"_cell_total\"). But they were not used in the processing of the U2OS data.","project":"SRP031849"}
{"number_samples":20,"species":"human","abstract":"Rett syndrome (RTT) is one of the most prevalent female mental disorders. De novo mutations in methyl CpG binding protein 2 (MeCP2) are a major cause of RTT. MeCP2 regulates gene expression as a transcription regulator as well as through long-range chromatin interaction. Because MeCP2 is present on the X chromosome, RTT is manifested in a X-linked dominant manner. Investigation using murine MeCP2 null models and post-mortem human brain tissues has contributed to understanding the molecular and physiological function of MeCP2. In addition, RTT models using human induced pluripotent stem cells derived from RTT patients (RTT-iPSCs) provide novel resources to elucidate the regulatory mechanism of MeCP2. Previously, we obtained clones of female RTT-iPSCs that express either wild type or mutant MECP2 due to the inactivation of one X chromosome. Reactivation of the X chromosome also allowed us to have RTT-iPSCs that express both wild type and mutant MECP2. Using these unique pluripotent stem cells, we investigated the regulation of gene expression by MeCP2 in pluripotent stem cells by transcriptome analysis. We found that MeCP2 regulates genes encoding mitochondrial membrane proteins. In addition, loss of function in MeCP2 results in de-repression of genes on the inactive X chromosome. Furthermore, we showed that each mutation in MECP2 affects a partly different set of genes. These studies suggest that fundamental cellular physiology is affected by mutations in MECP2 from very early fetal development and that a therapeutic approach targeting to unique forms of mutant MeCP2 is needed. Overall design: RNA samples from normal ESCs/iPSCs, RTT-iPSCs and MeCP2 KD iPSCs were obtained. Gene expression of those cells were analyzed.","project":"SRP031858"}
{"number_samples":20,"species":"human","abstract":"We performed RNA-seq to identify differentially expressed genes between left and right hemispheres. This datased contains raw mRNA sequencing data of superior temporal cortices from both left and right hemispheres of eleven human brains.","project":"SRP031868"}
{"number_samples":24,"species":"human","abstract":"Purpose: The purpose of this experiment is to identify a C9-ALS/FTD specific genomic profile in fibroblast lines that is distinct from sporadic ALS without C9orf72 expansion and non-neurologic control cells. The study will then evaluate the effect on this identified profile of ASO treatment targeting the sense strand RNA transcript of the C9orf72 gene. Methods: Expression profiling was performed on RNAs from fibroblasts of four C9orf72 patients, four control individuals and four sporadic ALS patients using Multiplex Analysis of PolyA-linked Sequences method. Results: Hierarchical clustering of expression values for all genes showed that the four C9orf72 patient lines had an expression profile distinct from control and sporadic ALS lines. Statistical comparison of expression values between the four C9orf72 lines and the four control lines revealed that 122 genes were upregulated (defined by a False Discovery Rate FDR<0.05) and 34 genes were downregulated (defined by a False Discovery Rate FDR <0.05) in C9orf72 patient fibroblasts. Conclusions: A genome wide RNA signature can be defined in fibroblasts with C9orf72 expansion. ASO-mediated reduction of C9orf72 RNA levels in fibroblasts with the hexanucleotide expansion efficiently reduced accumulation of GGGGCC RNA foci. This did not, however, generate a reversal of the C9orf72 RNA profile. Overall design: Use of Multiplex Analysis of PolyA-linked Sequences to identify expression changes in fibroblasts from amyotrophic lateral sclerosis and frontotemporal dementia patients harboring an hexanucleotide expansion in the C9orf72 gene.","project":"SRP032165"}
{"number_samples":17,"species":"human","abstract":"HERV abundance in Human brain","project":"SRP032168"}
{"number_samples":2,"species":"human","abstract":"Multidrug resistance (MDR) frequently develops in cancer patients exposed to chemotherapeutic agents and is usually brought about by over-expression of P-glycoprotein (P-gp) which acts as a drug efflux pump. MiRNAome profiling using next-generation sequencing identified differentially expressed microRNAs (miRs) between parental K562 cells and MDR K562 cells (K562/ADM) induced by chronic adriamycin treatment. Overall design: MiRNAome profiling in untreated K562 cells and K562 cells exposed to long-term adriamycin treatment","project":"SRP032275"}
{"number_samples":1,"species":"human","abstract":"In this study we investigated the suitability of blood to identify HD transcriptomic biomarkers, validated the outcome in an independent cohort and derived a first empiric panel of biomarkers capable of predicting HD motor scores. Finally we examined whether patient gene expression profiles could provide information about HD affected biological pathways. Overall design: Examination of differentially expressed genes in peripheral whole blood using linear modelling of  gene expression data (3' DGE) against UHDRS total motor scores of Huntington's Disease mutation carriers and controls.","project":"SRP032279"}
{"number_samples":1,"species":"human","abstract":"RNAseq to determine baseline expression of kinome in MDA-MB-231 claudin-low breast cancer cell line","project":"SRP032280"}
{"number_samples":1,"species":"human","abstract":"RNAseq of SUM159PT claudin-low breast cancer cell line to determine baseline kinome expression","project":"SRP032344"}
{"number_samples":9,"species":"human","abstract":"RNA sequencing (RNA-seq) analysis revealed 31 novel lncRNAs in HCASMC, including a vascular cell-enriched lncRNA called SENCR (for Smooth muscle and Endothelial cell long Non-Coding RNA). RT-PCR and hybridization studies show SENCR exists in two isoforms and is transcribed antisense from the 5’ end of the FLI1 gene. Knockdown of SENCR has no effect on FLI1 mRNA or protein expression. Biochemical fractionation and RNA fluorescence in situ hybridization (FISH) studies indicate SENCR is a cytoplasmic lncRNA. RNA-seq experiments in HCASMC where SENCR is attenuated disclose decreased expression of Myocardin and many SMC contractile genes; conversely a pro-migratory gene signature is increased. RT-PCR and Western blotting validated several differentially expressed genes following SENCR knockdown. Loss-of-function studies in scratch wound and Boyden chamber assays support SENCR as an inhibitor of vascular cell migration. Overall design: Total RNAs of 3 replicates of normal human coronary artery smooth muscle cells (Mock1, Mock2 and Mock3) were sequenced and analyzed for identification of novel lncRNAs. One of identified novel lncRNAs from that experiment is SENCR. To study its function, SENCR knock-down experiment were performed and then RNA-seq profiles of 3 replicates of both SENCR-knockdown samples and corresponding controls were compared.","project":"SRP032363"}
{"number_samples":1,"species":"human","abstract":"We used PacBio data to identify more reliable transcripts from hESC, based on which we can estimate gene/transcript abundance better from Illumina data. Overall design: PacBio long reads and Illumina short reads were generated from the same hESC cell line H1. PacBio reads were error-corrected by Illumina reads to identify transcripts. rSeq is used to estimate gene/transcript abundance of the identified transcriptome.","project":"SRP032367"}
{"number_samples":8,"species":"human","abstract":"We find significant evidence of the OVOL, AP1, STAT1, STAT3, and NFKB1 TFs having important roles in MET. We prioritize known gene/drug targets for follow-up in the clinic, and show that the AP1/MYC TF pair is a strong candidate for intervention. Overall design: Examination of the effects of OVOL1 and OVOL2 overexpression common to prostate cancer and breast cancer models.","project":"SRP032456"}
{"number_samples":14,"species":"human","abstract":"RNA sequencing data for patients with Embryonal tumor with multilayered rosettes and Primitive neuroectodermal tumor as controls.","project":"SRP032476"}
{"number_samples":11,"species":"human","abstract":"Background: The Aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that is activated by xenobiotic chemicals that function as AHR ligands.  The response to xenobiotic AHR ligands is toxicity and the induction of drug metabolizing enzymes. The impact of AHR knockdown on gene expression and pathways in human breast cancer cells in absence of xenobiotic AHR ligands has not been investigated on a genome-wide scale. Methods: MCF-7 cells were used as a model of human breast cancer.  The AHR was silenced with short interfering RNA against AHR (siAHR).  RNA-sequencing coupled with Ingenuity pathway analysis (IPA) was used to determine the impact of AHR knockdown on gene expression and pathways in the absence of xenobiotic AHR ligands.  Western blot analysis and recombinant tumor necrosis factor (TNF) was used to investigate the impact of AHR knockdown on TNF induction of MNSOD expression. Results: We found that the AHR is transcriptionally active in MCF-7 breast cancer cells in the absence of xenobiotic AHR ligands.  In total, the expression of 634 genes was significantly changed in AHR knockdown cells compared to controls at a false discovery rate of < 10%. The analysis confirmed that drug metabolizing enzymes were AHR targets; however, we found that AHR also promoted the expression of genes that were not directly related to the metabolism of xenobiotics, such as those involved in lipid and eicosanoid synthesis.  Gene pathway analysis of the AHR regulated gene dataset predicted TNF activity to be reduced in AHR knockdown cells.  Our finding that AHR knockdown inhibited TNF-stimulated increases in MNSOD expression confirmed this IPA prediction. Conclusions: This is the first gene expression profiling study of AHR knockdown breast cancer cells.  Several known and novel AHR targets were identified.  The results suggest that endogenous AHR regulation impacts eicosanoid synthesis by regulating gene expression.  The IPA prediction of reduced TNF activity in AHR knockdown cells was confirmed by showing that TNF-induced increases in MNSOD expression was inhibited in AHR knockdown cells.   The requirement of AHR for MNSOD activation is novel and provides a new link by which AHR may impact reactive oxidative species (ROS) signaling. Overall design: Expression profiles by mRNA sequencing were generated for human MCF-7 cells transfected with cRNAi (6 replicates) or AHR-siRNA (5 replicates) for 36hr. Sequencing was performed on an Illumina HiSeq 1000 using a 2x100 base paired-end strategy.","project":"SRP032510"}
{"number_samples":11,"species":"human","abstract":"Here, seeking to gain insight into the array of transcripts engaged with miRNAs in human brain, we performed HITS-CLIP to profile transcriptome-wide Ago2:RNA interactions in a panel of eleven post-mortem adult human brain samples harvested from adult motor cortex and cingulate gyrus, regions associated with movement and psychiatric disorders . Overall design: High-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP) Eleven post-mortem adult human brain tissues were subjected to ultraviolet radiation to crosslink proteins with nucleic acids, and HITS-CLIP was performed as previously described by Chi et al Nature. 2009 Jul 23;460(7254):479-86 using an anti-Ago2 polyclonal antibody and some additional modifications.","project":"SRP032539"}
{"number_samples":11,"species":"human","abstract":"Here, seeking to gain insight into the array of transcripts engaged with miRNAs in human brain, we performed HITS-CLIP to profile transcriptome-wide Ago2:RNA interactions in a panel of eleven post-mortem adult human brain samples harvested from adult motor cortex and cingulate gyrus, regions associated with movement and psychiatric disorders . Overall design: High-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP) Eleven post-mortem adult human brain tissues were subjected to ultraviolet radiation to crosslink proteins with nucleic acids, and HITS-CLIP was performed as previously described by Chi et al Nature. 2009 Jul 23;460(7254):479-86 using an anti-Ago2 polyclonal antibody and some additional modifications.","project":"SRP032540"}
{"number_samples":2,"species":"human","abstract":"Analysis of Allelic bias in clonal lymphoblastoid cells. Abstract: In mammals, numerous autosomal genes are subject to mitotically stable monoallelic expression (MAE), including genes that play critical roles in a variety of human diseases. Due to challenges posed by the clonal nature of MAE, very little is known about its regulation; in particular, no molecular features have been specifically linked to MAE. Here we report an approach that distinguishes MAE genes in human cells with great accuracy: a chromatin signature consisting of chromatin marks associated with active transcription (H3K36me3) and silencing (H3K27me3) simultaneously occurring in the gene body. The MAE signature is present in ~20% of ubiquitously expressed genes and over 30% of tissue-specific genes across cell types. Notably, it is enriched among key developmental genes that have bivalent chromatin structure in pluripotent cells. Our results open a new approach to the study of MAE that is independent of polymorphisms, and suggest that MAE is linked to cell differentiation. Overall design: Poly A purified total RNA was used for library construction using a method described by Parkhomchuk et. al. NAR 2009. The library was strand-specific but the pipeline for data analysis does not assume the library is strand-specific.","project":"SRP032542"}
{"number_samples":3,"species":"human","abstract":"We have identified a single miRNA, miR-181a, that can modulate TGF-ß signaling to induce and maintain EMT, and effect further downstream events of tumour cell survival, altered response to chemotherapy, migration, invasion and dissemination in vivo. Our present study provides an understanding of how enhanced expression of miR-181a can confer malignant and invasive traits through the modulation of a canonical signaling pathway and a consequent maintenance of a mesenchymal state. Furthermore, inhibition of miR-181a led to a reversion of EMT and subsequent events through decreased TGF-ß signaling. Our data confirmed Smad7 as a functional target through which TGF-ß-mediated EMT occurs; re-expression of Smad7 lacking its 3'UTR was able to rescue miR-181a-mediated phenotypes, deeming Smad7 as a critical mediator of miR-181a-induced EMT. Other recent studies support the crucial role(s) that miRNAs play in mediating EMT and consequent aggressive disease traits. For example, the miR-106b-25 cluster has also been shown to target Smad7 and mediate TGF-ß-induced EMT downstream to Six1 in breast cancer34. miR-9 directly targets E-cadherin and inhibition of miR-9 had led to an inhibition of metastasis35. Conversely, the miR-200 and -205 family was shown to target transcriptional repressors of E-cadherin, ZEB1 and SIP1, and re-expression of these miRNAs led to a mesenchymal-to-epithelial transition and prevented TGF-ß -induced EMT36. Overall design: A2780 ovarian cancer cell lines stably expressing either pBABE (control vector), p181a#1( clone 1 expressing miR-181a) or p181a#2( clone 2 expressing miR-181a)","project":"SRP032559"}
{"number_samples":27,"species":"human","abstract":"To identify transcripts altered upon LIN-41 knockdown, we transfected either a control siRNA or one of two different LIN-41 siRNAs into human embryonic stem cells and collected total RNA 72 hours after transfection. Overall design: We compared transcript levels between control siRNA or LIN-41 siRNA treated cells.","project":"SRP032743"}
{"number_samples":6,"species":"human","abstract":"SOX11 (Sex determining region Y-box 11) expression is specific for MCL as compared to other Non-Hodgkin’s lymphomas. However, the function and direct binding targets of SOX11 in MCL are largely unknown. We used high-resolution ChIP-Seq to identify the direct target genes of SOX11 in a genome-wide, unbiased manner and elucidate its functional significance. Pathway analysis identified WNT, PKA and TGF-beta signaling pathways as significantly enriched by SOX11 target genes. qCHIP confirmed that SOX11 directly binds to individual genes in these pathways in both MCL cell lines and patients. Interrogation of an eighty-two patient  gene-expression dataset demonstrated that SOX11 mRNA expression was inversely proportional to Ki-67, a marker of cell proliferation. Functional studies using RNA interference demonstrate that SOX11 directly regulates WNT signaling and modulates chemotherapy sensitivity to cytarabine in MCL. We analyzed SOX11 expression in three independent well-annotated tissue microarrays from the University of Wisconsin (UW), Karolina Institute and British Columbia Cancer Agency (BCCA). Our findings suggest that high SOX11 expression is associated with improved survival in a subset of MCL patients, particularly those treated with intensive chemotherapy incorporating cytarabine. Transcriptional regulation of WNT and other biological pathways affects by SOX11 target genes may help explain the impact of SOX11 expression on patient outcomes. Overall design: RNA-seq experiments studying SOX11-mediated regulation of gene transcription by examining genes differentially expressed following SOX11 depletion in 3 MCL cell lines, Granta-519, Z138 and JEKO-1","project":"SRP032754"}
{"number_samples":232,"species":"human","abstract":"Immunity to malaria can be acquired through natural exposure to Plasmodium falciparum (Pf), but only after years of repeated infections. Typically, this immunity is acquired by adolescence and confers protection against disease, but not Pf infection per se. Efforts to understand the mechanisms of this immunity are integral to the development of a vaccine that would mimic the induction of adult immunity in children. The current study applies transcriptomic analyses to a cohort from the rural village of Kalifabougou, Mali, where Pf transmission is intense and seasonal. Signatures that correlate with protection from malaria may yield new hypotheses regarding the biological mechanisms through which malaria immunity is induced by natural Pf infection. The resulting datasets will be of considerable value in the urgent worldwide effort to develop a malaria vaccine that could prevent more than a million deaths annually. Overall design: 108 samples; paired pre- and post-challenge for 54 individuals 198 samples; paired pre- and post-challenge for 99 individuals","project":"SRP032775"}
{"number_samples":20,"species":"human","abstract":"Goal: To define the digital transcriptome of three breast cancer subtypes (TNBC, Non-TNBC, and HER2-positive) using RNA-sequencing technology. To elucidate differentially expressed known and novel transcripts, alternatively spliced genes and differential isoforms and lastly expressed variants in our dataset. Method: Dr. Suzanne Fuqua (Baylor College of Medicine) provided the human breast cancer tissue RNA samples. All of the human samples were used in accordance with the IRB procedures of Baylor College of Medicine. The breast tumour types, TNBC, Non-TNBC and HER2-positive, were classified on the basis of immunohistochemical and RT-qPCR classification. Results: Comparative transcriptomic analyses elucidated differentially expressed transcripts between the three breast cancer groups, identifying several new modulators of breast cancer. We discovered subtype specific differentially spliced genes and splice isoforms not previously recognized in human transcriptome. Further, we showed that exon skip and intron retention are predominant splice events in breast cancer. In addition, we found that differential expression of primary transcripts and promoter switching are significantly deregulated in breast cancer compared to normal breast.  We also report novel expressed variants, allelic prevalence and abundance, and coexpression with other variation, and splicing signatures. Additionally we describe novel SNPs and INDELs in cancer relevant genes with no prior reported association of point mutations with cancer Overall design: mRNA profiles of 17 breast tumor samples of three different subtypes (TNBC, non-TNBC and HER2-positive) and normal human breast organoids (epithelium) samples (NBS) were sequenced using Illumina HiSeq.","project":"SRP032789"}
{"number_samples":16,"species":"human","abstract":"Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative condition characterized by loss of motor neurons in the brain and spinal cord. Expansions of a hexanucleotide repeat (GGGGCC) in the noncoding region of the C9ORF72 gene are the most common cause of the familial form of ALS (C9-ALS), as well as frontotemporal lobar degeneration and other neurological diseases. How the repeat expansion causes disease remains unclear, with both loss of function (haploinsufficiency) and gain of function (either toxic RNA or protein products) proposed. We report a cellular model of C9-ALS with motor neurons differentiated from induced pluripotent stem cells (iPSCs) derived from ALS patients carrying the C9ORF72 repeat expansion. No significant loss of C9ORF72 expression was observed, and knockdown of the transcript was not toxic to cultured human motor neurons. Transcription of the repeat was increased, leading to accumulation of GGGGCC repeat–containing RNA foci selectively in C9-ALS iPSC-derived motor neurons. Repeat-containing RNA foci colocalized with hnRNPA1 and Pur-a, suggesting that they may be able to alter RNA metabolism. C9-ALS motor neurons showed altered expression of genes involved in membrane excitability including DPP6, and demonstrated a diminished capacity to fire continuous spikes upon depolarization compared to control motor neurons. Antisense oligonucleotides targeting the C9ORF72 transcript suppressed RNA foci formation and reversed gene expression alterations in C9-ALS motor neurons. These data show that patient-derived motor neurons can be used to delineate pathogenic events in ALS. Overall design: Transcriptome profiling from iPSC derived motor neurons compared to controls","project":"SRP032798"}
{"number_samples":20,"species":"human","abstract":"Chromatin-based functional genomic analyses and genomewide association studies (GWASs) together implicate enhancers as critical elements influencing gene expression and risk for common diseases. Here, we performed systematic chromatin and transcriptome profiling in human pancreatic islets. Integrated analysis of islet data with those generated by the ENCODE project in nine cell types identified specific and significant enrichment of type 2 diabetes and related quantitative trait GWAS variants in islet enhancers. Our integrated chromatin maps reveal that most enhancers are short (median = 0.8 kb). Each cell type also contains a substantial number of more extended (=3 kb) enhancers. Interestingly, these stretch enhancers are often tissue-specific and overlap locus control regions, suggesting that they are important chromatin regulatory beacons. Indeed, we show that (i) tissue specificity of enhancers and nearby gene expression increase with enhancer length; (ii) neighborhoods containing stretch enhancers are enriched for important cell type-specific genes; and (iii) GWAS variants associated with traits relevant to a particular cell type are more enriched in stretch enhancers compared with short enhancers. Reporter constructs containing stretch enhancer sequences exhibited tissue-specific activity in cell culture experiments and in transgenic mice. These results suggest that stretch enhancers are critical chromatin elements for coordinating cell type-specific regulatory programs and that sequence variation in stretch enhancers affects risk of major common human diseases. Overall design: Integrated analysis of islet chromatin modification and transcriptome data with those generated by the ENCODE project. NISC Comparative Sequencing Program","project":"SRP032812"}
{"number_samples":18,"species":"human","abstract":"Adenocarcinoma in situ (AIS) is considered an intermediate step in the progression of normal lung tissue to invasive adenocarcinoma.  However, the molecular mechanisms underlying this progression remain to be fully elucidated due to difficulties in obtaining and preserving clinical samples for downstream analyses.  Formalin fixation and paraffin embedding (FFPE) is a tissue preservation system that is widely used as a means for long-term storage.  Until now, challenges in working with FFPE have precluded using new RNA sequencing technologies (RNA-seq), which would help clarify some of the key pathways affected in the transition from normal to AIS to invasive adenocarcinoma.  Recent technological advances have made it possible to sequence RNA from archival tissues.  Also, isolation techniques including laser-capture micro-dissection provide the ability to select histopathologically distinct tissues, allowing researchers to study transcriptional variations between tightly juxtaposed cell and tissue types.  Utilizing these technologies and new alignment tools we examined differential expression of long intergenic non-coding RNAs and mRNAs across normal, AIS and invasive adenocarcinoma samples from six patients to identify possible markers of lung cancer progression. RNA extracted and sequenced from these 18 samples generated an average of 198 million reads per sample.  After alignment and filtering, uniquely aligned reads represented an average 35% of the total reads.  We detected differential expression of a number of lincRNAs and mRNAs when comparing normal to AIS, or AIS to invasive adenocarcinoma.  Of these, 5 lincRNAs and 31 mRNAs were consistently up- or down-regulated from normal to AIS and more so to invasive carcinoma.  We validated the up-regulation of two mRNAs and one lincRNA by RT-qPCR as proof of principle. Our findings indicate a potential role of not only mRNAs, but also lincRNAs in invasive adenocarcinoma.  We anticipate that our current findings will lay the groundwork for future experimental studies of candidate RNAs from FFPE samples to identify their functional roles in lung cancer. Overall design: RNA-Seq in 6 patients in norma lung, adenocarcinoma in situ of the lung, and invasive lung carcinoma","project":"SRP032833"}
{"number_samples":1,"species":"human","abstract":"Conventioal cytogenetic on 2 sinonasal sarcoma (SNS) cases showed t(2;4)(q37.1;q31.3) translocation previously. The aim of this whole transcriptome analysis is to determine potential candidate genes involved in this translocation. Overall design: Total RNA extracted from frozen cultured primary tumor cells from a SNS characterized at the cytogenetics level was subjected to Illumina whole transcriptome RNA-sequencing anslysis.","project":"SRP032922"}
{"number_samples":7,"species":"human","abstract":"Down syndrome (trisomy 21) is the most common viable chromosomal disorder with intellectual impairment and several other developmental abnormalities. Here, we report the generation and characterization of induced pluripotent stem cells (iPSCs) derived from monozygotic twins discordant for trisomy 21 in order to eliminate the effects of the variability of genomic background. The alterations observed by genetic analysis at the iPSC level and at first approximation in early development illustrate the developmental disease transcriptional signature of Down syndrome. Moreover, we observed an abnormal neural differentiation of Down syndrome iPSCs in vivo when formed teratoma in NOD-SCID mice, and in vitro when differentiated into neuroprogenitors and neurons. These defects were associated with changes in the architecture and density of neurons, astroglial and oligodendroglial cells together with misexpression of genes involved in neurogenesis, lineage specification and differentiation. Furthermore, we provide novel evidence that dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A) on chromosome 21 likely contribute to these defects. Importantly, we found that targeting DYRK1A pharmacologically or by shRNA results in a considerable correction of these defects. Overall design: mRNA-seq profiling of iPS cells (4 euploid and 3 trisomy 21) derived from fibroblasts of monozygotic twins discordant for trisomy 21","project":"SRP032928"}
{"number_samples":8,"species":"human","abstract":"Our data throw light upon the effect of WRN deficiency on gene expression and epigenomic modification, which indicates aging-associated changes from both genomic and epigenomic level. Overall design: It was compared between WRN+/+ and WRN-/- in hESCs and hMSCs that the gene expression landscapes and epigenetic modifications(H3K4me3, H3K27me3, H3K9me3 and  5-methylcytosine).","project":"SRP032942"}
{"number_samples":12,"species":"human","abstract":"Type 2 diabetes mellitus (T2DM) is a multi-factorial disease characterized by the inability of beta-cells in the endocrine pancreas to produce sufficient amounts of insulin to overcome insulin resistance in peripheral tissue. To investigate the function of miRNAs in T2DM, we sequenced the small RNAs of human islets cells from diabetic and non-diabetic organ donors and identified a cluster of miRNAs in an imprinted locus on human chromosome 14 to be dramatically down-regulated in T2DM islets. These miRNAs are highly and specifically expressed in human beta-cells. The down-regulation of this imprinted locus strongly correlates with increased methylation of its promoter in T2DM islets, providing evidence for an epigenetic modification that contributes to the pathogenesis of T2DM. Targets of the Chr 14q32 cluster of miRNAs were identified by high-throughput sequencing of cross-linked and immunoprecipitated RNA (HITS-CLIP) of Argonaute. We have also identified a unique class of sequences, termed chimeric reads, that represent an in vivo ligation of miRNAs and their targets while in complex with Argonaute, and which allow for the direct identification of miRNA:target relationships in vivo. Overall design: There are three experiments in this submission. All are in human islets or islet cell types. The first is a  comparison of miRNA levels in sorted alpha versus beta cells. There is one replicate for this experiment. The second  experiment is to measure the expression of miRNAs in whole islets as a function of glucose levels. There are three  levels and one replicate for each condition. The third exeriment is a comparison of whole islets taken from human  donors that were suspected/confirmed Type 2 diabetic or considered controls. There are 3 controls and 4 T2D samples.","project":"SRP032953"}
{"number_samples":2,"species":"human","abstract":"Profile of RNA expression in a C-33A cell line derived from an HPV negative cervical carcinoma in the presence or absence of HPV1 E2 expression. Overall design: mRNA profiles of C-33A cells in presence or absence of HPV1 E2 expression were generated by deep sequencing using Illumina GAIIx. Two samples (no replicates). One control and one experimental.","project":"SRP032989"}
{"number_samples":6,"species":"human","abstract":"We employed next generation sequencing to examine whether knocking down the steroid receptor RNA activator (SRA) gene significantly affect the expression levels of certain genes in MCF-7 cells. MCF-7 cells were transfected with either a pool of four non-target control siRNAs or a pool of four SRA siRNAs for 32 hrs. 157 million reads were generated from triplicate samples of the control group; 151 million reads were generated from triplicate samples of the SRA knockdown group. Six genes were identified as significantly changed in the expression levels with the cutoff of q value = 0.05, fold change = 0.5 or = 2, and reads per kilobase per million mapped reads (RPKM) = 1. However, except for SRA itself, the other five genes were shown by real-time PCR to be only affected by one siRNA in the SRA siRNA pool. Further analysis of this dataset with different cuttoff setting may reveal true SRA-regulated genes in MCF-7. Overall design: MCF-7 cells were cultured in high glucose DMEM with 10% fetal bovine serum, 2 mM Glutamax-1, 100 units/ml penicillin and 100 µg/ml streptomycin. ON-TARGETplus SMARTpool for human SRA (Thermo Scientific, L-027192-00-0005) was used to knockdown SRA (siSRA) and ON-TARGETplus Non-targeting Pool Thermo Scientific, D-001810-10-05) was used as a negative control (siCtrl). A total of 25 nM siRNA was transfected in 6-well dishes using Lipofectamine™ RNAiMAX Reagent (Life Technologies, Invitrogen) following the manufacturer’s recommendations. Polyadenylated RNA was purified from the cells 32 hrs after transfection. cDNA libraries were prepared and double-stranded cDNA was fragmented using DNase I according to Illumina specifications, prior to adaptor ligation. Sequencing libraries were amplified and sequenced using an Illumina HiSeq 2000 sequencer.","project":"SRP032999"}
{"number_samples":2,"species":"human","abstract":"To understand the funtion of Colorectal cancer GWAS results, we perform a comprehensive analysis using biofeatures of HCT116 colon cancer cell line and got a list of risk-asscociated SNP. Risk-associated SNP are likely exerting their effects through promoters or enhancer. In order to understand the importance of the genes with risk-associated SNP in their promoters and enhancers' putatively targeted genes,  we did a comparison of these genes between HCT116 colon cancer cell and normal colon and try to understand their function Overall design: Two biological replicates of HCT116 were compared to the data of two normal colon samples already deposited in GEO (GSM1010974 and GSM1010942).","project":"SRP033057"}
{"number_samples":8,"species":"human","abstract":"Reprogramming somatic cells to induced pluripotent stem cells (iPSCs) sets their identity back to an embryonic age. This presents a fundamental hurdle for modeling late-onset disorders using iPSC-derived cells. We therefore developed a strategy to induce age-like features in multiple iPSC-derived lineages and tested its impact on modeling Parkinson’s disease (PD). We first describe markers that predict fibroblast donor age and observed the loss of these age-related markers following iPSC induction and re-differentiation into fibroblasts. Remarkably, age-related markers were readily induced in iPSC-derived fibroblasts or neurons following exposure to progerin including dopamine neuron-specific phenotypes such as neuromelanin accumulation. Induced aging in PD-iPSC-derived dopamine neurons revealed disease phenotypes requiring both aging and genetic susceptibility such as frank dendrite degeneration, progressive loss of tyrosine-hydroxylase expression and enlarged mitochondria or Lewy body-precursor inclusions. Our study presents a strategy for inducing age-related cellular properties and enables the modeling of late-onset disease features. Overall design: Induced pluripotent stem cell-derived midbrain dopamine neurons from a young and old donor overexpressing either GFP or Progerin.","project":"SRP033078"}
{"number_samples":15,"species":"human","abstract":"Objectives: Idiopathic pulmonary fibrosis (IPF) is a complex disease in which a multitude of proteins and networks are disrupted. Interrogation of genome-wide transcription through RNA sequencing (RNA-Seq) enables the determination of genes whose differential expression is most significant in IPF, as well as the detection of alternative splicing events which are not easily observed with traditional microarray experiments. Methods: Messenger RNA extracted from 8 IPF lung samples and 7 healthy controls was sequenced on an Illumina HiSeq. Analysis of differential expression and exon usage was performed using Bioconductor packages. The gene periostin was selected for validation of alternative splicing by quantitative PCR, and pathway analysis was performed to determine enrichment for differentially expressed and spliced genes. Results: There were 873 genes differentially expressed in IPF (FDR 5%), and 440 unique genes had significant differential splicing events (FDR 5%). In particular, cassette exon 21 of the gene periostin was significantly more likely to be spliced out in IPF samples (adj pval = 2.06e-09), and this result was confirmed by qPCR (Wilcoxon pval = 3.11e-4). We also found that genes close to SNPs in the discovery set of a recent IPF GWAS were enriched for genes differentially expressed in our data, including genes like mucin5B and desmoplakin which have been previously associated with IPF. Conclusions: There is significant differential splicing and expression in IPF lung samples as compared with healthy controls. We found a strong signal of differential cassette exon usage in periostin, an extracellular matrix protein whose increased gene-level expression has been associated with IPF and its clinical progression, but for which differential splicing has not been studied in the context of IPF. Our results suggest that alternative splicing of periostin and other genes may be involved in the pathogenesis of IPF. Overall design: mRNA sequencing of 8 IPF and 7 control lung tissue samples.","project":"SRP033095"}
{"number_samples":10,"species":"human","abstract":"The goal was to catalogue mutations in mantle cell lymphoma cell lines.","project":"SRP033098"}
{"number_samples":14,"species":"human","abstract":"SF3B1 mutations in Breast Cancer","project":"SRP033115"}
{"number_samples":14,"species":"human","abstract":"We used 2', 3'-cyclic phosphate cDNA synthesis and Illumina sequencing to identify and quantify metal-ion-independent endoribonuclease cleavage sites in host and viral RNAs. Overall design: To make cDNA libraries, we exploited the 2’, 3’-cyclic phosphate at the end of RNA fragments produced by RNase L and other metal-ion-independent endoribonucleases. Using Arabidopsis thaliana tRNA ligase, RNA fragments with 2’, 3’-cyclic phosphates were covalently attached to defined RNA linkers containing an 8-base long unique molecular identifier (UMI) sequence. Libraries prepared in this manner contain cDNA derived exclusively from RNA fragments with 2', 3'-cyclic phosphates. The UMI sequence allows for detailed quantitation. We optimized and validated 2’, 3’-cyclic phosphate cDNA synthesis and Illumina sequencing methods using viral RNAs cleaved with purified RNase L, viral RNAs cleaved with purified RNase A, and RNA from uninfected and poliovirus-infected HeLa cells.","project":"SRP033116"}
{"number_samples":13,"species":"human","abstract":"Using strand specific RNA-seq to assess the EBV transcriptome during reactivation of Akata cells, we found extensive bidirectional transcription extending across nearly the entire genome. Overall design: Illumina strand-specific RNA-seq of BCR-activated Akata cells at 9 time points","project":"SRP033117"}
{"number_samples":29,"species":"human","abstract":"Purpose: We performed RNA-Immunoprecipitation in Tandem (RIPiT) experiments against human Staufen1 (Stau1) to identify its precise RNA binding sites in a transcriptome-wide manner. To monitor the consequences of Stau1 binding in terms of target mRNA levels and ribosome occupancy, we modified the levels of endogenous Stau1 in cells by siRNA or overexpression and performed RNA-sequencing and ribosome-footprinting experiments. Staufen1 (Stau1) is a double-stranded RNA (dsRNA) binding protein implicated in mRNA transport, regulation of translation, mRNA decay and stress granule homeostasis. Here we combined RNA-Immunoprecipitation in Tandem (RIPiT) with RNase footprinting, formaldehyde crosslinking, sonication-mediated RNA fragmentation and deep sequencing to map Staufen1 binding sites transcriptome-wide. We find that Stau1 binds complex secondary structures containing multiple short helices, many of which are formed by inverted Alu elements in annotated 3'UTRs or in \"strongly distal\" 3'UTRs extending far beyond the canonical polyadenylation signal. Stau1 also interacts with both actively translating ribosomes and with mRNA coding sequences (CDS) and 3'UTRs in proportion to their GC-content and internal secondary structure-forming propensity.  On mRNAs with high CDS GC-content, higher Stau1 levels lead to greater ribosome densities, suggesting a general role for Stau1 in modulating the ability of ribosomes to elongate through secondary structures located in CDS regions. Overall design: We used HEK293 cells expressing near endogenous levels of wild-type Flag-Stau1 (65KDa isoform with an N-Terminal Flag tag). As a control we used a mutant version of Stau1 that is not functional for dsRNA binding. Formaldehyde crosslinking experiments and RNase footprinting experiments were done in two biological replicates. All RNASeq, Ribosome footprinting and PAS-Seq were done in two biological replicates.","project":"SRP033119"}
{"number_samples":24,"species":"human","abstract":"RNA-seqs followed by miRNA transfections (miR-124 and miR-155) into four different cell lines( HeLa, HEK293, Huh7, and IMR90). Overall design: There are two biological replicates of RNA-seqs per each miRNA transfection per each sample and there are corresponding mock transfections.","project":"SRP033131"}
{"number_samples":384,"species":"human","abstract":"Single-cell expression profiling by RNA-Seq promises to exploit cell-to-cell variation in gene expression to reveal regulatory circuitry governing cell differentiation and other biological processes.  Here, we describe Monocle, a novel unsupervised algorithm for ordering cells by progress through differentiation that dramatically increases temporal resolution of expression measurements. This reordering unmasks switch-like changes in expression of key regulatory factors, reveals sequentially organized waves of gene regulation, and exposes regulators of cell differentiation. A functional screen confirms that a number of these regulators dramatically alter the efficiency of myoblast differentiation, demonstrating that single-cell expression analysis with Monocle can uncover new regulators even in well-studied systems. Overall design: We selected primary human myoblasts as a model system of cell differentiation to investigate whether ordering cells by progress revealed new regulators of the process. We sequenced RNA-Seq libraries from each of several hundred cells taken over a time-course of serum-induced differentiation. Please note that this dataset is a single-cell RNA-Seq data set, and each cell comes from a capture plate.  Thus, each well of the plate was scored and flagged with several QC criteria prior to library construction, which are provided as sample characteristics; CONTROL indicates that this library is a off-chip tube control library constructed from RNA of approximately 250 cells and 'DEBRIS' indicates that the well contained visible debris (and may or may not include a cell). Libraries marked DEBRIS thus cannot be confirmed to come from a single cell.","project":"SRP033135"}
{"number_samples":6,"species":"human","abstract":"rnaseq of mcf7 overexpressing ron/msp and ron/msp-shmbd4","project":"SRP033199"}
{"number_samples":2,"species":"human","abstract":"Recent small RNA sequencing data has uncovered extensive modification of the 3’ end of mature microRNAs (miRNAs). This non-templated nucleotide addition can impact miRNA gene regulatory networks through the control of miRNA stability or by interfering with the repression of target mRNAs. The miRNA modifying enzymes responsible for this regulation remain largely uncharacterized. Here we describe the ability for two related terminal uridyl transferases (TUTases), Zcchc6 (TUT7) and Zcchc11 (TUT4), to 3’ mono-uridylate a specific subset of miRNAs involved in cell differentiation and Hox gene control. Zcchc6/11 selectively uridylate these miRNAs in vitro, and we biochemically define a bipartite sequence motif that is necessary and sufficient to confer Zcchc6/11 catalyzed uridylation. Depletion of these TUTases in cultured cells causes the selective loss of 3’ mono-uridylation of many of the same miRNAs. Interestingly, upon TUTase dependent loss of uridylation we observe a concomitant increase in non-templated 3’ mono-adenylation. Our results uncover the molecular basis for sequence specific miRNA mono-uridylation by Zcchc6/11, highlight the precise control of different 3’ miRNA modifications in cells, and have implications for miRNA regulation during development. Overall design: small RNA profiles in TUTases knock-down and control HeLa cells were generated by Illumina deep sequencing","project":"SRP033206"}
{"number_samples":2,"species":"human","abstract":"Type 2 diabetes mellitus (T2DM) is a complex disease characterized by the inability of the insulin-producing ß-cells in the endocrine pancreas to overcome insulin resistance in peripheral tissues. To determine if microRNAs are involved in the pathogenesis of human T2DM, we sequenced the small RNAs of human islets from diabetic and non-diabetic organ donors. We identified a cluster of miRNAs in an imprinted locus on human chromosome 14q32 that is highly and specifically expressed in human ß-cells and dramatically down-regulated in islets from T2DM organ donors. The down-regulation of this locus strongly correlates with hyper-methylation of its promoter. Using HITS-CLIP for the essential RISC-component Argonaute, we identified disease-relevant targets of the chromosome 14q32 microRNAs, such as IAPP and TP53INP1 that cause increased ß-cell apoptosis upon over-expression in human islets. Our results support a role for microRNAs and their epigenetic control by DNA methylation in the pathogenesis of T2DM. Overall design: Identification of miRNA-target interaction in human islets using HITS-CLIP, one mRNA library and one miRNA library","project":"SRP033239"}
{"number_samples":68,"species":"human","abstract":"The aim of our study was to discover a miR marker panel prognostic of 5-year survival in OSCC patients that may be utilized in parallel with the current clinical covariates. We assessed differential expression of miRNAs genome-wide via deep sequencing in 20 tumor tissue samples. We also attempted to identify deregulated miR expression signatures that may serve as the prognostic marker of cancer survival. Selected miR marker-based panel then may serve as a guide for selection of appropriate follow-up chemo/radiation treatment, significantly improving the clinical management of OSCC and the overall survival rate. Overall design: Identify miRs differentially expressed in the poor prognosis group compared to the good prognosis group","project":"SRP033248"}
{"number_samples":8,"species":"human","abstract":"SnowShoes-FTD, a fusion transcript discovery tool, was used to identify fusions in breast cancer cell lines using the RNA-Seq data Overall design: Total RNA extracted from cell lines. The total RNA was used for construction of RNA-Seq library for RNA-Sequencing.","project":"SRP033250"}
{"number_samples":2,"species":"human","abstract":"Vascular endothelial growth factor A (VEGF-A) is a master regulator of vascular development and function. Consequently, VEGF-A regulated gene expression programs have been under heavy research. In this study we study the transcriptional regulation of VEGF-A-responsive genes in primary aortic endothelial cells (HAEC) and vein endothelial cells (HUVEC) using genome wide global run-on sequencing (GRO-Seq). We show that half of VEGF-A induced genes display a paused phenotype under basal conditions and are thus poised for rapid gene activation. Promoters of paused genes are distinguished from those of non-paused by higher basal enrichment for active histone marks H3K4me3, H3K9ac and H3K27ac. We also show that nearly 100% of the VEGF-upregulated genes are induced at the level of elongation, whereas 38-53% are also induced at the level of initiation. We also report a comprehensive chromatin interaction map generated in HUVECs using tethered conformation capture (TCC) and characterize chromatin interactions in relation to transcriptional activity. We demonstrate that sites of active transcription are more likely to engage in chromatin looping and identify chromatin compartments enriched for VEGF-regulated genes. Cell-type specific transcriptional activity was also shown to reflect boundaries of chromatin interactions within the HOXA cluster. Collectively, these findings provide new insight into mechanisms behind VEGF-A-regulated transcriptional programs in endothelial cells. Overall design: ChIP-Seq (H3K4me2), GRO-Seq and HiC profiling was performed in HUVECs and HAECs.","project":"SRP033253"}
{"number_samples":144,"species":"human","abstract":"RNA sequencing of human leukemia Overall design: The goals of this project are to obtain a comprehensive study of mutations and gene expression in human acute myeloid leukemia (AML). Methods: AML cells were thawed. DNA and RNA (polyA) was extracted and sequences were obtained with an illumina HiSeq 2000 sequencer. Results are pending.","project":"SRP033266"}
{"number_samples":6,"species":"human","abstract":"Unraveling complex signaling programs animating developmental lineage-decisions is pivotal to differentiate human pluripotent stem cells (hPSC) into pure populations of desired lineages for regenerative medicine. Developmental signals are strikingly temporally dynamic: BMP and Wnt initially specify primitive streak (progenitor to endoderm) yet 24 hours later suppress endoderm and induce mesoderm. At lineage bifurcations we show mutually-exclusive embryonic lineages are segregated through cross-repressive signals: TGFß and BMP/MAPK duel to respectively specify pancreas versus liver from endoderm. Unilateral endodermal differentiation requires blockade of alternative fates at every stage, revealing a universal developmental strategy for efficient differentiation and anterior-posterior patterning of diverse hPSC lines into highly-pure endodermal populations. This culminated in hPSC-derived hepatic progenitors that, for the first time, engraft long-term in genetically-unconditioned mouse livers and secrete human albumin. Finally, thirty transcriptional and chromatin state maps capturing endoderm commitment revealed endodermal enhancers reside in an unanticipated diversity of \"pre-enhancer\" chromatin states before activation. Overall design: Endoderm RNA-seq and ChIP-seq data sets","project":"SRP033267"}
{"number_samples":13,"species":"human","abstract":"In contrast to the paucity of clinic GIST (Gastrointestinal stromal tumor; 0.0014 % happening rate), there is a more than twenty thousand times higher rate for miniGISTs, about ~30% of human population could be identified to have miniGISTs. Exploring the molecular differences between miniGISTs and overt GISTs will help us to understand the transformation mechanisms along with GIST progression, but C-kit and PDGFRA mutations analysis failed to identify the differences between these two groups.   Conclusion: miniGISTs have significant distinct gene expression profile with overt GISTs by both unsupervised and supervised analysis. Our finding indicated that c-kit and DOG1 expression are the very early events in the genesis of miniGIST and overt GIST, and miniGIST is a molecular distinct group with overt GIST. Overall design: Within our study, we collected six miniGISTs with diameter less than 1cm from autopsies and seven overt GISTs from clinic, performed the first system-wide gene expression profiling of miniGISTs and overt GISTs by 3’end RNA Sequencing, and further compared the expression profiles between these two groups.","project":"SRP033276"}
{"number_samples":9,"species":"human","abstract":"Metastasis is the major cause of death in cancer patients, yet the genetic/epigenetic programs that drive metastasis are poorly understood. Here, we report a novel epigenetic reprogramming pathway that is required for breast cancer metastasis. Concerted differential DNA methylation is initiated by activation of the RON receptor tyrosine kinase by its ligand, macrophage stimulating protein (MSP). Through PI3K signaling, RON/MSP promotes expression of the G:T mismatch-specific thymine glycosylase MBD4. RON/MSP and MBD4-dependent aberrant DNA methylation results in misregulation of a specific set of genes. Knockdown of MBD4 reverses methylation at these specific loci, and blocks metastasis. We also show that the MBD4 glycosylase catalytic residue is required for RON/MSP-driven metastasis. Analysis of human breast cancers using a set of specific genes that are regulated by RON/MSP through MBD4-directed aberrant DNA methylation revealed that this epigenetic program is significantly associated with poor clinical outcome. Furthermore, inhibition of Ron kinase activity with a new pharmacological agent prevents activation of the RON/MBD4 pathway and blocks metastasis of patient-derived breast tumor grafts in vivo. Overall design: Examination of 3 cell types.","project":"SRP033282"}
{"number_samples":6,"species":"human","abstract":"In this study we present the first genome-wide expression profiling of peripheral B cells by massive parallel RNA sequencing in patients with allergic asthma validating the discovery potential of this approach in allergy. Overall design: RNA-seq was used to asses expression differences in B CD19 Lymphocytes from house dust mite allergic patients and healthy controls.","project":"SRP033335"}
{"number_samples":16,"species":"human","abstract":"DAZAP1 was depleted in culterd HEK 293T cells using shRNA and the resulting poly A RNA were isolated  c-DNA library constructed and paired end sequenced on illumina Hi-seq 2000 platform the data was compared to a control shRNA depleted cell Overall design: Gene expression and splicing switches upon DAZAP1 knockdown","project":"SRP033336"}
{"number_samples":16,"species":"human","abstract":"Rationale: Asthma is a chronic inflammatory airway disease. The most common medications used for its treatment are ß2-agonists and glucocorticosteroids, and one of the primary tissues that these drugs target in the treatment of asthma is the airway smooth muscle. We used RNA-Seq to characterize the human airway smooth muscle (HASM) transcriptome at baseline and under three asthma treatment conditions. Methods: The Illumina TruSeq assay was used to prepare 75bp paired-end libraries for HASM cells from four white male donors under four treatment conditions: 1) no treatment; 2) treatment with a ß2-agonist (i.e. Albuterol, 1µM for 18h); 3) treatment with a glucocorticosteroid (i.e. Dexamethasone (Dex), 1µM for 18h); 4) simultaneous treatment with a ß2-agonist and glucocorticoid, and the libraries were sequenced with an Illumina Hi-Seq 2000 instrument. The Tuxedo Suite Tools were used to align reads to the hg19 reference genome, assemble transcripts, and perform differential expression analysis using the protocol described in https://github.com/blancahimes/taffeta Overall design: mRNA profiles obtained via RNA-Seq for four primary human airway smooth muscle cell lines that were treated with dexamethasone, albuterol, dexamethasone+albuterol or were left untreated.","project":"SRP033351"}
{"number_samples":1,"species":"human","abstract":"Human Transcriptome or Gene expression","project":"SRP033365"}
{"number_samples":5,"species":"human","abstract":"Poly(A) tails enhance the stability and translation of most eukaryotic messenger RNAs, but difficulties in globally measuring poly(A)-tail lengths have impeded greater understanding of poly(A)-tail function. Here we describe poly(A)-tail length profiling by sequencing (PAL-seq) and apply it to measure tail lengths of millions of individual RNAs isolated from yeasts, cell lines, Arabidopsis thaliana leaves, mouse liver, and zebrafish and frog embryos. Poly(A)-tail lengths were conserved between orthologous mRNAs, with mRNAs encoding ribosomal proteins and other 'housekeeping' proteins tending to have shorter tails. As expected, tail lengths were coupled to translational efficiencies in early zebrafish and frog embryos. However, this strong coupling diminished at gastrulation and was absent in non-embryonic samples, indicating a rapid developmental switch in the nature of translational control. This switch complements an earlier switch to zygotic transcriptional control and explains why the predominant effect of microRNA mediated deadenylation concurrently shifts from translational repression to mRNA destabilization. Overall design: 64 samples from a variety of species","project":"SRP033369"}
{"number_samples":73,"species":"human","abstract":"To understand the role of trans factors in the regulation of splicing and alternative splicing, we knocked-down 22 RNA-binding proteins previously shown or suspected to be involved in the regulation of splicing (“splicing factors”). We performed knockdown experiments in triplicate in the Hapmap lymphoblastoid cell line GM19238 using small interfering RNAs (siRNAs). RNA was extracted 72 hours after knockdown for RNA-seq library preparation. The following splicing factors were analyzed: ACIN1, ADAR, HNRPA2B1, HNRPF, HNRPH1, HNRPH2, HNRPK, HNRPL, HNRPR, PCBP2, PTBP1, RBM23, RBM39, RBMX, RNPS1, SRSF1, SRSF3, SRSF4, SRSF8, SRSF9, SRSF10 and SYNCRIP. Seven controls were generated with non-specific siRNA probes transfection. RNA-seq libraries were multiplexed and sequenced on 3 different flow-cells on an Illumina HiSeq 2000 sequencer (single-end 107 bp long reads). Overall design: 73 samples, including 3 replicates for each of the 22 knockdowns and 7 controls","project":"SRP033393"}
{"number_samples":4,"species":"human","abstract":"IMR90 cells were infected with pLNC-RAS:ER (from Jesus Gil lab) with retroviral gene transfer. Infected cells were drug selected G418. The cells were induced either with ethanol as control or with 100nM final conc 4-hydroxytamoxifen (sigma H7904) for ectopic expression of protein Overall design: We used RNA-Seq to detail the global programme of gene expression in human IMR90 oncogene induced senescence","project":"SRP033401"}
{"number_samples":9,"species":"human","abstract":"Neural stem cells (NSC) derived from human parthenogenic stem cells (hpSC) have been observed to show stronger positive functional effects than hpSC-derived dopaminergic neuron precursors (DAP) in treatment of induced Parkinson Disease in animal models. RNAseq of the two types of cells were normalized and analyzed to compare gene expression profiles. Overall design: cDNA library of hpsC, NSC and DAP triplicates were sequenced using Illumina HiSeq 2000. The sequence reads were mapped to hg19 reference genome and hits that passed quality filters were analyzed for differential expression.","project":"SRP033432"}
{"number_samples":1,"species":"human","abstract":"In mammals, the carboxy-terminal domain (CTD) of RNA polymerase (Pol) II consists of 52 conserved heptapeptide repeats containing the consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Post-translational modifications of the CTD coordinate the transcription cycle and various steps of mRNA maturation. Here we describe Tyr1 phosphorylation (Tyr1P) as a hallmark of initiating Pol II in mammalian cells, in contrast to what was described in yeast. Tyr1P is predominantly found in antisense orientation at promoters but is also specifically enriched at active enhancers. Mutation of Tyr1 to phenylalanine (Y1F) prevents the formation of the hyper-phosphorylated Pol IIO form, induces degradation of Pol II to the Pol IIB form and results in a lethal phenotype. Our results suggest that Tyr1P has evolved specialized and essential functions in higher eukaryotes associated with antisense promoter and enhancer transcription, and Pol II stability. Overall design: This study was performed in a human Raji cell line. It contains ChIP-seq data for H3K36me3 (two replicates), H3K4me1 (two replicates), H3K4me3 (two replicates), Pol II (three replicates), Ser2P (two replicates), Ser5P (two replicates), Ser7P (two replicates), Tyr1P 3D12 (two replicates) and Tyr1P 8G5 (one replicate).  MNase-experiment for nucleosomes was performed in paired-end sequencing on one replicate, 4 replicates for the input genomic DNA was used and one replicate was generated for the short strand specific RNA experiment.","project":"SRP033435"}
{"number_samples":20,"species":"human","abstract":"Amyotrophic lateral sclerosis (ALS) is a paralytic degenerative disease of the nervous system. In the SOD1 mouse model of ALS we found loss of the molecular and functional microglia signature associated with pronounced expression of miR-155 in SOD1 mice. We also found increased expression of miR-155 in the spinal cord of ALS subjects. Genetic ablation of miR-155 increased survival in SOD1 mice and reversed the abnormal microglial and monocyte molecular signature. In addition, dysregulated proteins in the spinal cord of SOD1 mice that we identified in human ALS spinal cords and CSF were restored in SOD1G93A/miR155-/- mice. Treatment of SOD1 mice with anti-miR-155 SOD1 mice injected systemically or into the cerebrospinal fluid prolonged survival and restored the microglial unique genetic and microRNA profiles. Our findings provide a new avenue for immune based therapy of ALS by targeting miR-155. Overall design: Total RNA was isolated from whole lumbar spinal cord homogenate from healthy control donors without known neurologic diseases and sporadic and familial ALS.","project":"SRP033464"}
{"number_samples":36,"species":"human","abstract":"Purpose: study the role of MALT1 auto-proteolysis in T cell receptor mediated activation of NF-kB. Methods: Jurkat cells were generated that express wild type MALT1, the auto-cleavage deficient MALT1-R149A mutant, the catalytic inactive MALT1-C464A mutant or the R149A-C464A double mutant (RACA). Expression of endogenous MALT1 was inactivated using TALEN technology for the Jurkat cells expressing MALT1-R149A (JDM-RA) and MALT1-C464A (JDM-CA). Illumina HISeq 2000 deep sequencing was performed to determine the mRNA profiles for MALT1, JDM-RA, JDM-CA and RACA cells in unstimulated conditions or after treatment with 75ng/ml PMA and 150 ng/ml ionomycin for 3 or 18 hrs. Results: PMA ionomycin stimulation of the MALT1 auto-cleavage defective JDM-RA cells fails to activate NF-kB-dependent transcription like for the MALT1 catalytic inactive JDM-CA cells and the double RACA mutant cells. Conclusion: MALT1 autoproteolysis is essential for transcription of NF-kB target genes Overall design: mRNA profiles of Jurkat expressing MALT1, MALT1-R149A, MALT1-C464A and MALT1-R149A-C464A after 0, 3 and 18 hours of stimulation with PMA and Ionomycin were generated by deep sequencing, in duplicate, using  Illumina HISeq 2000","project":"SRP033466"}
{"number_samples":10,"species":"human","abstract":"We sequenced the WGS/WGBS/Transcriptome/mircoRNA of a type 2 diabetes family","project":"SRP033491"}
{"number_samples":8,"species":"human","abstract":"We report that AUF1 modulates global mRNA stability and translation, in turn promoting the maintenance of DNA integrity. Overall design: Please see individual series. For AUF1 PAR-CLIP, the four isoforms of AUF1 (p37, p40, p42, and p45) tagged with a Flag epitope were expressed in HEK293 cells. For total RNA-Seq HEK293 cells were transfected with Control siRNA, AUF1 siRNA, Empty Vector, Flag-AUF1 p37, p40, p42, or p45 as well as WI-38 cells were collected at PDL 15 and 55 and also transfected with Control siRNA, AUF1 siRNA, HuR siRNA. For Ribo-Seq HeLa cells were transfected with Control siRNA, AUF1 siRNA, or HuR siRNA.","project":"SRP033498"}
{"number_samples":5,"species":"human","abstract":"We report that AUF1 modulates global mRNA stability and translation, in turn promoting the maintenance of DNA integrity. Overall design: Please see individual series. In short, for AUF1 PAR-CLIP, the four isoforms of AUF1 (p37, p40, p42, and p45) tagged with a Flag epitope were expressed in HEK293 cells. For total RNA-Seq HEK293 cells were transfected with Control siRNA, AUF1 siRNA, Empty Vector, Flag-AUF1 p37, p40, p42, or p45 as well as WI-38 cells were collected at PDL 15 and 55 and also transfected with Control siRNA, AUF1 siRNA, HuR siRNA. For Ribo-Seq HeLa cells were transfected with Control siRNA, AUF1 siRNA, or HuR siRNA.","project":"SRP033502"}
{"number_samples":3,"species":"human","abstract":"We report the sequencing of small RNAs present in the plasma of three normal subjects. In addition to microRNAs we identified abundant non-human small RNA sequences. The organisms from which these were derived were identified by BLAST searches with contigs assembled from the short sequences. The taxonomic profiles were very consistent between individuals, including plants and microbes reported previously in the microbiome, but in proportions distinct from other sites. The majority of bacterial reads were from the phylum Proteobacteria, whilst for 5 of 6 individuals over 90% of the more abundant fungal reads were from the phylum Ascomycota; of these over 90% were from the order Hypocreales. Most reads mapped to rRNA sequences and the presence of specific common sequences was confirmed by RT-PCR. In addition, extremely abundant small RNAs derived from human Y RNAs were detected. We conclude that a characteristic profile of a subset of the human microbiome can be obtained by sequencing small RNAs present in the blood. The origin and potential function of these molecules remains to be determined, but the specific profile is likely to reflect health status. This facile test has immense potential to provide biomarkers for the diagnosis and prognosis of human disease. Overall design: The profile of small RNAs present in the plasma of three normal subjects was determined","project":"SRP033505"}
{"number_samples":2,"species":"human","abstract":"Lysine Specific Demethylase 1 (LSD1, KDM1A) functions as a transcriptional corepressor through demethylation of histone 3 lysine 4 (H3K4), but has coactivator function on some genes through unclear mechanisms. We show that LSD1, interacting with CoREST, associates with and coactivates androgen receptor (AR) on a large fraction of androgen-stimulated genes. A subset of these AR/LSD1-associated enhancer sites have histone 3 threonine 6 phosphorylation (H3T6ph), and these sites are further enriched for androgen-stimulated genes. Significantly, despite its coactivator activity, LSD1 still mediates H3K4me2 demethylation at these androgen-stimulated enhancers. FOXA1 is also associated with LSD1 at AR regulated enhancer sites, and a FOXA1 interaction with LSD1 enhances binding of both proteins at these sites. These findings show LSD1 functions broadly as a regulator of AR function, that it maintains a transcriptional repression function at AR-regulated enhancers through H3K4 demethylation, and has a distinct AR-linked coactivator function mediated by demethylation of other substrates. Overall design: Determine the role of LSD1 in androgen signaling.","project":"SRP033511"}
{"number_samples":3,"species":"human","abstract":"Background: Eukaryotic cells express a complex layer of noncoding RNAs. An intriguing family of regulatory RNAs includes transcripts from the opposite strand of protein coding genes, so called natural antisense transcripts (NATs). Here, we test the hypothesis that antisense transcription triggers RNA interference and gives rise to endogenous short RNAs (endo-siRNAs). Methods/Results: We used cloned human embryonic kidney cells (HEK293) followed by short RNAseq to investigate the small genic RNA transcriptome. 378 genes gave rise to short RNA reads that mapped to exons of RefSeq genes. The length profile of short RNAs showed a broad peak of 20-24 nucleotides, indicative of endo-siRNAs. Collapsed reads mapped predominantly to the first and the last exon of genes (74%). RNAs reads were intersected with sequences occupied by RNAPolII or bound to Argonaute (AGO1  by crosslinking, ligation, and sequencing of hybrids, CLASH). In the first exon, 94% of the reads correlated with PolII occupancy with an average density of 130 (relative units); this decreased to 65%/20 in middle exons and 54%/12 in the last exon. CLASH reads mapping to multi-exon genes showed little distribution bias with an average of about 5 CLASH reads overlapping with 60% of the endo-siRNA reads. However, endo-siRNAs (21-25 nt) intersecting with CLASH reads were enriched at the 5'end and decreased towards the 3'end. We then investigated the 378 genes with particular focus on features indicative for short RNA production; however, found that endo-siRNA numbers did not correlate with gene structures that favor convergent transcription. In contrast, our gene set was found notably over-represented in the NATsDB sense/antisense group as compared to non-overlapping and non-bidirectional groups.  Moreover, read counts showed no correlation with the steady-state levels of the related mRNAs and the pattern of endo-siRNAs proved reproducible after an induced mutagenic insult. Conclusions: Our results suggest that antisense transcripts contribute to low levels of endo-siRNAs in fully differentiated human cells. A characteristic endo-siRNA footprint is being produced at sites of RNAPolII transcription which is also related to AGO1. This endo-siRNA signature represents an intriguing finding and its reproducibility suggests that the production of endo-siRNAs is a regulated process with potential homoeostatic impact. Overall design: Size selected RNASeq of 3 human embryonic kidney cell (HEK293) samples. 1 control and 2 samples exposed to 100 µg/ml ethyl methanesulfonate for 24 hrs.","project":"SRP033515"}
{"number_samples":185,"species":"human","abstract":"Heart failure is associated with high morbidity and mortality and its incidence increases worldwide. MicroRNAs (miRNAs) are potential markers and targets for diagnostic and therapeutic applications, respectively. We determined myocardial and circulating miRNA abundance and its changes in patients with stable and end-stage heart failure before and at different time points after mechanical unloading by a left ventricular assist device (LVAD) by small-RNA-sequencing. MiRNA changes in failing heart tissues partially resembled that of fetal myocardium. Consistent with prototypical miRNA–target-mRNA interactions, target mRNA levels were negatively correlated to changes in abundance for highly expressed miRNAs in heart failure and fetal hearts. The circulating small RNA profile was dominated by miRNAs, and fragments of tRNAs and small cytoplasmic RNAs. Heart- and muscle-specific circulating miRNAs (myomirs) increased up to 140-fold in advanced heart failure, which coincided with a similar increase in cardiac troponin I protein, the established marker for heart injury. These extracellular changes nearly completely reversed 3 months following initiation of LVAD support. In stable heart failure, circulating miRNAs showed less than 5-fold differences compared to normal, and myomir and cardiac troponin I levels were only captured near the detection limit. These findings provide the underpinning for miRNA-based therapies and emphasize the usefulness of circulating miRNAs as biomarkers for heart injury performing similar to established diagnostic protein biomarkers. Overall design: Total RNA isolated from human left ventricular myocardium of failing hearts due to dilated or ischemic cardiomyopathy before and after mechanical unloading by a left ventricular assist device, and fetal myocardium compared to non-failing postnatal myocardium was subjected to multiplexed small RNA-sequencing on the Illumina platform. mRNA gene expression data using Illumina HumanHT-12v4 beadarrays for a subset of the myocardial samples is available (GSE52601).","project":"SRP033566"}
{"number_samples":24,"species":"human","abstract":"Human pluripotent stem cells hold great potential for regenerative medicine, but available cell types have important limitations. While embryonic stem cells derived from fertilized embryos (IVF-ESCs) are considered the \"gold standard\" of pluripotency, they are allogeneic to potential recipients. Autologous induced pluripotent stem cells (iPSCs) are prone to epigenetic and transcriptional aberrations. To determine whether accumulation of such aberrations is intrinsic to somatic cell reprogramming or secondary to the reprogramming method, we generated a genetically matched collection of human IVF-ESCs, iPSCs, and ESCs derived by somatic cell nuclear transfer (SCNT; NT-ESCs), and subjected them to genome-wide genetic, epigenetic and transcriptional analyses. SCNT-based reprogramming is mediated by the full complement of oocyte cytoplasmic factors, thus closely recapitulating early embryogenesis. NT-ESCs and iPSCs derived from the same somatic donor cells contained comparable numbers of de novo copy number variations (CNVs), suggesting that the two reprogramming methods may not differ significantly in mutagenic or selective pressure. On the other hand, the DNA methylation and transcriptome profiles of NT-ESCs corresponded very closely to those of IVF-ESCs, while iPSCs differed markedly from IVF-ESCs and harbored residual DNA methylation patterns typical of parental fibroblasts, suggesting incomplete reprogramming. We conclude that human somatic cells can be faithfully reprogrammed to pluripotency by SCNT and are therefore ideal candidates for cell replacement therapies. Overall design: Duplicate cDNA libraries of two IVF-ESCs, three sendai produced iPSC lines, two retro-virus produced iPSC lines, four NT-ESCs, and the parental fibroblast line were sequenced using Illumina HiSeq 2000. The sequence reads were mapped to hg19 reference genome and hits that passed quality filters were analyzed for differential expression.","project":"SRP033569"}
{"number_samples":1,"species":"human","abstract":"To show the expression level of a potential oncogene or a tumor suppressor in colon cancer cells","project":"SRP033582"}
{"number_samples":1,"species":"human","abstract":"The histone variant macroH2A has been implicated in transcriptional repression, but the molecular mechanisms that contribute to global macroH2A-dependent genome regulation remain elusive. Using ChIP-seq coupled with transcriptional profiling in macroH2A knock-down cells (GSE53103) we demonstrate that singular macroH2A nucleosomes occupy transcription start sites of subsets of both expressed and repressed genes with opposing regulatory consequences. Specifically macroH2A nucleosomes mask repressor binding sites in expressed genes, but activator binding sites in repressed genes thus generating distinct chromatin landscapes limiting genetic or extracellular inductive signals. We show that composite nucleosomes containing macroH2A and NRF-1 are stably positioned on gene regulatory regions and can buffer the transcriptional noise typifying antiviral responses. In contrast, macroH2A nucleosomes without NRF-1 bind promoters weakly and mark genes with noisier gene expression patterns. Thus, the strategic position and stabilization of macroH2A nucleosomes in human promoters defines robust gene expression patterns. Overall design: ChIP-seq experiments were performed in human Namalwa B cells and Hela epithelial cells in order to investigate the function of specialized nucleosomes containing the macroH2A histone variant and the transcription factor NRF-1. Native ChIP, Native RecHIP or crosslink ChIP  were performed in replicates, sequenced and aligned to the reference genome (hg18). mRNA-seq experiments were performed in human Namalwa B cells using a strand-specific protocol. Please see manuscript for details.","project":"SRP033612"}
{"number_samples":2,"species":"human","abstract":"Ten eleven translocation (TET) enzymes catalyse the oxidative reactions of 5-methylcytosine (5mC) to promote the demethylation process. The reaction intermediate 5-hydroxymethylcytosine (5hmC) has been shown to be abundant in embryonic stem cells and tissues, but strongly depleted in human cancers. Genetic mutations of TET2 gene were associated with lleukemia, whereas TET1 downregulation has been shown to promote malignancy in breast cancer.  Here, we report that TET1 is downregulated in colon tumours from the initial stage. TET1 silencing in primary epithelial colon cells increase their cellular proliferation while its   re-­­expression in colon cancer cells inhibits their proliferation and the growth of tumour xenografts even at later stages. We found that TET1 binds and maintains hypomethylated the promoter of the DKK genes inhibitors of the WNT signalling to promote their expression. Downregulation of TET1 during colon cancer initiation leads to repression, by DNA methylation the promoters of the inhibitors of the WNT pathway resulting in a constitutive activation of the WNT pathway. Thus the DNA hydroxymethylation mediated by TET1 controlling the WNT signalling is a key player of tumour growth. These results provide new insights for understanding how tumours escape cellular controls Overall design: Transcriptome analysis of Caco-2 cell line expressing TET1 protein.","project":"SRP033646"}
{"number_samples":10,"species":"human","abstract":"RNA-seq of in vitro virus infections","project":"SRP033647"}
{"number_samples":6,"species":"human","abstract":"Comparison of gene expression profiles in the peripheral blood samples were collected from patients with H7N9 infection and healthy people","project":"SRP033696"}
{"number_samples":8,"species":"human","abstract":"miR-137 plays critical roles in the nervous system and tumor development. An increase in its expression is required for neuronal differentiation while its reduction is implicated in gliomagenesis. To evaluate the potential of miR-137 in glioblastoma therapy, we conducted genome-wide target mapping in glioblastoma cells by measuring levels of associations between PABP and mRNAs in cells transfected with miR-137 mimics vs. controls via RIPSeq. Impact on mRNA levels was also measured by RNASeq. By combining the results of both experimental approaches, 1468 genes were determined to be negatively impacted by miR-137; among them, 595 (40%) contain miR-137 predicted sites.  The most relevant targets include oncogenic proteins and players in neurogenesis like c-KIT, YBX1, AKT2, CDC42, CDK6 and TGFß2. Interestingly, we observed that several identified miR-137 targets are also predicted to be regulated by miR-124, miR-128 and miR-7, which are equally implicated in neuronal differentiation and gliomagenesis. We suggest that the concomitant increase of these four miRNAs in neuronal stem cells or their repression in tumor cells could produce a robust regulatory effect with major consequences to neuronal differentiation and tumorigenesis. Overall design: Identification of genes affected by miR137 transfection via RIP-Seq and RNA-Seq in U251 and U343 cells","project":"SRP033711"}
{"number_samples":62,"species":"human","abstract":"See \"Akula et al., Molecular Psychiatry in Press\". RNA-sequencing (RNA-seq) is a powerful technique to investigate the complexity of gene expression in the human brain. We used RNA-seq to survey the brain transcriptome in high-quality post-mortem dorsolateral prefrontal cortex from 11 individuals diagnosed with bipolar disorder (BD) and from 11 age- and gender-matched controls.  Deep sequencing was performed, with over 350 million reads per specimen. At a false-discovery rate of <5%, we detected 5 differentially-expressed (DE) genes and 12 DE transcripts, most of which have not been previously implicated in BD.  Among these, PROM1/CD133 and ABCG2 play important roles in neuroplasticity. We also show for the first time differential expression of long non-coding RNAs (lncRNAs) in BD.  DE transcripts include those of SRSF5 and RFX4, which along with lncRNAs play a role in mammalian circadian rhythms. The DE genes were significantly enriched for several Gene Ontology (GO) categories. Of these, genes involved with GTPase binding were also enriched for BD-associated SNPs from previous genome-wide association studies, suggesting that differential expression of these genes is not simply a consequence of BD or its treatment.  Many of these findings were replicated by microarray in an independent sample of 60 cases and controls. These results highlight common pathways for inherited and non-inherited influences on disease risk that may constitute good targets for novel therapies. Overall design: Brain transcriptome in bipolar disorder","project":"SRP033725"}
{"number_samples":5,"species":"human","abstract":"We identified miRNAs differentially regulated upon Salmonella infection by comparative deep-sequencing analysis of cDNA libraries prepared from the small RNA population (10–29 nt) of HeLa cells infected with Salmonella (20 hpi) and mock-treated cells. Considering that at a MOI of 25 Salmonella is internalized in only 10-15% of the HeLa cells, we separated the fraction of cells which had internalized Salmonella (Salmonella+) from the bystander fraction (Salmonella-) by fluorescence-activated cell sorting (FACS), and extended the analysis of miRNA changes to these samples. Interestingly, we observed that Salmonella infection induces a significant decrease in the expression of all the detected members of the miR-15 family Overall design: miRNA profiles of HeLa cells infected with Salmonella Typhimurium were generated by deep sequencing, using Illumina HiSeq2000.","project":"SRP034007"}
{"number_samples":4,"species":"human","abstract":"To have a global picture of the targets of the miR-15 family, we assessed transcriptome changes, by deep-sequencing, of HeLa cells transfected with 3 members of the miR-15 family (miR-15a, miR-16 or miR-503) or a control miRNA (cel-miR-231). We observed a very extensive overlap between the genes down-regulated by these 3 miRNAs, as expected for miRNAs belonging to the same family. Overall design: transcriptmic profiles of HeLa cells treated miR-15a, miR-16, miR-503 and control-miR  were generated by deep sequencing, using Illumina HiSeq2000.","project":"SRP034008"}
{"number_samples":6,"species":"human","abstract":"We evaluated the transcriptome changes induced by infection with Salmonella (20 hpi, MOI 100). Overall design: Transcriptmic profiles of HeLa cells infected with Salmonella Typhimurium were generated by deep sequencing, using Illumina HiSeq 2000.","project":"SRP034009"}
{"number_samples":4,"species":"human","abstract":"Liver stage of malaria parasite exports SLTRiP and PB268 to the cytosol of parasite infected host cell. To know the host genes perturbed by WT-PBANKA, SLTRiP-KO and PB268-KO parasite growth, we did transcriptomic sequencing of infected host cells.  We did mRNA sequencing of four samples for comparative analysis of WT and PB-knockout parasites infected host cells at 22 hours of post sporozoites infection. Overall design: mRNA profiles of Plasmodium PBANKA, PBSLTRiP-KO, PB268-KO parasite infected and uninfected HepG2 cells after  22hrs of sporozoites infections  were generated by deep sequencing using Illumina GAIIx.","project":"SRP034011"}
{"number_samples":3,"species":"human","abstract":"Micro (mi)RNAs are small non-coding RNAs with key regulatory functions. Recent advances in the field allowed researchers to identify their targets. However, much less is known regarding the regulation of miRNA themselves. The accumulation of these tiny regulators can be modulated at various levels during their biogenesis from the transcription of the primary transcript (pri-miRNA) to the stability of the mature miRNA. Here, we studied the importance of the pri-miRNA secondary structure for the regulation of mature miRNAs accumulation. To this end, we used the Kaposi’s sarcoma herpesvirus, which encodes a cluster of twelve pre-miRNAs. Using small RNA profiling and quantitative northern blot analysis, we measured the absolute amount of each mature miRNAs in different cellular context. We found that the difference in expression between the least and most expressed viral miRNA could be as high as 60-fold. Using high-throughput selective 2’-hydroxyl acylation analyzed by primer extension (hSHAPE), we then determined the secondary structure of the long primary transcript. We found that highly expressed miRNAs derived from optimally structured regions within the pri-miRNA. Finally, we confirmed the importance of the local structure by swapping stem-loops for highly and lowly expressed miRNAs, which resulted in a perturbed accumulation of the mature miRNA. Overall design: Examination of sRNA profiles in 3 independent B cell lines expressing KSHV miRNAs or infected with KSHV, without replicate","project":"SRP034013"}
{"number_samples":2,"species":"human","abstract":"Activation of the PI3K pathway in estrogen receptor a (ER)-positive (+) breast cancer is associated with reduced ER expression and activity, luminal B subtype, and poor outcome.  PTEN is a negative regulator of the PI3K pathway typically lost in ER-negative (-) breast cancer.  To clarify the effect of PTEN down-regulation on the response of ER+/HER2- breast cancer to endocrine therapy, we established reduced PTEN cell models using inducible knockdown.  We found that only moderate PTEN reduction is sufficient to enhance PI3K signaling, generate a gene signature associated with luminal B subtype, and cause endocrine resistance.  Combining endocrine therapy with mTOR, AKT, or MEK inhibitors improves antitumor activity, but the efficacy varies by type of endocrine therapy and the specific inhibitor.  Fulvestrant plus an AKT inhibitor is the most potent combination when PTEN is reduced, inducing apoptosis and tumor regression.  This combination deserves further study in patients with PI3K pathway activation. Overall design: Gene expression profiles (by RNA-seq) were taken of a cell model with or without reduced PTEN (using inducible knockdown).","project":"SRP034017"}
{"number_samples":8,"species":"human","abstract":"Gene splicing requires three basal genetic elements; the 3’ and 5’ splice sites and the branchpoint to which the 5’ intron termini is ligated to form a closed lariat during the splicing reaction. The 5’ and 3’ splice sites that define exon boundaries have been widely identified, revealing pervasive transcription and splicing of human genes. However, the locations of the third requisite element, the branchpoint, are still largely unknown. Here we employ two complementary approaches, targeted RNA sequencing and exoribonuclease digestion, to distil sequenced reads that traverse the lariat junction and, via non-conventional alignment, locate human branchpoint nucleotides. Alignments identify 88,748 branchpoints that correspond to 20% of known introns, with 76% supported by diagnostic sequence mismatch errors. This affords a first genome-wide analysis of branchpoints, describing their distribution, selection, and the existence of a diverse array of overlapping sequence motifs with distinct usage, evolutionary histories, and co-variation with distal splicing elements. The overlap of branchpoints with noncoding human genetic variation also indicates a notable contribution to disease. This annotation and analysis incorporates branchpoints into transcriptomic research and reflects a core role for this element in the regulatory code that governs gene splicing and expression. Overall design: RNaseR validation of branchpoint nucleotides","project":"SRP034158"}
{"number_samples":2,"species":"human","abstract":"RNAseq replicate in Proliferating and Senescent IMR90s Overall design: We used RNAseq to examine RNA levels in human IMR90 replicative senescence (RS)","project":"SRP034163"}
{"number_samples":10,"species":"human","abstract":"RNA m5C methylation profile of MCF10A and MDA486 by using MeRIP-Seq protocol Overall design: Immunoprecipitation of Methylated mRNA at Cytosine (m5C) residues: Affinity purified of anti-methyl cytosine (m5C) polyclonal antibody 7ug (Zymo Research, Catalog#A3001-50) was conjugated with protein-A magnetic beads for 2 h at 4°C in end to end rotator. After that, conjugated beads were extensively washed with RNA immunoprecipitation (RIP) wash buffer to remove unbound antibody. Fragmented 25 ug polyA RNA (mRNA) was incubated with m5C conjugated beads for overnight at 4°C in in the rotating platform in RIP buffer. RIP was done using Megna RNA Immunoprecipitation kit (Millipore, Catalog#17-700). m5C mRNA-immune bead complex was treated with proteinase K buffer to release m5C mRNA from the conjugated antibody. To isolate m5C, mRNA was treated with phenol:chloroform:isoamyl and mixed with 400 ul of chloroform, which was centrifuged at 14000 rpm for 10 minutes to separate aqueous phase. The aqueous phase was ethanol precipitated at -80°C for overnight, to get m5C mRNA. This precipitated m5C mRNA pellet was washed twice with 70% ethanol and air dried. Finally, m5C mRNA pellet was dissolved in nuclease free Water. The m5C mRNA integrity and conentration was quantified by bioanalyzer (Agilent) and Qubit 2.0 flurometer (Invitrogen). The fragmented mRNA was used by following TruSeq RNA Sample Preparation Guide to develop RNA-Seq library for sequencing.","project":"SRP034528"}
{"number_samples":4,"species":"human","abstract":"IMR90 cells were passaged until replicative senescence and compared with proliferating cells. Overall design: We used RNA-Seq to detail the global programme of gene expression in human IMR90 replicative induced senescence","project":"SRP034541"}
{"number_samples":30,"species":"human","abstract":"We reported a new method of single-cell universal poly(A)-independent RNA sequencing (SUPeR-seq) to obtain single cell whole transcriptome analysis that covers both poly(A) plus and poly(A) minus RNA species. We built libraries of single mouse ESC , 10pg, 100pg and 1ng diluted mouse ES total RNA, single HEK293T cell and mouse early embryos before 4-cell stage. We analyzed these data using bioinformatics and find a good coverage of non-polyadenylated RNAs, circular RNA for example. to make a comparison,we also used mouse ES cells and HEK293T cells to build libraries using the protocol publish by F.Tang in 2009(sample name Tang2009_*). We also de novo assembled new genes in mouse ESCs and early embryo samples, further validated known and novel zygotic genes with a-Aminatine treated 2-cell embryos(sample name SUPeR-seq_a-AM_treated_2-cell*). Overall design: 3 Single mouse ES cells, 7 single HEK293T cells, 8 diluted mouse ES total RNA samples and 19 mouse embryo samples are performed using SUPeR-seq to evaluate its ability to detect genes in single RNAs and to cover poly(A)- RNA species.","project":"SRP034543"}
{"number_samples":4,"species":"human","abstract":"We elucidate a neurological syndrome affecting both the PNS and CNS defined by CLP1 mutations that impair tRNA splicing Overall design: Identification and biochemical characterization of mutant CLP1 in human patients","project":"SRP034547"}
{"number_samples":24,"species":"human","abstract":"microRNAs (miRNAs) are small noncoding RNAs that post-transcriptionally regulate gene expression by targeting specific mRNAs. Altered expression of circulating miRNAs have been associated with age-related diseases including cancer and cardiovascular disease. Although we and others have found an age-dependent decrease in miRNA expression in peripheral blood mononuclear cells (PBMCs), little is known about the role of circulating miRNAs in human aging. Here, we examined miRNA expression in human serum from young (mean age 30 years) and old (mean age 64 years) individuals using next generation sequencing technology and real-time quantitative PCR. Of the miRNAs that we found to be present in serum, three were significantly decreased in 20 older individuals compared to 20 younger individuals: miR-151a-5p, miR-181a-5p and miR-1248. Consistent with our data in humans, these miRNAs are also present at lower levels in the serum of elderly rhesus monkeys. In humans, miR-1248 was found to regulate the expression of mRNAs involved in inflammatory pathways and miR-181a was found to correlate negatively with the pro-inflammatory cytokines IL-6 and TNFa and to correlate positively with the anti-inflammatory cytokines TGFb and IL-10.  These results suggest that circulating miRNAs may be a biological marker of aging and could also be important for regulating longevity. Identification of stable miRNA biomarkers in serum could have great potential as a noninvasive diagnostic tool as well as enhance our understanding of physiological changes that occur with age. Overall design: Examination of microRNAs isolated from human serum from 11 young (mean age 30 yrs) and 11 old (mean age 64 yrs) individuals and from peripheral blood mononuclear cells from one young (30 yr) and one old (64 yr) individual.","project":"SRP034586"}
{"number_samples":10,"species":"human","abstract":"we performed RNA sequencing analysis using 10 tissue samples from human prostate and evaluated efficiency and accuracy of eRNA on mRNA-seq data analysis. Overall design: We sequenced mRNAs from the 10 human tissue samples. After that, we identified mRNAs in these samples against known human genes.","project":"SRP034592"}
{"number_samples":4,"species":"human","abstract":"The three-dimensional (3D) organization of the genome within the cell nucleus contributes to cell-specific gene expression in different cell types1. High-throughput 3C–derived methods have revealed that the genome is segmented into contiguous topologically associating domains (TADs), which help to orchestrate gene expression changes during differentiation and development2-5. Using ChIP-Seq, Hi-C and 3D modelling techniques, we reveal that TADs regulate the rapid gene expression changes induced by progestin in T47D breast cancer cells. In response to the hormone, TADs maintain their borders and operate as discrete regulatory units in which the majority of the genes are either transcriptionally activated or repressed. Additionally, the epigenetic signatures of the TADs are coordinately modified by hormone in correlation with the transcriptional changes. Hormone-induced changes in gene activity and chromatin remodelling are accompanied by structural changes that are distinct for activated or repressed TADs. Integrative 3D modelling revealed that TADs are structurally expanded if active and compacted if repressed, and that this is accompanied by differential changes in accessibility. We thus propose that TADs function as “regulons” to enable spatially proximal genes to be coordinately transcribed in response to hormones. Overall design: T47D-MTVL human breast cancer cells were incubated with the progestin R5020 for different times at 37ºC and prepared for ChIP-Seq or Hi-C according published protocols","project":"SRP034599"}
{"number_samples":9,"species":"human","abstract":"Knockdowns of c-JUN and JUND had opposite effects on PC3 prostate cell migration. We predicted that c-JUN and JUND control the same set of cell migration genes, but in opposite directions. To test this hypothesis, mRNA with expression changes in c-JUN and JUND knockdown PC3 cell lines were compared to mRNA levels in control (luciferase knockdown) PC3 cells by RNA-seq. Overall design: mRNA profiles of luciferase knockdown (WT),  c-Jun knockdown, and Jun-D knockdown in PC3 cells were generated using deep sequencing, in triplicate, using Illumina HiSeq. Knockdowns were stable shRNA expression from a lentiviral construct selected with puromycin.","project":"SRP034601"}
{"number_samples":12,"species":"human","abstract":"Heterogeneity in pluripotent cells marks a metastable state where cells may drift between native and lineage-primed populations.  While the role for these heterogeneities are unclear, they may reflect the dynamic equilibriums of signaling networks and have a direct effect on differentiation potentialities.  Here, we report the role of the cell cycle in establishing heterogeneity of human pluripotent stem cells.  By utilizing the FUCCI cell cycle indicator system coupled to fluorescent activated cell sorting (FACS), we have uncovered that the cell cycle drives heterogeneity at the epigenetic, transcriptional and post-transcriptional levels.  Our data show widespread dynamics in 5-hydroxymethylcytosine (5hmC) during the cell cycle.  Furthermore, transcript profiling by RNA-sequencing identified >500 genes that were cell cycle-regulated, of which the largest cohort of genes were transcriptional regulators. In sum, we demonstrate the role of the cell cycle in coordinating cellular transitions between metastable states in pluripotent stem cells. Overall design: mRNA sequencing of the cell cycle phases; early & late G1, S and G2/S from human ES cells in triplicate.","project":"SRP034606"}
{"number_samples":14,"species":"human","abstract":"LED, a long non-coding RNA induced by Nutlin-3a, was found to activate enhancer RNAs, one of which was found in an intron of CDKN1A (p21). Overall design: Impact of expression of LED was assessed in MCF7 cells using RNAseq and GROseq. Chromatin binding of LED was assessed using ChIRPseq.","project":"SRP034626"}
{"number_samples":11,"species":"human","abstract":"We profile the expression pattern of human pulmonary microvascular endothelial cells (HPMECs) at different time points of hypoxic stress. Through mRNA-seq, we identify functional waves of minor gene up-regulation at 8 and 24h hypoxia exposure followed by a massive wave of transcriptional activation after 48 hours. By weighted gene co-expression network analysis, we identify hub genes that likely play central roles in hypoxia transcription program. Strikingly, these hub genes included a prominent group of lincRNAs, suggesting non-coding RNAs may also have pivotal roles in the hypoxia regulatory circuit.  HPMECs share a core hypoxia signature profile, but with some notably differences, indicating a portion of HPMECs hypoxia response is cell-specific. Collectively, our study comprehensive surveys the hypoxia transcriptome, and provides insight into the temporal dynamics of hypoxia transcriptional response. Overall design: Time-course expression profiling of HPMECs exposed to hypoxia","project":"SRP034634"}
{"number_samples":2,"species":"human","abstract":"ERa is essential for the anti-proliferative response of breast cancer cells not only to estrogen antagonists, but also to estrogen withdrawal by means of aromatase inhibitors. We explored here one of the simplest explanation for this, consisting in the possibility that ERa may have a wide genomic function in absence of ligands. The genomic binding of ERa in the complete absence of estrogen was then studied using hormone-dependent MCF7 cells, by chromatin immunoprecipitation sequencing. From these data, 4.2K highly significant binding events were identified, which were further confirmed by comparing binding events in cells expressing ERa to cells silenced for ERa. Apo-ERa binding sites were distributed close to genes with functions associated to cell growth and epithelial maintenance and show significant overlap with binding of other transcription factors important for luminal epithelial breast cancer. Interestingly, we found that upon ERa silencing cognate gene transcription in absence of estrogen is downregulated and this is accompanied by increased H27Kme3 at ERa binding sites. RNA-Seq experiments showed that unliganded ERa controls basal transcription widely, including both coding and noncoding transcripts. Genes affected by ERa silencing can be easily functionally related to mammary epithelium differentiation and maintenance, especially when considering downregulated genes. Additional functions related to inflammatory and immune response was observed. Our data unravel unexpected actions of ERa in breast cancer cells and provide a novel framework to understand success and failure of hormone therapy in breast cancer. Overall design: Examination of unligandend estrogen receptor alpha (aERa) DNA interactions in control and aERa siRNA treated MCF7 cells.","project":"SRP034657"}
{"number_samples":3,"species":"human","abstract":"This experiment sought to determine the genome-wide interactome of CTCF in human cells. Overall design: PAR-CLIP seq for CTCF was performed in U2OS cells in 2 biological replicates","project":"SRP034666"}
{"number_samples":2,"species":"human","abstract":"Our goal is to identify high-confidence candidate genes associated with sub-threshold QT interval loci. We predicted enhancer-promoter targets using two different methods. Overall design: 1) We used the correlation in activity between enhancers and transcribed genes across 59 human cell lines and tissues to identify significantly correlated enhancer-promoter pairs that represent potential regulatory interactions. 2) We used chromosome conformation capture followed by high-throughput sequencing (4C-seq) to probe physical enhancer-promoter interactions.","project":"SRP034672"}
{"number_samples":2,"species":"human","abstract":"As the leading cause of food-borne illness in the world, Salmonella have evolved a sophisticated machinery to alter host cell function to promote virulence and survival.In this study, we compare production of non-coding RNAs between Salmonella-infected cells and mock infection cells. Using Solexa deep sequencing, we detected a panel of 19-24nt Salmonella-derived non-coding RNA fragments with considerable large copy numbers in human colonic epithelial HT-29 cells following Salmonella infection.The fragment with the highest copy number, Sal-1, was further validated by both quantitative reverse transcription polymerase chain reaction (qRT-PCR) and northern blot. The generation of Sal-1 requires the infection of host cells by Salmonella, and the processing of the Sal-1 “primary” or “precursor” to the mature Sal-1 in Salmonella-infected cells is Dicer-independent but Argonaute 2 (Ago2)-dependent. Functionally, Sal-1 suppresses the expression of colonic epithelial cell endogenous inducible nitric oxide synthase (iNOS) via targeting its open reading frame and thus reduces the bacterial resistance of host cells. Overall design: Screening and identification of Salmonella-encoded microRNA-like RNA fragments","project":"SRP034691"}
{"number_samples":8,"species":"human","abstract":"MicroRNAs have been implicated in various skin cancers, including melanoma, squamous cell carcinoma, and basal cell carcinoma; however, the expression of microRNAs and their role in Merkel cell carcinoma (MCC) have yet to be explored in depth. To identify microRNAs specific to MCC (MCC-miRs), next-generation sequencing (NGS) of small RNA libraries was performed on different tissue samples including MCCs, other cutaneous tumors, and normal skin. Comparison of the profiles identified several microRNAs upregulated and downregulated in MCC. For validation, their expression was measured via qRT-PCR in a larger group of MCC and in a comparison group of non-MCC cutaneous tumors and normal skin. Eight microRNAs were upregulated in MCC: miR-502-3p, miR-9, miR-7, miR-340, miR-182, miR-190b, miR-873, and miR-183. Three microRNAs were downregulated: miR-3170, miR-125b, and miR-374c. Many of these MCC-miRs, with the miR-183/182/96a cistron in particular, have connections to tumorigenic pathways implicated in MCC pathogenesis. In situ hybridization confirmed that the highly expressed MCC-miR, miR-182, is localized within tumor cells. Furthermore, NGS and qRT-PCR reveals that several of these MCC-miRs are highly expressed in the patient-derived MCC cell line, MS-1.  These data indicate that we have identified a set of MCC-miRs with high implications for MCC research. Overall design: To identify microRNAs specific to Merkel cell carcinoma (MCC) next-generation sequencing (NGS) of small RNA libraries was performed on different tissue samples including MCCs, other cutaneous tumors, and normal skin","project":"SRP034698"}
{"number_samples":15,"species":"human","abstract":"Human erythroblasts purified from cord blood were cultured in vitro and FACS-sorted into five highly purified populations representing distinct differentiation stages:  proerythroblasts, early basophilic erythroblasts, late basophilic erythroblasts, polychromatophilic erythroblasts, and orthochromatophilic erythroblasts. The methods for culture and sorting experiments are given in Hu et al. 2013. For each RNA-seq library, RNA was isolated from 1x 106 sorted human erythroblasts using RNeasy Plus Mini kits (Qiagen). Libraries were then prepared using Illumina TruSeqTM RNA kits to obtain 50 nt reads. Collaborators at the New Your Blood Center were responsible for erythroblast culture, FACS purification of erythroblast populations, and acquisition of RNA-seq data. Collaborators at U.C. Berkeley and Lawrence Berkeley National Laboratory performed data analysis and experimental validation of alternative splicing in erythroblasts. Results: Differentiating erythroblasts execute a dynamic alternative splicing program that is enriched in genes affecting cell cycle, organelle organization, chromatin function, and RNA processing. Alternative splicing plays a major role in regulating gene expression to ensure synthesis of appropriate proteome at each stage as the cells remodel in preparation for production of mature red cells. Overall design: Erythroid differentiation stage-specific transcriptome analysis was performed by RNA-seq analysis of highly purified erythroblast populations","project":"SRP034711"}
{"number_samples":52,"species":"human","abstract":"Directed differentiation of cells in vitro is a powerful approach for dissection of developmental pathways, disease modeling and regenerative medicine, but analysis of such systems are complicated by heterogeneous and asynchronous cellular responses to differentiation-inducing stimuli. To enable deep characterization of heterogeneous cell populations, we developed an efficient digital gene expression profiling protocol that enables surveying of mRNA in thousands of single cells at a time. We then applied this protocol to profile 11,116 cells collected during directed differentiation of human adipose-derived stem/stromal cells. The resulting data reveals the major axes of cell-to-cell variation within and between time points and suggests a link between incomplete adipogenesis in vitro and adipocyte dysfunction in vivo. Overall design: High-throughput single cell RNA-seq method applied to human adipose tissue-derived stromal/stem cells during differentiation towards an adipogenic fate","project":"SRP034712"}
{"number_samples":47,"species":"human","abstract":"In this study we aimed to assess technical variability associated with globin depletion in addition to assessing general technical variability in RNA-Seq from whole blood derived samples. We compared technical and biological replicates having undergone globin depletion or not and found that globin depletion removed approximately 80% of globin transcripts, improved the correlation of technical replicates, allowed for reliable detection of thousands of additional transcripts and generally increased transcript abundance measures. Overall design: Peripheral whole blood transcriptome assessed by RNA-Seq on Illumina HiSeq 2000 in 6 healthy individuals and 6 pooled samples, either globin depleted or not.","project":"SRP034732"}
{"number_samples":14,"species":"human","abstract":"High expression of the ETS family transcription factor ERG is associated with poor clinical outcome in acute myeloid leukemia (AML) and acute T-cell lymphoblastic leukemia (T-ALL). In murine models, high ERG expression induces both T-ALL and AML. However, no study to date has defined the effect of high ERG expression on primary human hematopoietic cells. In the present study, human CD34+ cells were transduced with retroviral vectors to elevate ERG gene expression to levels detected in high ERG AML. RNA sequencing was performed on purified populations of transduced cells to define the effects of high ERG on gene expression in human CD34+ cells. Integration of the genome-wide expression data with other data sets revealed that high ERG drives an expression signature that shares features of normal hematopoietic stem cells, high ERG AMLs, early T-cell precursor-ALLs and leukemic stem cell signatures associated with poor clinical outcome. Functional assays linked this gene expression profile to enhanced progenitor cell expansion. These results support a model whereby a stem cell gene expression network driven by high ERG in human cells enhances the expansion of the progenitor pool, providing opportunity for the acquisition and propagation of mutations and the development of leukemia. Overall design: RNA sequencing in ERG overexpressing human CD34+ cells","project":"SRP034736"}
{"number_samples":4,"species":"human","abstract":"Schizophrenia (SZ) and autism spectrum disorders (ASD) are highly heritable neuropsychiatric/neurodevelopmental disorders, although environmental factors, such as maternal immune activation (MIA), play a role as well. Inflammatory cytokines appear to mediate the effects of MIA on neurogenesis and behavior in animal models. However, drugs and cytokines that trigger MIA can also induce a febrile reaction, which could have independent effects on neurogenesis through heat shock (HS)-regulated cellular stress pathways. However, this has not been well-studied. As a first step towards addressing the role of fever in MIA, we used a recently described model of human brain development in which induced pluripotent stem cells (iPSCs) differentiate into 3-dimensional neuronal aggregates that resemble a first trimester telencephalon. RNA-seq was carried out on aggregates that were heat shocked at 39oC for 24 hours, along with their control partners maintained at 37oC.  Overall, 186 genes showed significant differences in expression following HS (p<0.05), including known HS-inducible genes, as expected, as well as those coding for NGFR and a number of SZ and ASD candidates, including SMARCA2, DPP10, ARNT2, AHI1 and ZNF804A. The degree to which the expression of these genes decrease or increase during HS is similar to that found in copy loss and copy gain CNVs, although the effects of HS are likely to be more transient. Overall design: RNA-seq was carried out on neuronal aggregates as described by Mariani et al. with slight modification (PMID:22761314). For the heat shock experiment, a group of 49 day old aggregates was placed in an incubator set at 39C for 24 hours, while control sets of aggregates were maintained at 37C. The incubator conditions were otherwise unchanged. After detaching the aggregates, total cellular RNA was isolated using the miRNeasy Kit (Qiagen) according to the manufacturer’s protocol. Lastly, RNAseq profiles of HS and Control were compared","project":"SRP034737"}
{"number_samples":15,"species":"human","abstract":"Through integrative analysis of clinical breast cancer gene expression datasets, cell line models of breast cancer progression, and mutation data from cancer genome resequencing studies, we have identified RNA binding motif protein 47 (RBM47) as a candidate suppressor of breast cancer metastasis. RBM47 inhibited breast cancer progression in experimental models. Transcriptome-wide analysis of RBM47 localization by HITS-CLIP revealed widespread binding to mRNAs, preferentially at the 3' UTRs. RBM47 altered the abundance of a subset of its target mRNAs. Some of the mRNAs stabilized by RBM47, as exemplified by dickkopf WNT signaling pathway inhibitor 1 (DKK1), mediate tumor suppressive effects downstream of RBM47. This work identifies RBM47 as a suppressor of breast cancer progression and highlights the potential of global RNA modulatory events as a source of metastasis-promoting phenotypic traits. Overall design: Cancer cells transduced with doxycycline-inducible wildtype RBM47 or the RBM47-I281fs mutant, treated with increasing concentrations of doxycycline.","project":"SRP034873"}
{"number_samples":33,"species":"human","abstract":"RNA-Seq of retina and RPE/choroid/sclera from 8 normal human eyes","project":"SRP034875"}
{"number_samples":5,"species":"human","abstract":"The accurate characterization of transcripts and levels across species is critical for understanding transcriptome evolution. As available RNA-seq data accumulate rapidly, there is a great demand for tools that build gene annotations for cross-species RNA-seq analysis. Prevailing methods of ortholog annotation for RNA-seq analysis, do not take inter-species variation in mappability into consideration. Here we developed a computational framework that integrates previous approaches with multiple filters to improve the accuracy of inter-species transcriptome comparisons. The implementation of this approach in comparing RNA-seq data of human, chimpanzee, and rhesus macaque brain transcriptomes has reduced the false discovery of differentially expressed genes, while keeping the false negative rate low.","project":"SRP034929"}
{"number_samples":6,"species":"human","abstract":"Background: Undifferentiated pleomorphic sarcoma (UPS), used to be called malignant fibrous histiocytoma (MFH), is a malignant soft tissue tumor of uncertain origin, and is characterized by morphology. UPS often share similar morphological characters with other sarcomas, especially Leiomyosarcoma.  Leiomyosarcoma (LMS) is another malignant soft tissue sarcoma with complex genomic abnormalities, origin from smooth muscle.  As a result, development of gene signature and/or biomarkers distinguishing UPS and LMS will definitely help the pathologist to precisely diagnose those patients. However, in the past, UPS was reported to be indistinguishable with LMS by genomic profiles.  Methods and Results: In this study, 3’ end RNA Sequencing (3SEQ) was used to expression profile 6 UPS and 99 LMS cases.  Overall, UPS was undistinguished with LMS by 3SEQ data, however, when we stratified LMS into three subtypes, UPS was shown to share similar expression pattern with Subtype II LMS, but had distinct molecular expression patterns with Subtype I and Subtype III LMS.  Additional Immunohistochemistry staining by using LMS Subtype I and Subtype II markers validated that UPSs were positive for Subtype II marker ARL4C, but negative for Subtype I marker LMOD1. Furthermore, CD4 was shown to be significantly more highly expressed in UPS than LMS in both mRNA and protein levels.  Conclusion: This study first reported that UPS shared similar gene expression pattern with subtype II LMS and UPS recapitulated the expression profiles of subtype II LMS. Overall design: In this study, 3’ end RNA Sequencing (3SEQ) was used to expression profile 6 UPS and 99 LMS cases. In order to explore the molecular differences between UPS and LMS, We analyzed the expression data by SAMseq  to identify the genes which were significantly differently expressed between UPS and LMS, between UPS and each LMS subtype.","project":"SRP034953"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for A549 cells treated with bufalin for 3 hours","project":"SRP035224"}
{"number_samples":3,"species":"human","abstract":"PT_Leukemia","project":"SRP035252"}
{"number_samples":1,"species":"human","abstract":"We try to study RNA expression by comparative K562 cells RNA-Seq.","project":"SRP035255"}
{"number_samples":10,"species":"human","abstract":"Pancreatic beta-cell dysfunction and death are central in the pathogenesis of type 2 diabetes. Saturated fatty acids cause beta-cell failure and contribute to diabetes development in genetically predisposed individuals. Here we used RNA-sequencing to map transcripts expressed in five palmitate-treated human islet preparations, observing 1,325 modified genes. Palmitate induced fatty acid metabolism and endoplasmic reticulum (ER) stress. Functional studies identified novel mediators of adaptive ER stress signaling. Palmitate modified genes regulating ubiquitin and proteasome function, autophagy and apoptosis. Inhibition of autophagic flux and lysosome function contributed to lipotoxicity. Palmitate inhibited transcription factors controlling beta-cell phenotype including PAX4 and GATA6. 59 type 2 diabetes candidate genes were expressed in human islets, and 11 were modified by palmitate. Palmitate modified expression of 17 splicing factors and shifted alternative splicing of 3,525 transcripts. Ingenuity Pathway Analysis of modified transcripts and genes confirmed that top changed functions related to cell death. DAVID analysis of transcription binding sites in palmitate-modified transcripts revealed a role for PAX4, GATA and the ER stress response regulators XBP1 and ATF6. This human islet transcriptome study identified novel mechanisms of palmitate-induced beta-cell dysfunction and death. The data point to crosstalk between metabolic stress and candidate genes at the beta-cell level. Overall design: 5 human islet of Langerhans preparations examined under 2 conditions (control and palmitate treatment)","project":"SRP035268"}
{"number_samples":12,"species":"human","abstract":"The analysis has revealed that paroxetine has a weak estrogen-like activity because it modulates the expression of a number of estrogen-regulated genes using MCF7aroERE breast cancer cell lines studies","project":"SRP035276"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for A549 cells treated with BF211 for 3 hours","project":"SRP035285"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for A549 cells treated with 211HCl for 3 hours","project":"SRP035287"}
{"number_samples":15,"species":"human","abstract":"Purpose:The purpose of this study is to create unbiased, stage-specific transcriptomes by RNA-seq analyses of pure populations of both murine and human erythroblasts at distinct developmental stages. Methods: Recently developed FACS-based methods (Chen et al, PNAS, Liu et al, Blood, Hu et al Blood) were employed to purify morphologically and functionally discrete populations of cells, each representing specific stages of terminal erythroid differentiation. RNA was prepared from these cells and subjected to RNA-seq analyses. Results: There were vast temporal changes in gene expression across the differentiation stages, with each stage exhibiting unique transcriptomes.Clustering and network analyses revealed that differing stage-specific patterns of expression across differentiation were transcriptionally enriched for genes of differing function. Numerous differences were present between human and murine transcriptomes, with significant variation in the global patterns of gene expression. Conclusions: These data provide a significant resource for studies of normal and perturbed erythropoiesis, allowing a deeper understanding of mechanisms of erythroid development, differentiation, and inherited and acquired disease. Overall design: Both murine and human erythroblasts at distinct developmental stage mRNA profiles were generated by deep sequencing, in triplicate, using IlluminaHiSeq 2000.","project":"SRP035312"}
{"number_samples":10,"species":"human","abstract":"To systematically investigate early host transcriptional response to HIV-1 infection, including polyadenylated and non-polyadenylated transcripts, we separately sequenced Total RNAs (Total RNA-seq) from HIV-1-infected SupT1 cells, a human CD4+ T cell line. We show many non-polyadenylated RNAs were also differentially expressed upon HIV-1 infection, as expected only detected by Total RNA-seq. Interestingly, we identified 8 times more differentially expressed genes at 12 hour post-infection by Total RNA-seq than by mRNA-seq, mostly protein-coding genes. Overall design: 6 Total RNA sample sets were sequenced. Samples were either Mock infected, HIV-1 infected, or UV-inactivated viron infected for either 12 hours or 24 hours. Each sample set contains replicates. mRNA-seq data was processed previously and is part of a different study. It was referenced in this study. The mRNA-seq data can be found in GSE38006. This data set refers to the Whole Transcriptome data from similar samples.","project":"SRP035316"}
{"number_samples":8,"species":"human","abstract":"We generated induced human hepatocyte by transduction of several lineage specific transcription factors and analyzed the gene expression profile of generated hepatocytes and freshly isolated human hepatocytes through RNA sequencing. Overall design: Global gene expression profile analysis","project":"SRP035387"}
{"number_samples":6,"species":"human","abstract":"Sequencing of 5'ends of RNA molecules from Caco-2 cells +/- TNFa stimulation Overall design: CAGE library construction from RNA extracted from Caco-2 cells","project":"SRP035391"}
{"number_samples":8,"species":"human","abstract":"The goal of this project is to study transcriptome change by knocking down ZNF804A, a schizophrenia and bipolar disorder candidate gene, in early neurons derived from iPSCs. Overall design: Neural progenitor cells (NPCs) were developed from human induced pluripotent stem cells (iPSCs) and transduced by two independent shRNA vectors targeting ZNF804A, a schizophrenia and bipolar disorder candidate gene. After recovery and selection in puromycin, neuronal differentiation was induced. After 14 days, RNA was recovered and analyzed by RNA-seq. The expression profiles were compared with NPCs that were transduced with scrambled control vectors. This corresponds to controls 1-3 and KD 1-3, which was carried out on a male iPSC line. Scramble 1 and 2 and KD1 and 2 are technical replicates. Scrambled 3 and KD 3 were carried out on an independent NPC culture. For control 4 and KD4, neuronal differentiation was induced, and on day 10 the cells were transduced with the same ZNF804A KD and scrambled control vectors used for scrambled control 3 and KD3. In addition, this last set was carried out on a female iPSC line","project":"SRP035417"}
{"number_samples":4,"species":"human","abstract":"MicroRNA induces deadenylation of its targets according to the current model in the scientific community, but this model is based on the studies of a few individual genes. We tested the model by examining the global effect of miRNA on poly(A) tail of the targets. Synthetic miR-1 mimic was transfected into HeLa cells, and the poly(A) length was measured by TAIL-seq. Deadenylation of miR-1 targets was evident as early as 3 hours post-transfection without a significant change in mRNA level. After 6 or 9 hours, target mRNA level was substantially downregulated. Therefore, although there are some exceptions, our result confirms that, in general, miRNA indeed induces deadenylation. Furthermore, our kinetic global analysis confirms that deadenylation precedes mRNA decay. Overall design: Four culture dishes of HeLa cells were treated with miR-1 all together, then collected and prepared for sequencing after different time of incubation (0, 3, 6, and 9 hours post-transfection)","project":"SRP035419"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for SK cells treated with Bufalin for 3 hours","project":"SRP035452"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for SK-hep-1 cells treated with Bf211-HCl for 3 hours","project":"SRP035453"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for Hep3B cells","project":"SRP035454"}
{"number_samples":1,"species":"human","abstract":"RNA-seq for SK-hep-1 cells","project":"SRP035455"}
{"number_samples":2,"species":"human","abstract":"Human breast cancer cells MDA-MB-435 had higher metastatic potential and other aggressive characteristics than MCF-7 cells.  Next-generation RNA sequencing (RNA-Seq) was used to clear this concern through comparison the transcriptomic expression profiles of these two cells. Overall design: Total RNA were extracted from MCF-7 and MDA-MB-435S cells, and the polyA+ mRNA was sequenced using Illumina Genome Analyzer IIx. The reads were mapped to RefSeq RNA reference sequences.","project":"SRP035461"}
{"number_samples":2,"species":"human","abstract":"Purpose is to examine the effect of SmN expression to gene expression in HeLa Overall design: Compare the differences between the transcriptome of untreated and doxycycline-treated HeLa/TO-SmN in which expression of SmN is under doxycycline control","project":"SRP035464"}
{"number_samples":4,"species":"human","abstract":"Nasopharyngeal carcinoma (NPC) is a prevalent malignancyt disease in Southeast Asia among the Chinese population. Aberrant regulation of transcripts has been implicated in many types of cancers including NPC. Herein, we characterized mRNA and miRNA transcriptomes by RNA sequencing (RNASeq) of NPC model systems. Overall design: Matched total mRNA and small RNA of undifferentiated Epstein-Barr virus (EBV)-positive NPC xenograft X666 and its derived cell line C666, well-differentiated NPC cell line HK1, and the immortalized nasopharyngeal epithelial cell line NP460 have been sequenced by Solexa technology.","project":"SRP035477"}
{"number_samples":4,"species":"human","abstract":"Nasopharyngeal carcinoma (NPC) is a prevalent malignancyt disease in Southeast Asia among the Chinese population. Aberrant regulation of transcripts has been implicated in many types of cancers including NPC. Herein, we characterized mRNA and miRNA transcriptomes by RNA sequencing (RNASeq) of NPC model systems. Overall design: Matched total mRNA and small RNA of undifferentiated Epstein-Barr virus (EBV)-positive NPC xenograft X666 and its derived cell line C666, well-differentiated NPC cell line HK1, and the immortalized nasopharyngeal epithelial cell line NP460 have been sequenced by Solexa technology.","project":"SRP035478"}
{"number_samples":2,"species":"human","abstract":"We've developed a new Method to Analyze RNA following Intracellular Sorting (MARIS) allowing us to carry out gene expression studies on cells sorted based on intracellular immunoflourescence. The purpose of this study is to determine the degree of bias that MARIS introduces on gene expression. We report RNA-seq gene expression data from human embryonic stem cells differentiated to a stage in which insulin-expressing cells are present. Gene expression data using RNA isolated from live cells is compared to gene expression data using RNA isolated from MARIS processed cells (fixed, permeabilized, antibody stained and mock sorted) to determine the degree of correlation in gene expression between these two biologically identical samples. Overall design: Human embryonic stem cells are differentiated to a stage in which insulin-expressing cells are present and split into two biologically identical samples. RNA is immediately isolated from one sample using the RNeasy protocol (live sample). RNA is isolated from the second sample following MARIS (processed sample) with all cells collected after the sort in order to keep the cell type composition between the live and processed samples the same.","project":"SRP035482"}
{"number_samples":2,"species":"human","abstract":"Rituximab alone or in combination with chemotherapeutics is the first-line therapy for variety of lymphoproliferative disorders including low- and high grade non-Hodgkin’s lymphomas (NHL). Although the complete response rate is quite impressive, vast majority of patient presents recurrent disease. The association between CD20 expression and clinical outcome in patients strongly suggests that reduced CD20 expression leads to inferior response to RCHOP (rituximab, cyclophosphamide, vincristine, doxorubicin and prednisone). In order to understand how loss of CD20 leads to development of RCHOP resistance, we developed rituximab resistant DOHH2 model in vivo by chronic exposure to rituximab. Characterization of several resistant in vivo xenografts revealed one model that maintained resistance to an acute dose of rituximab and demonstrated loss of CD20. Further characterization of the model demonstrated a loss of CD20 is associated with over expression of BCL2 and BIM. In vivo efficacy studies showed resistant line is insensitive to acute dose of RCHOP and treatment with an inhibitor of BCL2 (ABT199) in combination with chemotherapy resulted in better efficacy than RCHOP alone. We have identified an in vivo model of DLBCL where loss of CD20 and over expression of anti-apoptotic protein BCL2 leads to RCHOP resistance. These data suggest the addition of BCL2 inhibitor to chemotherapy might be effective in treating CD20 negative lymphomas. Overall design: mRNA profiles of parental and rituximab resistant DOHH2 xenograft were generated by deep sequencing using Illumina HiSeq","project":"SRP035503"}
{"number_samples":35,"species":"human","abstract":"The DNA methylome and transcriptome of different brain regions in schizophrenia and bipolar disorder","project":"SRP035524"}
{"number_samples":24,"species":"human","abstract":"Vaccinia virus (VACV) is a large cytoplasmic dsDNA virus that causes dramatic alterations to many cellular pathways including microRNA biogenesis. Here we use small RNA sequencing to comprehensively quantify the impact of VACV infection on the expression of endogenous small non-coding RNAs (sncRNAs) at both early (6 h) and late (24 h) times post infection. Overall design: Non-coding RNAs (sncRNAs) were assessed at 6 hours and 24 hours, in naive and VACV-infected HeLa cells, in the presence or absence of the inhibitor AraC.  There were three replicates for each of the eight groups.","project":"SRP035554"}
{"number_samples":2,"species":"human","abstract":"Nuclear lamins contact the genome at the nuclear periphery through large domains and are involved in chromatin organization. Among broad peak calling algorithms available to date, none are suited for mapping lamin-genome interactions genome-wide. We disclose a novel algorithm, Enriched Domain Detector (EDD), for analysis of broad enrichment domains from ChIP-seq data. EDD enables discovery of genomic domains interacting with broadly distributed chromatin-associated proteins such as lamins. The main advantage of EDD over existing broad peak callers is sensitivity to domain width rather than enrichment strength at a particular site, and robustness against local variations. EDD is downloadable from http://github.com/eivindgl/edd. Overall design: RNA-seq experiments in human normal dermal fibroblasts (Lonza CC-2511; LDFs) and human normal primary dermal fibroblasts (Norwegian Stem Cell Center AD04DFs).","project":"SRP035617"}
{"number_samples":1,"species":"human","abstract":"somatic tumor exomes from colon cancer cell lines","project":"SRP035634"}
{"number_samples":2,"species":"human","abstract":"MicroRNAs (miRNAs), including host miRNAs and viral miRNAs, play vital roles in regulating host-virus interactions. DNA viruses encode miRNAs that regulate the viral life cycle. However, it is generally believed that cytoplasmic RNA viruses do not encode miRNAs, owing to inaccessible cellular miRNA processing machinery. Here, we provide a comprehensive genome-wide analysis and identification of miRNAs that were derived from hepatitis A virus (HAV; Hu/China/H2/1982), which is a typical cytoplasmic RNA virus. Using deep-sequencing and in silico approaches, we identified 2 novel virally encoded miRNAs, named hav-miR-1-5p and hav-miR-2-5p. Both of the novel virally encoded miRNAs were clearly detected in infected cells. Analysis of Dicer enzyme silencing demonstrated that HAV-derived miRNA biogenesis is Dicer dependent. Furthermore, we confirmed that HAV mature miRNAs were generated from viral miRNA precursors (pre-miRNAs) in host cells. Notably, naturally derived HAV miRNAs were biologically and functionally active and induced post-transcriptional gene silencing (PTGS). Genomic location analysis revealed novel miRNAs located in the coding region of the viral genome. Overall, our results show that HAV naturally generates functional miRNA-like small regulatory RNAs during infection. This is the first report of miRNAs derived from the coding region of genomic RNA of a cytoplasmic RNA virus. These observations demonstrate that a cytoplasmic RNA virus can naturally generate functional miRNAs, as DNA viruses do. These findings also contribute to improved understanding of host-RNA virus interactions mediated by RNA virus-derived miRNAs. Overall design: Examination of small RNA populations in KMB17 cells infected by human hepatitis A virus genotype IA (isolate H2) (21 days post-infected cells) and mock-infected KMB17 cells (21 days mock-infected control cells).","project":"SRP035638"}
{"number_samples":8,"species":"human","abstract":"Purpose: We find that Wnt7a-PAX6 axis determine corneal epithelial cell fate. To obtain global evidence for successful cell fate conversion, we performed gene expression profiling by RNA-seq on CECs, SECs, and LSCs after knocking down PAX6 and on SESCs transduced with PAX6 upon 3-D differentiation. Methods: Under 3-D culture condition, limbal stem cell (LSCs) can be differentiated to Cornea epithelial cells (CECs), and skin epithelial stem cells (SESCs) can be differentiated to skin epithelial cells (SECs). Total RNA was isolated from CECs, SECs, and LSCs after knocking down PAX6 (3-D shPAX6 LSCs) and on SESCs transduced with PAX6 (3-D PAX6+ SESCs) upon 3-D differentiation. Libraries were prepared following published standard protocol (Fox-Walsh K et al., 2011, genomics, 266-71). mRNA profiles were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000. Results: Following optimized decoding and mapping workfollow, we mapped about 5 million sequence reads to the human genome and identified more than 23659 transcripts per sample. Conclusions: Hierarchical clustering analysis of differentially expressed gene signatures revealed that the gene expression pattern of SESCs with PAX6 transduction was strikingly similar to that of CECs, whereas the profile of LSCs with PAX6 knockdown was highly related to that in SECs upon differentiation. These data therefore provided global evidence for a decisive role of the WNT7A/PAX6 axis in cell fate conversion from SESCs to CECs. Overall design: RNA-seq on CECs, SECs, and LSCs after knocking down PAX6 and on SESCs transduced with PAX6 upon 3-D differentiation, using Illumina HiSeq 2000","project":"SRP035641"}
{"number_samples":2,"species":"human","abstract":"The human genome harbors a large number of sequences encoding for RNAs that are not translated but control cellular functions by distinct mechanisms. The expression and function of the longer transcripts namely the long non-coding RNAs (lncRNAs) in the vasculature is largely unknown. Here, we characterized the expression of lncRNAs in human endothelial cells and elucidated the function of the highly expressed metastasis-associated lung adenocarcinoma transcript 1 (MALAT1; also known as MALAT-1 or NEAT2). Endothelial cells of different origin express high levels of the conserved lncRNAs MALAT1, TUG1, MEG, linc00657 and linc00493. MALAT1 was significantly increased by hypoxia and controls a phenotypic switch in endothelial cells. Silencing of MALAT1 by siRNAs or GapmeRs induced a pro-migratory response and increased basal sprouting and migration, whereas proliferation of endothelial cells was inhibited. If angiogenesis was further stimulated by VEGF, MALAT1 siRNAs induced discontinuous sprouts indicative of defective proliferation of stalk cells. In vivo studies confirmed that genetic ablation of MALAT1 inhibited proliferation of endothelial cells and reduced neonatal retina vascularization. Gene expression profiling followed by confirmatory qRT-PCR demonstrated that silencing of MALAT1 impaired the expression of various cell cycle regulators. Silencing of MALAT1 tips the balance from a proliferative to a migratory endothelial cell phenotype in vitro and its genetic deletion reduces neonatal vascular growth in vivo. Overall design: Human endothelial polyA+ RNA expression analysis. We used two replicate samples of human umbilical vein endothelial cells.","project":"SRP035665"}
{"number_samples":2,"species":"human","abstract":"Oct4, a key transcription factor for maintaining the pluripotency and self-renewal of stem cells has been reported previously. It also plays an important role in tumor proliferation and apoptosis, but the role of Oct4 been in tumor metastasis is still not very clear. Here, we found that ectopic expression of Oct4 in breast cancer cells can inhibit their migration and invasion. Detailed examinations revealed that Oct4 up-regulates expression of E-cadherin, indicative of its inhibitory role in epithelial-mesenchymal transition (EMT). RNA-sequence assay showed that Oct4 down-regulates expression of Rnd1. As an atypical Rho protein, Rnd1 can affect cytoskeleton rearrangement and regulate cadherin-based cell-cell adhesion by antagonizing the typical Rho protein, RhoA. Ectopic expression of Rnd1 in MDA-MB-231 cells changes cell morphology which influences cell adhesion and increases migration. It is reported that EMT is accompanied by cytoskeleton remodeling, we hypothesized that Rnd1 may play a role in regulating EMT. Over-expression of Rnd1 can partly rescue the inhibitory effects induced by Oct4, not only migration and invasion, but also in E-cadherin level and cellular morphology. Furthermore, silencing of Rnd1 can up-regulate the expression of E-cadherin in MDA-MB-231 cells. These results present evidence that ectopic expression of Oct4 increases E-cadherin and inhibits metastasis, effects which may be related to Rnd1 associated cell-cell adhesion in breast cancer cells. Overall design: Examination of mRNA profiles in MDA-MB-231 cells with OCT4 overexpressing","project":"SRP035670"}
{"number_samples":2,"species":"human","abstract":"To identify the functional non-coding RNAs in human DC development, we explored the gene expression profiles during DC development from peripheral monocytes. Using RNA-seq, we identified many annotated lncRNAs with markedly altered expressions during human DC differentiation and maturation. Overall design: RNA samples from human peripheral blood monocytes and monocyte-derived DCs were collected. RNAs from 3 donors were pooled together to form monocyte sample or DC sample and then these two samples were subjected to RNA-seq and analysis.","project":"SRP035679"}
{"number_samples":5,"species":"human","abstract":"Although many distinct mutations in a variety of genes are known to cause Amyotrophic Lateral Sclerosis (ALS), it remains poorly understood how they selectively impact motor neuron biology and whether they converge on common pathways to cause neural degeneration. Here, we have combined reprogramming and stem cell differentiation approaches with genome engineering and RNA sequencing to define the transcriptional changes that are induced in human motor neurons by mutant SOD1. Mutant SOD1 protein induced a transcriptional signature indicative of increased oxidative stress, reduced mitochondrial function, altered sub-cellular transport as well as activation of the ER stress and unfolded protein response pathways. Functional studies demonstrated that perturbations in these pathways were indeed the source of altered transcript levels. Overall design: 5 samples, 2 patient-derived SOD1A4V and 3 isogenic control samples where the mutation has been corrected. All samples are motor neurons derived from induced pluripotent stem cells (iPSCs), and isolated after lentiviral infection with an Hb9:RFP construct and FACS purification. Each sample is a separate biological replicate.","project":"SRP035862"}
{"number_samples":9,"species":"human","abstract":"Ultraviolet-B (UVB) irradiation of the skin was performed on rat (skin and DRG) and human (skin) tissue. The resulting changes in gene expression were then profiled by using RNA-seq to compare gene expression between irradiated and non-irradiated samples Overall design: Poly(A) selected RNA was sequenced for 5 irradiated and 4 non-irradiated humans (skin), and for 6 irradiated and 6 non-irradiated rats (DRG and skin)","project":"SRP035864"}
{"number_samples":12,"species":"human","abstract":"The identification of circulating endothelial progenitor cells has led to speculation regarding their origin as well as their contribution to neovascular development. Two distinct types of endothelium make up the blood and lymphatic vessel system. However, it has yet to be determined whether there are distinct lymphatic-specific circulating endothelial progenitor cells. We isolated circulating endothelial colony forming cells (ECFCs) from whole peripheral blood. These cells are endothelial in nature, as defined by their expression of endothelial markers and their ability to undergo capillary morphogenesis in three-dimensional culture. A subset of isolated colonies express markers of lymphatic endothelium, including VEGFR-3 and Prox-1, with low levels of VEGFR-1, a blood endothelial marker, while the bulk of the isolated cells express high VEGFR-1 levels with low VEGFR-3 and Prox-1 expression. The different isolates have differential responses to VEGF-C, a lymphatic endothelial specific cytokine, strongly suggesting that there are lymphatic specific and blood specific ECFCs. Global analysis of gene expression revealed key differences in the regulation of pathways involved in cellular differentiation between blood and lymphatic-specific ECFCs. These data indicate that there are two distinguishable circulating ECFC types, blood and lymphatic, which are likely to have discrete functions during neovascularization. Overall design: RNA was isolated from 2 blood-specific ECFC cell lines and 2 lymphatic-specific ECFC cell lines 3 separate times each","project":"SRP035883"}
{"number_samples":6,"species":"human","abstract":"The leukemogenic fusion protein RUNX1/ETO impairs myeloid differentiation and drives malignant self-renewal. Here we use digital footprinting and ChIP-sequencing to identify the core RUNX1/ETO responsive transcriptional network of t(8;21) cells. We show that the transcriptional programme underlying leukemic propagation is regulated by a dynamic equilibrium between RUNX1/ETO and RUNX1 complexes, which bind in a mutually exclusive fashion to identical genomic sites. Perturbation of this equilibrium by RUNX1/ETO knockdown results in a global reassembly of transcription factor complexes within pre-existing open chromatin and the formation of a new C/EBP-dominated transcriptional network driving myeloid differentiation. Our work demonstrates that the block in myeloid differentiation in t(8;21) is caused by the dynamic balance between RUNX1/ETO and RUNX1 activities and the repression of C/EBP and highlights the core targets of epigenetic reprogramming in t(8;21) AML. Overall design: ChIP-sequencing, DNAse1-sequencing and RNA-sequncing was used for the identification of a dynamic core transcriptional network in t(8;21) AML This submission represents the RNA-Seq component of the study","project":"SRP035930"}
{"number_samples":7,"species":"human","abstract":"In the present study adenovirus type 2 RNA splicing events were quantitatively mapped by using deep cDNA sequencing. The majority of the previously identified splice sites could be confirmed. The lack of complete consistency between the present and previous results appears to be that some sites have been incorrectly mapped in previous studies, such as the splice sites for pVII, pVIII and E3-11.6K. everal previously predicted splice sites such as that for E3-14.5K and E4ORF3/4 were not detected.","project":"SRP035934"}
{"number_samples":179,"species":"human","abstract":"To increase our understanding of psoriasis, we utilized RNA-seq to assay the transcriptomes of lesional psoriatic and normal skin. We sequenced polyadenylated RNA-derived cDNAs from 92 psoriatic and 82 normal punch biopsies, generating an average of ~38 million single-end 80-bp reads per sample. Comparison of 42 samples* examined by both RNA-seq and microarray [GSE13355] revealed marked differences in sensitivity, with transcripts identified only by RNA-seq having much lower expression than those also identified by microarray. RNA-seq identified many more differentially expressed transcripts enriched in immune system processes.  Weighted gene co-expression network analysis (WGCNA) revealed multiple modules of coordinately expressed epidermal differentiation genes, overlapping significantly with genes regulated by the long non-coding RNA TINCR, its target gene, staufen-1 (STAU1), the p63 target gene ZNF750, and its target KLF4. Other coordinately expressed modules were enriched for lymphoid and/or myeloid signature transcripts and genes induced by IL-17 in keratinocytes. Dermally-expressed genes were significantly down-regulated in psoriatic biopsies, most likely due to expansion of the epidermal compartment. These results demonstrate the power of WGCNA to elucidate gene regulatory circuits in psoriasis, and emphasize the influence of tissue architecture in both differential expression and co-expression analysis. *The list of 42 samples examined by both RNA-seq and microarray is provided in the 'MAoverlappedsamples.txt'. Overall design: 92 psoriatic and 82 normal skin samples","project":"SRP035988"}
{"number_samples":1,"species":"human","abstract":"Transcriptome sequencing of Trichomonas vaginalis during infection of human cells in different timepoints.","project":"SRP036029"}
{"number_samples":6,"species":"human","abstract":"Epithelial-to-mesenchymal transition (EMT) is a dynamic process that relies on cellular plasticity; an EMT/MET axis is critical for metastatic colonization of carcinomas. Unlike epithelial programming, regulation of mesenchymal programming is not well understood in EMT. Here we describe the first microRNA that enhances exclusively mesenchymal programming. We demonstrate that microRNA-424 is up-regulated early during a TWIST1/SNAI1-induced EMT, and that it causes cells to express mesenchymal genes without affecting epithelial genes, resulting in a mixed/intermediate EMT. Further, microRNA-424 increases motility, decreases adhesion and induces a growth arrest, changes associated with a complete EMT. Patient microRNA-424 levels positively associate with TWIST1/2 and EMT-like gene signatures and is increased in primary tumors versus matched normal breast. However, microRNA-424 is down-regulated in metastases versus matched primary tumors. Correspondingly, microRNA-424 decreases tumor initiation and is post-transcriptionally down-regulated in macrometastases in mice. RNA-seq identified microRNA-424 regulates numerous genes associated with EMT and breast cancer stemness including the novel miR-424 target, TGFBR3, which regulates mesenchymal phenotypes without influencing miR-424 effects on tumor-initiating phenotypes; instead, we show that ERK signaling is critical for such tumor-initiating effects of miR-424. These findings suggest microRNA-424 plays distinct roles downstream of EMT-inducing factors, facilitating earlier stages, but repressing later stages, of metastasis. Overall design: Examination of mRNA levels in MCF12A human breast cell lines that stably over-expressed miR-424 or an empty vector (EV) control. Each group has three replicates.","project":"SRP036035"}
{"number_samples":10,"species":"human","abstract":"Leiomyosarcoma (LMS) is a malignant neoplasm of smooth muscle and is an aggressive soft tissue tumor, have complex genetic abnormalities and could be defined as three molecular subtypes. Since that the molecular heterogeneity of LMS, the pathogenesis analysis per subtype will be highly necessary and helpful to understand the etiology of this more common sarcoma. Overall design: Within this study, we collected four Myometrium, three Leiomyoma, three LMS cell lines and 99 LMSs (GSE45510), performed the system-wide gene expression profiling by 3'end RNA Sequencing, and found that there are significant different molecular pathways along the pathogenesis for those three molecular subtypes.","project":"SRP036053"}
{"number_samples":8,"species":"human","abstract":"We profiled the dynamic, comprehensive transcriptome during human erythroid differentiation in vitro. The erythroid cells at day 4, 8, 11 and 14 differentiation stages were harvested and sequenced by Illumia 72 bp paired-end sequencing format, respectively. Overall design: Expression profiling of erythroid cells on differentiation days 4, 8, 11 and 14 and performed mRNA-seq on two biological replicates at each stage.","project":"SRP036133"}
{"number_samples":1,"species":"human","abstract":"These data are transcriptomes from the HapMap trio (CEU cell-lines Gm12878, Gm12891 and Gm12892)","project":"SRP036136"}
{"number_samples":3,"species":"human","abstract":"We carried out a genome-wide investigation of the primary transcriptional targets of 1a,25(OH)2D3 in breast epithelial cancer cells using RNA-Seq technology. We identified early transcriptional targets of 1a,25(OH)2D3 involved in adhesion, growth regulation, angiogenesis, actin cytoskeleton regulation, hexose transport, inflammation and immunomodulation, apoptosis, endocytosis and signaling. Furthermore, we found several transcription factors to be regulated by 1a,25(OH)2D3 that subsequently amplify and diversify the transcriptional output driven by 1a,25(OH)2D3 leading finally to a growth arrest of the cells. Moreover, we could show that 1a,25(OH)2D3 elevates the trimethylation of histone H3 lysine 4 at several target gene promoters. Our present transcriptomic analysis of differential expression after 1a,25(OH)2D3 treatment provides a resource of primary 1a,25(OH)2D3 targets that might drive the antiproliferative action in breast cancer epithelial cells. ","project":"SRP036145"}
{"number_samples":8,"species":"human","abstract":"In order to dtermine how well a mouse genetic model of alveolar soft part sarcoma (ASPS) mimics the human disease, five human ASPS tumor samples and three normal skeletal muscle samples were profiled by RNAseq and compared to samples from five mouse tumors induced by expression of ASPSCR1-TFE3 and three normal mouse skeletal muscle samples, also profiled by RNAseq. Overall design: The reference was really comparing 5 human ASPS tumors to 5 mouse tumors that histologically mimic ASPS, but using skeletal muscle controls (3 from each species) as a sounding board for differential expression.","project":"SRP036769"}
{"number_samples":3,"species":"human","abstract":"Human iPS cell line, immortalized B-cells, and primary fibroblasts frome Personal Genome Project voluteer 1 Overall design: Each sample per cell type.","project":"SRP036790"}
{"number_samples":45,"species":"human","abstract":"Soft tissue sarcomas (STTs) are neoplasms of mesenchymal cells, and are composed of more than 60 subtypes. Although many of these STTs could be distinguished from each other by morphology and limited IHC biomarkers, the precise diagnosis of STTs with similar morphology is still difficult to achieve. Therefore developing novel diagnosis biomarker and molecular signatures will be very helpful to pathologist in clinic. Overall design: Within this study, 3’ end RNA Sequencing (3SEQ) was used to expression profile different subtypes of soft tissue sarcomas.","project":"SRP036821"}
{"number_samples":4,"species":"human","project":"SRP036840"}
{"number_samples":3,"species":"human","abstract":"We knocked down TR4 expression by 2 lentiviral mediated vectors at day 11 of erythroid differentiation in order to identify TR4 downstream targets. Overall design: Two lentiviruses and one empty control were used to knockdown TR4 expression. The cells were harvested at day 11 during erythroid differentiation.","project":"SRP036843"}
{"number_samples":6,"species":"human","abstract":"The histone variant macroH2A1 and the poly(ADP-ribose) polymerase PARP-1 both regulate gene transcription by modulating chromatin structure and function. Of the two macroH2A1 splice variants, macroH2A1.1 and macroH2A1.2, the former is often suppressed in cancer and has the unique ability to interact with poly(ADP-ribose). Using ChIP-seq in primary lung fibroblasts, we demonstrate that macroH2A1 is incorporated into either of two spatially and functionally distinct types of chromatin; the first is marked by H3 K27 trimethylation, while the second contains a set of nine histone acetylations. MacroH2A1-regulated genes are involved in cancer progression are specifically found in macroH2A1-containing acetylated chromatin. Through the recruitment of PARP-1, macroH2A1.1 promotes the acetylation of H2B K12 and K120 which plays a key role in the regulation of macroH2A1 target genes in primary cells. The macroH2A1/PARP-1 pathway regulating H2B K12 and K120 acetylation is disrupted in cancer cells, in part, explaining macroH2A1’s role in cancer suppression. Overall design: Three biological replicates of RNA-seq from cells expressing shRNA directed against macroH2A1 or luciferase as a control","project":"SRP037550"}
{"number_samples":8,"species":"human","abstract":"Single 4 mm punch biopsies were obtained from lesional forearm skin of four patients with early, diffuse SSc within 6 months of first onset of non-Raynaud’s symptoms, and normal skin of four controls without disease. Total RNA was extracted from skin biopsies using QIAgen RNeasy Plus Mini kit. Ribosomal RNA was depleted from the large RNA fraction using Invitrogen Ribominus kit. RNA-Seq library was synthesized by NuGen Ovation RNA-Seq System v2 using cDNA prepared by random hexamer priming. Libraries were multiplexed and sequenced on an Illumina HiSeq 2000 platform and 187-242 million 50 bp paired-end reads were obtained per sample.","project":"SRP037553"}
{"number_samples":2,"species":"human","abstract":"NGN3-eGFP could be used to mark endocrine progenitor cells and their progeny in differentiating hESCs, suggesting that profiling analysis of NGN3-eGFP+ cells could lead to the discovery of novel gene participation. Six batches of NGN3-eGFP+ and NGN3-eGFP -cells purified by FACS from a reporter hES cell line cell cultures at stage 4 were pooled together, respectively. RNA-seq-based analysis of the two cell fractions identified a total of 3976 differentially expressed genes (P-value<0.05; Fold >2), of which 72% are enriched in NGN3-GFP+ cells. As expected, all the genes expressed in pancreatic progenitor cells are enriched in NGN3-eGFP-cells; while all endocrine-associated genes are enriched in NGN3-eGFP+ cells. Among the 3976 genes, 195 have potential transcriptional factor activity and 453 are potential non-coding RNA genes, most of which have not been well investigated in pancreatic endocrine cells. The gene ontology term ‘integral to membrane’ GO: 0016021 identified 1003 differentially expressed genes that encoded ion channels, adhesion molecules and transporters. Overall design: NGN3-eGFP Postive and negtive cells were purified at stage 4 and subjected to RNA-sequncing","project":"SRP037579"}
{"number_samples":6,"species":"human","abstract":"HDAC inhibitors are thought to regulate gene expression by post-translational modification of histone as well as non-histone proteins. Often studied at single loci, increased histone acetylation is the paradigmatic mechanism of action, however, little is known of the extent of genome-wide changes of the mammalian genome when stimulated by the hydroxamic acids, TSA and SAHA. In primary human vascular endothelial cells we map the chromatin modifications, histone H3 acetylation of lysine 9 and 14 (H3K9/14ac) using chromatin immunoprecipitation (ChIP) coupled with massive parallel sequencing (ChIP-seq). Since acetylation mediated gene expression is often associated with modification of other lysine residues we also examined H3K4me3 and H3K9me3 as well as changes in CpG methylation (CpG-seq). Genome-wide mRNA sequencing indicates the differential expression of about 30% of genes, with almost equal numbers being up- and down- regulated. We observe deacetylation conferred by TSA and SAHA that are associated with decreased gene expression. Histone deacetylation is associated with the loss of p300/CBP binding at gene promoters. This study provides an important framework for HDAC inhibitor function in vascular biology and a comprehensive description of genome-wide deacetylation. Overall design: HAEC mRNA profiles of SAHA treated and control samples were generated by deep sequencing, in triplicate, using Illumina GAIIx.","project":"SRP037718"}
{"number_samples":6,"species":"human","abstract":"The human genome shares a remarkable amount of genomic sequence with our closest living primate relatives. Researchers have long sought to understand what regions of the genome are responsible for unique species-specific traits. Previous studies have shown that many genes are differentially expressed between species, but the regulatory elements contributing to these differences are largely unknown. Here we report a genome-wide comparison of active gene regulatory elements in human, chimpanzee, and macaque, and we identify hundreds of regulatory elements that have been gained or lost in the human or chimpanzee genomes since their evolutionary divergence. These elements contain evidence of natural selection and correlate with species-specific changes in gene expression. Polymorphic DNA bases in transcription factor motifs that we found in these regulatory elements may be responsible for the varied biological functions across species. This study directly links phenotypic and transcriptional differences between species with changes in chromatin structure. Overall design: DGE-seq","project":"SRP037719"}
{"number_samples":27,"species":"human","abstract":"We developed a customized RNA-Seq strategy to identify the 3' termini of degradation intermediates of histone mRNAs. Using this strategy, we identified two types of oligouridylated degradation intermediates: RNAs ending at different sites of the 3' side of the stemloop that resulted from initial degradation by 3'hExo and intermediates near the stop codon and within the coding region. We established a method for identifying degradation intermediates for replication-depdendent histone mRNAs.  Histone mRNAs are present in large amounts in S-phase cells.  Inhibition of DNA replication results in a rapid degradation of histone mRNA.  We developed an approach to selectively amplify the 3' ends of histone mRNA molecules, including degradation intermediates, which we then sequenced.  Using a novel bioinformatics platform we identified degradation intermediates including many that had untemplated nts added to the 3' end.  We identified a large variety of degradation intermediates that contained oligo(U) tails.  We were able to deduce the 3' to 5' pathway of histone mRNA degradation. Overall design: RNA-seq with a custom 3' linker: The samples analyzed included two biological replicates of RNAs before and 20 min after HU treatment (samples 1, 26, 27) capped RNAs from cells treated with HU for 20 min (samples 2,3,16,17) RNA isolated from polyribosomes 20 min after HU treatment, from untreated cells (samples 4-9) and cells treated with pactamycin for 5 min (samples 10-15); from cells in which the exosome subunit Pml/SC100 had been knocked down (samples 22-23) and cells treated with the control siRNA (samples 20-21). Please note that, for Sample 2 and Sample 3, the sequencer was unable to recover both ends in the mate pairs for these two sequencing runs, but we chose to use these data anyway since the reads that were recovered included the 3' linker that we used to determine the presence of untemplated additions.","project":"SRP037722"}
{"number_samples":2,"species":"human","abstract":"Alternative polyadenylation is an important cellular mechanism that enables generation of  mRNA isoforms that differ in their 3' untranslated regions (3' UTRs) and consequently in their susceptibility to miRNA and RNA binding protein mediated regulation. A dramatic change in polyadenylation site usage, leading to the systematic expression of short 3’ UTR isoforms is known to occur upon induction of proliferation in resting cells. To understand the functional consequences of short 3’ UTR isoform expression we used 3' end sequencing and quantitative mass spectroscopy to determine polyadenylation site use, mRNA and protein levels in murine and human naive and activated T cells. We found that while the process and its impact on the susceptibility to miRNA and RNA binding protein mediated regulation are evolutionarily conserved, the conservation is poor at the level of individual orthologous genes. Contrary to the common belief, we did not find that transcriptome-wide 3' UTR shortening leads to a matched increase in mRNA and protein levels of genes with tandem polyadenylation sites. Overall design: 3' ends of transcripts were profiled by high-throughput sequencing in murine and human naive and activated T cells.","project":"SRP037735"}
{"number_samples":6,"species":"human","abstract":"Long non-coding RNAs (lncRNAs) have emerged as critical regulators of gene expression and chromatin modifications, with important functions in development and disease. Here we sought to identify and functionally characterize lncRNAs critical for vascular vertebrate development with significant conservation across species. Genome-wide transcriptomic analyses during human vascular lineage specification enabled the identification of three conserved novel lncRNAs: TERMINATOR, ALIEN and PUNISHER that are specifically expressed in pluripotent stem cells, mesoderm and endothelial cells, respectively. Gene expression profiling, alongside RNA immunoprecipitation coupled to mass spectrometry, revealed a wide range of new molecular networks and protein interactors related to post-transcriptional modifications for all three lncRNAs. Functional experiments in zebrafish and murine embryos, as well as differentiating human cells, confirmed a developmental-stage specific role for each lncRNA during vertebrate development. The identification and functional characterization of these three novel non-coding provide a comprehensive transcriptomic roadmap and shed new light on the molecular mechanisms underlying human vascular development. Overall design: Time course RNA-Seq analysis H1 ESCs differentiated into endothelial cells","project":"SRP037762"}
{"number_samples":63,"species":"human","abstract":"The intention was to detect genes that are determining trastuzumab efficiency in HER2-positive breast cancer cell lines with different resistance phenotypes. While BT474 should be sensitive to the drug treatment, HCC1954 is expected to be resistant due to a PI3K mutation. The cell line BTR50 has been derived from BT474 and was cultured to be resistant as well. Based on RNA-Seq data, we performed differential expression analyses on these breast cancer cell lines with and without trastuzumab treatment. In detail, five separate tests were performed, namely resistant cells vs. wild type, i.e. HCC1954 and BTR50 vs. BT474, respectively, and untreated vs. drug treated cells. The significant genes of the first two tests should contribute to resistance. The significant genes of the test BT474 vs. its drug treated version should contribute to the trastuzumab effect. To exclude false positives from the combined gene set (#64), we removed ten genes that were also significant in the test BTR50 vs. its drug treated version. This way we ended up with 54 genes that are very likely to determine trastuzumab efficiency in HER2-positive breast cancer cell lines. Overall design: mRNA profiles of human breast cancer cell lines were generated by deep sequencing using Illumina HiSeq 2000. The cell lines BT474 and HCC1954 were analyzed with and without trastuzumab treatment. HCC1954 is known to be trastuzumab resistant. Additionally, the cell line BTR50 was generated as resistant version of BT474, and was analyzed with and without trastuzumab as well.","project":"SRP037775"}
{"number_samples":46,"species":"human","abstract":"Purpose: To profile the transcriptomes of omental adipose tissues from obese and lean humans. Methods: Omental adipose tissues from obese and lean patients were subjected to RNA-Seq. Results: Differential expression analysis identified 206 dysregulated genes (p-value < 0.05 by moderated t-test and fold change = 2 in obesity) that are known to be involved in a multitude of functions, including response to stress, inflammatory response and leukocyte adhesion. Differential splicing analysis uncovered the possible role of TLR4 RNA splicing in obesity. Our findings suggest that, as a person experiences weight gain/obesity, the adipose splicing pattern of TLR4 transcripts changes in favor of activation of TLR4 signaling, which in turn may contribute to the progression of obesity-related inflammation and complications. Conclusion:  This study provides a look into the transcriptome of disease-state adipose tissue in obesity, and demonstrates the potential importance of aberrant RNA splicing and expression in obesity-associated immune dysregulation. Overall design: Study design is of cross-sectional nature. Seven samples (three obese and four lean) were analyzed.","project":"SRP037778"}
{"number_samples":1,"species":"human","abstract":"We show that in vivo MBD2 is mainly recruited to CpG island promoters that are highly methylated. We also report that MBD2 binds to a subset of CpG island promoters that are characterized by the presence of active histone marks and RNA polymerase II (Pol2). At such sites, MBD2 binds downstream of the transcription start site. Active promoters bound by MBD2 show low to medium gene expression levels and H3K36me3 deposition suggesting a putative role for MBD2 in blocking polymerase II (Pol2) elongation at these promoters. Overall design: To gain further insight into the function of and epigenetic regulation by MBD2 we generated a tagged version of the protein and stably expressed it in the MCF-7 cell line. We mapped genome wide binding of MBD2 by ChIP sequencing (ChIP-seq) and together with base resolution whole genome bisulfite sequencing (WGBS) we were able to determine the methylation content and the role of methylation density at MBD2 enriched regions. We further dissected MBD2 binding properties, taking advantage of a large set of ChIP-seq data including active histone marks, RNA polymerase II (POL2) and strand specific RNA-seq.","project":"SRP037971"}
{"number_samples":8,"species":"human","abstract":"we report transcritomic characterization of changes in CTNNB1 mutation positive HCCs in comparison to paratumoral tissues Overall design: 4 pair of HCC and paratumoral liver tissues are sequenced and compared","project":"SRP037982"}
{"number_samples":8,"species":"human","abstract":"Domain analysis of NKX2-2's role in the regulation of transcription in Ewing sarcoma.","project":"SRP038006"}
{"number_samples":6,"species":"human","abstract":"AML3 cells were treated with Azacytidine and compared against untreated cells Overall design: We used RNA-Seq to detail the global programme of gene expression in human untreated and Azacytidine treated AML3 cells","project":"SRP038101"}
{"number_samples":3,"species":"human","abstract":"Whole transcriptome sequencing for three common pancreatic cancer cell lines.","project":"SRP038143"}
{"number_samples":12,"species":"human","abstract":"Eukaryotic cells rapidly reduce protein synthesis in response to various stress conditions. This can be achieved by the phosphorylation-mediated inactivation of a key translation initiation factor, eIF2. However, the persistent translation of certain mRNAs is required for deployment of an adequate stress response. We carried out ribosome profiling of cultured human cells under conditions of severe stress induced with sodium arsenite. Although this led to a ~4.5-fold general translational repression, the protein coding ORFs of certain individual mRNAs exhibited resistance to the inhibition. Nearly all resistant transcripts possess at least one efficiently translated uORF that repress translation of the main coding ORF under normal conditions. Site specific mutagenesis of two identified stress resistant mRNAs (PPP1R15B and IFRD1) demonstrated that a single uORF is sufficient for eIF2-mediated translation control in both cases. Phylogenetic analysis suggests that at least two regulatory uORFs (namely in SLC35A4 and MIEF1) encode functional protein products. Overall design: Ribosome profiling of sodium arsenite treated cells for examination of translational response to induction of eIF2 phosphoylation","project":"SRP038695"}
{"number_samples":4,"species":"human","abstract":"To identify QKI targets, we performed QKI knockdown in BEAS2B cells and analyzed alternative splicing patterns by high-throughput RNA sequencing. Overall design: The mRNA profiles of control- and QKI-knockdown BEAS2B cells were generated by deep sequencing using Illumina GAIIx sequencer.","project":"SRP038702"}
{"number_samples":9,"species":"human","abstract":"The detachment of epithelial cells, but not cancer cells, causes anoikis due to reduced energy production. Invasive tumor cells generate three splice variants of the metastasis gene osteopontin. The cancer-specific form osteopontin-c supports anchorage-independence through inducing oxidoreductases and upregulating intermediates/enzymes in the hexose monophosphate shunt, glutathione cycle, glycolysis, glycerol phosphate shuttle, and mitochondrial respiratory chain. Osteopontin-c signaling upregulates glutathione (consistent with the induction of the enzyme GPX-4), glutamine and glutamate (which can feed into the tricarboxylic acid cycle). Consecutively, the cellular ATP levels are elevated. The elevated creatine may be synthesized from serine via glycine and also supports the energy metabolism by increasing the formation of ATP. Metabolic probing with N-acetyl-L-cysteine, L-glutamate, or glycerol identified differentially regulated pathway components, with mitochondrial activity being redox dependent and the creatine pathway depending on glutamine. The effects are consistent with a stimulation of the energy metabolism that supports anti-anoikis. Our findings imply a synergism in cancer cells between osteopontin-a, which increases the cellular glucose levels, and osteopontin-c, which utilizes this glucose to generate energy. Overall design: mRNA profiles of MCF-7 cells transfected with  osteopontin-a, osteopontin-c and vector control were generated by RNA-Seq, in triplicate, by Illumina HiSeq.","project":"SRP038726"}
{"number_samples":4,"species":"human","abstract":"We have used RNA-seq to examine gene expression from paired human CRC/control mucosa samples and identified CRC-specific expressed long noncoding RNAs Overall design: To identify CRC-specific expressed genes, including long noncoding RNAs","project":"SRP038759"}
{"number_samples":3,"species":"human","abstract":"Major- and minor-group rhinoviruses enter their host by binding to the cell surface molecules ICAM-1 and LDL-R, respectively, which are present on both macrophages and epithelial cells. Although epithelial cells are the primary site of productive HRV infection, previous studies have implicated macrophages in establishing the cytokine dysregulation that occurs during rhinovirus-induced asthma exacerbations. Even though major- and minor-group rhinoviruses are nearly genetically identical, these viruses do not replicate with equal success in monocyte-lineage cell lines. In human primary macrophages, differential mitochondrial activity and signaling pathway activation was observed between major- and minor-group rhinovirus upon initial HRV binding, indicating discordant receptor-dependent response to these rhinovirus types. As well, variances in phosphorylation of kinases (p38, JNK, ERK5) and transcription factors (ATF-2, CREB, CEBP-alpha) were observed between the major- and minor- group HRV treatments. The difference between major- and minor- group HRV activation of signaling pathways was confirmed through RNA-sequencing and observation of differential production of the asthma-relevant cytokines CCL20, CCL2, and IL-10. This is the first report of genetically similar viruses eliciting dissimilar cytokine release, transcription factor phosphorylation, and MAPK activation from macrophages. These results suggest that receptor dependence plays a role in establishing the inflammatory microenvironment initiated in part by monocytic-lineage cells in the human airway upon exposure to rhinovirus. Overall design: RNA sequencing of monocyte-derived macrophages after mock infection or infection by HRV16 or HRV1A","project":"SRP038761"}
{"number_samples":4,"species":"human","abstract":"We report the derivation of 2 different methods of generating cardiac myocytes from human ESCs. The traditional route is via cardiac progenitor cells and the second, new approach is through re-directing hemogenic endothelium into the cardiac lineage using inhibition of Wnt/b-catenin signaling Overall design: Examination of 2 different cardiac populations using RNA-seq","project":"SRP038767"}
{"number_samples":31,"species":"human","abstract":"We report on explant osteoblast cultures from human patients, demonstrating that there are at least three sub-types of non-syndromic craniosynostosis defined by similarity of gene expression profiles. Overall design: Osteoblast growth in culture, 23 craniosynostosis skull samples (7 metopic; 8 coronal; 3 lambdoid; 5 sagittal) and 8 normal (4 cranial bones and 4 long bones)","project":"SRP038863"}
{"number_samples":6,"species":"human","abstract":"Dicer is a deeply conserved endoribonuclease with key functions in small RNA biogenesis. Here we employed PAR-CLIP/iPAR-CLIP to identify direct Dicer binding sites in the transcriptomes of human cells and human. We found hundreds of novel miRNAs and non-canonical Dicer substrates with high sensitivity. Small RNA production depended on structure of the binding site and is globally biased towards the 5' arm of hairpins. Unexpectedly, in both species Dicer bound numerous hairpins inside mRNAs without observable small RNA production. Our data revealed ~100 mRNAs of protein coding genes to be targeted in both human and worm. These mRNAs significantly overlapped with the RNAi pathway. We also, unexpectedly, found that mitochondrial transcripts are Dicer targets in both species. We demonstrate functional consequences of Dicer binding by perturbation analysis. Taken together,we provide the first genome-wide catalog of direct Dicer targets. Our results suggest widespread function outside of miRNA biogenesis. Overall design: PAR-CLIP basically as described previously (Hafner et al. 2010).","project":"SRP038919"}
{"number_samples":3,"species":"human","abstract":"Dicer is a deeply conserved endoribonuclease with key functions in small RNA biogenesis. Here we employed PAR-CLIP/iPAR-CLIP to identify direct Dicer binding sites in the transcriptomes of human cells and human. We found hundreds of novel miRNAs and non-canonical Dicer substrates with high sensitivity. Small RNA production depended on structure of the binding site and is globally biased towards the 5' arm of hairpins. Unexpectedly, in both species Dicer bound numerous hairpins inside mRNAs without observable small RNA production. Our data revealed ~100 mRNAs of protein coding genes to be targeted in both human and worm. These mRNAs significantly overlapped with the RNAi pathway. We also, unexpectedly, found that mitochondrial transcripts are Dicer targets in both species. We demonstrate functional consequences of Dicer binding by perturbation analysis. Taken together,we provide the first genome-wide catalog of direct Dicer targets. Our results suggest widespread function outside of miRNA biogenesis. Overall design: Regulatory impact of Dicer binding was assessed by knock down experiments in human HEK293 cells. Drosha knockdown and mock transfections were used as controls. In total 3 samples.","project":"SRP038921"}
{"number_samples":2,"species":"human","abstract":"Dicer is a deeply conserved endoribonuclease with key functions in small RNA biogenesis. Here we employed PAR-CLIP/iPAR-CLIP to identify direct Dicer binding sites in the transcriptomes of human cells and human. We found hundreds of novel miRNAs and non-canonical Dicer substrates with high sensitivity. Small RNA production depended on structure of the binding site and is globally biased towards the 5' arm of hairpins. Unexpectedly, in both species Dicer bound numerous hairpins inside mRNAs without observable small RNA production. Our data revealed ~100 mRNAs of protein coding genes to be targeted in both human and worm. These mRNAs significantly overlapped with the RNAi pathway. We also, unexpectedly, found that mitochondrial transcripts are Dicer targets in both species. We demonstrate functional consequences of Dicer binding by perturbation analysis. Taken together,we provide the first genome-wide catalog of direct Dicer targets. Our results suggest widespread function outside of miRNA biogenesis. Overall design: Argonaute loaded small RNAs were extracted from FLAG:AGO2 and FLAG:AGO3 expressing HEK293 cells. Small RNA was purified and length selected (see supplementary methods).","project":"SRP038925"}
{"number_samples":2,"species":"human","abstract":"ADARs are the primary factors underlying A-to-I editing in metazoans. We conducted the first global study of ADAR1-RNA interaction in human cells using CLIP-Seq. In contrast to the expected predominant binding of ADAR1 to Alu repeats, thousands of CLIP sites were located in non-Alu regions. This unexpectedly frequent non-Alu binding enabled discovery of transcriptome-wide functional and biophysical targets of ADAR1 in the regulation of mRNA processing including alternative 3' UTR usage and alternative splicing. In addition, a global analysis of ADAR1 binding to non-Alu regions also revealed its primary interaction with microRNA (miRNA) transcripts in the nucleus, which subsequently affected expression levels of mature miRNAs. A complex global picture was revealed regarding the dependence of this function on the double-stranded RNA binding domains or deaminase activity. Our study unfolded a broad landscape of the diverse functional roles of ADAR1. Overall design: To charaterize ADAR1 binding profiles in U87 cells, we performed CLIP-seq using two different ADAR1 monoclonal antibodies.","project":"SRP038963"}
{"number_samples":10,"species":"human","abstract":"ADARs are the primary factors underlying A-to-I editing in metazoans. We conducted the first global study of ADAR1-RNA interaction in human cells using CLIP-Seq. In contrast to the expected predominant binding of ADAR1 to Alu repeats, thousands of CLIP sites were located in non-Alu regions. This unexpectedly frequent non-Alu binding enabled discovery of transcriptome-wide functional and biophysical targets of ADAR1 in the regulation of mRNA processing including alternative 3' UTR usage and alternative splicing. In addition, a global analysis of ADAR1 binding to non-Alu regions also revealed its primary interaction with microRNA (miRNA) transcripts in the nucleus, which subsequently affected expression levels of mature miRNAs. A complex global picture was revealed regarding the dependence of this function on the double-stranded RNA binding domains or deaminase activity. Our study unfolded a broad landscape of the diverse functional roles of ADAR1. Overall design: To identify ADAR binding dependent miRNA defferential expression profiles, U87MG cells were transfected with ADAR1 overexpression vector, RNA binding mutant (EAA and E912A), siRNA of ADAR1 or controls.","project":"SRP038964"}
{"number_samples":4,"species":"human","abstract":"This proof-of-principle experiment was designed to demonstrate the feasibility of proximity labeling for RNA–protein interactions Overall design: IPL-seq on 293T-Rex expressing MSA-SNRPN70 (sample) or NFH-SNRPN70 (control)","project":"SRP038969"}
{"number_samples":2,"species":"human","abstract":"Analysis of 5-hydroxymethylcytosine changes in hypoxia in neuroblastoma cells Overall design: 5-hmC distribution was analyzed from cells exposed to hypoxia (1% O2) or normoxia (20% O2); mRNA was sequenced in parallel for expression analysis.","project":"SRP038987"}
{"number_samples":4,"species":"human","abstract":"RNA-seq on cryopreserved human hepatocytes from a 19 year old Caucasian male donor with no history of medications","project":"SRP039039"}
{"number_samples":61,"species":"human","abstract":"Airway epithelial cells (AEC) are critical components of the inflammatory and immune response during exposure to pathogens. AECs in monolayer culture and differentiated epithelial cells in air-liquid interface (ALI) represent two distinct and commonly used in vitro models, yet differences in their response to pathogens have not been investigated. In this study, we compared the transcriptional effects of flagellin on AECs in monolayer culture versus ALI culture using exon microarrays and RNAsequencing. We found that AECs cultured in monolayer and ALI have strikingly different transcriptional states at baseline. When challenged with flagellin, monolayer AEC cultures greatly increased transcription of numerous genes mapping to wounding response, immunity and inflammatory response. In contrast, AECs in ALI culture had an unexpectedly muted response to flagellin, both in number of genes expressed and relative enrichment of inflammatory and immune pathways. In conclusion, In vitro culturing methods have a dramatic effect on the transcriptional profile of AECs at baseline and after stimulation with flagellin. These differences suggest that epithelial responses to pathogen challenges are distinctly different in culture models of intact and injured epithelium. Overall design: A total of eight independent RNAseq experiments were conducted. Four RNAseq experiments (n = 2 unstimulated, n = 2 stimulated with flagellin) were performed using AECs grown in monolayer. Four RNAseq experiments (n =2 unstimulated, n = 2 stimulated with flagellin) were conducted using AECs grown in ALI cultures","project":"SRP039077"}
{"number_samples":8,"species":"human","abstract":"Transcriptome of human HL-60 and HEK-293 cells depending on culture cell density","project":"SRP039085"}
{"number_samples":4,"species":"human","abstract":"By mapping the genomic enrichments  of H3K4me3 and H3K27me3 modifications in pure populations of hESCs during the G2, mitotic and G1 phases of the cell cycle, we characterize cell cycle-dependent variations in the epigenetic landscape of bivalent genes, altering the current view of mitotic inheritance in pluripotent cells. We identified novel classes of bivalent domains that are highly enriched with H3K4me3 during mitosis, depleted during G1 only, and ubiquitously bivalent. These bivalent domains are associated with specific genes and expression patterns during differentiation. These cell cycle-dependent epigenetic profiles are unique to hESCs and are not observed following initiation of phenotype commitment. Our results establish a new dimension in cell cycle-dependent chromatin regulation that advances understanding of contributions the pluripotent epigenetic landscape to hESC identity. Overall design: Study of two histone modifications, H3K4me3 and H3K27me3, during three cell cycle phases in two cell types. Study of gene expression from these two cell types in asynchronous cells.","project":"SRP039346"}
{"number_samples":32,"species":"human","abstract":"Trisomy 21 (T21) is the most frequent genetic cause of cognitive impairment. To assess the perturbations of gene expression in T21, and to eliminate the noise of the genomic variability, we studied the transcriptome of fetal fibroblasts from a pair of monozygotic twins discordant for T21. Here we show that the differential expression between the twins is organized in domains along all chromosomes that are either up- or downregulated. These gene expression dysregulation domains (GEDDs) can be defined by the expression level of their gene content, and are well conserved in induced pluripotent stem cells derived from the twins’ fibroblasts. Comparison of the transcriptome of the Ts65Dn mouse model of DS and wild-type, also showed GEDDs along the mouse chromosomes that were syntenic in human. The GEDDs correlate with the lamina-associated (LADs) and replication domains of mammalian cells. The overall LADs position was not altered in trisomic cells. However, the H3K4me3 profile of the trisomic fibroblasts was modified and accurately followed the GEDD pattern. These results suggest that the nuclear compartments of trisomic cells undergo modifications of the chromatin environment influencing the overall transcriptome and that GEDDs may therefore contribute to some T21 phenotypes. Overall design: mRNA-Seq profiling in Down syndrome: fibroblasts derived from a pair of monozygotic twins discordant for trisomy 21 (4 replicates), iPS cells from the same pair of discordant twins, fibroblasts from a pair of normal monozygotic twins, fibroblasts from 16 unrelated individuals (8 trisomic and 8 euploid controls), fibroblasts from the Ts65Dn mouse model of Down syndrome (1 trisomic mouse and 1 control wt).","project":"SRP039348"}
{"number_samples":4,"species":"human","abstract":"This work provides a comprehensive and global perspective of gene expression in human epidermal melanocytes using RNA-Seq. Our focus on cell signalling genes and differential expression between lightly and darkly pigmented melanocytes may help to identify novel pigmentation genes and potential pharmacological targets.","project":"SRP039354"}
{"number_samples":2,"species":"human","abstract":"Massively parallel sequencing of human urinary exosome/microvesicle RNA reveals a predominance of non-coding RNA","project":"SRP039357"}
{"number_samples":31,"species":"human","abstract":"The rate of transcription elongation plays important roles in the timing of expression of full-length transcripts as well as for the regulation of alternative splicing. In this study we coupled Bru-Seq technology with 5,6-dichlorobenzimidazole 1-ß-D-ribofuranoside (DRB) to estimate the elongation rates of over 2,000 individual genes in human cells. This technique, BruDRB-Seq, revealed gene-specific differences in elongation rates with a median rate of around 1.5 kb/min. We found that genes with fast elongation rates showed higher densities of H3K79m2 and H4K20me1 marks compared to slower elongating genes. Furthermore, fast elongation rates had a positive correlation with gene length, low complexity DNA sequence and distance from nearest active transcription unit. Features that negatively correlated with elongation rate included exon density and the number of LINE sequences in the gene. The BruDRB-Seq technique offers new opportunities to interrogate mechanisms of regulation of transcription elongation. Overall design: Measurement of RNA Pol II elogation rate. Normal fibroblasts (HF1 and TM), Cockayne syndrome group B fibroblasts, K562 and MCF-7 cells were exposed to DRB for 60 minutes, after which a washout was performed. Nascent RNA was labeled using bromouridine for 10 minutes immediately after the washout. The genomic region extending from actice Trancription Start Sites was used to determine the gene's elongation rate. Please note that the nf_0h_3* samples are duplicated sample records of GSM1062445 and GSM1062446, for the convenient retrieval of the complete raw data from SRA.","project":"SRP039359"}
{"number_samples":33,"species":"human","abstract":"Using RNA-Seq, we reported novel findings in the comparison of transcriptome profiles of isogenic HMDM and IPSDM during differentiation and polarization. First, IPSDM lost expression of pluripotency markers, had remarkably distinct gene expression profiles relative to precursor iPSCs, and had largely similar gene expression as HMDM. Second, macrophage polarization to M1 was associated with a dramatic change in the transcriptome; expression profiles of IPSDM- and HMDM-derived M1 lines were highly correlated with each other but much less so with their respective IPSDM and HMDM precursors. Third, M2-HMDM lines had limited difference in gene expression compared to their non-polarized precursors, likely due to the known M2-like phenotype of M-CSF differentiated macrophages and their similarity to the IL-4 derived M2 phenotype Finally, through RNA-Seq we identified many new genes modulated during polarization in both HMDM and IPSDM thus providing novel, and potentially regulatory, candidates that warrant further study. Overall design: iPS, IPSDM (including M1/M2) and HMDM (including M1/M2)cells were sequenced by Illumina HiSeq 2000 with poly-A selection","project":"SRP039361"}
{"number_samples":55,"species":"human","abstract":"N6-methyladenosine (m6A) is a common modification of mRNA, with potential roles in fine-tuning the RNA life cycle, but little is known about the pathways regulating this process and its physiological role. Here, we used mass-spectrometry to identify a dense network of proteins physically interacting with METTL3, a core component of the methyltransferase complex, and show that two of them, WTAP and KIAA1429, are required for methylation.  Combining high resolution m6A-Seq with knockdown of WTAP allowed us to define accurate maps, at near single-nucleotide resolution, of sites of mRNA methylation across four dynamic programs in human and mouse, including development, differentiation, reprogramming and immune response. Internal WTAP-dependent methylation sites were largely static across the different surveyed conditions and present in the majority of mRNAs. However, methylations were found at much lower levels within highly expressed mRNAs, and methylation is inversely correlated with mRNA stability, consistent with a role in establishing an overall basal, cell-type invariant, distribution of degradation rates. In addition, we identify thousands of WTAP-independent methylation sites at transcription initiation sites, forming part of the mRNA cap structure. We show that the methylations occur at the first transcribed nucleotide, and find that thousands of transcripts are present in different isoforms differing in the methylation state of the first transcribed nucleotide, a previously unappreciated complexity of the transcriptome. Together, our data sheds new light on the proteomic and transcriptional underpinnings of this epitranscriptomic modification in mammals. Overall design: Examination of m6A methylation in human Hek293 and A549 cell lines, in human embryonic stem cells (ESCs) undergoing differentiation to neural progenitor cells (NPCs),  in OKMS inducible fibroblasts reprogrammed into iPSC, and upon knockdown of factors using siRNAs or shRNAs.","project":"SRP039397"}
{"number_samples":2,"species":"human","abstract":"Rectal cancer stage II carcinoma and adjacent normal tissue transcriptome","project":"SRP039399"}
{"number_samples":1,"species":"human","abstract":"identify the target of CELF1 protein","project":"SRP039460"}
{"number_samples":4,"species":"human","abstract":"We identify here host genomic regions that are more likely to be damaged during an infection with the human gastirc pathogen Helicobacter pylori and characterize underlying features of them. By ChIP-seq we obtained global information about the localization of the DNA-damage marker H2AXpS139. We used 20 ng of immunoprecipitated DNA of the human gastric cell line AGS to produce 240 mio sequence reads in total and compared the damage-pattern of 6 h and 19 h infected cells with that of 10 Gy irradiated ones. While the overall levesl of DNA damage wetre similar in terms of broken DNA and accumulation of H2AXpS139, infection-induced damage clearly accumulates towards the ends of chromosomal arms, namely telomere-proximal and close to centromeres, it also enriches mainly in genic and especially in transcribed regions. We show, taht this damage pattern is unique to the infection, occurs also in human non-transformed gastric epithelial cells and shows some correlation with previously identified cancer-related genes. Overall design: Examination of a DNA-damage induced histone modification from four different conditions.","project":"SRP039552"}
{"number_samples":2,"species":"human","abstract":"Rbfox target network (HITS-CLIP in mouse brain)","project":"SRP039559"}
{"number_samples":8,"species":"human","abstract":"Hepatosplenic T-cell lymphoma (HSTL) is an aggressive lymphoma cytogenetically characterized by isochromosome 7q   [i(7)(q10)], of which the molecular consequences remain unknown. We report here results of an integrative genomic and transcriptomic (expression microarray and RNA-sequencing) study of six HSTL cases with i(7)(q10) and three cases with ring 7   [r(7)], a rare variant aberration. Using high resolution array CGH, we prove that HSTL is characterized by the common loss of a 34.88 Mb region at 7p22.1p14.1 (3506316-38406226 bp) and duplication/amplification of a 38.77 Mb region at 7q22.11q31.1 (86259620-124892276 bp). Our data indicate that i(7)(q10)/r(7)-associated loss of 7p22.1p14.1 is a critical event in the development of HSTL, while gain of 7q sequences drives progression of the disease and underlies its intrinsic chemoresistance. Loss of 7p22.1p14.1 does not target a postulated tumor suppressor gene but unexpectedly enhances the expression of CHN2 from the remaining 7p allele, resulting in dysregulation of a pathway involving RAC1 and NFATC2 with a cell proliferation response. Gain of 7q leads to increased expression of critical genes, including RUNDC3B, PPP1R9A and ABCB1, a known multidrug resistance gene. RNA-sequencing did not identify any additional recurrent mutations or fusion, suggesting that i(7)(q10) is the only driver event in this tumor. Our study confirms the previously described gene expression profile of HSTL and identifies a set of 24 genes, including three located on chromosome 7 (CHN2, ABCB1 and PPP1R9A), distinguishing HSTL from other malignancies Overall design: The fastq files of all samples were mapped to the reference human genome (assembly GRCh37.68). The mapping was performed  allowing detection of insertions and deletions (indels). The mapped reads were used to calculate read counts and FPKM (Fragment Per Kilobase of exon model per Million of mapped read) per gene. The DESeq algorithm was applied to identify differentially expressed genes. The mapped reads were used to predict mutations and gene fusions.","project":"SRP039591"}
{"number_samples":8,"species":"human","abstract":"The tumor suppressor p53 has been studied extensively as a direct transcriptional activator of protein-coding genes.  Recent studies, however, have shed light on novel regulatory functions of p53 within noncoding regions of the genome.  Here, we use a systematic approach that integrates transcriptome-wide differential expression analysis, genome-wide p53 binding profiles, chromatin state maps, and additional genomic features to characterize the global regulatory roles of p53 in response to DNA damage in both human and mouse fibroblast models.  In addition to known p53 targets, we identify many previously unappreciated mRNAs and long noncoding RNAs that are regulated by p53.  Moreover, we find that p53 binding events occur predominantly within enhancer elements in both human and mouse systems.  The ability to modulate enhancer activity offers an additional layer of complexity to the p53 network and greatly expands the diversity of genomic elements that are directly regulated by p53. Overall design: Human and Mouse fibroblasts cultured in the presence or absence of doxorubicin followed by RNA-Seq (Human:2 cell lines, each condition in duplicate; Mouse:MEF cell line,each condition in triplicate) and p53 ChIP-Seq (Human:2 cell lines, input and IP for each; Mouse:MEF cell line, input and IP)","project":"SRP039598"}
{"number_samples":16,"species":"human","abstract":"Hepatocellular carcinoma (HCC) is one of the most common and lethal cancers worldwide and has a poor prognosis. Promoters represent an essential regulatory element of gene transcription in human genome. In order to understand the promoter methylation in relation with gene transcription in HCCs, We applied a liquid hybridization capture-based bisulfite sequencing (LHC-BS) approach to examine the promoter methylome of HCCs, for which we customized 150,407 capture probes and enabled coverage of 91.8% of the RefSeq gene promoters within human genome. We found the differential promoter DNA methylation between HCCs and peripheral normal tissues. Then we integrated promoter methylomic and transcriptomic profiling and described gene expression and regulation in HCCs. Lastly, We validated the key genes in larger number of samples and screened candidate genes aberrantly regulated by DNA methylation in human HCCs. Overall design: RNA-seq for 8 pairs of HCC tumor and non-tumor liver (NTL) samples.","project":"SRP039694"}
{"number_samples":4,"species":"human","abstract":"Identification of the RNA recognition element of the RBPMS family of RNA-binding proteins and their transcriptome-wide mRNA targets","project":"SRP040014"}
{"number_samples":134,"species":"human","abstract":"Transcriptomic analysis of the Novel Middle East Respiratory Syndrome Coronavirus (MERS-CoV)","project":"SRP040070"}
{"number_samples":4,"species":"human","abstract":"Here we report an approach that has permitted us to uncover the sites and mechanisms of action of a drug, referred to as SD70, initially identified by phenotypic screening for inhibitors of ligand and genotoxic stress-induced translocations in prostate cancer cells. Based on synthesis of a derivatized form of SD70 that permits its application for a ChIP-seq-like approach, referred to as Drug-seq, we were next able to efficiently map the genome-wide binding locations of this small molecule, revealing that it largely co-localized with androgen receptor (AR) on regulatory enhancers. Based on these observations, we performed the appropriate global analyses to ascertain that SD70 inhibits the androgen-dependent AR program, and prostate cancer cell growth, acting, at least in part, by functionally inhibiting the jumonji (JMJ) domain-containing demethylase, KDM4C. Drug-seq represents a powerful strategy for new drug development by mapping genome-wide location of small molecules, a powerful adjunct to contemporary drug development strategies. Overall design: Global Run-On (GRO) assay followed by high-throughput sequencing (GRO-seq).","project":"SRP040117"}
{"number_samples":12,"species":"human","abstract":"The estrogen receptor-a (ERa) is a transcription factor which plays a critical role in controlling cell proliferation and tumorigenesis by recruiting various cofactors to estrogen response elements (EREs) to induce or repress gene transcription. A deeper understanding of these transcriptional mechanisms may uncover novel therapeutic targets for ERa-dependent cancers. Here we show for the first time that BRD4 regulates ERa-induced gene expression by affecting elongation-associated phosphorylation of RNA Polymerase II (RNAPII P-Ser2) and histone H2B monoubiquitination (H2Bub1). Consistently, BRD4 activity is required for estrogen-induced proliferation of ER+ breast and endometrial cancer cells and uterine growth in mice. Genome-wide occupancy studies revealed an enrichment of BRD4 on transcriptional start sites as well as EREs enriched for H3K27ac and demonstrate a requirement for BRD4 for H2B monoubiquitination in the transcribed region of estrogen-responsive genes. Importantly, we further demonstrate that BRD4 occupancy correlates with active mRNA transcription and is required for the production of ERa-dependent enhancer RNAs (eRNAs). These results uncover BRD4 as a central regulator of ERa function and potential therapeutic target. Overall design: mRNA expression profiles of MCF7 cells treated with +/- estrogen treatment under negative control siRNA, BRD4 siRNA or JQ1 treatment, in duplicates.","project":"SRP040136"}
{"number_samples":13,"species":"human","abstract":"Induced pluripotent stem cells (iPSCs) offer opportunity for insight into the genetic requirements of the X chromosome for somatic and germline development. Turner syndrome is caused by complete or partial loss of the second sex chromosome; while more than 90% of Turner cases result in spontaneous fetal loss, survivors display an array of somatic and germline clinical characteristics. Here, we derived iPSCs from Turner syndrome and control individuals and examined germ cell development as a function of X chromosome composition. We analyzed gene expression profiles of derived iPSCs and in vitro differentiated cells by single cell qRT-PCR and RNA-seq. We whoed that two X chromosomes are not necessary for reprogramming or pluripotency maintenance. Genes that escape X chromosome inactivation (XCI) between control iPSCs and those with X chromosome aneuploidies revealed minimal expression differences relative to a female hESC line. Moreover, when we induced germ cell differentiation via murine xenotransplantation of iPSC lines into the seminiferous tubules of busulfan-treated mice, we observed that undifferentiated iPSCs, independent of X chromosome composition, when placed within the correct somatic environment, are capable of forming early germ cells in vivo. Results indicate that two intact X chromosomes are not required for germ cell formation; however, clinical data suggest that two sex chromosomes are required for maintenance of human germ cells. Overall design: RNA-seq of  H9 cells, iPSCs from Turner syndrome and control individuals and in vitro differentiated cells","project":"SRP040145"}
{"number_samples":15,"species":"human","abstract":"The paired-end next-generation sequencing of all small RNAs of less than 200 nucleotides in length from four different human cell lines (SKOV3ip1, MCF-7, BJ-Tielf, INOF) allowed us to determine the exact sequence(s) and variations of human box C/D snoRNAs (small nucleolar RNAs), revealing processing patterns of this class of molecules.   Two distinct groups of box C/D snoRNAs were identified based on the position of their ends with respect to their characteristic boxes and the terminal base pairing potential. Short box C/D snoRNAs start sharply 4 or 5 nucleotides upstream of their box C and end 2 or 3 nucleotides downstream of their box D. In contrast, long box C/D snoRNAs start 5 or 6 nucleotides upstream of their box C and end 4 or 5 nucleotides downstream of their box D, increasing the likelihood of formation of a k-turn between their boxes C and D. Sequencing of SKOV3ip1 cells following the depletions of NOP58, a core box C/D snoRNA-binding protein and of RBFOX2, a splicing factor, shows that the short box C/D snoRNA forms are significantly more affected by the depletion of RBFOX2 while the long snoRNA forms, which display more canonical box C/D snoRNA features, are significantly more affected by the depletion of NOP58. Together the data suggest that box C/D snoRNAs are divided into at least two groups of RNA with distinct maturation and functional preferences. Overall design: Small RNAs (<200 nucleotides) were isolated from different human cell lines that were either untreated or depleted of NOP58 or RBFOX2 using specific siRNAs. The resulting libraries were multiplexed and paired-end sequenced using Illumina HiSeq.","project":"SRP040236"}
{"number_samples":4,"species":"human","abstract":"In this study, we have uncovered novel proteolytic processing of the histone H3 tail in senescence models in primary fibroblasts and melanocytes. Cleavage of H3 tail occurs at two distinct residues and is mediated by Cathepsin L. We show that variant H3.3 is preferentially cleaved, and that cleaved histones are associated with chromatin and incorporated into nucleosomes.  We also found that the histone chaperone ASF1a is required for chromatin incorporation of the cleaved histone species. Further, we show that overexpression of cleaved H3.3 induces a senescence program in fibroblasts in the absensence of oncogenic signaling. Overall design: For the RNA-seq studies, growing IMR90 fibroblasts were compared to cells induced to senesce via oncogene activation or cleaved H3.3 overexpression. Growing controls consist of IMR90 cells infected with empty retroviral construct pBabe and grown under normal conditions for 13 days prior to RNA isolation. For oncogene-induced senescence samples, IMR90s carrying a tamoxifen-inducible H-RasV12 retroviral construct were induced to senesce by addition of 10nM tamoxifen to the media for 8 days. Finally, IMR90s were infected with a retroviral construct expressing the cleaved form of H3.3 with a C-terminal Flag tag. RNA samples form this group were isolated at days 3 (early) and 13 (late) post-infection. In all cases, total RNA samples were isolated using RNeasy kit (Qiagen) and prepared at the Icahn School of Medicine at Mount Sinai Genomics Core Facility for poly A library construction and sequencing on IlluminaHiSeq 2500.","project":"SRP040243"}
{"number_samples":18,"species":"human","abstract":"We show that N6-methyladenosine (m6A), the most abundant internal modification in mRNA/lncRNA with still poorly characterized function, alters RNA structure to facilitate the access of RBM for heterogeneous nuclear ribonucleoprotein C (hnRNP C). We term this mechanism m6A-switch. Through combining PAR-CLIP with Me-RIP, we identify 39,060 m6A-switches among hnRNP C binding sites transcriptome-wide. We show that m6A-methyltransferases METTL3 or METTL14 knockdown decreases hnRNP C binding at 16,582 m6A-switches. Taken together, 2,798 m6A-switches of high confidence are identified to mediate RNA-hnRNP C interactions and affect diverse biological processes including cell cycle regulation. These findings reveal the biological importance of m6A and provide insights into the sophisticated regulation of RNA-RBP interactions through m6A-induced RNA structural remodeling. Overall design: Measure the m6A methylated hnRNP C binding sites transcriptome-wide by PARCLIP-MeRIP; measure the differential hnRNP C occupancies upon METTL3/METTL14 knockdown by PAR-CLIP; measure RNA abundance and splicing level changes upon HNRNPC, METTL3 and METTL14 knockdown","project":"SRP040278"}
{"number_samples":96,"species":"human","abstract":"K562 single cell RNA-seq study","project":"SRP040288"}
{"number_samples":1,"species":"human","abstract":"We performed mRNA-seq of a PRKACA-mutant adrenal tumor and demonstrated that the mutation is expressed at the mRNA level. Overall design: Total RNA obtained from a cortisol-producing adrenal tumor with a PRKACA p.Leu206Arg mutation.","project":"SRP040292"}
{"number_samples":10,"species":"human","abstract":"Glucocorticoids (GC) have been widely used as coadjuvants in the treatment of solid tumors, but GC treatment may be associated with poor pharmacotherapeutic response and/or prognosis. The genomic action of GC in these tumors is largely unknown. Here we find that dexamethasone (Dex, a synthetic GC) regulated genes in triple-negative breast cancer (TNBC) cells are associated with drug resistance. Importantly, these GC-regulated genes are aberrantly expressed in TNBC patients and associated with unfavorable clinical outcomes. Interestingly, in TNBC cells, Compound A (CpdA, a selective GR modulator) only regulates a small number of genes not involved in carcinogenesis and therapy resistance. Mechanistic studies using a ChIP-exo approach reveal that Dex- but not CpdA-liganded glucocorticoid receptor (GR) binds to a single glucocorticoid response element (GRE), which drives the expression of pro-tumorigenic genes. Our data suggest that development of safe coadjuvant therapy should consider the distinct genomic function between Dex- and CpdA-liganded GR. Overall design: To study GR-regulated genes and define GRE in human genome, RNA-seq and GR ChIP-exo are performed in MDA-MB-231 cells before/after dex and CpdA stimulation. Each experiment includes two replicates.","project":"SRP040300"}
{"number_samples":19,"species":"human","abstract":"Study single cell transcriptional heterogeneity","project":"SRP040309"}
{"number_samples":2,"species":"human","abstract":"Alterations in chromatin modifications, including DNA methylation and histone modification patterns, have been characterized under exposure of several environmental pollutants, including nickel. As with other carcinogenic metals, the mutagenic potential of nickel compounds is low and is not well correlated with its carcinogenic effects. Nickel exposure, however, is associated with alterations in chromatin modifications and related transcriptional programs, suggesting an alternative pathway whereby nickel exposure can lead to disease. To investigate the extent to which nickel exposure disrupts chromatin patterns, we profiled several histone modifications, including H3K4me3, H3K9ac, H3K27me3 and H3K9me2 as well as the insulator binding protein CTCF and the transcriptomes of control BEAS-2B cells and cells treated with nickel for 72 hours. Our results show significant alterations of the repressive histone modification H3K9me2 in nickel-exposed cells with spreading of H3K9me2 into new domains associated with gene silencing. We furthermore show that local regions of active chromatin can protect genes from nickel-induced H3K9me2 spreading. Interestingly, we show that nickel exposure selectively disrupts weaker CTCF sites, leading to spreading of H3K9me2 at these regions. These results have major implications in the understanding of how environmental carcinogens can affect chromatin dynamics and the consequences of chromatin domain disruption in disease progression. Overall design: Treat BEAS-2B cells with NiCl2 for 72 hours and compare histone modification, CTCF binding to control BEAS-2B cells to see how they regulated gene expression by RNA-seq","project":"SRP040327"}
{"number_samples":19,"species":"human","abstract":"Mucosal-associated invariant T (MAIT) cells are abundant in humans and recognize conserved bacterial antigens derived from riboflavin precursors, presented by the non-polymorphic MHC class I-like molecule MR1. Here, we show via transcriptomic analysis that human MAIT cells are remarkably oligoclonal in both blood and liver, display high inter-individual homology, and exhibit a restricted length CDR3ß domain of the TCRVß chain. We extend this analysis to a second sub-population of MAIT cells expressing a semi-invariant TCR conserved between individuals. Overall design: Study of CDR3 regions of TCRalpha and beta sequences","project":"SRP040328"}
{"number_samples":30,"species":"human","abstract":"The MCF-7 were infected with either control adenovirus expressing B-galactosidase (Ad) or adenovirus expressing ERB (AdERbeta) for 72 h.  For knockdown of the endogenous ERa in MCF-7 cells, cells were treated with siRNA for 24h (AdERbeta+SiERalpha).  Then cells were treated with Veh (0.1% EtOH), 10 nM E2 or 1 uM BEs (botanical extracts) for 24h. Overall design: Duplicate samples run; treatment after knockdown included a control treatment (V), estradiol (E2) or botanical extracts; genistein (Gen), S-equol, liquiritigenin (Liq)","project":"SRP040418"}
{"number_samples":12,"species":"human","abstract":"Preparation of exosomes isolated from semen contain a substantial amount of RNA, mostly from 20 to 100 nucleotides in length. We sequenced separately 20-40 and 40-100 nucleotide fractions of  RNA from exosomes isolated from semenal fluid from six healthy donors. We found various classes of small non-coding RNA, including mature microRNA and piwi-RNA,  as well as abundant Y RNAs and tRNAs present in both full length and fragmented forms. Specific RNAs were consistently present in all donors. For example, fifteen (of ~2,600 known) microRNAs constituted over 80% of mature microRNA in SE. Additionally, tRNA fragments were strongly enriched for 5’-ends of 18-19 or 30-34 nucleotides in length. Overall design: Size-fractionated small RNA profiles from exosomes isolated from the seminal fluid of six healthy donors","project":"SRP040421"}
{"number_samples":36,"species":"human","abstract":"Mammals have evolved an XY sex chromosome system, resulting in dosage imbalance not only between sexes, but also between X-chromosome and autosome. Overall design: mRNA profiles of 9 pairs of human endometrial carcinoma and adjacent tissues were generated by Illumina 100-nucleotide paired-end sequencing","project":"SRP040442"}
{"number_samples":9,"species":"human","abstract":"Rhabdomyosarcoma transcriptome sequencing","project":"SRP040454"}
{"number_samples":286,"species":"human","abstract":"Development of heterogeneous RNA-Seq (hRNA-Seq) to simultaneously capture prokaryotic and eukaryotic expression profiles of bacteria-infected cells.","project":"SRP040472"}
{"number_samples":22,"species":"human","abstract":"PIWI-interacting RNAs (piRNAs) are a novel class of small ncRNAs initially isolated from germ line cells; although recent studies report that they are expressed also in somatic cells. To elucidate the role of piRNAs in somatic cells, in particular from breast cancer, we performed the first extensive next generation sequencing expression analysis of small RNA transcriptomes of hormone responsive breast cancer cell lines in different culture conditions. In addition, to understand the behavior of piRNAs with respect to miRNAs in breast tumor tissues, small RNA sequence data set available in Gene Expression Omnibus (GSE39162)  database was used. Results led to the identification of ~100 and ~150 human piRNAs in the breast cancerous cell lines and tumors respectively, several of which differentially expressed in cell lines under different experimental conditions tested or in response to ERß and in tumor tissues. Western blotting and Q-PCR analysis also revealed the presence in breast cell lines of PIWIL (PIWI Like) subfamily members proteins encoded in the human genome (PIWIL2/HILI and PIWIL4/HIWI2) and of other components of the piRNA biogenesis pathways, suggesting that this might indeed be functional in somatic cells. These results show that piRNAs are expressed in human somatic cells, in particular in cancer, where their expression is influenced by neoplastic transformation, growth conditions and estrogen receptor beta. More important, we demonstrate for the first time a distinct pattern of piRNAs expression in cancerous vs normal breast tissues, which suggests a potential role of these epigenetic modulators in mammary carcinogenesis and maintenance of the cancer cell phenotype. Overall design: In addition, to understand the behavior of piRNAs with respect to miRNAs in breast tumor tissues, small RNA sequence data set available in Gene Expression Omnibus (GEO; GSM957192 TAX577740T ,GSM957194 TAX577740N, GSM957195 TAX577453T, GSM957197 TAX577453N, GSM957198 TAX577745T, GSM957200 TAX577745N, GSM957201 TAX577579T, GSM957203 TAX577579N) was used.","project":"SRP040505"}
{"number_samples":24,"species":"human","abstract":"Adenosine deaminases acting on RNA (ADARs) are involved in adenosine (A) to inosine (I) RNA editing and human ADARs are implicated in neurological diseases. Here we generated human embryonic stem cells (hESCs) lacking ADAR1 to investigate its role in neural development in a human context. We found that ADAR1 deficiency significantly retarded neural induction with widespread mRNA and miRNA expression changes. We have genome widely examined the changes of  A-to-I editing and miRNA expression after ADAR1 knockdown. Such aberrant mRNA and miRNA expression was not due to reduced A-to-I editing after ADAR1 knockdown. We further revealed that ADAR1 regulates miRNA biogenesis independent of its editing activity. Overall design: In order to study the function role of ADAR1 in neural induction, we used shRNA to stably knocked down ADAR1 in human embryonic stem cell (hESC) h9 line and then differentiated ADAR1 deficiency hESC h9 cells to specific types of neurons. We then collect total RNAs at different time points at undifferentiated stage (day 0, d0 for short), day 10 (d10), day 17 (d17) and day 35 (d35), and obtained RNA-seq and small RNA-seq datasets for further analyses.","project":"SRP040525"}
{"number_samples":201,"species":"human","abstract":"The goal of this project is to infect relevant human host cells (MonoMac6, adult human neural progenitor (AHNP) cells and AHNP-derived neurons) with Toxoplasma gondii parasites of differing lineages to generate transcriptional mRNA and miRNA profiles. These datasets will allow the characterization of how genetically different parasites that cause distinct types of human toxoplasmosis alter the expression of protein-encoding and miRNA-encoding genes in both the human host and the parasite.","project":"SRP040547"}
{"number_samples":2,"species":"human","abstract":"Purpose: Ribosome profiling and RNA-Seq were used to map the location and abundance of translating ribosomes on human skeletal muscle transcripts from a patient with Becker muscular dystrophy. Methods: Tissue homogenates were prepared from frozen sections of a muscle biopsy obtained from a patient with an NM_004006:c.40_41delGA dystrophin mutation and a normal control.  Ribosome-protected fragments and total RNA were prepared from a single homogenate, so starting RNA populations for both libraries were closely matched.  Homogenates were not clarified before RNase digestion to avoid loss of ribosomes associated with large molecular weight complexes and RNA-Seq libraries were prepared after rRNA subtraction to avoid positional loss of 5’ reads.   RPF-Seq libraries were built using the TruSeq Small RNA Sample Preparation Kit (Illumina) and RNA-Seq libraries were built using the TruSeq RNA Sample Preparation v2 Kit (Illumina). RPF-Seq and RNA-Seq libraries were subjected to 50 cycles of single-end sequencing on an Illumina HiSeq 2000 instrument.  Trimmed and filtered RPF- and RNA-Seq reads were mapped to RefSeq fasta sequences downloaded from the UCSC genome browser (hg19 assembly). Results: Most mutations that truncate the reading frame of the DMD gene result in loss of dystrophin expression and lead to Duchenne muscular dystrophy.  However, amelioration of disease severity can result from alternate translation initiation beginning in DMD exon 6 that results in the expression of a highly functional N-truncated dystrophin.  This novel isoform results from usage of an internal ribosome entry site (IRES) within exon 5 that is glucocorticoid-inducible. IRES activity is confirmed in patient muscle by both peptide sequencing and ribosomal profiling. Conclusions: Our results provide a molecular explanation for the rescue of 5’ truncating mutations via a heretofore undescribed mechanism of post-transcriptional regulation of dystrophin expression. The presence of a glucocorticoid-inducible IRES within a highly conserved region of the DMD sequence strongly suggests a programmed role for alternate translation initiation, and ongoing efforts to understand the relevant cell lineage-specific and/or conditional activation signals will shed light on underlying mechanisms of IRES control and elucidate potentially novel functions of dystrophin. Overall design: Skeletal muscle ribosome-protected fragment and RNA-Seq profiles from a patient with an NM_004006:c.40_41delGA dystrophin mutation and a normal control were generated by deep sequencing using the Illumina HiSeq 2000.","project":"SRP040550"}
{"number_samples":6,"species":"human","abstract":"The PR-domain family e(PRDMs) encodes transcriptional regulators, several of which are deregulated in cancer. We found that loss of Prdm11 accelerates MYC-driven lymphomagenesis in the Eµ-Myc mouse model. Moreover, we show that patients with PRDM11-deficient diffuse large B cell lymphomas (DLBCLs) have poorer overall survival and belong to the non-Germinal Center B cell (GCB)-like subtype. Mechanistically, genome-wide mapping of PRDM11 binding sites coupled with transcriptome sequencing in human DLBCL cells evidenced that PRDM11 associates with transcriptional start sites of target genes and regulates important oncogenes such as FOS and JUN.  Hence, we characterize PRDM11 as a novel tumor suppressor controlling the expression of key oncogenes and add new mechanistic insight into B-cell lymphomagenesis. Overall design: RNA-seq performed after knockdown of Prdm11","project":"SRP040577"}
{"number_samples":30,"species":"human","abstract":"To assess the consequence of a lower quantity of input material on the quality of the resulting sequence data, RNA sequencing combined with exome capture sequencing - a technique we refer to as cDNA-Capture - was performed on varying amounts (60, 50, 10, 2 and 0.8ng) of MVP Disease Match Set Human Adult Colon RNA derived from normal and adenocarcinoma margin (Stratagene catalog number 540077). Each sample was sequenced in triplicate.","project":"SRP040586"}
{"number_samples":13,"species":"human","abstract":"It has been unclear whether ischemic stroke induces neurogenesis or neuronal DNA-rearrangements in the human neocortex. We show here that neither is the case, using immunohistochemistry, transcriptome-, genome- and ploidy-analyses, and determination of nuclear bomb test-derived 14C-concentration in neuronal DNA. A large proportion of cortical neurons display DNA-fragmentation and DNA-repair short time after stroke, whereas neurons at chronic stages after stroke show DNA-integrity, demonstrating the relevance of an intact genome for survival. Overall design: Analyze of potential fusion transcripts in 13 samples, seven cortical ischemic stroke tissue and six control cortex, by deep sequencing using Illumina HiSeq 2000","project":"SRP040622"}
{"number_samples":5,"species":"human","abstract":"Patients receiving mechanical ventilation for acute lung injury (ALI) or acute respiratory distress syndrome (ARDS) are at risk for ventilator-associated lung injury (VILI). Our RNA-Seq data provides a comprehensive survey of the transcriptome of HMVLECs under pathological and physiological CS conditions, either with TNF-alpha or not. It provides a useful resource for our in-depth understanding of genes and pathways contributing the pathogenesis of ALI and VILI.","project":"SRP040647"}
{"number_samples":3,"species":"human","abstract":"Our computational approach identified E2F1 as a potential collaborator of EZH2 in androgen-independent prostate cancer. This experiment is to designed to validate the crosstalking of E2F1 and EZH2 pathways. We showed that majority of the EZH2-induced genes in androgen-independent prostate tumor cells are in downstream of E2F1, providing insight into the EZH2-E2F1 collaborative regulatory pathway. Overall design: We performed RNA-seq experiments to compare the gene expression profles before and after E2F1 siRNA silencing","project":"SRP040679"}
{"number_samples":3,"species":"human","abstract":"KLF5 possesses both tumor-suppressing and tumor-promoting activities, though the mechanism controlling these opposing functions is unknown. In cultured non-cancerous epithelial cells, KLF5 converts from pro-proliferative to anti-proliferative activity upon TGFß-induced acetylation, which sequentially alters the KLF5 transcriptional complex and the expression of genes such as p15 and MYC. In nude mice, KLF5 also suppressed tumor growth in an acetylation-dependent manner. Furthermore, deacetylation switched KLF5 to tumor-promoting activity, and blocking TGFß signaling attenuated the tumor suppressor activity of KLF5. The results from RNA-Seq indicated the different downstream genes of AcKLF5 and unAcKLF5 and the mechanisms that determine the opposing functions of AcKLF5 and unAcKLF5. These results provide novel insights into the mechanism by which KLF5 switches from anti-tumorigenic to pro-tumorigenic function, and also suggest the roles of AcKLF5 and unAcKLF5, respectively, in the tumor-suppressing and tumor-promoting functions of TGFß. Overall design: Xenografts of DU-145 cell lines expressing KLF5, KLF5 mutant K369R or the empty vector PLHCX were surgically isolated from mice at day 38 after injection. RNA-Seq was performed using these three groups of xenografts.","project":"SRP040692"}
{"number_samples":4,"species":"human","abstract":"Latency-associated nuclear antigen (LANA), a multifunctional protein expressed by the Kaposi sarcoma-associated herpesvirus (KSHV) in latently-infected cells, is required for stable maintenance of the viral episome. This is mediated by two interactions: LANA binds to specific sequences (LBS1 and 2) on viral DNA, and also engages host histones, tethering the viral genome to host chromosomes in mitosis. LANA has also been suggested to affect host gene expression, but both the mechanism(s) and role of this dysregulation in KSHV biology remain unclear. Here we have examined LANA interactions with host chromatin on a genome-wide scale using ChIP-seq, and show that LANA predominantly targets human genes near their transcriptional start sites (TSSs). These host LANA-binding sites are generally found within transcriptionally active promoters and display striking overrepresentation of a consensus DNA sequence virtually identical to the LBS1 motif in KSHV DNA. Comparison of the ChIP-seq profile with whole transcriptome (RNA-seq) data reveals that few of the genes that are differentially regulated in latent infection are occupied by LANA at their promoters. This suggests that direct LANA binding to promoters is not the prime determinant of altered host transcription in KSHV-infected cells. Most surprisingly, the association of LANA to both host and viral DNA is strongly disrupted during the lytic cycle of KSHV. This disruption can be prevented by the inhibition of viral DNA synthesis, suggesting the existence of novel and potent regulatory mechanisms linked to either viral DNA replication or late gene expression. Overall design: Profiling of KSHV LANA positioning on the host genome and examination of gene expression from promoters bound by KSHV LANA.","project":"SRP040694"}
{"number_samples":1,"species":"human","abstract":"Transcriptome of Human 45, XO fibroblast using RNA-seq suggest distinct genes and long noncoding RNAs associated with the pathophysiology of Turner’s syndrome.","project":"SRP040702"}
{"number_samples":1,"species":"human","abstract":"Transcriptome of Human 46, XX fibroblast cell line","project":"SRP040704"}
{"number_samples":6,"species":"human","abstract":"We utilized whole genome sequencing of mRNA (RNA-seq) to understand the extent to which the senescence-associated secretory phenotype is regulated by p38MAPK Overall design: Examination of replicates of young, senescent or p38MAPK-inhibited senescent BJ human foreskin fibroblasts.","project":"SRP040745"}
{"number_samples":3,"species":"human","abstract":"PBDEs are widely used in consumer and household products as flame retardants. Many studies have shown that PBDEs could disrupt thyroid hormone homeostasis and adversely affect brain development. Here, we explored the toxical effects of BDE209 on HEK 293 cells and found that BDE209 may have a role in nucleosome remodeling. Many gene sets involved in cancer are enriched at the BDE209-treated sample. This indicates the carcinogenicity of BDE209. Interestingly, the impacts of BDE209 dissoved in DMSO on gene expression are more pronounced than the simple additive effects of BDE209 and DMSO alone. Overall design: Gene expression profiles of human embryonic kidney 293 cells (HEK 293) cultured in normal medium, and medium containing BDE209 or DMSO were generated by deep sequencing, using Illumina HighSeq2000, respectively..","project":"SRP040764"}
{"number_samples":11,"species":"human","abstract":"miRNA high-throughput sequencing was used to investigate endometriosis lesion-specific miRNA expression profiles by comparing a set of paired samples of peritoneal endometriotic lesions and matched healthy surrounding tissue together with eutopic endometrium of the same patients. We found that miRNAs of surrounding peritoneal tissue mask most of the miRNA expression differences that could originate from endometriotic tissue and thus only miRNAs with significantly different levels in the endometriotic lesions compared to peritoneal tissue were detected. According to the results of this study, two miRNAs – miR-34c and miR-449a showed remarkably higher expression in lesions compared to healthy tissue. Overall design: Eleven tissue samples (two endometria, five peritoneal lesions and four matched adjacent normal-appearing tissues) were analysed from two patients with a histologically confirmed diagnosis of moderate-severe endometriosis (III-IV stage)","project":"SRP040772"}
{"number_samples":6,"species":"human","abstract":"Monocytes, a type of mononuclear white blood cells, leave the circulation and enter into tissues in response to tissue damage or infection. These cells can encounter a hypoxic environment in the tissue that they extravasate to. The goal of this study was to identify changes in the transcriptome of such cells following exposure to hypoxia.","project":"SRP040806"}
{"number_samples":4,"species":"human","abstract":"Gene fusions and chimeric transcripts occur frequently in cancers and in some cases drive the development of the disease. An accurate detection of these events is crucial for cancer research and in a long-term perspective could be applied for personalized therapy. RNA-seq technology has been established as an efficient approach to investigate transcriptomes and search for gene fusions and chimeric transcripts on a genome-wide scale. A number of computational methods for the detection of gene fusions from RNA-seq data have been developed. However, recent studies demonstrate differences between commonly used approaches in terms of specificity and sensitivity. Moreover their ability to detect gene fusions on the isoform level has not been studied carefully so far. Here we propose a novel computational approach called InFusion for fusion gene detection from deep RNA sequencing data. Validation of InFusion on simulated and on several public RNA-seq datasets demonstrated better detection accuracy compared to other tools. We also performed deep RNA sequencing of two well-established prostate cancer cell lines. Using these data we showed that InFusion is capable of discovering alternatively spliced gene fusion isoforms as well as chimeric transcripts that include non-exonic regions. In addition our method can detect anti-sense transcription in the fusions by incorporating strand specificity of the sequencing library. Overall design: Detection of fusion genes and chimeric transcripts from deep RNA-seq data","project":"SRP040966"}
{"number_samples":15,"species":"human","abstract":"We recently showed that some human induced pluripotent stem cell (iPSC) clones were defective in neural differentiation and were marked with the activation of long term repeats (LTRs) of human endogenous retroviruses (HERVs). We herein demonstrated that these LTRs were transiently overexpressed during the generation of iPSCs and contributed to reprogramming. When the generation of iPSCs was completed, LTRs were re-suppressed to levels similar to those in human ES cells. However, differentiation-defective iPSC clones maintained high LTR expression levels, which indicated that these clones failed to complete reprogramming. lincRNA-RoR, a long intergenic non-coding RNA (lincRNA) that was previously shown to support the induction and maintenance of pluripotency, was detected among the LTR-driven transcripts. Short hairpin RNAs against the conserved sequence in LTRs or lincRNA-RoR markedly reduced the efficiency of iPSC generation. Reprogramming factors including OCT3/4, SOX2, and KLF4 bound to most LTRs. The expression of KLF4 was low in normal iPSC clones, but remained high in differentiation-defective clones. The forced expression of KLF4 in human embryonic stem cells led to the activation of LTRs and defects in neural differentiation. These results demonstrated that the transient overexpression of KLF4/LTR/lincRNA-RoR played crucial roles in reprogramming toward pluripotency in humans, whereas a failure in its re-silence resulted in differentiation defects. Overall design: Nine samples were prepared as intermediate state of cells between human dermal fibroblast and iPSC. One iPSC clone and 4 subclones derived from defective iPSC exhibit normal differentiation ability.","project":"SRP041008"}
{"number_samples":60,"species":"human","abstract":"Purpose: We applied RNA sequencing technology for high-throughput analysis of transcriptional changes within human MM cell lines JJN3 and U266 due to individual and combination drug treatment. Methods: JJN3 and U266 cells were treated with pan-HDACi panbobinostat, DNMTi 5-Azacytidine, panobinostat+5-Azacytidine or NMP for 4h or 24h in triplicate and transcriptional changes assessed by RNAseq using Illumina HiSeq platform. Specifically, JJN3 cells were treated with 10nM panobinostat, 2.5µM 5-Azacytidine, panobinostat+5-Azacytidine (at given doses), or 10mM NMP. U266 cells were treated with 10nM panobinostat, 10µM 5-Azacytidine, panobinostat+5-Azacytidine (at given doses), or 10mM NMP. Results: We report unique and overlapping transcriptional signatures that lead to the induction of apoptosis in human MM cell lines in a cell-specific manner due to individual or combination treatments. Conclusions: A detailed analysis of differential transcriptional events in human MM cell lines due to HDACi, DNMTi, HDACi+DNMTi and NMP appear to define the molecular events leading to apoptosis and drug mechanism of action. Overall design: We tested triplicate experiments at 4h and 24hr time points in JJN3 and U266 cell lines against vehicle control treated cells.","project":"SRP041036"}
{"number_samples":17,"species":"human","abstract":"Research human brain function and evolution by using high depth transcriptome sequencing data from different human brain region.","project":"SRP041044"}
{"number_samples":2,"species":"human","abstract":"Dicer is a central enzymatic player in RNA interference (RNAi) pathways that acts to regulate gene expression in nearly all eukaryotes. Although the cytoplasmic function of Dicer is well documented in mammals, there is little known about any possible nuclear function.  Here we show that Dicer is present in both the nucleus and cytoplasm, but that its nuclear levels are tightly regulated. In its nuclear manifestation Dicer interacts with RNA polymerase II (Pol II) at actively-transcribed gene loci. Loss of Dicer causes the appearance of endogenous dsRNA, leading to induction of the interferon response pathway and consequent cell death. Our results suggest that Pol II-associated Dicer restricts endogenous dsRNA formation from overlapping non-coding RNA transcription units. Failure to do so has catastrophic effects on cell function. Taken together, we present here a micro (mi)RNA independent role for human nuclear Dicer. Overall design: DICER ChIP-seq profile in wt 293 HEK cells, and dsRNA-seq profile in wt and DICER-depleted 293 HEK cells","project":"SRP041057"}
{"number_samples":4,"species":"human","abstract":"Colorectal cancer (CRC) has the third highest incidence and mortality rates among the US population. According to the most recent concept of carcinogenesis, human tumors are organized hierarchically, and the top of this hierarchy is occupied by malignant stem cells, or cancer stem cells (CSCs), which possess unlimited self-renewal and tumor-initiating capacities and high resistance to conventional anticancer therapies. To reflect the complexity and diversity of human tumors and to provide clinically and physiologically relevant in vivo and in vitro models, a large banks of well characterized patient-derived low-passage cell lines, and especially CSC-enriched cell lines are urgently needed. Overall design: Using RNA-Seq, we have performed a functional genomic analysis in tumor-initiating fractions of CR4 (small) cells grown adherent to type I collagen versus grown as 3D spheroids, in comparison to the bulk tumor cells (long and dychotomized cells) grown under standard culture conditions.","project":"SRP041063"}
{"number_samples":4,"species":"human","abstract":"Background.  Androgen receptor splice variant-7 (AR-V7) is a truncated form of the androgen receptor protein which lacks the ligand-binding domain, the target of enzalutamide and abiraterone, but remains constitutively active as a transcription factor. We hypothesized that detection of AR-V7 in circulating tumor cells (CTCs) from men with advanced prostate cancer would be associated with resistance to enzalutamide and abiraterone. Methods.  We used quantitative reverse-transcription polymerase-chain-reaction (qRT-PCR) to interrogate CTCs for the presence or absence of AR-V7 from prospectively enrolled patients with metastatic castration-resistant prostate cancer initiating treatment with either enzalutamide or abiraterone. We examined associations between AR-V7 status and PSA response rates, PSA-progression-free-survival (PSA-PFS), clinical/radiographic-progression-free-survival (PFS), and overall survival (OS). Multivariable Cox regression analyses were performed to determine the independent effect of AR-V7 status on clinical outcomes. Results.  Thirty-one enzalutamide-treated patients and thirty-one abiraterone-treated patients were enrolled, of which 38.7% and 19.4% had detectable AR-V7 from CTCs, respectively. Among men receiving enzalutamide, AR-V7–positive patients had inferior PSA response rates (0% vs 52.6%, P=0.004), PSA-PFS (median: 1.4 vs 6.0 months, P<0.001), PFS (median: 2.1 vs 6.1 months, P<0.001), and OS (median: 5.5 months vs not reached, P=0.002) compared to AR-V7–negative patients. Similarly, among men receiving abiraterone, AR-V7–positive patients had inferior PSA response rates (0% vs 68.0%, P=0.004), PSA-PFS (median: 1.3 months vs not reached, P<0.001), PFS (median: 2.3 months vs not reached, P<0.001), and OS (median: 10.6 months vs not reached, P=0.006). The negative prognostic impact of AR-V7 was maintained after adjusting for full-length-AR expression. Conclusions.  Detection of AR-V7 in CTCs from patients with castration-resistant prostate cancer is associated with resistance to enzalutamide and abiraterone. Overall design: A total of four metastatic castration-resistant prostate tumor samples from four patients were subjected to RNA-seq. Two samples were positive for androgen receptor splice variant 7 and the other two were negative for this variant.","project":"SRP041094"}
{"number_samples":14,"species":"human","abstract":"T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we abrogated the expression of JMJD3 (KDM6B) and UTX (KDM6A) H3K27me3 demethylases in human T-ALL lines and assayed for genome-wide expression changes using RNA sequencing. This piece of data was further integrated to ChIP-Sequencing analysis of H3K27me3 from the same treatment as well as H3K27me3 and JMJD3 genome-wide analysis from treatment of T-ALL lines with the GSKJ4 inhibitor. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to identify a hitherto unknown role of JMJD3as an oncogenice facilitator in leukemia whereas UTX seems to play a tumor suppressor role. Overall design: Whole RNA was extracted from 1-5 million T-ALL (lines) cells or primary cells using the RNAeasy kit (Qiagen) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Invitrogen)) or Ribominus RNA was used for library preparation. cDNA preparation and strand-specific library construction was performed using the dUTP method. Libraries were sequenced on the Illumina HiSeq 2000 using 50bp single-read method. Differential gene expression analysis was performed for each matched knockdown vs control pairs, separately in each biological or technical replicate in each of two cell lines (CUTLL1, CEM). Three types of comparisons were tested: (a) JMJD3 knockdown vs Renilla, (b) JMJD3 knockdown vs UTX knockdown, and (c) UTX knockdown vs Renilla. Analysis was performed using both DEGseq and Cufflinks packages leading to very similar conclusions.","project":"SRP041100"}
{"number_samples":12,"species":"human","abstract":"T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we chemically inhibited the H3K27me3 demethylase JMJD3 using the GSKJ4 inhibitor and assayed for genome-wide changes in H3K27me3 and JMJD3 enrichment. This piece of data was further integrated to expression changes using RNA sequencing as well as ChIP-Sequencing analysis of H3K27me3 upon genomic knock-down of JMJD3 and UTX. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to identify a hitherto unknown role of JMJD3 as an oncogenice facilitator in leukemia whereas UTX seems to play a tumor suppressor role. Overall design: Whole RNA was extracted from 1-5 million primary cells from CUTLL1 human T cell leukemia cells untreated or treated with 2micromolar GSKJ4 using the RNAeasy kit (Qiagen) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Invitrogen)) or Ribominus RNA was used for library preparation. cDNA preparation and strand-specific library construction was performed using the dUTP method. Libraries were sequenced on the Illumina HiSeq 2000 using 50bp single-read method. Differential gene expression analysis was performed between knockout vs wild-type background samples. Analysis was performed using DEGseq package leading to very similar conclusions.","project":"SRP041102"}
{"number_samples":12,"species":"human","abstract":"RNA helicases are important regulators of gene expression that act by remodeling RNA secondary structures and as RNA-protein interactions. Here, we demonstrate that MOV10 has an ATP-dependent 5' to 3' in vitro RNA unwinding activity and determine the RNA-binding sites of MOV10 and its helicase mutants using PAR-CLIP. We find that MOV10 predominantly binds to 3' UTRs upstream of regions predicted to form local secondary structures and provide evidence that MOV10 helicase mutants are impaired in their ability to translocate 5' to 3' on their mRNA targets. MOV10 interacts with UPF1, the key component of the nonsense-mediated mRNA decay pathway. PAR-CLIP of UPF1 reveals that MOV10 and UPF1 bind to RNA in close proximity. Knockdown of MOV10 resulted in increased mRNA half-lives of MOV10-bound as well as UPF1-regulated transcripts, suggesting that MOV10 functions in UPF1-mediated mRNA degradation as an RNA clearance factor to resolve structures and displace proteins from 3' UTRs. Overall design: Flp-In T-REx HEK293 cells expressing FLAG/HA-tagged MOV10 WT, MOV10 K530A, MOV10 D645N and UPF1 were used to determine the protein-RNA interaction sites of RNA helicases MOV10 and UPF1 as well as MOV10 inactive variants using PAR-CLIP in combination with next generation sequencing. mRNA half-life changes of MOV10-targeted mRNA were determined by measuring mRNA half-lives by mRNA sequencing of mock and MOV10-depleted HEK293 cells.","project":"SRP041130"}
{"number_samples":20,"species":"human","abstract":"High resolution transcriptional profiling of H1-derived human neuronal precursor cells over a timecourse of differentiation in vitro. Overall design: Human NPC differentiation timecourse covers Days 0,1,2,4,5,11, and 18 after induction of neuronal differentiation as described in manuscript. Each time point was assayed in triplicate cultures with the exception of Day 5, in which one outlier culture has been removed.","project":"SRP041159"}
{"number_samples":45,"species":"human","abstract":"Potential vorinostat-resistance candidate genes were identified using RNA interference screening in vorinostat-resistant HCT116 cells (HCT116-VR) using a synthetic lethal approach.  In order to understand the mechanisms by which these genes contributed to vorinostat response, transcriptomic analysis was conducted on HCT116-VR cells and those with siRNA-mediated knockdown of each of the vorinostat resistance candidate genes. Overall design: There are 45 samples in total, from triplicate independent biological experiments of 15 samples each.  The negative control to which all gene knockdowns are compared is the mock transfection control (mock).","project":"SRP041162"}
{"number_samples":44,"species":"human","abstract":"Many neurological and psychiatric disorders affect the cerebral cortex, and a clearer understanding of the molecular processes underlying human corticogenesis will provide greater insight into such pathologies. To date, knowledge of gene expression changes accompanying corticogenesis is largely based on murine data. Here we present a searchable, comprehensive, temporal gene expression dataset encompassing cerebral cortical development from human embryonic stem cells (hESCs). Using a modified differentiation protocol and RNA-Seq technology with computational analysis, we identified sets of genes and long non-coding RNAs that significantly change during corticogenesis, and those enriched for disease-associations. Numerous alternatively-spliced genes with varying temporal patterns of expression are revealed, including TGIF1, involved in holoprosencephaly and MARK1, involved in autism. We have created a database (http://cortecon.neuralsci.org) that provides online, query-based access to changes in RNA expression and alternatively spliced transcripts during human cortical development. Overall design: Nine timepoints (days 0,7,12,19,26,33,49,63,77) covering human corticogenesis from embyronic stem cells.","project":"SRP041179"}
{"number_samples":4,"species":"human","abstract":"The roles of Argonaute proteins in cytoplasmic miRNA and RNAi pathways are well established. However, their implication in small RNA-mediated transcriptional gene silencing in the mammalian cell nucleus is less understood. We have recently shown that intronic siRNAs cause chromatin modifications that inhibit RNA polymerase II elongation and modulate alternative splicing in an Argonaute-1 (AGO1) dependent manner. Here we used chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) to investigate the genome-wide distribution of AGO1 nuclear targets. Unexpectedly we found that about 80% of AGO1 clusters are associated with cell-type specific transcriptional enhancers, most of them (73%) overlapping active enhancers. This association seems to be mediated by long, rather than short, enhancer RNAs; and to be more prominent in intragenic, rather than intergenic, enhancers. Paradoxically, crossing ChIP-seq with RNA-seq data upon AGO1 depletion revealed that enhancer bound AGO1 is not linked to the global regulation of gene transcription but to the control of constitutive and alternative splicing, which was confirmed by an individual gene analysis explaining how AGO1 controls inclusion levels of the cassette exon 107 in the SYNE2 gene. Overall design: Genome-wide analysis of AGO-1 and different histone modifications in 2 cell types","project":"SRP041210"}
{"number_samples":5,"species":"human","abstract":"Microprocessor is responsible for conversion of pri-miRNA transcripts into pre-miRNA hairpins in miRNA biogenesis. The in vivo properties of this process remain enigmatic. Here, we present the first study of in vivo transcriptome-wide pri-miRNA processing using next-generation sequencing of chromatin-associated pri-miRNAs. We identify a distinctive Microprocessor signature in the transcriptome profile, from which efficiency of the endogenous processing event can be accurately quantified. This analysis reveals differential susceptibility to Microprocessor cleavage as a key regulatory step in miRNA biogenesis. Processing is highly variable among pri-miRNAs and a better predictor of miRNA abundance than primary transcription itself. Processing is also largely stable across three cell lines, suggesting a major contribution of sequence determinants. Based on differential processing efficiencies we define functionality for short sequence features adjacent to the pre-miRNA hairpin. In conclusion, we identify Microprocessor as the main hub for diversified miRNA output and suggest a role for uncoupling miRNA biogenesis from host gene expression. Overall design: We performed polyA independent deep sequencing of the chromatin-isolated RNA fraction for 5 samples: 2 replicates in HeLa cells, 1 Drosha knock down in HeLa cells, 1 sample for A549 cells and 1 sample for HEK293 cells. We also performed deep sequencing of the small RNA fraction in the same cell lines.","project":"SRP041228"}
{"number_samples":22,"species":"human","abstract":"The translational control of oncoprotein expression is implicated in many cancers. Here we report an eIF4A/DDX2 RNA helicase-dependent mechanism of translational control that contributes to oncogenesis and underlies the anticancer effects of Silvestrol and related compounds. For example, eIF4A promotes T-ALL development in vivo and is required for leukaemia maintenance. Accordingly, inhibition of eIF4A with Silvestrol has powerful therapeutic effects in vitro and in vivo. We use transcriptome-scale ribosome footprinting to identify the hallmarks of eIF4A-dependent transcripts. These include 5'UTR sequences such as the 12-mer guanine quartet (CGG)4 motif that can form RNA G-quadruplex structures. Notably, among the most eIF4A-dependent and Silvestrol-sensitive transcripts are a number of oncogenes, super-enhancer associated transcription factors, and epigenetic regulators. Hence, the 5'UTRs of selected cancer genes harbour a targetable requirement for the eIF4A RNA helicase. Overall design: Comparison of ribosome-protected RNA for drug treated and DMSO treated KOPT-K1 cell, two replicates of ribosome-protected RNA sequencing and three replicates of RNA-seq.","project":"SRP041255"}
{"number_samples":2,"species":"human","abstract":"Ewing sarcoma is an aggressive pediatric small round cell tumor that predominantly occurs in bone.  Approximately 85% of Ewing sarcomas harbor the EWS/FLI fusion protein, which arises from a chromosomal translocation, t(11:22)(q24:q12). EWS/FLI interacts with numerous lineage-essential transcription factors to maintain mesenchymal progenitors in an undifferentiated state. We previously showed that EWS/FLI binds the osteogenic transcription factor RUNX2 and prevents osteoblast differentiation. In this study, we investigated the role of another Runt-domain protein, RUNX3, in Ewing sarcoma.  RUNX3 participates in mesenchymal-derived bone formation and is a context dependent tumor suppressor and oncogene.  RUNX3 was detected in all Ewing sarcoma cells examined, whereas RUNX2 was detected in only 73% of specimens. Like RUNX2, RUNX3 binds to EWS/FLI via its Runt domain. EWS/FLI prevented RUNX3 from activating the transcription of a RUNX-responsive reporter, p6OSE2.  Stable suppression of RUNX3 expression in the Ewing sarcoma cell line A673 delayed colony growth in anchorage independent soft agar assays and reversed expression of EWS/FLI-responsive genes.  These results demonstrate an important role for RUNX3 in Ewing sarcoma. Overall design: RNA-seq to compare transcriptiome of control A673 ewing sarcoma cells stably expression a non-target or RUNX3 shRNA","project":"SRP041263"}
{"number_samples":1,"species":"human","abstract":"4sUDRB-seq measure elongation and initiation rates","project":"SRP041272"}
{"number_samples":48,"species":"human","abstract":"Physiological and behavioral circadian rhythms are driven by a conserved transcriptional/translational negative feedback loop in mammals. Although most core clock factors are transcription factors, post-transcriptional control introduces delays that are critical for circadian oscillations. Little work has been done on circadian regulation of translation, so to address this deficit we conducted ribosome profiling experiments in a human cell model for an autonomous clock. We found that most rhythmic gene expression occurs with little delay between transcription and translation, suggesting that the lag in the accumulation of some clock proteins relative to their mRNAs does not arise from regulated translation. Nevertheless, we found that translation occurs in a circadian fashion for many genes, sometimes imposing an additional level of control on rhythmically expressed mRNAs and, in other cases, conferring rhythms on non-cycling mRNAs.  Most cyclically transcribed RNAs are translated at one of two major times in a 24h day, while rhythmic translation of most non-cyclic RNAs is phased to a single time of day. Unexpectedly, we found that the clock also regulates the formation of cytoplasmic processing (P) bodies, which control the fate of mRNAs, suggesting circadian coordination of mRNA metabolism and translation. Overall design: U2-OS cell time course over one day of 12 wildtype total RNA-seq samples, 12 wildtype ribosome profiling samples, 12 Bmal1 knockdown total RNA-seq samples and 12 Bmal1 knockdown ribosome profiling samples. All samples have duplicates.","project":"SRP041298"}
{"number_samples":17,"species":"human","abstract":"We have combined high-quality genome sequencing and RNA-sequencing data within a 17-individual, three generation family. Using these data, we have contrasted cis-acting expression, allele-specific expression and splicing quantitative trait loci (collectively termed eQTLs) within the family to eQTLs discovered within a cell-type and ethnicity-matched population sample. We identified that eQTL that exhibit larger effects in the family compared to the population are enriched for rare regulatory and splicing variants and were more likely to influence essential genes. In addition, we identify several large effect-size eQTLs within the family for genes involved in complex disease. Through analysis of eQTLs in a large family we also report the utility of non-coding genome annotation to predicting the effect of rare non-coding variants. We find that a combination of distance to the transcription start site, evolutionary constraint and epigenetic annotation is considerably more informative for predicting the consequence of rare non-coding variants than for common variants. In summary, through transcriptome analyses within a large family we are able to identify the contribution of rare non-coding variants to expression phenotypes and further demonstrate the predictive potential of diverse non-coding genome annotation for interpretation of the impact of rare non-coding variants. Overall design: RNA-Sequencing of CEPH/UTAH family 1463","project":"SRP041338"}
{"number_samples":3,"species":"human","abstract":"JAGuaR is an alignment protocol for RNA-seq reads that uses an extended reference to increase alignment sensitivity. It uses BWA to align reads to the genome and reference transcript models (including annotated exon-exon junctions) specifically allowing for the possibility of a single read spanning multiple exons. Reads aligned to the transcript models are then re-mapped on to genomic coordinates, transforming alignmentsthat span multiple exonsinto large-gapped alignments on the genome. While JAGuaR does not detect novel junctions, we demonstrate how JAGuaR generates fast and accurate transcriptome alignments, which allows for both sensitive and specific SNV calling.","project":"SRP041367"}
{"number_samples":24,"species":"human","abstract":"Genetically engineered human pluripotent stem cells (hPSCs) have been proposed as a source for transplantation therapies and are rapidly becoming valuable tools for human disease modeling. However, many of the potential applications are still limited by the lack of robust differentiation paradigms that allow for the isolation of defined functional tissues. These challenges could be overcome by the use of adult tissue stem cells derived from hPSCs, as their restricted potential could limit the differentiation towards other undesired linages, and allow in vitro expansion and long- term propagation of fully differentiated tissue. To isolate adult stem cells from hPSCs, we applied genome-editing to generate an LGR5-GFP reporter system and subsequently developed a differentiation protocol for human intestinal tissue comprising an adult stem cell niche and all major cell types of the adult intestine. This novel derivation protocol is highly robust and even permits the isolation of intestinal organoids without the LGR5 reporter. Transcriptional profiling, electron microscopy and functional analysis revealed that such human organoid cultures could be derived with high purity, and a composition and morphology similar to that of cultures obtained from human biopsies. Importantly, hPSC-derived organoids responded to the canonical signaling pathways that control self-renewal and differentiation in the adult human intestinal stem cell compartment. With our ability to genetically engineer hPSCs using site-specific nucleases, this adult stem cell system provides a novel platform by which to study human intestinal disease in vitro. Overall design: RNA from primary organoid samples was isolated from organoid lines that were both cultured for 1-6 months and derived from duodenum, ileum, or rectum biopsies of human subjects as described previously (Sato et al., Gastroenterology 2011) grown in media called WENR+inhibitors. RNA was also isolated from various steps in the culturing and differentiation protocol.","project":"SRP041377"}
{"number_samples":4,"species":"human","abstract":"Dramatic changes of gene expressions are known to occur in human endometrial stromal cells (ESC) during decidualization.  The changes in gene expression are associated with changes of chromatin structure, which are regulated by epigenetic mechanisms such as histone modifications.  Here, we investigated genome-wide changes in histone modifications and mRNA expressions associated with decidualization in human ESC using chromatin immunoprecipitation (ChIP) combined with next-generation sequencing.  ESC were incubated with estradiol and medroxyprogesterone acetate for 14 days to induce decidualization.  The ChIP-sequence data showed that induction of decidualization increased H3K27ac and H3K4me3 signals in many genomic regions but decreased in only a few regions.  Most (80%) of the H3K27ac-increased regions and half of the H3K4me3-increased regions were located in the distal promoter regions (more than 3 kb upstream or downstream of the transcription start site).  RNA-sequence showed that induction of decidualization up-regulated 881 genes, 223 of which had H3K27ac- or H3K4me3-increased regions in the proximal and distal promoter regions.  Induction of decidualization increased the mRNA levels of these genes more than it increased the mRNA levels of genes without H3K27ac- or H3K4me3-increased regions.  Pathway analysis revealed that up-regulated genes with the H3K27ac- or H3K4me3-increased regions were associated with insulin signaling.  These results show that histone modification statuses genome-widely change in human ESC by induction of decidualization.  The main changes of histone modifications are increases of H3K27ac and H3K4me3 in both the proximal and distal promoter regions, which are involved in the up-regulation of gene expression that occurs during decidualization. Overall design: mRNA profiles of human endometrial stromal cells with and without EP inductions for 2 individuals. (EP induction: induction with estradiol (10-8 M) and medroxyprogesterone acetate (10-6 M))","project":"SRP041387"}
{"number_samples":18,"species":"human","abstract":"The development of affinity purification technologies together with mass spectrometric analyses of the purified protein mixtures (AP-MS) has been used both to identify new protein-protein interactions and to define the subunit composition of protein complexes. Transcription factor protein interactions, however, have not been systematically analyzed using these approaches. Here, we have investigated whether ectopic expression of an affinity tagged transcription factor as bait in AP-MS experiments perturbs gene expression in cells resulting in false positive identification of bait associated proteins when typical experimental controls are used. Using quantitative proteomics and RNA-Seq, we determined that the increase in the abundance of a set of proteins caused by overexpression of the transcription factor RelA is not sufficient for these proteins to then copurify non-specifically and be misidentified as bait associated proteins. Therefore typical controls should be sufficient and a number of different baits can be compared with a common set of controls. This is of practical interest when identifying bait interactors from a large number of different baits. As expected, we found several known RelA interactors enriched in our RelA purifications (NFêB1, NFêB2, Rel, RelB, IêBá, IêBâ and IêBå). We also found several proteins not previously described in association with RelA, including the small mitochondrial chaperone Tim13. Using a variety of biochemical approaches, we further investigated the nature of the association between Tim13 and NFêB family transcription factors. The work here therefore provides a conceptual and experimental framework for analyzing transcription faction protein interactions. Overall design: Gene expression profiles were assayed in triplicate from HEK293 cells expressing either Halo-RelA, Halo-NFkB1, or Halo tag alone.","project":"SRP041396"}
{"number_samples":2,"species":"human","abstract":"A transcriptome-wide analysis of intron retentions.","project":"SRP041469"}
{"number_samples":313,"species":"human","abstract":"A new method to measure elongation and intitiation rates Overall design: Reversal inhibition of transcription with DRB and tagging newly transcribed RNA with 4-thiouridine (4sU)","project":"SRP041471"}
{"number_samples":1,"species":"human","abstract":"De novo transcriptome assembly for analysis and functional annotation of genes expressed in Alport syndrome iPSCs","project":"SRP041474"}
{"number_samples":1,"species":"human","abstract":"De novo transcriptome assembly for analysis and functional annotation of genes expressed in Alport syndrome iPSCs","project":"SRP041475"}
{"number_samples":5,"species":"human","abstract":"Summary: Adult germline stem cells (AGSCs) are multifunctional - they must self renew, maintain genome pluripotency, and prepare for gametogenesis – which involves meiotic and chromatin repackaging phases. To better understand AGSCs and gametogenesis, we derived high-resolution profiles of transcription, DNA methylation, 5hmC, and multiple histone modifications at key stages. First, AGSCs display chromatin ‘poising’ of enhancers and promoters of genes utilized in embryo development. Second, the pluripotency network in AGSCs is remarkably distinct from ESCs - lacking Nanog, Sox2, or Prdm14 expression.  Third, spermatogenesis involves stage-specific transcription and distinctive chromatin dynamics, but virtually no changes in DNAme.  Surprisingly, we observe co-incidence of RNA polymerase II, high H3K4me3, and DNA methylation at 20-35% of genes transcribed during gametogenesis - including piRNA clusters - but often observe attendant promoter 5hmC.  Our work reveals key differences between AGSCs and other germ/stem cells, and reveals both logical and unexpected chromatin-transcription relationships accompanying germline developmental transitions. Overall Design: mRNA and small RNA profiles of human sperm generated by deep sequencing using Illumina HiSeq 2000 and Illumina Genome Analyzer II.","project":"SRP041494"}
{"number_samples":7,"species":"human","abstract":"Long noncoding RNAs (lncRNAs) have emerged as key players in different cellular processes and are required for diverse functions in vivo. However, fundamental aspects of lncRNA biology remain poorly characterized, including their subcellular localization, abundance and variation at a single cell resolution. Here, we used single molecule, single-cell RNA fluorescence in situ hybridization (RNA FISH) to survey 61 lncRNAs, chosen by properties such as conservation, tissue specific expression, and expression abundance, and to catalog their abundance and cellular localization patterns in three human cell types. Our lncRNAs displayed diverse sub-cellular localization patterns ranging from strictly nuclear localization to almost exclusive cytoplasmic localization, with the majority localized primarily in the nucleus. The low abundance of these lncRNAs as measured in bulk cell populations cannot be explained by high expression in a small subset of 'jackpot' cells. Simultaneous analysis of lncRNAs and mRNAs from corresponding divergently transcribed loci showed that divergent lncRNAs do not present a distinct localization pattern and are not always co-regulated with their neighbor. Overall, our study highlights important differences and similarities between lncRNAs and mRNAs. The rich set of localization patterns we observe are consistent with a broad range of potential functions for lncRNA, and assists in hypothesis generation for mechanistic studies. Here we provide the RNA-Seq expression matrix, as well as RNA-Seq raw data, which we used for comparison with RNA FISH molecule counts. Overall design: We estimate FPKM of coding genes and lncRNAs across HeLa, human lung fibroblasts and human foreskin. This study includes data from human foreskin fibroblasts (hFF), human lung fibroblasts (hLF), and HeLa cells. An hFF sample (GSM1376178) and the hLF samples (GSM1376175-GSM1376177) were previously submitted and are available in GSE30554 as GSM759893 and GSM759890-GSM759892, respectively. The HeLa samples (GSM591670-GSM591671) were previously submitted and are available in GSE23316. The complete dataset representing: (1) the hFF Samples, including the re-analysis of the hFF Sample from GSE30554, (2) the re-analysis of the hLF Samples from GSE30554, and (3) the re-analysis of the HeLa Samples from GSE23316, is linked below as a supplementary file.","project":"SRP041508"}
{"number_samples":9,"species":"human","abstract":"Given that many soft tissue tumors is driven by genomic translocations, we performed whole transcriptome sequencing in 9 gastrointestinal stromal tumors to identify global expression patterns and fusion genes.","project":"SRP041531"}
{"number_samples":8,"species":"human","abstract":"The endometrial perivascular microenvironment is rich in mesenchymal stem-like cells that express type 1 integral membrane protein Sushi domain containing 2 (SUSD2) but the role of these cells in the decidual transformation of this tissue in pregnancy is unknown. We used an antibody directed against SUSD2 (W5C5) to isolate perivascular (W5C5+) and non-perivascular (W5C5-) fibroblasts from mid-luteal biopsies. We show that SUSD2 expression, and hence the ratio of W5C5+ to W5C5- cells, changes in culture depending on cell-cell contact and activation of the Notch signaling pathway. RNA sequencing revealed that cultures derived from W5C5+ progenitor cells remain phenotypically distinct by the enrichment of novel and established endometrial perivascular signature genes. In an undifferentiated state, W5C5+-derived cells produced lower levels of various chemokines and inflammatory modulators when compared to their W5C5- counterparts. This divergence in secretomes became more pronounced upon decidualization, which transformed perivascular W5C5+ cells into the dominant source of a range of trophic and immunomodulatory cytokines, including leukemia inhibitory factor (LIF). Our findings indicate that the decidual response is spatially organized with differentiating perivascular cells establishing distinct cytokine and chemokine gradients that could direct trophoblast towards maternal vessels and govern local immune responses in pregnancy. Overall design: Analysis of paired human endometrial stromal cultures, originating from either W5C5+ or W5C5- cells, from four biological replicates - a total of 8 samples - by Illumina RNAseq.","project":"SRP041573"}
{"number_samples":1,"species":"human","abstract":"Discarded live tumor tissue from a metastatic focus in the patient’s lung was collected under institutional review board approval through the NUT midline carcinoma registry (www.NMCRegistry.org). From this tissue the first known NUT-variant cell line, 1221, was established. To determine the putative partner gene to NUT, we performed comprehensive RNA-sequencing on RNA purified from 1221. We identified an in-frame transcript fusing the 5’ coding sequence of NSD3 (exons 1-7) to exons 2-7 of NUT. Expression of the NSD3-NUT fusion oncoprotein was verified by immunobloting with an antibody to NUT, revealing an approximately 200kDa band that is similar in size to BRD3-NUT, but smaller than BRD4-NUT Overall design: Identification of a NUT fusion partner using RNA extracted from live cultured 1221 cell line derived from a lung metastasis from the index case of a 13 year old female with NUT-positive NMC.","project":"SRP041594"}
{"number_samples":2,"species":"human","abstract":"We report the genome-wide profiling of FXR binding by ChIP-seq from GW4064 or DMSO treated primary human hepatocytes. We reported altered RNA expression profiles in primary human hepatocypes upon GW4064 treatment compared to DMSO control by RNA-seq. We also reported the altered RNA expression profiles in livers from WT C57BL/6J mice upon GW4064 treatment compared to vehicle control. Overall design: Primary human hepatocytes were treated with 5uM GW4064 or DMSO control, 1 hour later, cells were fixed and collected for chromatin isolation. 24 hours post treatment, cells were isolated for RNA isolation. This submission represents HTS component of study.","project":"SRP041597"}
{"number_samples":26,"species":"human","abstract":"Inflammasomes are intracellular innate immune sensors that respond to pathogen and damage-associated signals with the proteolytic cleavage of caspase-1, resulting in IL-1_ and IL-18 secretion and macrophage pyroptosis. The discovery that heterozygous gain-of-function mutations in NLRP3 lead to oversecretion of IL-1_ and cause the autoinflammatory disease spectrum Cryopyrin Associated Periodic Syndrome (CAPS), led to the successful use of IL-1 blocking therapies1. We found that a de novo missense mutation in the regulatory domain of the NLRC4 (IPAF, CARD12) inflammasome causes early-onset recurrent fever flares and Macrophage Activation Syndrome (MAS). Functional analyses demonstrated spontaneous production of the inflammasome-dependent cytokines IL-1² and IL-18 exceeding levels in CAPS patients. The NLRC4 mutation led to constitutive caspase-1 cleavage in transduced cells and enhanced spontaneous production of IL-18 by both patient and NLRC4 mutant macrophages. Thus, we describe a novel monoallelic inflammasome defect that expands the autoinflammatory paradigm to include MAS and suggests novel targets for therapy. Overall design: Whole blood RNA-seq from seven timepoints of one patient with NLRC4-MAS as compared to five healthy pediatric controls, 7 NOMID patients with active disease prior to anakinra treatment and the same 7 NOMID patients with inactive disease after anakinra treatment. Please note that seven time points are chronologic time point. They are ordinal, in that \"1\" was drawn before \"2\", but the distance in time between points is not constant. Thus, time points 4 through 7 correspond to samples drawn while the patient was well AND on treatment. However there may be differences between 4 and 7 pertaining to the length of treatment, and for that reason any of these samples were not considered replicates.","project":"SRP041620"}
{"number_samples":2,"species":"human","abstract":"RNA-binding proteins (RBPs) are critical regulators of gene expression, but only a small fraction have been studied for their role in malignancy.  Here we report a systematic analysis of RBP CUGBP Elav-Like Family member 1 (CELF1 alias CUGBP1) roles in mRNA alternative splicing, translation and turnover in oral cancer cells. CELF1 is overexpressed in carcinogen-induced oral cancer tumorigenesis mouse model and specific inhibition of CELF1 reduces  tumor growth in vivo. Deep transcriptomic analysis revealed that hundreds of mRNAs were differentially regulated as a function of CELF1 expression in oral cancer cells.  More importantly, the presence of CELF1 promoted alternative splicing of several target mRNAs which are known to be involved in various cancer biological processes. Using a pulse SILAC-based quantitative proteomic approach, we observed hundreds of proteins whose translation was controlled by CELF1 and those altered proteins were implicated in malignancy.  Altogether, these data provided a comprehensive view of the CELF1 mRNA regulatory network in OSCC and suggests that CELF1 is a viable target for therapeutic intervention. Significance  Post-transcriptional mechanisms that regulate cancer cells which are believed to constitute the driving force of many malignancies are poorly understood. We show that oral squamous cell carcinoma (OSCC) emerge as a result of an over expressed RNA-binding protein CELF1, a protein implicated in mRNA turnover, alternative splicing and translation. The resulting mRNA expression repertoire regulates networks of genes whose expression changes regulate oral cancer pathogenesis. Inhibition of CELF1 mitigated OSCC tumor-forming capacity and offers an attractive therapeutic option for oral malignancies. Overall design: Transcriptomic analysis of UMSCC-74B oral cancer cells as a function of CELF1 protein expression.","project":"SRP041647"}
{"number_samples":2,"species":"human","abstract":"The vertebrate and neural-specific SR-related protein nSR100/SRRM4 regulates an extensive program of alternative splicing with critical roles in nervous system development. However, the mechanism by which nSR100 controls its target exons is poorly understood. We demonstrate that nSR100-dependent neural exons are associated with a unique configuration of intronic cis-elements that promote rapid switch-like regulation during neurogenesis. A key feature of this configuration is the insertion of specialized intronic enhancers between polypyrimidine tracts and acceptor sites that bind nSR100 to potently activate exon inclusion in neural cells, while weakening 3' splice site recognition and contributing to exon skipping in non-neural cells. nSR100 further operates by forming multiple interactions with early spliceosome components bound proximal to 3' splice sites. These multifaceted interactions achieve dominance over neural exon silencing mediated by the splicing regulator PTBP1. The results thus illuminate a widespread mechanism by which a critical neural exon network is activated during neurogenesis. Overall design: RNA-Seq was used to obtain mRNA profiles of various N2A and 293T cell lines from human and mouse, respectively, to investigate the roles of nSR100, Ptbp1 and U2af65 in alternative splicing regulation. PAR-iCLIP and iCLIP experiments followed by high throughput sequencing were conducted to obtain RNA binding profiles of nSR100, PTBP1 and U2af65.","project":"SRP041656"}
{"number_samples":143,"species":"human","abstract":"mRNA profiling of CD34+ human cord blood-derived cell treated with UM171, SR1 or both Overall design: mRNA profiles of CD34+ human cord blood-derived cell treated with DMSO (control), SR1 [500nM], UM171  [35nM] or combination SR1 [500nM]+ UM171  [35nM] for 30min, 3hr, 12hr, 24hr, 48hr, 72hr were generated by deep sequencing","project":"SRP041675"}
{"number_samples":2,"species":"human","abstract":"Bcl-2-accociated transcription factor 1(BCLAF1) has been shown to be involved in multiple biological processes. Transcript variants encoding different isoforms that are generated by alternative splicing have been found for this gene, but little is known about the mechanisms governing its splicing regulation and whether the misregulation is associated with cancer development. Mechanistic analysis revealed that splicing factor SRSF10 specifically interacts with exon5a and activates its inclusion, as RNAi-mediated knockdown of SRSF10 induced a dramatic skipping of exon5a. To define a comprehensive programm of alternative splicing that is regulated by SRSF10 in RKO cells, we used RNA-seq coupled with a bioinformatic analysis to identify the extensive splicing network regulated by SRSF10 in RKO cells. Overall design: RNA-seq for control (si-NC) and SRSF10-knockdown (si-SRSF10) human RKO cells","project":"SRP041694"}
{"number_samples":6,"species":"human","abstract":"We have utilized the RNA-Seq technology to identify genes with distinct expression patterns between failing and non-failing hearts. In an era of next-generation sequencing studies, our study demonstrates how knowledge gained from a small set of samples with accurately measured gene expressions using RNA-Seq can be leveraged as a complementary strategy to discern the genetics of complex disorders. Overall design: Identify the signature genes based on RNA-seq come from six Heart Failure and healthy individuals. Validation is based on Affymetrix microarray of a total of 313 individuals with/without Heart Failure.","project":"SRP041706"}
{"number_samples":14,"species":"human","abstract":"Aicardi-Goutières syndrome (AGS) is a severe childhood inflammatory disorder that shows clinical and genetic overlap with systemic lupus erythematosus (SLE). AGS is thought to arise from the accumulation of incompletely metabolized endogenous nucleic acid species owing to mutations in nucleic acid degrading enzymes TREX1 (AGS1), RNase H2 (AGS2, 3 and 4) and SAMHD1 (AGS5). However, the identity and source of such immunogenic nucleic acid species remain undefined. Using genome-wide approaches, we show that fibroblasts from AGS patients with AGS1-5 mutations are burdened by excessive loads of RNA:DNA hybrids. Using MethylC-seq, we show that AGS fibroblasts display pronounced and global loss of DNA methylation and demonstrate that AGS-specific RNA:DNA hybrids often occur within DNA hypomethylated regions. Altogether, our data suggest that RNA:DNA hybrids may represent a common immunogenic form of nucleic acids in AGS and provide the first evidence of epigenetic perturbations in AGS, furthering the links between AGS and SLE. Overall design: DNA-RNA immunoprecipitation (DRIP-seq) was performed on control 1, control 3, AGS1 P1, AGS1 P2, AGS2 P1, AGS2 P2, AGS4 P1, AGS4 P2, AGS5 P1 and AGS5 P2. MethylC-seq was performed on control 1, control 2, AGS1 P1, AGS2 P1, AGS4 P1 and AGS5 P1. Reduced representation bisulfite sequencing (RRBS) was performed on GM12697 and AGS2 LCL cell lines. RNA-seq was performed on 2 biological replicates each of control 1, AGS1 P1, AGS2 P1, AGS2 P2, AGS4 P1, AGS4 P2 and AGS5 P1.","project":"SRP041718"}
{"number_samples":696,"species":"human","abstract":"Analyze the transcriptomes of 347 cells from 10 distinct populations in both of low-coverage (~0.27 million reads per cell) and high-coverage (~5 million reads per cell) to identify cell-type-specific biomarkers, and to compare gene expression across samples specifically for cells of a given type as well as to reconstruct developmental lineages of related cell types.","project":"SRP041736"}
{"number_samples":14,"species":"human","abstract":"Whole-exome sequencing studies have implicated chromatin modifiers and transcriptional regulators in autism spectrum disorder (ASD) through the identification of de novo loss of function mutations in affected individuals. Many of these genes are co-expressed in mid-fetal human cortex, suggesting ASD risk genes converge in regulatory networks that are perturbed in ASD during neurodevelopment. To elucidate such networks we mapped promoters and enhancers bound by the chromodomain helicase CHD8, which is strongly enriched in ASD-associated de novo loss of function mutations, using ChIP-seq in mid-fetal human brain, human neural stem cells (hNSCs), and embryonic mouse cortex. We find that CHD8 targets are strongly enriched for ASD risk genes that converge in ASD-associated co-expression networks in human midfetal cortex. CHD8 knockdown in hNSCs results in significant dysregulation of ASD risk genes targeted by CHD8, as well as additional genes important for neurodevelopment, including members of the Wnt/ß-catenin signaling pathway. Integration of CHD8 binding data with genetic and gene co-expression data in ASD risk models provides support for additional ASD risk genes. Together, our results suggest that loss of CHD8 function contributes to ASD through regulatory perturbation of other ASD risk genes during human cortical development. Overall design: Two biological replicates for each ChIP with appropriate Input control Four biological replicates for each condition in knockdown experiments (Ctrl construct, Chd8 target C, and Chd8 target G) No raw data are provided for human samples (GSM1381214-GSM1381221) due to patient confidentiality issue. Human alignments were anonymized by removing sequence information and provided as aligned bam files instead","project":"SRP041738"}
{"number_samples":1,"species":"human","abstract":"Negative and positive supercoiling is generated in “twin domains” in the template DNA respectively upstream and downstream of the translocating RNA polymerase II (RNAPII). The accumulation of this torsional stress may facilitate or impede transcription. By transiently breaking one strand, allowing axial rotation of the DNA and resealing the nick, Topoisomerase I (Top1) removes torsional stress. Besides the relaxation of the excess torsional stress generated during transcription elongation, Top1 has been also linked to early events during transcription initiation in vitro. However the mechanisms coordinating Top1 activity with RNA synthesis throughout the different stages of the transcription cycle remain obscure. To interrogate the coupling between transcription and Top1 function, we generated high-resolution maps of both Top1 occupancy and activity in vivo. The genomic distribution patterns of Top1 and RNAPII were overlaid to reveal the dynamics of Top1 at transcribed loci. This work is of interest to the broad scientific community, from biochemists and molecular biologists, to cancer biologists and oncologists, and onto physical chemists and physicists who explore the physico-elastic properties of polymers such as DNA.","project":"SRP041742"}
{"number_samples":54,"species":"human","abstract":"Advancing pluripotent stem cell technologies for modeling hematopoietic stem cell development and therapies requires identifying key regulators of hematopoietic commitment from human pluripotent stem cells (hPSCs). Here, by screening the effect of 27 candidate factors, we identified two groups of transcriptional regulators capable of inducing distinct hematopoietic programs from hPSCs: pan-myeloid (GATA2 and ETV2) and erythro-megakaryocytic (GATA2 and TAL1). In both cases, these transcription factors directly converted hPSCs to endothelium, which subsequently transformed into blood cells with pan-myeloid or eryhtro-megakaryocytic potential. These data demonstrate that two distinct genetic programs regulate the hematopoietic development from hPSCs and that both of these programs specify hPSCs directly to hemogenic endothelial cells. Additionally, this study provides a novel method for efficient induction of blood and endothelial cells from hPSCs via overexpression of modified mRNA for selected transcription factors. Overall design: mRNA profiles of 54 samples, including 4 samples in duplicate and 6 control samples were generated by deep sequencing, using Illumina HiSeq 2500.","project":"SRP041751"}
{"number_samples":8,"species":"human","abstract":"FBXW7 modulates stress response by post-translational modification of HSF1 HSF1 orchestrates the heat-shock response upon exposure to heat stress and activates a transcriptional program vital for cancer cells. Genes positively regulated by HSF1 show increeased expression during heat shock while their expression is reduced during recovery. Genes negatively regulated by HSF1 show the opposite pattern. In this study we utilized the HCT116 FBXW7 KO colon cell line and its wild type counterpart to monitor gene expression changes during heat shock (42oC, 1 hour) and recovery (37oC for 2 hours post heat shock) using RNA sequencing. These results revealed that the heat-shock response pathway is prolonged in cells deficient for FBXW7. Overall design: Whole RNA was extracted from 1 million HCT116 WT or FBXW7KO  cells using the RNAeasy kit (Qiagen) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Invitrogen)) or Ribominus RNA was used for library preparation. cDNA preparation and strand-specific library construction was performed using the dUTP method. Libraries were sequenced on the Illumina HiSeq 2000 using 50bp single-read method. Differential gene expression analysis was performed for each matched recovery versus heat-shock pairs, separately in each biological replicate and cell line (WT or KO). Two types of comparisons were tested: (a) WT recovery vs WT heat shock, (b) FBXW7 KO recovery vs heat shock.","project":"SRP041753"}
{"number_samples":9,"species":"human","abstract":"Eukaryotic RNAs with premature termination codons (PTCs) are eliminated by nonsense-mediated decay (NMD). While human nonsense RNA degradation can be initiated either by an endonucleolytic cleavage event near the PTC or through decapping, the individual contribution of these activities on endogenous substrates has remained unresolved. Here we unambiguously establish that SMG6-catalyzed endonucleolysis is the primary initiating step in human nonsense RNA decay. We also show that both protein-coding and ‘non-coding’ genes hosting snoRNAs in their introns produce considerable amounts of NMD-sensitive splice variants, suggesting that these RNAs are merely by-products of a primary snoRNA production process. Finally, genes encoding multiple snoRNAs generally yield elevated numbers of alternative transcript isoforms, enabling the differential expression of individual snoRNAs. These findings demonstrate a hitherto unappreciated potential for decoupling of the individual expression levels of functional exon- and intron-encoded species from such composite genes. Overall design: HEK293 Flp-In T-Rex cells were subjected to siRNA-mediated depletion of XRN1 and co-depletion of XRN1 with either SMG6 or UPF1. All the treated samples together with the controls were subjected to both RNA-seq and 5'-end-seq. RNA-seq was used for detecting NMD isoforms and their expression levels. 5'-end-seq was used for finding NMD decay intermediates (decapped and endocleaved molecules). CAGE was used to distinguish decapped from endocleaved RNA fragments.","project":"SRP041788"}
{"number_samples":8,"species":"human","abstract":"We investigated differential gene expression in response to treatment of multiple myeloma cells with EZH2 inhibitor Overall design: RNA-seq in two cell lines","project":"SRP041819"}
{"number_samples":2,"species":"human","abstract":"Hypomorphic mutations of the transcription factor PAX5 occur in one third of B-progenitor acute lymphoblastic leukemias (B-ALLs). To identify PAX5-regulated genes in B-ALL, here we employ inducible expression of PAX5 in a human B-ALL cell line (REH) that harbors a loss-of-function mutation in PAX5. In this model, inducing PAX5 expression is associated with competitive disadvantage. Overall design: Comparison of REH cell lines with Dox-inducible expression of PAX5-IRES-GFP, or control GFP alone. GFP positive cells were isolated by FACS.","project":"SRP041820"}
{"number_samples":24,"species":"human","abstract":"Our studies indicate that glucose and acetate can regulate histone acetylation by altering the acetyl-CoA concentrations in the cell. The purpose of this study was to to determine whether specific gene sets correlated with acetyl-CoA availability. We conclude that 10% of glucose-regulated genes are acetyl-CoA regulated genes (genes suppressed or induced by low glucose and reversed by acetate). Acetate usually regulated gene expression in the same direction as glucose, suggesting that acetyl-CoA is a key mediator of glucose-dependent gene expression. Overall design: The experiments were performed in quadruplicates for each condition with a total of 12 samples","project":"SRP041825"}
{"number_samples":40,"species":"human","abstract":"Human D14+ / CD16+  monocytes were treated with GPBAR1 agonists or controls, and were stimulated with interferon gamma and LPS.  At 6 and 24 hours, the cells were profiled by RNAseq Overall design: 40 total samples, 5 per group with eight groups.  Individual donors used for multiple comparisons, so paired analysis is possible.  Control samples include unstimulated cells, and stimulated cells treated with vehicle control (DMSO).","project":"SRP041826"}
{"number_samples":16,"species":"human","abstract":"RNA-Seq technique was applied to investigate the effects of four cDNA amplification kits and two RNA-Seq library preparation kits to the deep sequencing results at different perspectives. Overall design: The same set of semen samples were applied to investigate the qualitative and quantitative effect of four cDNA amplification methods and two RNA-Seq library preparation methods on sperm transcript profiling.","project":"SRP041833"}
{"number_samples":12,"species":"human","abstract":"Half of human prostate cancers overexpress the transcription factor ERG. The in vivo consequences of ERG expression and the mechanisms by which ERG activity modulates prostate carcinogenesis remain poorly defined. We show that prostate-specific overexpression of ERG, at levels comparable to human prostate cancer, results in upregulation of the Hippo pathway target genes and age-dependent development of prostate tumors. We show that ERG binds to chromatin sites occupied by TEAD4, increases histone H3K9/14 acetylation at these sites and transactivates Hippo target genes. In human prostate cancer cells, ERG binds to the promoter of YAP1 and is required for promoter histone H3K9/14 acetylation and YAP1 expression. We found that while in normal mouse prostate YAP1 is prominantly nuclear in both basal and luminal epithelial cells, in normal human prostate YAP1 is present in basal but absent in luminal cells. In contrast, in human prostate cancer, YAP1 is upregulated in luminal cancer cells in a subset of the tumors. Expression of ERG in human prostate tumors correlates with the appearance of nuclear YAP1, which, in turn, strongly correlates with tumor recurrence. Futhermore, we demonstrate that genetic activation of YAP1 in mouse prostate epithelium is sufficient for the appearance of age-dependent prostate tumors with histological phenotypes that are similar to tumors caused by ERG upregulation. These results provide direct genetic evidence of a causal role for ERG in prostate cancer and reveal a previously unrecognized connection between ERG and the Hippo signaling pathway.","project":"SRP041840"}
{"number_samples":6,"species":"human","abstract":"We assayed the effect of c-Jun overexpression on gene expression in the three DDLPS cell lines using RNA-Seq (Illumina). Overall design: 141, LPS12 and 510 has been overexpressed with c-Jun or control c-DNA and results were analyzed in high-througput sequencing metadata.","project":"SRP041846"}
{"number_samples":53,"species":"human","abstract":"RNASeq data for mPB or CB-derived CD34+ exposed to UM171 Overall design: human mobilized peripheral blood or cord blood-derived CD34(+) cells were cultured for 16 hours with vehicle (DMSO),  dose response of UM171 [11.9nM, 19nM, 30.5nM, 48.8nM, 78.1nM and 125nM], SR1 [500nM] and combination of( UM171  [48.8nM]+SR1 [500nM])","project":"SRP041885"}
{"number_samples":1,"species":"human","abstract":"Stringtie, developed at the Center for Computational Biology at Johns Hopkins University.","project":"SRP041943"}
{"number_samples":40,"species":"human","abstract":"The use of low quality RNA samples in whole-genome gene expression profiling remains controversial. It is unclear if transcript degradation in low quality RNA samples occurs uniformly, in which case the effects of degradation can be normalized, or whether different transcripts are degraded at different rates, potentially biasing measurements of expression levels. This concern has rendered the use of low quality RNA samples in whole-genome expression profiling problematic. Yet, low quality samples are at times the sole means of addressing specific questions – e.g., samples collected in the course of fieldwork.","project":"SRP041955"}
{"number_samples":9,"species":"human","abstract":"Despite the well-recognized role of IL-13–induced transcriptional responses in allergic inflammation, the epigenetic mechanisms driven by IL-13 have not been well defined. We interrogated the transcriptional and epigenetic signatures of IL-13-induced epithelial responses focusing on the chromatin activation marks H3K4me3, H3K9Ac, and H3K27Ac. ChIP-sequencing analysis revealed that IL-13–inducible genes were epigenetically poised for induction and continued to accumulate epigenetic changes in response to IL-13. By intersecting the transcriptome and the epigenome of the IL-13 response, we identified neurotrophic tyrosine kinase receptor 1 (NTRK1) as a major target of IL-13 in epithelial cells. Using eosinophilic esophagitis as a model system for human allergic inflammation, we found that NTRK1 was dramatically induced in inflamed esophageal biopsies, and downstream mediators of NTRK1 signaling were elevated in diseased tissue. The NTRK1 ligand nerve growth factor (NGF) was constitutively expressed in control and disease states, indicating that induction of the receptor by IL-13 limited pathway activation. In epithelial cells, NGF and IL-13 synergistically induced transcription and secretion of the key eosinophil chemoattractant CCL26 (eotaxin-3). In summary, we demonstrate that IL-13–mediated allergic responses are epigenetically driven and identify NTRK1 as a novel epigenetic and transcriptional target of IL-13 that uniquely contributes to allergic inflammation. Overall design: Human esophageal epithelial cell line TE-7 was stimulated with IL-13 at 100 ng/ml for 2 hr, 6 hr and 24 hr and subjected to RNA-sequencing. In parallel, TE-7 cells were induced with IL-13 for 6 hr and subjected to ChIP-sequencing analysis for H3K4me3, H3K9Ac and H3K27Ac activating chromatin marks.","project":"SRP041956"}
{"number_samples":4,"species":"human","abstract":"Tumor metastasis remains the major cause of cancer-related death, but its molecular basis is still not well understood. Here we uncovered a splicing-mediated pathway that is essential for breast cancer metastasis. We show that the RNA-binding protein hnRNPM promotes breast cancer metastasis by activating the switch of alternative splicing that occurs during epithelial-mesenchymal transition (EMT). Genome-wide deep sequencing analysis suggests that hnRNPM potentiates TGFb signaling and identifies CD44 as a key downstream target of hnRNPM. hnRNPM ablation prevents TGFb-induced EMT and inhibits breast cancer metastasis in mice, whereas enforced expression of the specific CD44s splice isoform overrides the loss of hnRNPM and permits EMT and metastasis. Mechanistically, we demonstrate that the ubiquitously expressed hnRNPM acts in a mesenchymal-specific manner to precisely control CD44 splice isoform switching during EMT. This restricted cell-type activity of hnRNPM is achieved by competition with ESRP1, an epithelial-splicing regulator that binds to the same cis-regulatory RNA elements and is repressed during EMT. Importantly, hnRNPM is associated with aggressive breast cancer and correlates with increased CD44s in patient specimens. These findings demonstrate a novel molecular mechanism through which tumor metastasis is endowed by the hnRNPM-mediated splicing program. Overall design: RNAseq for control, hnRNPM siRNA treated lung metastatic LM2 clonal line, derived from the mesenchymal MDA-MB-231 cells","project":"SRP041962"}
{"number_samples":16,"species":"human","abstract":"Generation of abundant engraftable hematopoietic cells from autologous tissues promises new therapies for hematologic diseases. Differentiation of pluripotent stem cells into hematopoietic cells results in emergence of cells that have poor engraftment potential.  To circumvent this hurdle, we have devised a vascular niche model to phenocopy the developmental microenvironment of hemogenic cells thereby enabling direct transcriptional reprogramming of human endothelial cells (ECs) into hematopoietic cells. In this approach, transduction of human umbilical vein ECs (HUVECs) or adult human dermal microvascular ECs (hDMECs) with transcription factors (TFs), FOSB, GFI1, RUNX1, and SPI1 (FGRS) and induction with a instructive vascular niche feeder layer in a xenobiotic- and serum-free microenvironment results in generation of long-term engraftable hematopoietic multilineage progenitors (rEC-HMLPs).  The rEC-HMLPs had robust proliferative and multilineage colony forming units (CFU) potential, including granulocytic/monocytic, megakaryocytic, erythroid and lymphoid lineages. When transplanted, hDMEC-derived rEC-HMLPs were capable of long-term multilineage primary and secondary hematopoietic engraftment. A subset of engrafted rEC-HMLPs phenotypically and functionally resembled cord blood cells. By conditionally expressing the FGRS TFs, we further optimized reprogramming of ECs into rEC-HMLPs manifesting features of self-renewing multi-potent progenitor populations (MPPs).  Our approach replicates critical aspects of hematopoietic development and essential role of vascular niche induction in orchestrating hematopoietic specification and may prove useful for engineering autologous engraftable hematopoietic cells for treatment of inherited and acquired blood disorders. . Overall design: Transcriptome sequencing of rEC-HMLPs, hDMECs, HUVECs and other cell types","project":"SRP041988"}
{"number_samples":1,"species":"human","abstract":"We were interested in identifying novel splice junctions in Hep3B cells, especially in genes involved in cholesterol metabolism like HMGCR, so we sequenced polyA-tailed RNA from Hep3B cells. Overall design: 1 polyA-selected RNA sample was sequenced from Hep3B cells","project":"SRP041990"}
{"number_samples":48,"species":"human","abstract":"Circulating cell-free RNA in the blood provides a potential window into the health, phenotype, and developmental programs of a variety of human organs. We used high-throughput methods of RNA analysis such as microarrays and next-generation sequencing to characterize the global landscape of circulating RNA in human subjects. By focusing on tissue-specific genes, we were able to identify the relative contributions of these tissues to circulating RNA and monitor changes during tissue development and neurodegenerative disease states.","project":"SRP042027"}
{"number_samples":4,"species":"human","abstract":"Here we explored how the human macrophage response to tumor necrosis factor (TNF) is regulated by human synovial fibroblasts, the representative stromal cell type in the synovial lining of joints that become activated during inflammatory arthritis. Genome-wide transcriptome analysis (RNAseq) showed that co-cultured synovial fibroblasts modulate the expression of approximately one third of TNF-inducible genes in macrophages, including expression of target genes in pathways important for macrophage survival and polarization towards an alternatively activated phenotype. This work furthers our understanding of the interplay between innate immune and stromal cells during an inflammatory response, one that is particularly relevant to inflammatory arthritis. Our findings also identify modulation of macrophage phenotype as a new function for synovial fibroblasts that may prove to be a contributing factor in arthritis pathogenesis. Overall design: Human CD14+ MCSF-differentiated macrophages were cultured with or without synovial fibroblasts in transwell chambers.  TNF was added at Day 0, macrophages were harvested at Day 2. Total of 4 samples: (1) macrophages alone (2) macrophages with fibroblasts (3) macrophages with TNF (4) macrophages with fibroblasts and TNF. Macrophage RNA was purified using RNeasy mini kit (Qiagen).  Tru-seq sample preparation kits (Illumina) were used to purify poly-A transcripts and generate libraries with multiplexed barcode adaptors.  All samples passed quality control on a Bioanalyzer 2100 (Agilent).  Paired-end reads (50 x 2 cycles, ~75x106 reads per sample) were obtained on an Illumina HiSeq 2500.  The TopHat program was used to align the reads to the UCSC Hg19 human reference genome, while the Cufflinks program allowed for measurements of transcript abundance (represented by Fragments Per Kilobase of exon model per Million mapped reads (FPKM)).","project":"SRP042031"}
{"number_samples":4,"species":"human","abstract":"Infection of resting primary B-lymphocytes by Epstein Barr virus (EBV) generates a population of cells that are effectively immortal. This represents the first step in the establishment of life-long viral latency in vivo and generates precursors that can develop into the lymphoid malignancies. However, virus spread requires a switch from latency to the lytic replication cycle, a process orchestrated by the virally encoded protein Zta, an AP1-like transcription factor that interacts with a 7 base-pair DNA sequence element.  As Zta has the potential to reprogram the patterns of gene expression in the host cell, we undertook global transcriptome analyses (RNA sequencing) in a Burkitt's lymphoma derived cell line in which the Zta is expressed from an inducible promoter, mimicking the switch from latency to lytic cycle. We identified 2,263 host genes whose expression levels were altered. In parallel, we performed chromatin precipitation and next-generation DNA sequencing (ChIP-Seq) to identify genes that are direct targets of Zta. Integrating these data sets revealed 277 host genes that appear to be directly regulated by Zta. Surprisingly, the frequency and distribution of the Zta binding peaks suggests that Zta regulates host genes through long-range enhancers, with a median distance of 25.8kb from the transcriptional start site, rather than equivalents of the promoter elements through which Zta regulates viral genes. Overall design: Akata cells transduced with a plasmid encoding Zta, NGFR and GFP from a doxycycline regulated promoter, and control cells in which the Zta sequences were in the opposite orientation, were treated with 500 ng/ml doxycycline for 24h.  The NGFR-expressing cells were isolated with anti-NGFR antibodies coupled to paramagnetic beads.  RNA was prepared from two independent populations of control and Zta-expressing cells and the poly A-selected transcripts were analyzed by direct sequencing. ZTA: also referred to as BZLF1.","project":"SRP042043"}
{"number_samples":18,"species":"human","abstract":"OBJECTIVE: Acromegaly is a rare endocrine disorder with excess growth hormone (GH) production. This disorder has important metabolic effects in insulin resistance and lipolysis. The objective of this study was to explore transcriptional changes induced by GH in adipose tissue. METHODS: The patients underwent clinical and metabolic profiling including assessment of HOMA-IR. Explants of adipose tissue were assayed ex-vivo for lipolysis and ceramide levels. Adipose tissue was analyzed by RNA sequencing (RNA-seq). RESULTS: There was evidence of reduced insulin sensitivity based on the increase in fasting glucose, insulin and HOMA-IR score. We observed several previously reported transcriptional changes (IGF1, IGFBP3) as well as several novel transcriptional changes, some of which may be important for GH signal regulation (PTPN3 and PTPN4) and the effect of GH on growth and proliferation. Several transcripts could potentially be important in GH-induced metabolic changes. Specifically, induction of LPL, ABHD5, and ACVR1C could contribute to enhanced lipolysis and may explain the suggestive enhancement of adipose tissue lipolysis in acromegaly patients as reflected by glycerol release from the explants of the two groups of patients (p=0.09). Higher expression of SCD and TCF7L2 could contribute to insulin resistance. Expression of HSD11B1 was reduced and GR was increased, predicting modified glucocorticoid activity in acromegaly. CONCLUSIONS: We identified the acromegaly gene expression signature in human adipose tissue. The significance of altered expression of specific transcripts will enhance our understanding of the metabolic and proliferative changes associated with acromegaly. Overall design: DESIGN: Patients with acromegaly (n=9) or non-functioning pituitary adenoma (n=11) were prospectively observed from March 2011 to June 2012. Sequencing was performed on RNA from 7 acromegaly patients and 11 controls.","project":"SRP042086"}
{"number_samples":1,"species":"human","abstract":"HKCI-3 Whole Transcriptome Sequencing","project":"SRP042104"}
{"number_samples":4,"species":"human","abstract":"We compared the epigenetic status of the mutant and disease-free iPSCs at the whole genome level. Whole epigenome profiling based on trimethylated H3K4 (H3K4me3) showed concordant epigenetic remodeling in the two corrected clones when compared with two mutant iPSC clones. Overall design: Examination of genome-wide gene expression in Fanconi anemia patient iPSCs before and after gene correction","project":"SRP042119"}
{"number_samples":2,"species":"human","abstract":"Ongoing improvements to next generation sequencing technologies are leading to longer sequencing read lengths, but a thorough understanding of the impact of longer reads on RNA sequencing analyses is lacking. To address this issue, we generated and compared two RNA sequencing datasets of differing read lengths -- 2x75 bp (L75) and 2x262 bp (L262) -- and investigated the impact of read length on various aspects of analysis, including the performance of currently available read-mapping tools, gene and transcript quantification, and detection of allele-specific expression patterns. Our results indicate that, while the scalability of read-mapping tools and the cost-effectiveness of long read protocol is an issue that requires further attention, longer reads enable more accurate quantification of diverse aspects of gene expression, including individual-specific patterns of allele-specific expression and alternative splicing. Overall design: Two RNA-Seq datasets of differing read lengths (2x262 bp and 2x75 bp)","project":"SRP042150"}
{"number_samples":28,"species":"human","abstract":"We reprogrammed human CD34+ cells from cord blood using a lentiviral vector encoding OCT4, SOX2 and KLF4.We collected RNA from parental CD34+ cells (3samples), reprogramming timepoints (9 timepoints), iPS clones derived from this experiment (6 clones), and human ES cell lines (9 samples). All samples were sequenced at 100bp reads. Overall design: Endogenous retroelement expression during reprogramming","project":"SRP042153"}
{"number_samples":875,"species":"human","abstract":"We report transcriptomes from 430 single glioblastoma cells isolated from 5 individual tumors and 102 single cells from gliomasphere cells lines generated using SMART-seq. In addition, we report population RNA-seq from the five tumors as well as RNA-seq from cell lines derived from 3 tumors (MGH26, MGH28, MGH31) cultured under serum free (GSC) and differentiated (DGC) conditions. This dataset highlights intratumoral heterogeneity with regards to the expression of de novo derived transcriptional modules and established subtype classifiers. Overall design: Operative specimens from five glioblastoma patients (MGH26, MGH28, MGH29, MGH30, MGH31) were acutely dissociated, depleted for CD45+ inflammatory cells and then sorted as single cells (576 samples). Population controls for each tumor were isolated by sorting 2000-10000 cells and processed in parallel (5 population control samples). Single cells from two established cell lines, GBM6 and GBM8, were also sorted as single cells (192 samples). SMART-seq protocol was implemented to generate single cell full length transcriptomes (modified from Shalek, et al Nature 2013) and sequenced using 25 bp paired end reads. Single cell cDNA libraries for MGH30 were resequenced using 100 bp paired end reads to allow for isoform and splice junction reconstruction (96 samples, annotated MGH30L). Cells were also cultured in serum free conditions to generate gliomasphere cell lines for MGH26, MGH28, and MGH31 (GSC) which were then differentiated using 10% serum (DGC). Population RNA-seq was performed on these samples (3 GSC, 3 DGC, 6 total). The initial dataset included 875 RNA-seq libraries (576 single glioblastoma cells, 96 resequenced MGH30L, 192 single gliomasphere cells, 5 tumor population controls, 6 population libraries from GSC and DGC samples). Data was processed as described below using RSEM for quantification of gene expression. 5,948 genes with the highest composite expression either across all single cells combined (average log2(TPM)>4.5) or within a single tumor (average log2(TPM)>6 in at least one tumor) were included. Cells expressing less than 2,000 of these 5,948 genes were excluded. The final processed dataset then included 430 primary single cell glioblastoma transcriptomes, 102 single cell transcriptomes from cell lines(GBM6,GBM8), 5 population controls (1 for each tumor), and 6 population libraries from cell lines derived from the tumors (GSC and DGC for MGH26, MGH28 and MGH31). The final matrix (GBM_data_matrix.txt) therefore contains 5948 rows (genes) quantified in 543 samples (columns). Please note that the samples which are not included in the data processing are indicated in the sample description field.","project":"SRP042161"}
{"number_samples":30,"species":"human","abstract":"The rising incidence of obesity and related disorders such as diabetes and heart disease has  focused considerable attention on the discovery of novel therapeutics. One promising  approach has been to increase the number or activity of brown-like adipocytes in white  adipose depots, as this has been shown to prevent diet-induced obesity and reduce the  incidence and severity of type 2 diabetes. Thus, the conversion of fat-storing cells into  metabolically active thermogenic cells has become an appealing therapeutic strategy to  combat obesity. Here, we report a screening platform for the identification of small molecules  capable of promoting a white-to-brown metabolic conversion in human adipocytes. We  identified two inhibitors of Janus Kinase (JAK) activity with no precedent in adipose tissue  biology that permanently confer brown-like metabolic activity to white adipocytes.  Importantly, these metabolically converted adipocytes exhibit elevated UCP1 expression and  increased mitochondrial activity. We further found that repression of interferon signalling and  activation of hedgehog signalling in JAK-inactivated adipocytes contributes to the metabolic  conversion observed in these cells. Our findings highlight a novel role for the JAK/STAT  pathway in the control of adipocyte function and establish a platform to identify compounds  for the treatment of obesity. Overall design: Human pluripotent stem-cell derived mesenchymal progenitor cells (PSC-MPCs), white adipose cells (PSC-WA), and brown adipose cells (PSC-BA) were treated with DMSO (as control), a JAK3-inhibitor compound, and a SYK-inhibitor compound respectively. Transcriptomic expression profiling was performed at 24 hours and 7 days respectively. Three biological replicates are available for each condition defined by cell type, compound, and time.","project":"SRP042186"}
{"number_samples":2,"species":"human","abstract":"Gain-of-Function CCR4 Mutations in Adult T-cell Leukemia/Lymphoma","project":"SRP042199"}
{"number_samples":15,"species":"human","abstract":"Three normal human osteoblast samples, acquired from PromoCell, were used as controls to compare to RNA-seq data from prepublished osteosarcoma samples (submitted to the European Bioinformatics Institute; EGAS00001000263) for the purpose of evaluating expression levels of genes identified as common insertions sites in a Sleeping Beauty screen of osteosarcomas in mice. Overall design: Three normal human osteoblast samples (pellet form in RNAlater) were acquired from PromoCell (Heidelberg, Germany), and RNA was isolated from them immediately upon receipt.","project":"SRP042212"}
{"number_samples":2,"species":"human","abstract":"Purpose: AURKA plays an important role in breast cancer development. Exploring the gene expression profiles regulated by AURKA will facilitate to understand the mechanism which is responsible for AURKA induced breast cancer development.  Results: We found that 350 genes were significantly up-regulated during AURKA overexpression in MCF-10A cells, 346 genes were significantly down-regulated during AURKA overexpression in MCF-10A cells.   Conclusions: Our study indicated that 696 differentially expressed genes might contribute to AURKA induced breast cancer development. Overall design: MCF-10A cells overexpressed AURKA or the empty vector were subjected to RNA extraction. The resulted RNA samples were performed RNA-sequencing analyses of gene expression profiles.","project":"SRP042218"}
{"number_samples":315,"species":"human","abstract":"We report the global pattern of ileal gene expression in a cohort of 359 treatment-naïve pediatric Crohn Disease, Ulcerative Colitis patients and controls. We focus on genes with consistent altered expression in inflamed and unaffected ileum of CD [ileal-involved CD (iCD) and non-invloved ileal CD (cCD)], but not in the ileum of ulcerative colitis or control. Overall design: Ileal biopsies were obtained during diagnostic colonoscopies of children and adolescents aged less than 17 years, who presented with IBD-like symptoms. All patients underwent baseline colonoscopy and histological characterization; non-IBD controls were those with suspected IBD, but with no microscopic or macroscopic inflammation and normal radiographic, endoscopic, and histologic findings. Biopsies were stored at -80 degrees.","project":"SRP042228"}
{"number_samples":36,"species":"human","abstract":"Cancer tissue-like structures were developed by using established human tumor cell lines in perfusion-based bioreactor systems. In colorectal cancer (CRC) cell lines, perfusion allowed more homogeneous scaffold seeding than tri-dimensional (3D) static cultures and significantly (13.7 fold, p<0.0001) higher proliferation. Resulting tissues exhibited morphology and phenotypes similar to xenografts generated in immunodeficient mice. Whole transcriptome analysis of 2D, 3D static and 3D perfusion cultures revealed the highest correlation between xenografts and 3D perfusion cultures (r=0.985). Clinically relevant concentrations of 5-FU, used in neo- and adjuvant CRC treatment, had no effect on numbers of HT-29 CRC cells cultured in 3D perfusion or xenografts, as compared with a 55.8% reduction in 2D cultures. Treatment induced apoptosis in 2D cultures, but only “nucleolar stress” in perfused cells and xenografts, consistent with partial responsiveness.  In 3D perfusion cultures BCL-2, TRAF1, and FLIP gene expression was marginally affected, as compared with significant down-regulation in 2D cell cultures. Accordingly, ABT-199 BCL-2 inhibitor, induced cytostatic effects in 3D perfusion but not in 2D cell cultures (p=0.003). Tumor cells from partially responsive (Dworak 2) patients undergoing neo-adjuvant treatment, typically (10/11) expressed BCL-2, as compared with 0/3 highly (Dworak 3-4) responsive and 4/15 fully resistant CRC (Dworak 0/1, p=0.03), closely matching 3D perfusion cultures data. These results indicate that 3D perfusion cultures efficiently mimic phenotypic and functional features observed in xenografts and clinical specimens. These models may be of critical translational relevance to address fundamental human tumor cell biology issues and to develop predictive pre-clinical tests of novel compounds. Overall design: Expression profiles of colorectal cancer cell lines cultured in 2D, 3D static, 3D perfusion or growing as xenografts were generated by deep sequencing, in triplicates, using Illumina HiSeq2000.","project":"SRP042249"}
{"number_samples":55,"species":"human","abstract":"Genome-wide mapping and characterization of novel Notch-regulated long non-coding RNAs in acute leukemia Overall design: Total RNA was extracted from samples using the RNeasy Plus mini kit (Life Technologies, Carlsbad, CA). Samples were then subject to PolyA  selection (Figures 1E, 5F and 5G only) using oligo-dT beads (Life Technologies, Carlsbad, CA) or rRNA removal (all other samples) using the Ribo-Zero kit (Epicentre, Madison, WI)  according to the manufacturers instructions. The resulting RNA samples were then used as input for library construction using the dUTP method as described by Parkhomchuck et al, 2009. RNA libraries were then sequenced on the Illumina HiSeq 2000 or 2500 using 50bp paired-end reads.","project":"SRP042286"}
{"number_samples":3,"species":"human","abstract":"Reverse transcription-derived sequences account for at least half of the human genome. Although these retroelements are formidable motors of evolution, they can occasionally cause disease, and accordingly are inactivated during early embryogenesis through epigenetic mechanisms. In the mouse, at least for endogenous retroviruses, important mediators of this process are the tetrapod-specific KRAB-containing zinc finger proteins (KRAB-ZFPs) and their cofactor TRIM28. The present study demonstrates that KRAB/TRIM28-mediated regulation is responsible for controlling a very broad range of human-specific endogenous retroelements (EREs) in human embryonic stem (ES) cells and that it exerts, as a consequence, a marked effect on the transcriptional dynamics of these cells. It further reveals reciprocal dependence between TRIM28 recruitment at specific families of EREs and DNA methylation. It finally points to the importance of persistent TRIM28-mediated control of ERE transcriptional impact beyond their presumed inactivation by DNA methylation. Overall design: Analyses of epigentic effectors and marks in KAP1 WT and KD human embryonic stem cells","project":"SRP042297"}
{"number_samples":5,"species":"human","abstract":"Inhibition of SET by siRNA or SET antagonist and CIP2A by siRNA can downregulate c-MYC and c-MYC target genes. Overall design: Cells were treated with a SET antagonist (1µMOP449) for 12 hours, or siRNA for 48 hours.","project":"SRP042303"}
{"number_samples":16,"species":"human","abstract":"We report the total RNA-seq results after CDK9, CDK12 and CDK13 depletion in human HCT116 cells for three days. RNA-seq was performed in cells using two non-targeting replicates and two different shRNAs for each CDK knockdown.  For each CDK knockdown, most of the differentially expressed genes were down-regulated with a very small subset of genes upregulated. Different CDK proteins control distinct subsets of genes in vivo, with CDK12 and CDK13 sharing more overlap in function compared to CDK9. Besides, CDK12 and CDK13 loss preferentially affects DNA damage response and snRNA gene expression, respectively. Overall design: Examine the changes of mRNA expression levels after CDK9, CDK12 and CDK13 depletion.","project":"SRP042402"}
{"number_samples":2,"species":"human","abstract":"RNA sequencing (RNA-seq) analysis of a genome-wide gene expression change in primary human neonatal dermal BECs when subjected to laminar flow-induced shear stress Overall design: Primary human neonatal dermal BECs were subjected to low level laminar flow (2 Dyne/cm2) -induced shear stress for 12 hours. Total RNA were isolated from these cells as well as from the equivalent cells cultured under the static condition as a control.","project":"SRP042579"}
{"number_samples":72,"species":"human","abstract":"Psychiatric disorders are characterized by major fluctuations in psychological function over the course of weeks and months, but the dynamic characteristics of human brain function over this timescale in healthy individuals are unknown.  Over a period of 18 months, we performed intensive phenome-wide assessment of a single human, including brain connectivity using resting fMRI, measurements of psychological and physical health, and transcriptomic and metabolomic profiling. Brain connectivity varied across sessions in relation to behavioral variables including mood, fatigue, and food/caffeine intake, as well as variables related to inflammation and gut health.   Pathway-based analysis of gene expression in peripheral lymphocytes identified associations with physical health and brain connectivity, including associations between specific brain networks and a broad set of immune-related pathways. Metabolomic measures were strongly associated with dietary variance. This study integrates dense neuroimaging and -omics profiling to provide a detailed picture of the joint dynamics of human brain and metabolic function over time, an approach that is critical for the understanding of brain disorders characterized by increased variability of brain function. Overall design: Repeated measures on a single individual; RNA was extracted from PBMCs from a single human repeatedly over the course of 18 months, under fasted conditions at a consistent time of day.  This was collected along side a large number of other variables including brain imaging. More information is available at http://www.myconnectome.org","project":"SRP042596"}
{"number_samples":14,"species":"human","abstract":"In vitro modeling of human disease has recently become feasible with the adoption of induced pluripotent stem cell (iPSC) technology. Here, we established patient-derived iPSCs from an Li-Fraumeni Syndrome (LFS) family and investigated the role of mutant p53 in the development of osteosarcoma (OS). Several members of this family carried a heterozygous p53(G245D) mutation and presented with a broad spectrum of tumors including OS. Osteoblasts (OBs) differentiated from iPSC-derived mesenchymal stem cells (MSCs) recapitulated OS features including defective osteoblastic differentiation (OB differentiation) as well as tumorigenic ability. Systematic analyses revealed that the expression of genes enriched in LFS-derived OBs strongly correlated with decreased time to tumor recurrence and poor patient survival. In silico cytogenetic region enrichment analysis (CREA) demonstrated that LFS-derived OBs do not have genomic rearrangements and hence are a particularly valuable tool for elucidating early oncogenic events prior to the accumulation of secondary alterations. LFS OBs exhibited impaired upregulation of the imprinted gene H19 during osteogenesis. Restoration of H19 expression in LFS OBs facilitated osteogenic differentiation and repressed tumorigenic potential. By integrating human imprinted gene network (IGN) and functional genomic analyses, we found that H19-mediates suppression of LFS-associated OS through the IGN component DECORIN (DCN). Downregulation of DCN impairs H19-mediated osteogenic differentiation and tumor suppression. In summary, these findings demonstrate the feasibility of studying inherited human cancer syndromes with iPSCs and also provide molecular insights into the role of the IGN in p53 mutation-mediated tumorigenesis. Overall design: mRNAseq profiling during mesenchymal stem cell differentiation to osteoblasts.","project":"SRP042597"}
{"number_samples":36,"species":"human","abstract":"To identify which genes were regulated by mRNA helicase activity, the effect of eIF4A1 knockdown on the MCF7 cell transcriptome and translatome was determined. eIF4A1-dependent mRNAs were highly enriched for several classes of genes with oncogenic potential, which leads to a model whereby dysregulation of mRNA unwinding contribues to the malignant phenotype in breast cancer cells via preferential translation of a subset of genes. Overall design: Total, subpolysomal and polysomal RNA was isolated from MCF7 cells treated with either control siRNAs or siRNAs directed against eIF4A1 (48h post-transfection).","project":"SRP042616"}
{"number_samples":168,"species":"human","abstract":"RNA-seq was performed on breast cancer cell lines and primary tumors Overall design: RNA-seq was performed on 28 breast cancer cell lines, 42 Triple Negative Breast Cancer (TNBC) primary tumors, and 42 Estrogen Receptor Positive (ER+) and HER2 Negative Breast Cancer primary tumors, 30 uninovlved breast tissue samples that were adjacent to ER+ primary tumors, 5 breast tissue samples from reduction mammoplasty procedures performed on patients with no known cancer, and 21 uninvolved breast tissue samples that were adjacent to TNBC primary tumors.","project":"SRP042620"}
{"number_samples":16,"species":"human","abstract":"Peripheral blood RNA-Seq from human coronary artery calcification cases and controls; Coronary artery calcification (CAC) is a heritable and definitive morphologic marker of atherosclerosis that strongly predicts risk for future cardiovascular events. To search for genes involved in CAC, we used an integrative transcriptomic, genomic, and protein expression strategy using next-generation DNA sequencing in the discovery phase with follow-up studies using traditional molecular biology and histopathology techniques. Overall design: Eight cases and eight controls (matched for gender, age and ancestry); RNA sequencing of peripheral blood from a discovery set of eight CAC cases and eight matched controls was used to identify dysregulated genes, which were validated using the NanoString® and Affymetrix GeneChip® Human Exon ST Array platforms.  The median CAC scores for cases in the screening and validation sets were 1531.5 and 668.5, respectively, while all controls had a score of zero.","project":"SRP042623"}
{"number_samples":12,"species":"human","abstract":"In a fluorescence polarization screen for MYC-MAX interaction, we have identified a novel small molecule inhibitor of MYC, KJ-Pyr-9, from a Kröhnke pyridine library. The Kd of KJ-Pyr-9 for MYC in vitro is 6.5 ± 1.0 nM as determined by backscattering interferometry; KJ-Pyr-9 also interferes with MYC-MAX complex formation in the cell as shown in a protein fragment complementation assay. KJ-Pyr-9 specifically inhibits MYC-induced oncogenic transformation in cell culture; it has no or only weak effects on the oncogenic activity of several unrelated oncoproteins. KJ-Pyr-9 preferentially interferes with the proliferation of MYC-overexpressing human and avian cells and specifically reduces the MYC-driven transcriptional signature. In vivo, KJ-Pyr-9 effectively blocks the growth of a xenotransplant of MYC-overexpressing  human cancer cells. Overall design: 4 treatment groups analyzed in triplicate: no treatment(control), 20uM KJ-Pyr-9, 0.1ug/mL doxycycline and KJ-Pyr-9 in combination with doxycycline","project":"SRP042630"}
{"number_samples":10,"species":"human","abstract":"Specific mutations in the XPD subunit of transcription factor IIH result in combined xeroderma pigmentosum (XP)/Cockayne syndrome (CS), a severe DNA repair disorder characterized at the cellular level by a transcriptional arrest following UV irradiation. This transcriptional arrest has always been thought to be the result of faulty transcription-coupled repair. In the present study, we investigate the transcriptional dysregulation that follows UV irradiation in XP-D/CS compared with “pure” XP-D cells or WT cells. We also study how this process is affected by the inhibition of the histone deacetylase Sirt1.","project":"SRP042647"}
{"number_samples":2,"species":"human","abstract":"An increasing amount of studies integrate mRNA sequencing data into MS-based proteomics to complement the translation product search space. We present the generation of a protein synthesis-based database from deep sequencing of ribosome-protected mRNA fragments. This approach increases the overall protein identification rates with 3% and 11% (improved and new identifications) for human and mouse respectively and enables proteome-wide detection of 5’-extended proteoforms, uORF translation and near-cognate translation start sites. Overall design: Ribosome profiling of lactimidomycin and cycloheximide treated HCT116 cells","project":"SRP042937"}
{"number_samples":27,"species":"human","abstract":"The goal of this study is to simultaneously interrogate the gene expression programs in human host cells (human foreskin fibroblasts) infected with the intracellular parasite Trypanosoma cruzi. We conducted high-resolution sequencing of the transcriptomes of T. cruzi and infected human foreskin fibroblasts (HFFs) using an RNA-seq approach. An array of computational tools was applied to map reads to the T. cruzi and human genomes and reconstruct full-length transcripts. mRNA abundance was determined for T. cruzi genes at at various time points post-infection enabling us to identify co-expression patterns that correlate with the biology of the parasite. We also conducted a time course of infection in host cells to obtain a preliminary analysis of the dynamic nature of parasite and host cell gene expression programs in the context of infection. These data provide the first glimpse of T. cruzi gene expression programs that are uniquely activated in the context of intracellular infection along with the transcriptional response of the human host cell. The study provides a solid framework for future functional and genomic studies of Chagas disease as well as intracellular pathogenesis in general.","project":"SRP043008"}
{"number_samples":12,"species":"human","abstract":"Human cell line HCT116 incubated with Myxothiazol for 5 or 17 hours","project":"SRP043021"}
{"number_samples":15,"species":"human","abstract":"To adapt a non-strandaed library preparation protocol into a strand specific one in an automated fashion. Use the data to emphasize the advantages of strand specific data.","project":"SRP043027"}
{"number_samples":20,"species":"human","abstract":"Summary: Monocyte differentiation into macrophages represents one of the cornerstone processes in innate host defense. In addition, immunological imprinting of either tolerance or trained immunity after an initial infection determines the functional fate of innate immune cells and the susceptibility of the host to secondary infections. Here we comprehensively characterize the epigenetic profiles of these functional states relative to healthy adult naïve monocytes. Inflammatory and metabolic pathways are strongly modulated in the derived macrophages, including decreased activation of inflammasome components. The cAMP-dependent signaling pathway is remodeled and adrenergic signaling was functionally implicated in trained innate immunity induction in vivo. Interestingly, ?-Glucan trains innate immune cells through extensive remodeling of distal regulatory region-bound histone acetylation, resulting in a sizeable exclusive epigenomic signature. Accordingly, genome-wide transcription factor footprint analysis reveals a specific transcription factor repertoire at trained cell-specific enhancers when recouped with epigenetic data, forming a rich hypothesis generator to manipulate innate immunity. Overall Design: Monocytes were pre-incubated either with cell culture medium (RPMI), ß-glucan (5µg/mL) or with LPS (100ng/mL), for 24 hours in a total volume of 10 mL. After a wash-out, cells were cultured in RPMI supplemented with 10% human pool serum. Monocytes were collected at different time points (0 h and 6 d after treatment) and counted before further treatment for chromatin immunoprecipitation, RNA or DNaseI treatment. Different donor Buffycoats (BC) were used as independent replicates. Replicates were generated for all the profiles including ChIPseq,RNAseq and DNaseIseq.","project":"SRP043033"}
{"number_samples":2,"species":"human","abstract":"Mobile elements are important evolutionary forces that challenge genomic integrity. Long interspersed element-1 (L1, also known as LINE-1) is the only autonomous transposon still active in the human genome. It displays an unusual pattern of evolution, with at any given time a single active L1 lineage amplifying to thousands of copies before getting replaced by a new lineage likely under pressure of host restriction factors, which act notably by silencing L1 expression during early embryogenesis. Here, we demonstrate that in human embryonic stem cells (hESC) KAP1, the master co-factor of KRAB-containing zinc finger proteins (KRAB-ZFP) previously implicated in the restriction of endogenous retroviruses, represses a discrete subset of L1 lineages predicted to have entered the ancestral genome between 26.8 and 7.6 million years ago. In the mouse, we documented a similar chronologically conditioned pattern, albeit with a much contracted time scale. We could further identify an L1-binding KRAB-ZFP, suggesting that this rapidly evolving protein family is more globally responsible for L1 recognition. KAP1 knockdown in hESC induced the expression of KAP1-bound L1 elements, but their younger, human-specific counterparts (L1Hs) were unaffected. Instead, they were stimulated by depleting DNA methyltransferases, consistent with recent evidence demonstrating that the PIWI-piRNA pathway regulates L1Hs in hESC. Altogether, these data indicate that the early embryonic control of L1 is an evolutionary dynamic process, and support a model whereby newly emerged lineages are first suppressed by DNA methylation-inducing small RNA-based mechanisms, before KAP1-recruiting protein repressors are selected. Overall design: HA-tagged Gm6871 ChIP-seq in mES cells, RNA-seq in control and Gm6871 KD mES cells, KAP1 ChIP-seq in WT mES cells, RNA-seq in  control and DNMTs KD hES cells.","project":"SRP043041"}
{"number_samples":6,"species":"human","abstract":"Background: The ZNF217 gene, encoding a C2H2 zinc finger protein, is located at 20q13 and found amplified and overexpressed in greater than 20% of breast tumors. Current studies indicate ZNF217 drives tumorigenesis, yet the regulatory mechanisms of ZNF217 are largely unknown. Because ZNF217 associates with chromatin modifying enzymes, we postulate that ZNF217 functions to regulate specific gene signaling networks. Here, we present a large-scale functional genomic analysis of ZNF217, which provides insights into the regulatory role of ZNF217 in MCF7 breast cancer cells. Results: ChIP-seq analysis reveals that the majority of ZNF217 binding sites are located at distal regulatory regions associated with the chromatin marks H3K27ac and H3K4me1. Analysis of ChIPseq transcription factor binding sites shows clustering of ZNF217 with FOXA1, GATA3 and ERalpha binding sites, supported by the enrichment of corresponding motifs for the ERalpha-associated cisregulatory sequences. ERalpha expression highly correlates with ZNF217 in lysates from breast tumors (n=15), and ERalpha co-precipitates ZNF217 and its binding partner CtBP2 from nuclear extracts. Transcriptome profiling following ZNF217 depletion identifies differentially expressed genes co-bound by ZNF217 and ERalpha; gene ontology suggests a role for ZNF217-ERalpha in expression programs associated with ER+ breast cancer studies found in the Molecular Signature Database. Data-mining of expression data from breast cancer patients correlates ZNF217 with reduced overall survival in multiple subtypes. Conclusions: Our genome-wide ZNF217 data suggests a functional role for ZNF217 at ERalpha target genes. Future studies will investigate whether ZNF217 expression contributes to aberrant ERalpha regulatory events in ER+ breast cancer and hormone resistance Overall design: Differential RNA-seq profiling from triplicate biological replicates of MCF7 cells treated with scrambled siRNA or siZNF217.","project":"SRP043043"}
{"number_samples":6,"species":"human","abstract":"Our data demonstrated that Bcl6 directly binds and represses trafficking receptors S1pr1 and Grp183 by recruiting Hdac2 through the RD2 domain.  Deregulation of these genes impairs B-cell migration and may contribute to the Germinal Center failure in Bcl6RD2MUT mice. Overall design: RNAseq was performed in endogenous BCL6-depleted OCI-LY1 cells rescued with either WT or RD mutant BCL6 (N=3 for each group).","project":"SRP043078"}
{"number_samples":24,"species":"human","abstract":"Substantial effort is currently devoted to identifying cancer-associated alterations using genomics. Here, we show that standard blood collection procedures rapidly change the transcriptional and post-transcriptional landscapes of hematopoietic cells, resulting in biased activation of specific biological pathways, up-regulation of pseudogenes, antisense RNAs, and unannotated coding isoforms, and RNA surveillance inhibition. Affected genes include common mutational targets and thousands of other genes participating in processes such as chromatin modification, RNA splicing, T and B cell activation, and NF-?B signaling. The majority of published leukemic transcriptomes exhibit signals of this incubation-induced dysregulation, explaining up to 40% of differences in gene expression and alternative splicing between leukemias and reference normal transcriptomes. The effects of sample processing are particularly evident in pan-cancer analyses. We provide biomarkers that detect prolonged incubation of individual samples, and show that keeping blood on ice markedly reduces changes to the transcriptome. In addition to highlighting the potentially confounding effects of technical artifacts in cancer genomics data, our study emphasizes the need to survey the diversity of normal as well as neoplastic cells when characterizing tumors. This study is complemented by GSE61410: transcriptomic profiling of bone marrow cells from healthy individuals. Overall design: Peripheral blood mononuclear cells (PBMCs) were isolated from four healthy individuals, following an ex vivo incubation of variable length at either room temperature or on ice. RNA transcriptomes were measured using the Illumina HiSeq.","project":"SRP043080"}
{"number_samples":20,"species":"human","abstract":"We have generated nine synthetic poly-adenylated RNA transcripts that correspond to previously reported oncogenic gene fusions. These synthetic RNAs were spiked at known concentrations over a wide range into total RNA prior to construction of next-generation sequencing mRNA libraries to generate RNA-seq data.","project":"SRP043081"}
{"number_samples":21,"species":"human","abstract":"Advanced basal cell carcinomas (BCCs) frequently acquire resistance to Smoothened (SMO) inhibitors through unknown mechanisms, providing a unique opportunity to study human tumor evolution. Here, we identify SMO mutations in 50% (22/44) of resistant BCCs compared with 5.6% (2/36) of untreated BCCs (p<0.0001), and show that these mutations maintain Hedgehog signaling in the presence of SMO inhibitors. Alterations include four ligand binding pocket (LBP) mutations that define sites of inhibitor binding and four variants that confer constitutive activity and inhibitor resistance, thus defining pivotal residues of SMO that ensure receptor autoinhibition. Finally, we show that both classes of SMO variants respond to the aPKC-?/? inhibitor PSI and GLI2 antagonist ATO that operate downstream of SMO Overall design: Genome-wide gene expression profiling of 8 normal skin biopsies, 9 resistant basal cell cancers and 4 sensitive basal cell cancers.","project":"SRP043085"}
{"number_samples":6,"species":"human","abstract":"Changes in alternative splicing in breast cancer cells expressing control, empty vector or Flag-tagged wild type RBM47 were analyzed using paired-end, 100bp RNAseq. Related data published together with these data are found in GSE53779 Overall design: Triplicate RNAseq libraries were prepared from non-clonal brain metastatic breast cancer cells stably expressing empty-vector, and a clonal cell line (wt#10) expressing Flag-tagged, wild-type RBM47 under a doxycline-inducible promoter, both treated for three days with doxycycline to induce transgene expression","project":"SRP043090"}
{"number_samples":80,"species":"human","abstract":"The amount of RNA sequencing data on skeletal muscle is very limited. We have analyzed a large set of human muscle biopsy samples and provide extensive information on the baseline skeletal muscle transcriptome, including completely novel protein-coding transcripts. Overall design: Analyze of transcriptome in 23 skeletal muscle biopsy samples from six individuals. Four biopsies from each subject, two biopsies from each leg (except subject 6 which has only three biopsies in total).","project":"SRP043108"}
{"number_samples":12,"species":"human","abstract":"SAMHD1  restricts  HIV-1  replication  in  dendritic  and  other  myeloid  cells. SAMHD1 has been shown to possess a dGTP-dependent dNTP triphosphatase (dNTPase) activity and is proposed to inhibit HIV-1 replication by depleting the intracellular dNTP pool. Arguing against a role for SAMHD1 dNTPase in HIV-1 restriction, the phosphorylation of SAMHD1 regulates the restriction activity toward HIV-1 without affecting its ability to decrease cellular dNTP levels. Here, we show that SAMHD1 is a phospho-regulated RNase and that the RNase function is required for HIV-1 restriction. Mutation of the SAMHD1 D137 residue in the allosteric site (SAMHD1D137N) abolishes dNTPase activity but has no effect on RNase activity. This dNTPase-defective SAMHD1D137N mutant is able to restrict HIV-1 infection to nearly the same extent as wild-type SAMHD1. SAMHD1 associates with and degrades the HIV-1 genomic RNA during the early phases of infection. SAMHD1 silencing in macrophages and CD4+ T cells from healthy donors increases HIV-1 RNA stability, thus rendering the cells permissive for HIV-1 infection. Furthermore, the phosphorylation of SAMHD1 at position T592 abolishes the RNase activity toward HIV-1 RNA, and consequently the ability of SAMHD1 to restrict HIV-1 infection, uncovering the phosphorylation of SAMHD1 T592 as a negative regulatory mechanism of RNase activity. Together, our results demonstrate that SAMHD1 is an essential RNase that prevents HIV-1 infection by directly degrading HIV-1 genomic RNA in a phosphorylation-regulated manner. The unique property of SAMHD1 that cleaves HIV-1 genomic RNA with no sequence preferences could be exploited to develop a new class of intervention for error-prone retroviruses. Overall design: Ribosomal RNA-depleted total RNA profiles of mock, SAMHD1 wild type and mutants infected with HIV-1 were examined at the time of 0, 1, 3 h by Illumina Hiseq2500.","project":"SRP043144"}
{"number_samples":53,"species":"human","abstract":"Rationale: Asthma is a chronic inflammatory airway disease. Children with severe asthma have lower levels of vitamin D than children with moderate asthma, and among children with severe asthma, airway smooth muscle (ASM) mass is inversely related to vitamin D levels. Beta2 agonists are a common asthma medication that act partly by targetting the ASM. We used RNA-Seq to characterize the human ASM transcriptome of fatal and asthma vs. contols at baseline and under two treatment conditions. Methods: The Illumina TruSeq assay was used to prepare 75bp paired-end libraries for ASM cells from white donors, 6 with fatal asthma and 12 control donors under three treatment conditions: 1) no treatment; 2) treatment with a ß2-agonist (i.e. Albuterol, 1µM for 18h); 3) treatment with vitamin D 100 nM for 18h). Llibraries were sequenced with an Illumina Hi-Seq 2000 instrument. The Tuxedo Suite Tools were used to align reads to the hg19 reference genome, assemble transcripts, and perform differential expression analysis using the protocol described in https://github.com/blancahimes/taffeta Overall design: mRNA profiles obtained via RNA-Seq for primary human airway smooth muscle cell lines from fatal asthma or control donors that were treated with vitamin D, albuterol, or were left untreated.","project":"SRP043162"}
{"number_samples":12,"species":"human","abstract":"Genes were identified which modified their expression in a dose-dependent manner upon exposure to doxycycline. The more represented cellular pathways included all mitochondrial genes, the endoplasmic reticulum stress response, integrins and extracellular matrix components, and growth factors. Overall design: Examination of 4 different doses of doxycycline in three human pterygium samples.","project":"SRP043166"}
{"number_samples":1,"species":"human","abstract":"Coilin iCLIP data revealed 42 novel human snoRNAs of intronic origin. To validate their expression and estimate abundance  of novel and annotated snoRNAs, we performed RNA-seq on polyA- and rRNA-depleted RNA isolated from HeLa cells. Results show that expression of novel snoRNAs is comparable to the previously annotated snoRNAs. Overall design: 1 replicate of RNA depleted of polyA and ribosomal RNA.","project":"SRP043188"}
{"number_samples":38,"species":"human","abstract":"Facioscapulohumeral dystrophy (FSHD) is caused by decreased epigenetic repression of the D4Z4 macrosatellite array and recent studies have shown that this results in the expression of low levels of the DUX4 mRNA in skeletal muscle. Several other mechanisms have been suggested for FSHD pathophysiology and it remains unknown whether DUX4 expression can account for most of the molecular changes seen in FSHD. Since DUX4 is a transcription factor, we used RNA-seq to measure gene expression in muscle cells transduced with DUX4, and in muscle cells and biopsies from control and FSHD individuals. We show that DUX4 target gene expression is the major molecular signature in FSHD muscle together with a gene expression signature consistent with an immune cell infiltration. In addition, one unaffected individual without a known FSHD-causing mutation showed expression of DUX4 target genes. This individual has a sibling with FSHD and also without a known FSHD-causing mutation, suggesting the presence of yet unidentified modifier locus for DUX4 expression and FSHD. These findings demonstrate that expression of DUX4 accounts for the majority of the gene expression changes in FSHD skeletal muscle together with an immune cell infiltration. Overall design: RNA-seq for muscle cells and biopsies from control and FSHD individuals.","project":"SRP043221"}
{"number_samples":4,"species":"human","abstract":"We report the application of high-throughput sequencing to performed the p53 regulated trancriptome in HCT116 colon cancer cells treated with the DNA damage 5FU. To study the direct targets of p53 we performed ChIP-seq to deterrmined the p53 biding sites and associated with the expression levels. With this study we identified the new genomic regions regulated by p53 and with special attention in those regions that are non coding and are differentially expressed by the DNA damage drug. Overall design: Description of the p53 transcriptome in HCT116 colon cancer cell line. The RNA-seq libraries were prepared from purified poly-A+ RNA from untreated and 5-Fluorouracil treated p53 +/+ HCT116 cells for 4 and 12h, including two independent samples for the time 12h. Paired-end and strand specific RNA sequencing libraries were prepared according to Illumina instructions and sequenced on HiSeq 2000 (Ilumina) with sequence length of 150 bp. Raw sequencing data were alignment to the human genome (hg19) using Tophat mapper.","project":"SRP043273"}
{"number_samples":12,"species":"human","abstract":"The patient-derived xenograft (PDX) model retains the heterogeneity of patient tumors, allowing a means to not only examine efficacy of a therapy across a population, but also study crucial aspects of cancer biology in response to treatment. Herein we describe the development and characterization of an ovarian-PDX model in order to study the development of chemoresistance. We demonstrate that PDX tumors are not simply composed of tumor-initiating cells, but recapitulate the original tumor’s heterogeneity, oncogene expression profiles, and clinical response to chemotherapy. Combined carboplatin/paclitaxel treatment of PDX tumors enriches the cancer stem cell populations, but persistent tumors are not entirely composed of these populations. RNA-Seq analysis of treated PDX tumors compared to untreated tumors demonstrates a consistently contrasting genetic profile after therapy, suggesting similar, but few, pathways are mediating chemoresistance. The pathways most significantly altered included Protein Kinase A signaling, GNRH signaling, and sphingosine-1-phosphate signaling. Pathways and genes identified by this methodology represent novel approaches to targeting the chemoresistant population in ovarian cancer Overall design: 6 pairs of Patient-Derived Xenografts (PDX) were ananlyzed using RNA-seq for a total of 12 samples. Each pair consists of a treated and untreated PDX of ovarian cancer. Treated Ovarian cancer PDXs were treated with 4 weeks of a combination of carboplatin and taxol. RNA was isolated and converted to cDNA. RNA-seq was conductred on the Illumina HiSeq 2000 with 50 bp paired end sequencing","project":"SRP043320"}
{"number_samples":2,"species":"human","abstract":"Gene expression and splicing switches upon RBM4 overexpression Overall design: 2 Samples","project":"SRP043336"}
{"number_samples":10,"species":"human","abstract":"T follicular helper (Tfh) cells are a subset of CD4+ T helper (Th) cells that migrate into germinal centers and promote B cell maturation into memory B and plasma cells. Tfh cells are necessary for promotion of protective humoral immunity following pathogen challenge, but when aberrantly regulated, drive pathogenic antibody formation in autoimmunity and undergo neoplastic transformation in angioimmunoblastic T-cell lymphoma and other primary cutaneous T-cell lymphomas. Limited information is available on the expression and regulation of genes in human Tfh cells. Using a fluorescence activated cell sorting-based strategy, we obtained primary Tfh and non-Tfh T effector (Teff) cells from tonsils and prepared genome-wide maps of active, intermediate, and poised enhancers determined by ChIP-seq, with parallel transcriptome analyses determined by RNA-seq. Tfh cell enhancers were enriched near genes highly expressed in lymphoid cells or involved in lymphoid cell function, with many mapping to sites previously associated with autoimmune disease in genome-wide association studies. A group of active enhancers unique to Tfh cells associated with differentially expressed genes was identified. Fragments from these regions directed expression in reporter gene assays. These data provide a significant resource for studies of T lymphocyte development and differentiation and normal and perturbed Tfh cell function. Overall design: Using a fluorescence activated cell sorting-based strategy, we obtained primary Tfh and non-Tfh T effector (Teff) cells from tonsils and prepared genome-wide maps of active, intermediate, and poised enhancers determined by ChIP-seq, with parallel transcriptome analyses determined by RNA-seq.","project":"SRP043339"}
{"number_samples":52,"species":"human","abstract":"We carried out RNA-sequencing (RNA-seq) of adult human postmortem neocortical brain tissue, and then correlated those expression values with the fMRI signal in each brain region Overall design: Ten cortical regions were included in the analysis: pre-motor cortex - PMV (BA6), dorsolateral prefrontal cortex – DLPFC (BA9), middle temporal gyrus – pMTG (BA21), superior temporal gyrus – pSTG (BA22), angular gyrus  - AG (BA39), supramarginal gyrus  - SMG (BA40), pars opercularis  - POP (BA44), pars triangularis  - PTr (BA45), middle frontal gyrus – MFG (BA46) and pars orbitalis  - POrB (BA47). For each brain region, three or more samples from left adult brain hemispheres were collected (ages range from 33 to 49) and only males were included to avoid the effect of sex","project":"SRP043364"}
{"number_samples":26,"species":"human","abstract":"The amount of RNA sequencing data on skeletal muscle is very limited. We have analyzed a large set of human muscle biopsy samples and provide extensive information on the baseline skeletal muscle transcriptome, including completely novel protein-coding transcripts. Overall design: Analyze of transcriptome in 24 skeletal muscle biopsy samples, 12 individuals and one biopsy per leg per individual","project":"SRP043368"}
{"number_samples":12,"species":"human","abstract":"The transcription factor FOXM1 binds to sequence-specific motifs on DNA (C/TAAACA) through its DNA binding domain (DBD), and activates proliferation- and differentiation-associated genes. Aberrant overexpression of FOXM1 is a key feature in oncogenesis and progression of many human cancers. Herein we identify novel inhibitors of FOXM1 that block DNA binding from a high-throughput screen applied to a library of 54,211 small molecules. One compound, FDI-6 (NCGC00099374) is studied in depth and is shown to bind directly to FOXM1 protein, displace the transcription factor from genomic targets in MCF-7 breast cancer cells, and induce concomitant transcriptional down-regulation. Global transcript profiling of MCF-7 cells by RNA-seq shows that FDI-6 specifically down regulates FOXM1-activated genes with FOXM1 occupancy confirmed by ChIP-seq. This small molecule mediated effect is selective for FOXM1-controlled genes with no effect on genes regulated by homologous forkhead family factors. Overall design: 12 samples, 100 single-ended RNAseq libraries: 3 replicates for untreated (0 hours); 3 replicates for 3 hours treatment; 3 replicates for 6 hours treatment; 3 replicates for 9 hours treatment. Treatment: compound FDI-6 inhibiting binding of FOXM-1 to DNA.","project":"SRP043378"}
{"number_samples":16,"species":"human","abstract":"We utilized RNA sequencing to expand and better define the molecular entities involved in the transcriptional programming within the eosphagus in eosinophilic esophagitis (EoE) Overall design: Examination of differentially expressed genes from RNA sequencing data performed on esophageal biopsy specimen from 6 healthy controls and 10 patients with active EoE","project":"SRP043388"}
{"number_samples":6,"species":"human","abstract":"Complex bacteriotherapy, such fecal microbiome transplantation (FMT) is an emerging therapeutic modality. Recent trials showed variable efficacy of FMT for treating ulcerative colitis (UC). However, only one case report utilized more than 5 FMTs when treating UC, thus far. We studied the clinical, metagenomic, and mucosal gene expression changes induced by serial FMTs in 3 pediatric UC patients during withdrawal of their immunosuppression. FMTs were safe and transiently supported immunotherapy withdrawal. This therapeutic effect associated with host gene expression changes relevant for enterocyte replication suppression. Our findings indicate the safety and therapeutic potential of serial FMTs in pediatric UC.","project":"SRP043391"}
{"number_samples":42,"species":"human","abstract":"Massively parallel DNA sequencing of thousands of samples in a single machine-run is now possible, but the preparation of the individual sequencing libraries is expensive and time-consuming. Tagmentation-based library construction, using the Tn5 transposase, is efficient for generating sequencing libraries but currently relies on undisclosed reagents, which severely limits development of novel applications and the execution of large scale projects. Here, we present simple and robust procedures for Tn5 transposase production and optimized reaction conditions for tagmentation-based sequencing library construction. We further show how molecular crowding agents both modulate library lengths and enable efficient tagmentation from sub-picogram amounts of cDNA. Comparison of single-cell RNA-sequencing libraries generated using produced and commercial Tn5 demonstrated equal performances in terms of gene detection and library characteristics. Finally, as naked Tn5 can be annealed to any oligonucleotide of choice, for example molecular barcodes in single-cell assays or methylated oligonucleotides for bisulfite sequencing, custom Tn5 production and tagmentation enables innovation in sequencing-based applications. Overall design: Sequencing libraries from cDNA was generated using in-house produced or commercial Tn5 and in a variety of tagmentation buffers and conditions.","project":"SRP043417"}
{"number_samples":2,"species":"human","abstract":"The goal of this experiment was to test whether human hepatocytes could give rise to biliary-like progenitor cells in an in vivo context.  Here Fah-/- Il2ry-/- Rag2-/-NOD mouse livers were humanized with human hepatocytes. Only hepatocytes engraft in the Fah-/- mouse at detectable levels in this model.  Then animals were given chronic liver injury with 0.1% ddc.  After injury we measured human-specific transcripts to determine whether the phenotype of the human cells had changed.  Specifically, we evaluated the relative levels of human biliary duct markers such as Spp1, Sox9, Krt7, etc.  and hepatocyte markers such as Alb, Ttr, Fah, etc. Overall design: 3 DDC treated chimeras and 6 untreated chimeras are included.  Additional controls include a normal human liver biopsy, FACS sorted primary intrahepatic human bile duct cells, mouse hepatocytes, and mouse intrahepatic biliary cells in ddc treated animal.","project":"SRP043434"}
{"number_samples":8,"species":"human","abstract":"We profiled primary breast cancer, nodal and liver metastatic tumours from three patients. At the time of initial diagnosis, all three patients presented with luminal breast cancer with adjacent nodal metastasis. They all received 5 years of enodrine therapy and all subsequently developed liver metastasis. Overall design: Examination of mRNA differences between primary, nodal and metastatic tumour samples.","project":"SRP043470"}
{"number_samples":2,"species":"human","abstract":"We report the chromatin modification dynamics at p53 binding sites upon treatment with DMSO or nutlin3-a (5uM) in IMR90 human lung fibroblasts using ChIP-seq and RNA-seq analyses. We assessed the genomewide changes in H3, H3K4me3, H3K4me2, H3K4me1, H3K27ac, H4K16ac, RNA polymerase II, and p53 in response to p53 activation. Overall design: ChIP-seq of 8 histone modifications or proteins for 2 conditions in 1 cell line.","project":"SRP043510"}
{"number_samples":2,"species":"human","abstract":"Single Cell RNAseq data and PacBio amplicon sequencing data","project":"SRP043513"}
{"number_samples":1,"species":"human","abstract":"We performed NGS-based transcript profiling (RNA-seq) to profile transcripts that are expressed in MCF10A cells.  12,332 genes with FPKM>1 were considered as expressed in MCF10A cells. Overall design: mRNA profiles of MCF10A cells were generated by deep sequencing using Illumina Hiseq 2000.","project":"SRP043578"}
{"number_samples":10,"species":"human","abstract":"MicroRNA (miRNA) play a major role in the post-transcriptional regulation of gene expression. In mammals most miRNA derive from the introns of protein coding genes where they exist as hairpin structures in the primary gene transcript, synthesized by RNA polymerase II (Pol II). These are cleaved co-transcriptionally by the Microprocessor complex, comprising DGCR8 and the RNase III endonuclease Drosha, to release the precursor (pre-)miRNA hairpin, so generating both miRNA and spliced messenger RNA1-4. However, a substantial minority of miRNA originate from Pol II-synthesized long non coding (lnc) RNA where transcript processing is largely uncharacterized5. Here, we show that most lnc-pri-miRNA do not use the canonical cleavage and polyadenylation (CPA) transcription termination pathway6, but instead use Microprocessor cleavage both to release pre-miRNA and terminate transcription. We present a detailed characterization of one such lnc-pri-miRNA that generates the highly expressed liver-specific miR-1227. Genome-wide analysis then reveals that Microprocessor-mediated transcription termination is commonly used by lnc-pri-miRNA but not by protein coding miRNA genes. This identifies a fundamental difference between lncRNA and pre-mRNA processing. Remarkably, inactivation of the Microprocessor can lead to extensive transcriptional readthrough of lnc-pri-miRNA, resulting in inhibition of downstream genes by transcriptional interference. Consequently we define a novel RNase III-mediated, polyadenylation-independent mechanism of Pol II transcription termination in mammalian cells. Overall design: Chromatin associated RNA-seq from sicntrl,siDrosha,siDGCR8 treated Hela cells. Same for sicntrl and siDGCR8 from Huh7 cells. Nuclear polyA + and polyA- RNA-seq from sicntrl and siDGCR8 in HeLa cells. Chromatin associated RNA-seq from siDicer treated Hela cells.","project":"SRP043593"}
{"number_samples":7,"species":"human","abstract":"Whole-exome sequencing studies have identified common mutations affecting genes encoding components of the RNA splicing machinery in hematological malignancies. Here, we sought to determine how mutations affecting the 3' splice site recognition factor U2AF1 altered its normal role in RNA splicing. We find that U2AF1 mutations influence the similarity of splicing programs in leukemias, but do not give rise to widespread splicing failure. U2AF1 mutations cause differential splicing of hundreds of genes, affecting biological pathways implicated in myeloid disease such as DNA methylation (DNMT3B), X chromosome inactivation (H2AFY), the DNA damage response (ATR, FANCA), and apoptosis (CASP8). We show that U2AF1 mutations alter the preferred 3' splice site motif in vivo, in cell culture, and in vitro. Mutations affecting the first and second zinc fingers give rise to different alterations in splice site preference and largely distinct downstream splicing programs. These allele-specific effects are consistent with a computationally predicted model of U2AF1 in complex with RNA. Our findings suggest that U2AF1 mutations contribute to pathogenesis by causing quantitative changes in splicing that affect diverse cellular pathways, and give insight into the normal function of U2AF1’s zinc finger domains. Overall design: mRNA profiles of K562 cells expressing U2AF1 WT, mutants and knockdown of U2AF1 generated by deep sequencing.","project":"SRP043621"}
{"number_samples":4,"species":"human","abstract":"Purpose:  Cellular senescence is a cell stress response resulting in permanent growth arrest and the production of an altered pro-inflammatory secretory profile known as the senescecnce-associated secretory phenotype (SASP). The induction of senescence in astrocytes, a cell type responsible for maintaining homeostasis within the central nervous system (CNS) and responding to CNS insults, has been implicated in neurodegenerative disease. However, little is known about the senescent transcriptome in CNS-derived cell types including astrocytes. Methods: To better understand senescence-associated gene expression changes in astrocytes, we investigated global changes in the astrocyte transcriptome using RNA-seq following the induction of oxidative stress-induced senescence with hydrogen peroxide. Results:  During senescence, we find evidence of a loss of brain expressed transcripts involved in diverse CNS processes including neuronal differentiation and development, gliogenesis, axonogenesis, and learning and memory as well as a loss of transcripts involved in MHC class II antigen processing and presentation. In addition, we find evidence for induction of the senescent phenotype including a loss of transcripts involved in cell division and an increase in the mRNA level of inflammatory mediators suggestive of a SASP. Conclusions: Overall, our findings suggest a loss of differentiated function in senescent astrocytes and a gain in neuroinflammatory function as part of the SASP as a potential mechanisms for dysfunction in the aging brain. Overall design: Examination of transcriptome changes by RNAseq in pre-senescent and senescent astrocytes using 2 biological replicates per condition","project":"SRP043644"}
{"number_samples":18,"species":"human","abstract":"Bipolar Disorder (BD) is a complex neuropsychiatric disorder that is characterized by intermittent episodes of mania and depression and, without treatment, 15% of patients commit suicide1. Hence, among all diseases, BD has been ranked by the WHO as a top disorder of morbidity and lost productivity2. Previous neuropathological studies have revealed a series of alterations in the brains of BD patients or animal models3, such as reduced glial cell number in the patient prefrontal cortex4, up-regulated activities of the PKA/PKC pathways5-7, and changes in dopamine/5-HT/glutamate neurotransmission systems8-11. However, the roles and causation of these changes in BD are too complex to exactly determine the pathology of the disease; none of the current BD animal models can recapitulate both the manic and depressive phenotypes or spontaneous cycling of BD simultaneously12,13. Furthermore, while some patients show remarkable improvement with lithium treatment, for yet unknown reasons, other patients are refractory to lithium treatment. Therefore, developing an accurate and powerful biological model has been a challenge for research into BD. The development of induced pluripotent stem cell (iPSC) technology has provided such a new approach. Here, we developed a human BD iPSC model and investigated the cellular phenotypes of hippocampal dentate gyrus neurons derived from the patient iPSCs. Using patch clamp recording, somatic Ca2+ imaging and RNA-seq techniques, we found that the neurons derived from BD patients exhibited hyperactive action potential (AP) firing, up-regulated expression of PKA/PKC/AP and mitochondria-related genes. Moreover, lithium selectively reversed these alterations in the neurons of patients who responded to lithium treatment. Therefore, hyper-excitability is one endophenotype of BD that is probably achieved through enhancement in the PKA/PKC and Na+ channel signaling systems, and our BD iPSC model can be used to develop new therapies and drugs aimed at clinical treatment of this disease. Overall design: total RNAseq from neurons generated from BD patient-specific iPS cells","project":"SRP043684"}
{"number_samples":3,"species":"human","abstract":"To explore infectious agents associated with nasopharyngeal carcinomas (NPCs), we have sequenced human NPC cell lines using Illumina RNA-sequencing.","project":"SRP043686"}
{"number_samples":10,"species":"human","abstract":"The study goal was to develop a new and robust protocol for blood preservation. It also involved a proof-of-concept study to preserve rare cell samples.","project":"SRP043688"}
{"number_samples":21,"species":"human","abstract":"Introduction: The identification of biomarkers in Barrett’s oesophagus (BO) that stratify groups at risk of progressing to oesophageal adenocarcinoma (OAC) would allow tailored surveillance strategies. We have applied a novel high throughput RNA sequencing analysis characterizing the BO mRNA transcriptome across the metaplasia-dysplasia sequence to identify potential markers of progression in an unbiased fashion and functionally validated these at the protein level. Methods: Matched biopsy samples for histology and RNA extraction were taken from BO patients of known histological grade. RNA was extracted from matched samples and sequenced to 60bp length (paired-end).21 samples were sequenced (HGD,7; LGD, 7 and SIM, 7). Reads obtained were mapped to NCBI build37.2 using TopHat. Read count generation, normalisation and differential expression (DE) analysis was performed using the HTSeq-DESeq pipeline. Significantly DE genes (>2 fold change in expression with B-H adjusted p-value <0.1) were further assessed for network and biological relevance using Ingenuity Pathway analysis. Candidate genes were selected and validated in a larger cohort (n=64) using RT-PCR.  Targets were further validated (by ELISA and immunohistochemistry) in independent cohorts of patients using serum and tissue microarrays. Results: DE analysis was performed in 3 groups with 2 conditions at a time using the lower grade cohort as control and the higher grade as comparator: SIM vs. LGD (demonstrated 218 DE genes, 131 up-regulated in LGD, 87 down-regulated compared to SIM), SIM vs. HGD (49 DE, 27 up, 22 down) and LGD vs. HGD (317 DE, 81 up, 216 down). Six network-central candidate genes (FOSB, IL-1B, SERPINA3, KLK7, GSTM5 &SCUBE2) were selected for RT-PCR validation following network and functional analysis. Circulating IL-1ß and SERPINA-3 demonstrated progressive significant increases in expression across the dysplasia sequence to OAC (p<0.005)). This was confirmed at the tissue level showing significant differences between SIM and dysplastic BO (p<0.05). Conclusion: The use of RNA-sequencing as a detailed and unbiased analysis method identifies IL-1ß and SERPINA-3 as novel candidates differentially expressed along the metaplasia-dysplasia-cancer sequence in BO. Overall design: RNA sequencing was performed on mRNA extracted from oesophgeal biopsies from patients with non dysplstic Barrett's esophagus, low grade dysplasia and high grade dysplasia using Illumina GAIIx","project":"SRP043694"}
{"number_samples":27,"species":"human","abstract":"The transcription factor GATA2 regulates chemotherapy resistance in prostate cancer. We report a novel GATA2 transcriptional program that has implications for chemotherapy resistance disease and aggressiveness in castration resistant prostate cancer. Overall design: Examination of the transcriptional network changes induced in human Ch-CRPC cell lines by two shRNA mediated knock down of GATA2 versus random shRNA control","project":"SRP043960"}
{"number_samples":6,"species":"human","abstract":"The investigation of the association of long non-coding RNAs (lncRNAs) with DNMT1 by RIP-seq reveals that DNMT1 interacts with DACOR1. We identified the genomic occupancy sites of DACOR1 through ChIRP-seq, which we found to significantly overlap with known differentially methylated regions (DMRs) in colon tumors.  Induction of DACOR1 in colon cancer cell lines significantly reduced their ability to form colonies in vitro, suggesting a growth suppressor function. Consistent with the observed phenotype, induction of DACOR1 led to the activation of tumor-suppressor pathways and attenuation of cancer-associated metabolic pathways. Notably, DACOR1 induction resulted in down-regulation of Cystathionine ß-synthase (CBS), which is known to lead to increased levels of S-adenosyl methionine (SAM) – the key methyl donor for DNA methylation. Overall design: The DNA methyltransferase DNMT1 was immunoprecipitated and all associated RNAs were identified by RNA-seq and subsequent bioinformatic analyses. Samples were prepared for DNA sequencing through the ChIRP-seq protocol which uses antisense DNA oligos to bind to DACOR1 and pull down associated DNA in complex. We sequenced one sample with DACOR1 specific probes and non-specific DNA probes as control. Three samples were collected for each control lentivirus transduced V852 colon cancer line and DACOR1 lentivirus transduced V852. Samples were processed for RNA-sequencing and analyzed through differential expression analysis.","project":"SRP043961"}
{"number_samples":9,"species":"human","abstract":"Sequencing of 5' ends of RNA molecules from control and exosome-depleted HeLa-S3 cells. Overall design: CAGE library construction from RNA extracted from control and exosome-depleted cells.","project":"SRP043962"}
{"number_samples":9,"species":"human","abstract":"ETS1 and RAS/ERK regulate a common gene expression program in establishing enviroment suitable for prostate cancer cell migration. Overall design: mRNA profiles of luciferase knockdown (WT), ETS1 knockdown, and U0126 treated DU145 cells were generated using deep sequencing, in triplicate, using Illumina HiSeq. Knockdowns were stable shRNA expression from a lentiviral construct selected with puromycin.","project":"SRP044013"}
{"number_samples":24,"species":"human","abstract":"The biphasic epithelioid (E-) and sarcomatoid(S-) components of sarcomatoid RCC and epithelioid (E-) and rhabdoid (R-) components of rhabdoid RCC shared a similar transcriptomic signature, despite morphologic differences; by contrast, the transcriptome of sarcomatoid and rhabdoid RCC was sharply distinct from non-sarcomatoid RCC. Overall design: Total RNA was processed for RNA-seq from the following patient samples:  7 sarcomatoid RCC (E- and S- pairs), 4 rhabdoid RCC (E- and R- pairs) and 15 non-sarcomatoid RCC.","project":"SRP044042"}
{"number_samples":15,"species":"human","abstract":"Polycomb Repressive Complex 2 (PRC2) plays crucial roles in transcriptional regulation and stem cell development. However, the context-specific functions associated with alternative subunits remain largely unexplored. Here we show that the related enzymatic subunits EZH1 and EZH2 undergo an expression switch during hematopoiesis. We examine the in vivo stoichiometry of the PRC2 complexes by quantitative proteomics and reveal the existence of an EZH1-SUZ12 sub-complex lacking EED. We provide evidence that EZH1 together with SUZ12 form a non-canonical PRC2 complex, occupy active chromatin domains in the absence of H3K27me3, and positively regulate gene expression. Loss of EZH2 expression leads to global repositioning of EZH1 chromatin occupancy to EZH2 targets. Moreover, we demonstrate that an erythroid-specific enhancer mediates transcriptional activation of EZH1, and a switch from GATA2 to GATA1 controls the developmental EZH1/2 switch by differential association with EZH1 enhancers during erythropoiesis. Thus, the lineage- and developmental stage-specific regulation of PRC2 expression and subunit composition leads to a switch from canonical silencing to non-canonical PRC2 functions during blood stem cell specification. Overall design: Transcriptional profiling in primary human fetal liver proerythroblasts upon lentiviral shRNA-mediated knockdown of EZH1, EZH2, EED, or SUZ12 by RNA-seq analysis.","project":"SRP044056"}
{"number_samples":8,"species":"human","abstract":"We investigated the ALDH2*2 genetic polymorphism and its underlying mechanisms for the first time in a human model system of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) generated from individuals carrying the most common heterozygous form of the ALDH2*2 genotype. We showed that the ALDH2*2 mutation confers elevated levels of reactive oxygen species (ROS) and toxic aldehydes such as 4HNE, thereby inducing cell cycle arrest and activation of apoptotic signaling pathways, especially during ischemic injury. ALDH2 exerts control of cell survival decisions via modulation of oxidative stress levels. This regulatory circuitry was found to be dysfunctional in the loss-of-function ALDH2*2 genotype, causing upregulation of apoptosis in cardiomyocytes following ischemic insult. These results reveal a novel function of the metabolic enzyme ALDH2 in modulation of cell survival decisions. Overall design: Molecular mechanism of increased ischemic damage in cardiomyocytes of ALDH2*2 genotype.","project":"SRP044084"}
{"number_samples":3,"species":"human","abstract":"Interferon responses play key roles in cellular defense against pathogens. Highly expressed interferon-induced proteins with tetratricopeptide repeats (IFITs) are proposed to function as RNA binding proteins, but the RNA binding and discrimination specificities of IFIT proteins remain unclear. Here we show that human IFIT5 has comparable affinity for RNAs with diverse phosphatecontaining 5’-ends, excluding the higher eukaryotic mRNA cap.  Systematic mutagenesis revealed that sequence substitutions in IFIT5 can alternatively expand or introduce bias in protein binding to RNAs with 5' monophosphate, triphosphate, cap0 (triphosphate-bridged N7-methylguanosine) or cap1 (cap0 with RNA 2’-O-methylation). We defined the breadth of cellular ligands for IFIT5 by using a thermostable group II intron reverse transcriptase for RNA sequencing. We show that IFIT5 binds precursor and processed tRNAs as well as other RNA polymerase III transcripts.  Our findings establish the RNA recognition specificity of the human innate immune response protein IFIT5.","project":"SRP044113"}
{"number_samples":16,"species":"human","abstract":"Deregulated expression of MYC is a driver of colorectal carcinogenesis, necessitating novel strategies to inhibit MYC function. The ubiquitin ligase HUWE1 (HECTH9, ARFBP1,  MULE) associates with both MYC and the MYC-associated protein MIZ1. We show here that HUWE1 is required for growth of colorectal cancer cells in culture and in orthotopic xenograft models. Using high throughput screening, we identify small  molecule inhibitors of HUWE1, which inhibit MYC-dependent transactivation in colorectal cancer cells, but not in stem and normal colon epithelial cells. Inhibition of HUWE1 stabilizes MIZ1. MIZ1 globally accumulates on MYC target genes and contributes to repression of MYC-activated target genes upon HUWE1 inhibition. Our data show that transcriptional activation by MYC in colon cancer cells requires the continuous degradation of MIZ1 and identify a novel principle that allows for inhibition of MYC function in tumor cells. Overall design: MIZ1 and MYC ChIPseq experiments in HUWE1 inhibitor-treated Ls174T cells as well as RNAseq experiments in HUWE1- or MIZ1-depleted Ls174T cells after HUWE1 inhibitor treatment. Sequencing was performed on an Illumina Genome Analyzer IIx.","project":"SRP044171"}
{"number_samples":12,"species":"human","abstract":"Combining the knock-down of miR-15a, miR-16 and miR-92a in human and macaque cells transiently, we performed RNA-seq to quantify the regulatory effect of miRNAs on their 3’UTR and CDS targets","project":"SRP044174"}
{"number_samples":12,"species":"human","abstract":"MCF-7 cells were treated with either ZNA (30 uM) or a vehice control for 3 or 12 hours.  Following RNA sequencing, the control-normalized data was used to analyze genes altered by ZNA treatment. Following RNA sequencing, the control-normalized data was used to analyze genes altered by ZNA treatment. Overall design: MCF-7 cells were treated with either ZNA (30 uM) or a vehice control for 3 or 12 hours.","project":"SRP044187"}
{"number_samples":14,"species":"human","abstract":"In this study we aim to determine the role of IL-4/STAT6 in gene expression in human keratinocytes using RNA-sequencing approach. Human keratinocytes were cultured for 2 or 5 days with calcium chloride to induce terminal differentiation as determined by the expression of epidermal differentiation complex genes. The cells were then stimulated with IL-4 for 3 and 24 hours, or along the 5 days culture period.  We observed that IL-4 inhibits fully differentiation of keratinocytes, induces genes involved with production of inflammatory mediators, and reduces the healing capacity of human keratinocytes. Moreover, STAT6 controlled important genes involved with calcium binding, inflammation and epidermis development. Overall design: Human keratinocytes were differentiated with calcium chloride for 2 days and incubated with media alone or 20ng/ml of recombinant human IL-4 for 3 and 24 hours. Human keratinocytes were differentiated with calcium chloride for 5 days with or wihout recombinant human IL-4 (20ng/ml). Keratinocytes transfected with control or STAT6 siRNA were differentiated with calcium chloride for 2 days and then stimulated with recombinant huma IL-4 for 24 hours.","project":"SRP044206"}
{"number_samples":4,"species":"human","abstract":"MiRNAs have been identfied to play an important role in cancer stem cells. MiR23b is differentially expressed in various forms of cancer including colorectal cancer as compared to their normal counterparts. MiR23b regulates various aspects of cell behaviour such as differentiation, apoptosis and motility. The goal of the study was to identify the novel role of miR23b in self-renewal property of colon cancer stem cells via regulation of its candidate target mRNAs. To address this aim, HT29 colon cancer cells were transfected with miR23b Precursor, Antimir and their respective controls. RNA SEQ analysis of the cells with altered levels of miR23b assisted in the identification of interesting mRNA targets influenced by miR23b expression and involved self-renewal pathways. Overall design: HT29 cells with altered levels of miR23b was subjected to RNA SEQ analysis to identify differential expresssion of mRNA targets of mIR23b","project":"SRP044229"}
{"number_samples":4,"species":"human","abstract":"Background: E-cadherin is an adherens junction protein that forms homophilic intercellular contacts in epithelial cells while also interacting with the intracellular cytoskeletal networks. It has roles including establishment and maintenance of cell polarity, differentiation, migration and signalling in cell proliferation pathways. Its downregulation is commonly observed in epithelial tumours and is a hallmark of the epithelial to mesenchymal transition (EMT). Methods: To improve our understanding of how E-cadherin loss contributes to tumorigenicity, we investigated the impact of its elimination from the non-tumorigenic breast cell line MCF10A. We performed cell-based assays and whole genome RNAseq to characterize an isogenic MCF10A cell line that is devoid of CDH1 expression due to an engineered homozygous 4bp deletion in CDH1 exon 11. Results: The E-cadherin-deficient line, MCF10A CDH1-/- showed subtle morphological changes, weaker cell-substrate adhesion, delayed migration, but retained cell-cell contact, contact growth inhibition and anchorage-dependent growth. Within the cytoskeleton, the apical microtubule network in the CDH1-deficient cells lacked the radial pattern of organization present in the MCF10A cells and F-actin formed thicker, more numerous stress fibres in the basal part of the cell. Whole genome RNAseq identified compensatory changes in the genes involved in cell-cell adhesion while genes involved in cell-substrate adhesion, notably ITGA1, COL8A1, COL4A2 and COL12A1, were significantly downregulated. Key EMT markers including CDH2, FN1, VIM and VTN were not upregulated although increased expression of proteolytic matrix metalloprotease and kallikrein genes was observed. Conclusions: Overall, our results demonstrated that E-cadherin loss alone was insufficient to induce an EMT or enhance transforming potential in the non-tumorigenic MCF10A cells but was associated with broad transcriptional changes associated with tissue remodelling. Overall design: Examination of the impact of E-cadherin (CDH1) loss in an isogenic pair of breast cell lines.","project":"SRP044241"}
{"number_samples":6,"species":"human","abstract":"Alternative splicing—the production of multiple mRNA isoforms from a single gene—is regulated in part by RNA-binding proteins (RBPs). While the RBPs Tra2a and Tra2ß have both been implicated in the regulation of alternative splicing, their relative contribution to this process are not well understood. Here we use iCLIP to identify Tra2ß target exons in MDA-MB-231 cells. We find that simultaneous—but not individual—depletion of Tra2a and Tra2ß induces substantial shifts in the splicing pattern of endogenous Tra2ß target exons identified by iCLIP. We next use RNA-seq following joint Tra2 protein depletion to comprehensively identify Tra2 protein-dependent exons in MDA-MB-231 cells. Overall design: Endogenous Tra2ß binding sites were mapped across the MDA-MB-231 cell transcriptome in biological triplicate iCLIP experiments. RNA-seq was performed using three biological replicates of negative control siRNA treated MDA-MB-231 cells and three biological replicates of TRA2A and TRA2B siRNA treated MDA-MB-231 cells.","project":"SRP044265"}
{"number_samples":14,"species":"human","abstract":"Dendritic-cell (DC) maturation involves substantial remodeling of their gene-expression program. Most research has focused on inducible gene-expression networks promoting the acquisition of new functions, such as cytokine production and enhanced T-cell-stimulatory capacity. In contrast, mechanisms that modulate DC-function by inducing gene silencing remain poorly understood. Here we describe a novel primary epigenetic-silencing response that makes major contributions to the DC-maturation process. The repressed genes function in pivotal processes - including antigen-presentation, extracellular-signal detection, signal-transduction and lipid-mediator biosynthesis - underscoring the central contribution of the silencing mechanism to rapid reshaping of DC-function. Interestingly, promoters of the repressed genes exhibit a surprisingly high frequency of PU.1-occupied sites, suggesting a novel role for this transcription factor in marking genes poised for inducible repression Overall design: Transcriptional changes induced by one hour LPS treatment in monocyte derived dendritic cells.","project":"SRP044286"}
{"number_samples":8,"species":"human","abstract":"We generated HeLa cell lines stably expressing either empty vector or MYC-UPF1-FLAG TEV and transfected each with either empty vector or TEV-protease encoding vector. The goal is to see what mRNAs are upregulated upon UPF1 cleavage.","project":"SRP044296"}
{"number_samples":10,"species":"human","abstract":"UBL5 is an atypical ubiquitin-like protein, whose function in metazoans remains largely unexplored. We show that UBL5 is required for sister chromatid cohesion maintenance in human cells. UBL5 primarily associates with spliceosomal proteins, and UBL5 depletion decreases pre-mRNA splicing efficiency, leading to globally enhanced intron retention. Defective sister chromatid cohesion is a general consequence of dysfunctional pre-mRNA splicing, resulting from the selective downregulation of the cohesion protection factor Sororin. As the UBL5 yeast orthologue, Hub1, also promotes spliceosome functions, our results show that UBL5 plays an evolutionary conserved role in pre-mRNA splicing, the integrity of which is essential for the fidelity of chromosome segregation. Overall design: Total RNA was extracted from HeLa cells treated with control (CTRL), UBL5 (#57, #58, or #82), or SART1 siRNAs for 48 h and processed for RNA-Seq analysis","project":"SRP044298"}
{"number_samples":4,"species":"human","abstract":"LPMC MNP subsets","project":"SRP044303"}
{"number_samples":10,"species":"human","abstract":"Transcriptional profiling of human quasi-haploid and haploid cells.","project":"SRP044391"}
{"number_samples":4,"species":"human","abstract":"Using neutrophils from a cohort of normal individuals, we generated transcriptomic profile of 4 individuals. Overall design: Here we generated gene expression profile of neutrophil cells of 4 normal individuals.This data was integreated with DNA methylation profiles of same individuals.","project":"SRP044593"}
{"number_samples":12,"species":"human","abstract":"The interplay between mitogenic and proinflammatory signaling pathways play key roles in determining the phenotypes and clinical outcomes of breast cancers.  We have used global nuclear run-on coupled with deep sequencing to characterize the immediate transcriptional responses of MCF-7 breast cancer cells treated with estradiol, TNFa, or both.  In addition, we have integrated these data with chromatin immunoprecipitation coupled with deep sequencing for estrogen receptor alpha (ERa), the pioneer factor FoxA1 and the p65 subunit of the NF-?B transcription factor. Our results indicate extensive transcriptional interplay between these two signaling pathways, which is observed for a number of classical mitogenic and proinflammatory protein-coding genes.  In addition, GRO-seq has allowed us to capture the transcriptional crosstalk at the genomic locations encoding for long non-coding RNAs, a poorly characterized class of RNAs which have been shown to play important roles in cancer outcomes.  The synergistic and antagonistic interplay between estrogen and TNFa signaling at the gene level is also evident in the patterns of ERa and NF-?B binding, which relocalize to new binding sites that are not occupied by either treatment alone.  Interestingly, the chromatin accessibility of classical ERa binding sites is predetermined prior to estrogen treatment, whereas ERa binding sites gained upon co-treatment with TNFa require NF-?B and FoxA1 to promote chromatin accessibility de novo.  Our data suggest that TNFa signaling recruits FoxA1 and NF-?B to latent ERa enhancer locations and directly impact ERa enhancer accessibility. Binding of ERa to latent enhancers upon co-treatment, results in increased enhancer transcription, target gene expression and altered cellular response.  This provides a mechanistic framework for understanding the molecular basis for integration of mitogenic and proinflammatory signaling in breast cancer. Overall design: Using GRO-seq and ChIP-seq (ER, FoxA1 and p65) to assay the molecular crosstalk of MCF-7 cells treated with E2, TNFa or both E2+TNFa.","project":"SRP044608"}
{"number_samples":6,"species":"human","abstract":"The emergence of anti-estrogen resistance in breast cancer is an important clinical phenomenon affecting long-term survival in this disease. Identifying factors that convey cell survival in this setting may guide improvements in treatment. Estrogen (E2) can induce apoptosis in breast cancer cells that have been selected for survival after E2 deprivation for long periods (MCF-7:5C cells), but the mechanisms underlying E2-induced stress in this setting have not been elucidated. Here, we report that the c-Src kinase functions as a key adapter protein for the estrogen receptor (ER, ESR1) in its activation of stress responses induced by E2 in MCF-7:5C cells. E2 elevated phosphorylation of c-Src, which was blocked by 4-hydroxytamoxifen (4-OHT), suggesting that E2 activated c-Src through the ER. We found that E2 activated the sensors of the unfolded protein response (UPR), IRE1a (ERN1) and PERK kinase (EIF2AK3), the latter of which phosphorylates eukaryotic translation initiation factor-2a (eIF2a). E2 also dramatically increased reactive oxygen species production and upregulated expression of heme oxygenase HO-1 (HMOX1), an indicator of oxidative stress, along with the central energy sensor kinase AMPK (PRKAA2). Pharmacologic or RNA interference-mediated inhibition of c-Src abolished the phosphorylation of eIF2a and AMPK, blocked E2-induced ROS production, and inhibited E2-induced apoptosis. Together, our results establish that c-Src kinase mediates stresses generated by E2 in long-term E2-deprived cells that trigger apoptosis. This work offers a mechanistic rationale for a new approach in the treatment of endocrine-resistant breast cancer. Overall design: MCF-7:5C cells were treated with vehicle (0.1% EtOH) as control, E2 (10-9mol/L), 4-OHT (10-6mol/L), E2 (10-9mol/L) plus 4-OHT (10-6mol/L), PP2 (5x10-6mol/L), and E2 (10-9mol/L) plus PP2 (5x10-6mol/L) respectively for 72 hours.","project":"SRP044610"}
{"number_samples":3,"species":"human","abstract":"A c-Src inhibitor blocks estrogen (E2)-induced stress and converts E2 responses from inducing apoptosis to growth stimulation in E2-deprived breast cancer cells. A reprogrammed cell line, MCF-7:PF, results with features of functional estrogen receptor (ER) and over-expression of insulin-like growth factor-1 receptor beta (IGF-1Rß). We addressed the question of whether the selective ER modulator 4-hydroxytamoxifen (4-OHT) could target ER to prevent E2-stimulated growth in MCF-7:PF cells. Selected expression of mRNA was measured through real-time RT-PCR. Global gene expression was analyzed by microarray and RNA-seq analysis. Unexpectedly, both 4-OHT and E2 stimulated cell growth in a concentration-dependent manner. Global gene expression analysis showed a remarkable overlap in genes regulated in the same direction by E2 and 4-OHT. Pathway enrichment analysis of the 280 genes commonly deregulated by 4-OHT and E2 revealed functions mainly related to membrane, cytoplasm, and metabolic processes. Further analysis of 98 up-regulated genes by both 4-OHT and E2 uncovered a significant enrichment in genes associated with membrane remodeling, cytoskeleton reorganization, cytoplasmic adapter proteins, cytoplasm organelles proteins, and related processes. 4-OHT was more potent than E2 to up-regulate some membrane remodeling molecules, such as EHD2, FHL2, HOMER3 and RHOF. In contrast, 4-OHT acted as an antagonist to inhibit expression of the majority of enriched membrane-associated genes in wild-type MCF-7 cells. Long-term selection pressure has changed the cell population responses to 4-OHT. Membrane-associated signaling is critical for 4-OHT-stimulated cell growth in MCF-7:PF cells. This study provides a rationale for the further investigation of targeted therapy for tamoxifen resistant patients. Overall design: Wild-type MCF-7 cells were treated with vehicle control (0.1% ethanol), E2 (10-9 mol/L) and 4-OHT (10-6 mol/L) respectively for 24 hours.","project":"SRP044611"}
{"number_samples":4,"species":"human","abstract":"The RNA biding protein, LARP1, has been proposed to function downstream of mTORC1 to positively regulate the translation of 5’TOP mRNAs such as ribosome protein (RP) mRNAs. However, its regulatory roles in mTORC1-mediated translation remain unclear. PAR-CLIP of LARP1 revealed its direct and dynamic interactions with RP mRNAs through pyrimidine-enriched sequences in the 5’UTR of RP mRNAs when mTOR activity is inhibited. Importantly, this LARP1 is a direct substrate of mTORC1 and S6K1/Akt, and phosphorylated LARP1 scaffolds mTORC1 on translation-competent mRNAs to facilitate 4EBP1 and S6K1 phosphorylation. Ablation of LARP1 causes multiple defects in the processes of translation including abnormal eIF4G1 interaction with RP mRNAs and inefficient RP mRNA elongation thereby reducing ribosome biogenesis and cell proliferation. These observations illustrate that LARP1 functions both an effector and a regulator for local mTORC1 activity, and acts as a molecular switch for ribosome biogenesis by sensing growth factor/nutrient signaling. Overall design: LARP1-bound RNA regions were sequenced from HEK293T cells under growing or mTOR-inactive conditions. In parallel, mRNA abundance was quantified, in biological duplicate, from HEK293T cells under the same conditions. Sequencing reads were mapped to human transcriptome reference from hg19 to determine LARP1 binding sites and mRNA abundances.","project":"SRP044653"}
{"number_samples":3,"species":"human","abstract":"NPC lncRNAs and mRNAs Transcriptome or Gene expression related to radioresistance and chemoresistance","project":"SRP044661"}
{"number_samples":94,"species":"human","abstract":"We obtained radiographically-localized biopsies during glioma resection surgeries to sample the tumor core and margins from multiple glioma patients. We also procured fresh, non-neoplastic brain tissue specimens from multiple patients having procedures to relieve epilespy symptoms or to place shunts to treat normal pressure hydrocephalus.  We then used RNA-Seq to compare expression patterns between geographically distinct regions of gliomas and computational deconvolution to estimate cell type-specific expression patterns in different disease subtypes. Overall design: RNA-Seq analysis in 39 contrast-enhancing glioma core samples, 36 non-enhancing FLAIR glioma margin samples, and 17 non-neoplastic brain tissue samples.","project":"SRP044668"}
{"number_samples":13,"species":"human","abstract":"In this study we report the discovery of an unexpected nuclear activity of small RNAs that target an ~8.6 megabase chromosomal domain located on the telomeric q-arm of chromosome 5 from an unbiased genome-wide RNAi screen. Short-hairpin RNAs that target genes within this large domain induce trans-activation of CREB signaling transcriptional reporters and induce an endogenous gene expression signature that includes CREB-target and neuronal-fate genes. Furthermore, CRISPR-guided deletion of AGO-1 and -2 demonstrate their mutually redundant roles in this mechanism, which we named Chromosomally Induced Trans Activation by RNAi (CITAR). Overall design: We sequenced the transcriptome of HEK293 cells undergoing Chromosomally-Induced Trans Activation by RNAi (CITAR), as measured by transcriptional activation of a CRE-Luciferase reporter (pGL4.29), by shRNAs targeting Chr5q35 loci (TSPAN17 at Chr5:176,074,388, TMED9 at  Chr5:177,019,213 and N4BP3 at Chr5:177,540,556), as well as two negative control shRNAs to account for infection (non-hairpin) and for engagement of the RNAi pathway (ASNA1 encoded on Chr19).","project":"SRP044673"}
{"number_samples":16,"species":"human","abstract":"Uridylation occurs pervasively on mRNAs in mammals, yet its mechanism and significance remain unknown. Here we identify TUT4 and TUT7 (also known as ZCCHC11 and ZCCHC6, respectively) as the enzymes that uridylate mRNAs. Uridylation readily occurs on deadenylated mRNAs that are not associated with poly(A) binding protein (PABPC1) in cells. Consistently, purified TUT4 and TUT7 (TUT4/7) selectively uridylate RNAs with short A tails (< ~25 nt) while PABPC1 antagonizes uridylation of polyadenylated mRNAs in vitro. In cells depleted of TUT4/7, the vast majority of mRNAs lose the U tails, and their half-lives are extended. Suppression of mRNA decay factors leads to the accumulation of uridylated mRNAs. In line with this, microRNA induces uridylation of its targets, and TUT4/7 is required for enhanced decay of microRNA targets. Our study explains the mechanism underlying selective uridylation of deadenylated mRNAs, and demonstrates a fundamental role of the U tail as a molecular mark for global mRNA decay. Overall design: HeLa cells were knocked down of control or TUT4/7, then total RNAs were prepared for RNA-seq on 0, 1, 2, 4h after actinomycin D treatment. The whole processes of experiments were repeated two times.","project":"SRP044679"}
{"number_samples":22,"species":"human","abstract":"Anaplastic Large Cell Lyomphoma","project":"SRP044708"}
{"number_samples":4,"species":"human","abstract":"A multi-omic integration of multiple high throughput technologies applied to a single, purified, primary cell population -- Naive CD4+ T cells.","project":"SRP044715"}
{"number_samples":6,"species":"human","abstract":"We performed RNA-Seq on three different induced pluripotent stem cell (iPSC)-derived neurons carrying copy number variants of chromosome 15q11-q13.1. Strand-specific sequencing of RNA of poly(A) selected RNA was performed using the Illumina platform.","project":"SRP044749"}
{"number_samples":12,"species":"human","abstract":"Here we have characterized the transcriptional processes underlying the formation of human brown in white (i.e. brite) adipocytes using a genome-wide approach. We show that the browning process is associated with reprogramming of peroxisome proliferator-activated receptor ? (PPAR?) binding to form brite adipocyte-selective PPAR? super-enhancers that appear to play a key role in activation of brite adipocyte-selective genes. We identify the KLF11 gene based on its association with a PPAR? super-enhancer and show that KLF11 is a novel browning factor directly induced by rosiglitazone and required for the activation of brite adipocyte-selective gene program by rosiglitazone. Overall design: Genome-wide profiling of Dnase I hypersenstive (DHS) sites, epigenomic marks, transcription factor and co-factor binding, and gene expression in hMADS white and brite adipocytes","project":"SRP044756"}
{"number_samples":11,"species":"human","abstract":"Oncogenic transformation by adenovirus small e1a depends on simultaneous interactions with the host lysine acetylases p300/CBP and the tumor suppressor RB. How these interactions influence cellular gene expression remains unclear. We find that e1a displaces RBs from E2F transcription factors and promotes p300 acetylation of RB1 K873/K874 to lock it into a repressing conformation that interacts with repressive chromatin-modifying enzymes. These repressing p300-e1a-RB1 complexes specifically interact with host genes that have unusually high p300 association within the gene body. The TGF?-, TNF-, and interleukin-signaling pathway components are enriched among such p300-targeted genes. The p300-e1a-RB1 complex condenses chromatin in a manner dependent on HDAC activity, p300 lysine acetylase activity, the p300 bromodomain, and RB K873/K874 and e1a K239 acetylation to repress host genes that would otherwise inhibit productive virus infection. Thus, adenovirus employs e1a to repress host genes that interfere with viral replication. Overall design: Examination of transcriptome by mRNA sequencing before and after infection by adenoviral e1a expressing vectors in growth arrested IMR90","project":"SRP044763"}
{"number_samples":28,"species":"human","abstract":"Primary human foreskin fibroblasts (HFF) were infected with wild-type simplex virus 1 (HSV-1) strain 17 at a multiplicity of infection (MOI) of 10. Newly transcribed RNA was labelled by adding 500µM 4-thiouridine (4sU) to the cell culture media for 1h. Total cellular RNA was isolated using Trizol. Newly transcribed RNA was purified following the protocol described in Raedle et al. JoVE 2013. Overall design: Newly transcribed RNA was labelled in one hour intervals during the first eight hours of HSV-1 infection. All nine 4sU-RNA samples as well as total cellular RNA of every second hour of infection were sent for sequencing. Two independent biological replicates were analysed.","project":"SRP044766"}
{"number_samples":3,"species":"human","abstract":"Compare the response of human (HEK293T) and bat (PakiT03) kidney cells infected with Hendra virus using proteomics informed by transcriptomics","project":"SRP044809"}
{"number_samples":24,"species":"human","abstract":"EGFR and MEK pathways were activated alone or in combination in human mammary epithelial cells. We profiled the pathway gene expression signatures using RNA-Seq. Overall design: mRNA was extracted from human mammary epithelial cells overexpressing EGFR gene, MEK gene, or EGFR and MEK genes in combination (or GFP control) for RNA-Seq analysis. Experiment was performed in six replicates per condition.","project":"SRP044854"}
{"number_samples":8,"species":"human","abstract":"The epithelium lining the epididymis in the male reproductive tract maintains a luminal environment that promotes sperm cell maturation. This process is dependent on the coordinated expression of many genes that encode proteins with a role in epithelial transport. We previously generated genome-wide maps of open chromatin in primary human fetal epididymis epithelial cells to identify potential regulatory elements controlling coordinated gene expression in the epididymis epithelium. Subsequent in silico analysis identified transcription factor binding sites (TFBS) that were over-represented in the HEE open chromatin, include the motif for paired box 2 (PAX2). PAX2 is a critical transcriptional regulator of urogenital tract development, which is well studied in the kidney but is unexplored in the epididymis. Due to the limited lifespan of primary HEE cells in culture we investigated the role of PAX2 in an immortalized HEE cell line (REP). First, REP cells were evaluated by DNase-seq and their open chromatin map overlapped that of primary HEE cells at ~ 65% of sites. Moreover, the PAX2-binding motif was again identified as an overrepresented TFBS within intergenic open chromatin, though on fewer chromosomes than in the primary HEE cells. To identify PAX2-target genes in REP cells, RNA-seq analysis was performed after siRNA-mediated depletion of PAX2 in comparison to a non-targeting siRNA. In response to PAX2-represssion, 3142 transcripts were differentially expressed (1334 up-regulated and 1808 down-regulated). Novel PAX2 targets included multiple genes encoding proteins with a predicted function in the epididymis epithelium. Overall design: mRNA profile of control and PAX2 knockdown REP cells","project":"SRP044867"}
{"number_samples":27,"species":"human","abstract":"Next generation sequencing was used to identify Notch mutations in a large collection of diverse solid tumors. NOTCH1 and NOTCH2 rearrangements leading to constitutive receptor activation were confined to triple negative breast cancers (TNBC, 6 of 66 tumors). TNBC cell lines with NOTCH1 rearrangements associated with high levels of activated NOTCH1 (N1-ICD) were sensitive to the gamma-secretase inhibitor (GSI) MRK-003, both alone and in combination with pacitaxel, in vitro and in vivo, whereas cell lines with NOTCH2 rearrangements were resistant to GSI. Immunohistochemical staining of N1-ICD in TNBC xenografts correlated with responsiveness, and expression levels of the direct Notch target gene HES4 correlated with outcome in TNBC patients. Activating NOTCH1 point mutations were also identified in other solid tumors, including adenoid cystic carcinoma (ACC). Notably, ACC primary tumor xenografts with activating NOTCH1 mutations and high N1-ICD levels were sensitive to GSI, whereas N1-ICD low tumors without NOTCH1 mutations were resistant. Overall design: Gene expression profiling for Notch-sensitive cancer cell lines using RNA-seq, each sample with triplicates","project":"SRP044917"}
{"number_samples":8,"species":"human","abstract":"Tightly controlled gene expression orchestrated by the transcription factor p63 during epithelial differentiation is important for development of epithelial-related structures such as epidermis, limb and craniofacial regions. How p63 regulates spatial and temporal expression of its target genes during these developmental processes is however not yet clear. By epigenomics profiling in stem cells established from one of these epithelial structures, the epidermis, we provide a global map of p63-bound regulatory elements that are categorized as single enhancers and clustered enhancers during epidermal differentiation. Transcriptomics analysis shows dynamic gene expression patterns during epidermal differentiation that correlates with the activity of p63-bound enhancers rather than with p63 binding itself. Only a subset of p63-bound enhancers is active in epidermal stem cells, and inactive p63-bound enhancers appear to function in gene regulation during the development of other epithelial tissues. Our data suggest a paradigm that p63 bookmarks genomic loci during the commitment of the epithelial lineage and regulates gene expression in different epithelial tissues through tissue-specific active enhancers. The catalogue of differentially expressed epidermal genes including non-coding RNAs and epithelial enhancers reported here provides a rich resource for studies of epithelial development and related diseases. Overall design: Comparison of gene expression at different stages of keratinocyte differentiation","project":"SRP044925"}
{"number_samples":9,"species":"human","abstract":"Background: To understand the transcriptional consequences of a TP53 modulation in the GIST cell context, we treated GIST430 with increasing doses of the MDM2-inhibitor nutlin-3 (racemate). Overall design: In this study, 3’ end RNA Sequencing (3SEQ) was used to expression profile GIST430 cell lines treated with DMSO and Nutlin-3, 2.5µM or 10µM for 18h. Then the gene expression profiles between these concentrations were compared.","project":"SRP044956"}
{"number_samples":12,"species":"human","abstract":"The experiment was designed to display differential gene expression profiling in three human intrahepatic cholangiocarcinoma (ICC) cells upon knockdow of LKB1 tumor suppressor, by using RNAseq technology. Overall design: LKB1 was first attenuated in three ICC cells: HuH-28, RBE, and SSP-25 by siRNA-mediated knockdown. Total RNA was extracted from duplicated cancers cells transfected with control siRNA and LKB1 specific siRNA at 48h posttransfection for RNAseq analysis. Differentially expressed genes in LKB1-attenuated ICC cells were identified in comparison to that in ICC cells transfected with control siRNA.","project":"SRP045048"}
{"number_samples":12,"species":"human","abstract":"Memory T cells are primed for rapid responses to antigen; however, the molecular mechanisms responsible for priming remain incompletely defined. CpG methylation in promoters is an epigenetic modification, which regulates gene transcription. Using targeted bisulfite sequencing, we examined methylation of 2100 genes (56,000 CpG) mapped by deep sequencing to T cell activation in human naïve and memory CD4 T cells. 466 CpGs (132 genes) displayed differential methylation between naïve and memory cells. 21 genes exhibited both differential methylation and gene expression before activation, linking promoter DNA methylation states to gene regulation; 6 genes encode proteins closely studied in T cells while 15 genes represent novel targets for further study. 39 genes exhibited reduced methylation in memory cells coupled with increased gene expression with activation compared to naïve cells, revealing specific genes more rapidly expressed in memory compared to naïve cells and potentially regulated by DNA methylation. These findings define a DNA methylation signature unique to memory CD4 T cells and correlated with activation-induced gene expression. Overall design: RNA sequencing of primary human naïve and memory CD4 T cells at rest and 48 hours post-activation.","project":"SRP045052"}
{"number_samples":8,"species":"human","abstract":"RNA-seq analysis of human 293 Tet-off cells depleted of PTBP1 and UPF1 alone and in tandem with specific siRNAs. Overall design: siRNA-based depletion of PTBP1, UPF1, and PTBP1/UPF1 together, with a validated non-silencing siRNA as a control.","project":"SRP045065"}
{"number_samples":16,"species":"human","abstract":"Single-end 36bp RNAseq in control and NKX2-1 knockdown samples to uncover genes regulated by NKX2-1. Single-end 36bp ChIPseq using NKX2-1 antibody (Santa Cruz, sc-13040) to indentify direct NKX2-1 binding sites across the NSCLC genome.","project":"SRP045118"}
{"number_samples":4,"species":"human","abstract":"RNA-seq Identification of a Novel Fusion Gene in a Mesenchymal Tumor","project":"SRP045126"}
{"number_samples":6,"species":"human","abstract":"Primary cells enter replicative senescence after a limited number of cell divisions. This process is associated with reproducible changes in DNA methylation (DNAm) at specific sites in the genome. The mechanism that drives senescence-associated DNAm changes remains unknown and may arise through drift in DNAm or through regulated, senescence dependent modifications at specific sites in the genome. In this study, we analyzed the reorganization of nuclear architecture and DNA methylation during long-term culture of human fibroblasts and mesenchymal stromal cells (MSCs). [RNA-seq] Overall design: RNA was isolated from 1,000,000 cells of three MSC donors (59, 64, and 73 years old) at passage 4 and passage 13 using the miRNeasy Mini Kit (Qiagen). Gene expression profiles were analzyed by  deep sequencing with IlluminaHiSeq 2000 technology with a read length of 50 bases at EMBL gene core facility (Heidelberg, Germany).","project":"SRP045154"}
{"number_samples":6,"species":"human","abstract":"Artificial transcription factors (ATFs) and genomic nucleases based on a DNA binding platform consist- ing of multiple zinc finger domains are currently be- ing developed for clinical applications. However, no genome-wide investigations into their binding speci- ficity have been performed. We have created six- finger ATFs to target two different 18 nt regions of the human SOX2 promoter; each ATF is constructed such that it contains or lacks a super KRAB do- main (SKD) that interacts with a complex contain- ing repressive histone methyltransferases. ChIP-seq analysis of the effector-free ATFs in MCF7 breast cancer cells identified thousands of binding sites, mostly in promoter regions; the addition of an SKD domain increased the number of binding sites ~5- fold, with a majority of the new sites located out- side of promoters. De novo motif analyses suggest that the lack of binding specificity is due to sub- sets of the finger domains being used for genomic interactions. Although the ATFs display widespread binding, few genes showed expression differences; genes repressed by the ATF-SKD have stronger bind- ing sites and are more enriched for a 12 nt motif. Interestingly, epigenetic analyses indicate that the transcriptional repression caused by the ATF-SKD is not due to changes in active histone modifications. Overall design: ATF plasmids were designed and stably integrated into MCF7 cells as described in (Stolzenburg et al. 2012).  Stable lines were grown at 30–80% confluency in Dulbecco’s Modified Eagle’s Medium (Corning, Corning, NY) supplemented with 10% heat-inactivated fetal bovine serum (Invitrogen, Life Technologies, Grand Island, NY) and 1% penicillin/streptomycin; cells were selected using 5 µg/ml puromycin (VWR, Radnor, PA) and 200 µg/ml G418 (VWR, Radnor, PA). ATF expression was induced by treatment with media containing 1µg/ml doxycycline (VWR, Radnor, PA) at 0 h, doxycycline media was refreshed at 48 h, and cells were harvested at 72 h. ATF expression was confirmed by hemagglutinin (HA) tag western blot prior to HA ChIP-seq, histone ChIP-seq and RNA-seq analysis.  For HA-tag ChIP-seq, stable MCF7 cell lines were induced using 100 ng/ml doxycycline (Sigma) at 0 h, doxycycline media was refreshed at 48 h, and cells were harvested at 72 h by crosslinking in a final concentration of 1% formaldehyde. Crosslinking was stopped after 5 min by adding glycine to a final concentration of 125 mM. Crosslinked cells were washed in cold phosphate buffered saline, lysed using 1 mL low-salt IP buffer (150 mM NaCl, 50 mM Tris-HCl (pH7.5), 5 mM EDTA, NP-40 (0.5%), Triton X-100 (1%) containing protease inhibitors) and aliquoted at 1 × 10?7 cells/mL. Cells are then sonicated to a fragment size range of 500–800 bp. Samples were then diluted in 1 mL low-salt buffer and incubated with 3 µL of anti-HA antibody (Covance, Princeton, NJ). Three-hundred microliter Sepharose A beads (GE Healthcare Life Science) were used for pull-down. Samples were sequenced at the UNC-CH Genome Analysis Facil- ity (Chapel Hill, NC) on an HiSeq (Illumina, San Diego, CA) to read counts of 4.1–67.3 M total reads. For histone ChIP-seq, antibodies to H3K4me3 (Cell Signaling Tech- nologies CST9751), H3K9Ac (Cell Signaling Technologies CST9649) and H3K9me3 (Diagenode pAb-056-050) were used; samples were prepared as previously described (O’Geen et al. 2011).","project":"SRP045155"}
{"number_samples":4,"species":"human","abstract":"In order to explore the status of DNA methylation in hypoxia response, we show that TET1, a DNA dioxygenase converting 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC), regulates hypoxia-responsive gene expression. Hypoxia/HIF-2a regulates the expression of TET1. Knockdown of TET1 mitigated hypoxia-induced EMT. RNA sequencing and 5hmC sequencing identified the set of TET1-regulated genes. Overall design: Four samples (Four samples, Hypoxia (scrambled control), Hypoxia (TET1-si), Normoxia (scrambled control) and Normoxia (TET1-si), are performed by RNA-Seq and hMeDIP-Seq  RNA-Seq and hMeDIP-Seq","project":"SRP045156"}
{"number_samples":4,"species":"human","abstract":"HITS-CLIP of control and transfected cells to find direct targetting of miR-200 family to mRNA","project":"SRP045204"}
{"number_samples":11,"species":"human","abstract":"The goal of this study is to identify deferentially expressed genes among three groups of individuals of the same family. These groups are : affected, unaffected wild, unaffected carrier.","project":"SRP045207"}
{"number_samples":36,"species":"human","abstract":"Primary human foreskin fibroblasts (HFF) were infected with wild-type simplex virus 1 (HSV-1) strain 17 at a multiplicity of infection (MOI) of 10. Ribosome profiling was performed at various times during infection with minor modification to the protocol described in Stern-Ginossar N et al., Science 2012 Overall design: Ribosome profiling was performed a 0, 1, 2, 4, 6 and 8 h post infection. Two biological replicates were analysed.","project":"SRP045214"}
{"number_samples":28,"species":"human","abstract":"Pseudouridine is the most abundant modification occurring on RNA, yet with the exception of a few well-studied RNA molecules little is known about the modified positions and their function(s). Here, we develop ?-seq, a method for transcriptome-wide quantitative mapping of pseudouridine. We validate ?-seq with synthetic spike-ins and de novo identification of the vast majority of previously reported pseudouridylated positions. ?-seq permits discovery of hundreds of novel pseudouridine modifications in human and yeast mRNAs and snoRNAs. Knockdown and knockout of pseudouridine synthases uncovers the cognate PUSs mediating pseudouridine catalysis at these individual novel sites and their target sequence features. In both human and yeast pseudouridine formation on mRNA depends on both site-specific PUSs – often guided by a specific sequence motif - and snoRNA-guided PUSs. Importantly, upon heat shock in yeast, Pus7-mediated pseudouridylation is induced at >200 sites in diverse mRNAs. Pus7 deletion in yeast leads to decreased recovery from heat shock and decreased RNA levels at otherwise pseudouridylated messages, suggesting a role for pseudouridine in enhancing transcript stability. Pseudouridine stoichiometries in rRNA are highly conserved from yeast to mammals, but are reduced in cells derived from dyskeratosis congenita patients, where the pseudouridine synthase DKC1 is mutated, compared to age matched controls. Our results establish pseudouridine as a ubiquitous and dynamic modification in mRNA, and provide a sensitive, quantitative and transcriptome-wide methodology to address its underlying mechanisms and function. Overall design: Examination of m6A methylation in human Hek293 and A549 cell lines, in human embryonic stem cells (ESCs) undergoing differentiation to neural progenitor cells (NPCs),  in OKMS inducible fibroblasts reprogrammed into iPSC, and upon knockdown of factors using siRNAs or shRNAs.","project":"SRP045222"}
{"number_samples":8,"species":"human","abstract":"Analysis of lin-CD34+CD45+ (iCD34+) cell population from two normal bone marrow-derived (BM1K and BM9) iPSCs  and two CML (CML15 and CML17) iPSCs . CML iCD34+ cells have characteristics similar to primary CML leukemia stem cell in patients. Results provide insight into molecular profile characterized CML iCD34 and mechanism of its maintenance and drug resistance. Overall design: iCD34+ cell samples obtained from two control BM1K and BM9 iPSCs (both for the same normal donor) and CML15 and CML17 iPSCs (both from the same patient in chronic phase of CML). Each group was treated with DMSO (control) or 5 µM imatinib. The complete phenotype for iCD34+ cells: lin-CD34+CD45+CD90+CD117+CD45RA-. This population also inclyde Rhodaminelow and ALDKhigh cells.","project":"SRP045234"}
{"number_samples":4,"species":"human","abstract":"Ribosome profiling suggests that ribosomes occupy many regions of the transcriptome thought to be non-coding, including 5' UTRs and lncRNAs. Apparent ribosome footprints outside of protein-coding regions raise the possibility of artifacts unrelated to translation, particularly when they occupy multiple, overlapping open reading frames (ORFs). Here we show hallmarks of translation in these footprints: co-purification with the large ribosomal subunit, response to drugs targeting elongation, trinucleotide periodicity, and initiation at early AUGs. We develop a metric for distinguishing between 80S footprints and nonribosomal sources using footprint size distributions, which validates the vast majority of footprints outside of coding regions. We present evidence for polypeptide production beyond annotated genes, including induction of immune responses following human cytomegalovirus (HCMV) infection. Translation is pervasive on cytosolic transcripts outside of conserved reading frames, and direct detection of this expanded universe of translated products enables efforts to understand how cells manage and exploit its consequences. Overall design: Ribosome profiling to verify that true ribosome footprints shift in response to different elongation inhibitors (CHX vs Emetine) and co-purify with an affinity-tagged large ribosomal subunit (bound vs input)","project":"SRP045257"}
{"number_samples":2,"species":"human","abstract":"We used HCT116 colorectal cancer cells with and without mutations in DNA methyltransferases (resulting in a 95% reduction in global DNA methylation levels) to study the relationship between DNA methylation, histone modifications, and gene expression. (The double knockout cell line is called DKO1) Overall design: Examination of DNA methylation, two histone modifications, RNA polymerase II ChIP-seq and RNA expression in two cell line (HCT116 and DKO1). One of relipates of HCT116 RNA P II ChIP-seq is from GSM970210. Histone modification data(H3K27ac, H3K4me3) of HCT116 are from GSM945304, GSE31755. Two replicates of RNA-seq data in HCT116 are from GSM1266733 and GSM1266734","project":"SRP045269"}
{"number_samples":2,"species":"human","abstract":"Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived normal human kidney transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis Overall design: The kidney tissue was immediately placed and stored in RNAlater® (Ambion), according to the manufacturer’s instruction. The tissue was manually microdissected under microscope in RNAlater® pool for glomerular and tubular compartment. Dissected tissue was homogenized and RNA was prepared using RNAeasy mini columns (Qiagen, Valencia, CA, US), according to the manufacturer’s instructions. RNA quality and quantity were determined using the Laboratory-on-Chip Total RNA PicoKit Agilent BioAnalyzer. Only samples without evidence of degradation were further used (RNA Integrity Number >6).","project":"SRP045276"}
{"number_samples":16,"species":"human","abstract":"Specification of primordial germ cells (PGCs) marks the beginning of the totipotent state. However, without a tractable experimental model, the mechanism of human PGC (hPGC) specification remains unclear. Here, we demonstrate specification of hPGC-like cells (hPGCLCs) from germline competent pluripotent stem cells. The characteristics of hPGCLCs are consistent with the embryonic hPGCs and a germline seminoma that share a CD38 cell-surface marker, which collectively defines likely progression of the early human germline. Remarkably, SOX17 is the key regulator of hPGC-like fate, whereas BLIMP1 represses endodermal and other somatic genes during specification of hPGCLCs. Notable mechanistic differences between mouse and human PGC specification could be attributed to their divergent embryonic development and pluripotent states, which might affect other early cell-fate decisions. We have established a foundation for future studies on resetting of the epigenome in hPGCLCs and hPGCs for totipotency and the transmission of genetic and epigenetic information. Overall design: RNA-Seq analysis to investigate transcriptomes of hPGC-like cells (hPGCLCs), fetal hPGCs, TCam-2 and hESCs","project":"SRP045294"}
{"number_samples":4,"species":"human","abstract":"BET-regulated transcriptome and BRD4, BRD2, BRD3 and Pol II ChIP-seq datasets in human ESCs before and after BET inhibition. Transcription factors and chromatin remodeling complexes are key determinants of embryonic stem cell (ESC) identity. In this study, we investigate the role of BRD4, a member of the bromodomain and extra-terminal domain (BET) family of epigenetic reader proteins, in control of ESC identity. We performed RNA-seq analyiss in the presense of small molecule inhibitors of BET proteins to show that BRD4 positively regulates the ESC transcriptome. We also integrated RNA-seq analysis with ChIP-sequencing datasets s for BRD4 (and for other BRD2 and BRD3) to demonstrate that BRD4 binds SEs and regulates the expression of SE-associated pluripotency genes. We have also conducted ChIP-seq analysis for Pol II binding to demonstrate that SE-associated genes depend on BRD4-dependent Pol II binding at TSS and gene body for their productive transcriptional elongation. Overall design: Total RNA was extracted from samples using the RNeasy Qiagen kit according to the manufacturer’s instructions. Deep sequencing of RNA (1ug) from hESCs FGF- or MS436-treated at day 1 and day 5 was performed as described in (Higgin et al., 2010c). Samples were subjected to PolyA  selection using magnetic oligo-dT beads. The resulting RNA samples were then used as input for library construction  as described by the manufacturer (Illumina, CA, USA). RNA libraries were then sequenced on the GAIIx system using 50bp single reads. Chromatin for ChIP-sequencing was obtained from FGF-maintained hESCs, vehicle or MS417-treated (at 250nM concentration for 6h) (10 to 20x106 cells/IP). ChIP-Seq libraries were generated using standard Illumina kit and protocol as described in (Ntziachristos et al., 2012). We performed cluster amplification and single read 50 sequencing-method using the Illumina HiSeq 2000, following manufacturer’s protocols.","project":"SRP045308"}
{"number_samples":4,"species":"human","abstract":"We compared the expression in embryonic stem cells of KRAB ZNF proteins and retrotransposon elements in the human genome under several conditions. Native human stem cells. Mouse stem cells containing a transchromosomic copy of human chromosome 11, with and without the introduction of plasmids containing KRAB Zinc Finger sequences. We show that certain KZNFs are responsible for the repression of certain retrotransposons in embryonic stem cells, preventing their spread across the genome. Overall design: RNA-seq of hESCs, rhESCs and mouse TC11 ESCs with ZNF91 plasmids (2 replicates), with empty vector plasmids (2 replicates), or no plasmids.","project":"SRP045318"}
{"number_samples":12,"species":"human","abstract":"Breast cancer is a major health problem affecting millions of women worldwide. Over 200,000 new cases are diagnosed annually in the USA, with approximately 40,000 of these cases resulting in death. HER2-positive (HER2+) breast tumors, representing 20–30 % of early-stage breast cancer diagnoses, are characterized by the amplification of the HER2 gene. However, the critical genes and pathways that become affected by HER2 amplification in humans are yet to be specifically identified. Furthermore, it is yet to be determined if HER2 amplification also affects the expression of long intervening non-coding (linc)RNAs, which are involved in the epigenetic regulation of gene expression. We examined changes in gene expression by next generation RNA sequencing in human tumors pre- and post- HER2 inhibition by trastuzumab in vivo, and changes in gene expression in response to HER2 knock down in cell culture models. We integrated our results with gene expression analysis of HER2+ tumors vs matched normal tissue from The Cancer Genome Atlas. The integrative analyses of these datasets led to the identification of a small set of mRNAs, and the associated biological pathways that become deregulated by HER2 amplification. Furthermore, our analyses identified three lincRNAs that become deregulated in response to HER2 amplification both in vitro and in vivo. Our results should provide the foundation for functional studies of these candidate mRNAs and lincRNAs to further our understanding of how HER2 amplification results in tumorigenesis. Also, the identified lincRNAs could potentially open the door for future RNA-based biomarkers and therapeutics in HER2+ breast cancer. Overall design: We compared changes in gene expression of both mRNAs and lincRNAs in BT474 cells that are treated with HER2 siRNAs vs cells treated with negative control siRNAs by RNA-seq.","project":"SRP045322"}
{"number_samples":6,"species":"human","abstract":"Mpn1 proteins are evolutionarily conserved exonucleases that modify spliceosomal U6 small nuclear RNAs (snRNAs) post-transcriptionally. Mutations in the human MPN1 gene are associated to the genodermatosis Clericuzio-type poikiloderma with neutropenia (PN). Mpn1 deficiency leads to aberrant U6 3’ end processing and accelerated U6 decay through unknown molecular mechanisms. Here we show that in mpn1? fission yeast cells U6 is barely bound by the protective Lsm2-8 complex, undergoes extensive oligoadenylation and is degraded by the nuclear RNA exonuclease Rrp6 independently of the poly(A) polymerase Cid14/Trf4. Mpn1 processes U6 in a spliceosome-dependent manner, as mutant U6 molecules that fail to join the spliceosome are not substrates for Mpn1. Moreover, human U6atac, the U6-like snRNA of the minor spliceosome, is a novel substrate for hMpn1. We unveil mechanistic details of a new U6 degradation pathway and further corroborate the notion that inefficient canonical and minor pre-mRNA splicing promotes PN. Overall design: the 3' termini of human U6, U6atac and vtRNA1-1 transcripts from PN patient derived cells and from PN patient cells, compensated with hMPN1 were sequenced.","project":"SRP045329"}
{"number_samples":2,"species":"human","abstract":"Mpn1 proteins are evolutionarily conserved exonucleases that modify spliceosomal U6 small nuclear RNAs (snRNAs) post-transcriptionally. Mutations in the human MPN1 gene are associated to the genodermatosis Clericuzio-type poikiloderma with neutropenia (PN). Mpn1 deficiency leads to aberrant U6 3’ end processing and accelerated U6 decay through unknown molecular mechanisms. Here we show that in mpn1? fission yeast cells U6 is barely bound by the protective Lsm2-8 complex, undergoes extensive oligoadenylation and is degraded by the nuclear RNA exonuclease Rrp6 independently of the poly(A) polymerase Cid14/Trf4. Mpn1 processes U6 in a spliceosome-dependent manner, as mutant U6 molecules that fail to join the spliceosome are not substrates for Mpn1. Moreover, human U6atac, the U6-like snRNA of the minor spliceosome, is a novel substrate for hMpn1. We unveil mechanistic details of a new U6 degradation pathway and further corroborate the notion that inefficient canonical and minor pre-mRNA splicing promotes PN. Overall design: The 3' termini of total RNA containing  2'3' cyclic phosphate from Mpn1-proficient and deficient human and yeast cells was sequenced","project":"SRP045330"}
{"number_samples":56,"species":"human","abstract":"Human neonates and older adults frequently exhibit a reduced capacity to control microbial infections. A variety of mechanisms involving both the innate and adaptive immune systems have been proposed to contribute to these deficiencies. The emergence of RNA sequencing (RNA-seq) as an accurate and quantitative method for examining mRNA levels provides an opportunity to compare transcriptional responses to a stimulus at a global scale in neonates, adults, and older adults. An examination of ex vivo monocyte responses to lipopolysaccharide stimulation or Listeria monocytogenes infection (with cord blood monocytes representing neonatal monocytes) revealed extensive similarities between all three age groups, with only a small number of genes exhibiting statistically significant differences. Using transcription factor motif analyses and RNA-seq data sets from a variety of mouse mutants, the most significant neonatal deficiencies corresponded to genes that require interferon response factor-3 or type 1 interferon signaling for their activation. In older adults, the most striking difference was broad, low-level activation of inflammatory genes prior to stimulation, consistent with prior evidence of a chronic inflammatory state in older adults. These results demonstrate the value of quantitative RNA-seq analyses and the feasibility of cross-species comparisons between well-defined mouse networks and human data sets. Overall design: RNA-seq of primary cells from three independent donors in three different age-groups across 3 time-points stimulated with either LPS or Listeria monocytogenes.","project":"SRP045352"}
{"number_samples":8,"species":"human","abstract":"Adipose stem cells (ASCs) and adipocytes play a crucial role in maintaining energy balance. We aim to examine the temporal relationship between gene expression and histone modification transitions during in vitro differentiation of human ASCs into adipocytes. Here, we examine by RNAseq proliferating ASCs (Day -2 prior to adipogenic induction), confluent ASCs (Day 0, adipogenic induction), pre-adipocytes (Day 3) and maturing adipocytes (Day 9). We find 1060, 5452 and 2216 genes differentially expressed between D-2/D0, D0/D3 and D3/D9 respectively. We identify gene clusters with distinct and dynamic expression patterns. In particular, adipogenic induction is marked by temporal waves of gene induction and downregulation. We report two types of transcriptional waves: (i) those showing transient induction or inactivation at D0, D3 or D9, and involved in sensory perception and immune response functions; and (ii) those showing long-lived induction or repression at these time points. Our data reveal a dynamic network of gene regulation during adipogenesis, involving signaling, immune and developmental processes. We identify 15 unique epigenetic states using Hidden Markov Modeling which reflects an epigenetically highly organised genome showing enhancer states are commonly consecutive. A heatmap for the abundance of epigenetic states for the expression clusters reveals a link between expression and epigenetic marking of the state suggesting an increase in the number of number of chromatin states with increase in expression. Our data point to a model of increased epigenetic complexity associated with gene expression. Overall design: Examination of expression of profiles of ASCs across differentiation","project":"SRP045364"}
{"number_samples":14,"species":"human","abstract":"Enhancers provide critical information directing cell-type specific transcriptional programs, regulated by binding of signal-dependent transcription factors and their associated cofactors. Here we report that the most strongly activated estrogen (E2)-responsive enhancers are characterized by trans-recruitment and in situ assembly of a large 1-2MDa complex of diverse DNA-binding transcription factors by ERa at ERE-containing enhancers. We refer to enhancers recruiting these factors as mega transcription factor-bound in trans (MegaTrans) enhancers. The MegaTrans complex is a signature of the most potent functional enhancers and is required for activation of enhancer RNA transcription and recruitment of coactivators, including p300 and Med1. The MegaTrans complex functions, in part, by recruiting specific enzymatic machinery, exemplified by DNA-dependent protein kinase. Thus, MegaTrans-containing enhancers represent a cohort of functional enhancers that mediate a broad and important transcriptional program and provide a molecular explanation for transcription factor clustering and hotspots noted in the genome. Overall design: Global Run On (GRO) assay followed by high throughput sequencing (GRO-seq)","project":"SRP045391"}
{"number_samples":8,"species":"human","abstract":"This study aims to determine, in detail, the changes in gene expression that occur in response to interferon alpha stimulation in human primary hepatocytes. Using a transcriptome sequencing approach, the study aimed to identify novel transcripts that were induced in response to interferons and may play a role in innate immunity.","project":"SRP045406"}
{"number_samples":24,"species":"human","abstract":"We extracted RNA from whole cells and RNA from the cytoplasm and performed RNA sequening to compare differences in gene expression level and investigate what is the most appropriate estimate of the amount of mRNA present in a given cell population. The study was based on three human cell lines. Overall design: Analyze of transcriptome in 3 human cell lines (U-2 OS, A-431, U-251MG). Each cell line was prepared with four biological replicates for total RNA and four for cytoplasmic RNA.","project":"SRP045421"}
{"number_samples":24,"species":"human","abstract":"RNA-seq was performed on LFS MDAH041 cells that were young (PD10-12), aging (PD17-19) and replicatively senescent (PD28-30), as well as spontaneously immortal cells and cells that were induced into senescence or quiescence,  in order to profile the pathways common in all 4 types of sensecence and the pathways affected as a cell approaches senescence. Overall design: RNA was sequenced in biological triplicates of each sample using the Illumina HiSeq2000","project":"SRP045441"}
{"number_samples":6,"species":"human","abstract":"RNA-seq analysis of samples from Non-silencing, RBFOX2 and CPSF2 knockdown cells Overall design: We analyzed 6 samples using the Illumina HiSeq 2000. Sequence data was processed by DexSeq. Two biological replicates were performed for Non-silencing, RBFOX2 and CPSF2 knockdown cells","project":"SRP045481"}
{"number_samples":134,"species":"human","abstract":"This study compared whole transcriptome signatures of 6 immune cell subsets and whole blood from patients with an array of immune-associated diseases. Fresh blood samples were collected from healthy subjects and subjects diagnosed type 1 diabetes, amyotrophic lateral sclerosis, and sepsis, as well as multiple sclerosis patients before and 24 hours after the first treatment with IFN-beta. At the time of blood draw, an aliquot of whole blood was collected into a Tempus tube (Invitrogen), while the remainder of the primary fresh blood sample was processed to highly pure populations of neutrophils, monocytes, B cells, CD4 T cells, CD8 T cells, and natural killer cells. RNA was extracted from each of these cell subsets, as well as the whole blood samples, and processed into RNA sequencing (RNAseq) libraries (Illumina TruSeq). Sequencing libraries were analyzed on an Illumina HiScan, with a target read depth of ~20M reads. Reads were demultiplexed, mapped to human gene models (ENSEMBL), and tabulated using HTSeq. Read count data were normalized by the TMM procedure (edgeR package). Overall design: We performed whole genome RNAseq profiling of immune cell subsets and whole blood from subjects with an array of immune-associated diseases.","project":"SRP045500"}
{"number_samples":16,"species":"human","abstract":"MicroRNAs (miRNAs) regulate target mRNAs through a combination of translational repression and mRNA destabilization, with mRNA destabilization dominating at steady state in the few contexts examined globally. Here, we extend the global steady-state measurements to additional mammalian contexts and find that regardless of the miRNA, cell type, growth condition, or translational state, mRNA destabilization explains most (66%–>90%) miRNA-mediated repression. We also determine the relative dynamics of translational repression and mRNA destabilization for endogenous mRNAs as a miRNA is induced. Although translational repression occurs rapidly, its effect is relatively weak, such that by the time consequential repression ensues, the effect of mRNA destabilization dominates. These results imply that consequential miRNA-mediated repression is largely irreversible and provide other insights into the nature of miRNA-mediated regulation. They also simplify future studies, dramatically extending the known contexts and time points for which monitoring mRNA changes captures most of the direct miRNA effects. Overall design: 85 samples from a variety of cell types and species","project":"SRP045501"}
{"number_samples":6,"species":"human","abstract":"RNA-seq is a sensitive and accurate technique to compare steady state levels of RNA between different cellular states. However, as it does not provide an account of transcriptional activity per se, other technologies are needed to more precisely determine acute transcriptional responses. Here, we have developed an easy, sensitive and accurate novel method, iRNA-seq, for genome-wide assessment of transcriptional activity based on analysis of intron coverage from total RNA-seq data. To test our method, we have performed total RNA-seq and RNA polymerase II (RNAPII) ChIP-seq profiling of the acute transcriptional response of human adipocytes to TNFa treatment and analyzed these using iRNA-seq in addition to different publically availbale dataset.  Comparison of the results derived from iRNA-seq analyses with results derived using current methods for genome-wide determination of transcriptional activity, i.e. Global Run-On (GRO)-seq and RNA polymerase II (RNAPII) ChIP-seq, demonstrate that iRNA-seq provides very similar results in terms of number of regulated genes and their fold change. However, unlike the current methods that are all very labor-intensive and demanding in terms of sample material and technologies, iRNA-seq is cheap and easy and requires very little sample material. In conclusion, iRNA-seq offers an attractive novel alternative to current methods for determination of changes in transcriptional activity at a genome-wide level. Overall design: Genome-wide assesment of the acute transcriptional response to TNFa in human SGBS adiposytes using total RNA-seq data end RNAPII ChIP-seq","project":"SRP045545"}
{"number_samples":2,"species":"human","abstract":"We have used RNA-seq to examine circular RNAs from poly(A)-/ribo- RNAs in human and mouse embryonic stem cells, and from from RNase R treated poly(A)-/ribo- RNAs in mouse embryonic stem cells. Overall design: In order to identify novel circular RNAs from different species","project":"SRP045561"}
{"number_samples":5,"species":"human","abstract":"Fusion proteins involving the ETS factor ERG have been associated with multiple cancers such as Ewing's sarcoma and prostate cancer. In acute myeloid leukemias harboring  t(16;21) another ERG fusion protein is expressed, FUS-ERG. Here, we found that this FUS-ERG oncofusion protein acts in the context of a heptad of proteins (ERG, FLI1, GATA2, LYL1, LNMO2, RUNX1 and TAL1) central to proper expression of genes involved in maintaining a stem cell hematopoietic phenotype. Moreover, in t(16;21) FUS-ERG co-occupies genomic regions bound by the nuclear receptor heterodimer RXR-RARA inhibiting target gene expression and interfering with hematopoietic differentiation. All-Trans Retinoic Acid treatment of t(16;21) cells as well as FUS-ERG knock down alleviate the myeloid differentiation block. Together, the results suggest that FUS-ERG acts as a transcriptional repressor of the retinoic acid signaling pathway. Overall design: Cell lines were used for RNA extraction for RNA-seq and ChIP experiments for ChIP-seq.","project":"SRP045565"}
{"number_samples":12,"species":"human","abstract":"T Cell stretch-enhancers are vulnerable to Jak inhibitor tofacitinib Overall design: Treatment of T cells with the Janus kinase (JAK) inhibitor, tofacitinib, disproportionately altered the expression of RA risk genes with stretech-enhancer (SE) structures.","project":"SRP045570"}
{"number_samples":6,"species":"human","abstract":"Mapping MBNL-regulated genome-wide alternative polyadenylation: We report that depletion of Mbnl proteins in mouse embryo fibroblasts (MEFs), DM mouse model quadriceps muscle, and DM-autopsy muscle tissue leads to mis-regulation of alternative polyadenylation Overall design: We compared WT, Mbnl1/2KO, Mbnl1/2KO/3siRNA, and Mbnl1/2KO/scrambled siRNA MEFs (n=2 for each group) to evaluate alternative polyadenylation shifts that occur due to progressive loss of Mbnl proteins. We also compared WT (1 day old, and 4 months old, n=2 each) and HSALR mouse model (4 months old, n=2) of myotonic dystrophy for developmental alternative polyadenylation defects in myotonic dystrophy. Finally, we compared control and DM1 autopsy muscle tissues (n=3) for changes in alternative polyadenylation. We performed HITS-CLIP analysis of binding sites of Mbnl1, Mbnl2 and Mbnl3 in MEFs (n=3 each). We also performed HITS-CLIP analysis for major skeletal muscle Mbnl protein, Mbnl1 in FVB WT adult muscle (4 months, n=3). Finally we performed HITS-CLIP analysis for CPSF6 in WT and Mbnl1/2 KO MEFs (n=3 each) Please note that the 'readme_Table.txt' describes the contents of 'Table S*.xlsx' files, and the readme_method.txt include additional details about experiemenal procedures.","project":"SRP045573"}
{"number_samples":7,"species":"human","abstract":"(h)MeDIP-seq and transcriptome data of human glioblastoma cells and neural progenitor cells","project":"SRP045590"}
{"number_samples":12,"species":"human","abstract":"Examination of polyA+ long RNA population from MCF-7 and MCF-10 cells in cytoplasm, nucleus & total sample to study the changes in alternative splicing in breast cancer.","project":"SRP045592"}
{"number_samples":1,"species":"human","abstract":"Canonical Wnt signaling in endothelial cells (ECs) is required for vascularization of the central nervous system (CNS) and for formation and maintenance of barrier properties unique to CNS vasculature. Gpr124 is an orphan member of the adhesion G-protein-coupled receptor family that is expressed in ECs and is essential for CNS angiogenesis and barrier formation via an unknown mechanism. Using canonical Wnt signaling assays in cell culture and genetic loss- and gain-of-function experiments in mice, we show that Gpr124 functions as a co-activator of Wnt7a- and Wnt7b-stimulated canonical Wnt signaling via a Frizzled receptor and Lrp co-receptor, and that Gpr124-stimulated signaling functions in concert with Norrin/Frizzled4 signaling to control CNS vascular development.  These experiments identify Gpr124 as a ligand-specific co-activator of canonical Wnt signaling. Overall design: Total mRNA from HEK-293/STF cells was subjected to RNAseq","project":"SRP045610"}
{"number_samples":12,"species":"human","abstract":"Purpose: We identified KPC1 as the ubiquitin ligase that binds to the p105 precursor of NF-kB, ubiquitinates it and mediates its proteasomal processing to generate the p50 active subunit of the transcription factor. Using U87-MG human glioblastoma xenografts, we observed that overexpression of KPC1 results in strong inhibition of tumor growth mediated via excessive generation of p50.The goal of this RNASeq study was to analyze the profile of gene expression in xenografts overexpressing control (V0), KPC1 or p50 vectors, and to further understand how the altered gene expression patterns can explain the tumor suppressive effect we observed. Results:Transcript analysis of U87-MG xenografts overexpressing control (V0), KPC1 or p50 vector mapped  to the human genome  revealed: • A strong similarity between overexpression of p50 and KPC1 (correlation of 0.51, p-value<10-300 ) • A specific signature of NF-kB targets [21 of the consistently changed genes are known to be regulated by NF-kB (p-value<3.4×10-9 )] • A significant (p-value<1.4×10-18) increase in the expression of 40 tumor suppressor genes, with no significant change in other classes. • A significant down regulation of a cluster of genes including LIN28B, IL-6, HMAGA2 and VEGFA. This finding links well to an established regulatory axis involving LIN28B, Let-7 microRNA, and IL-6 in inflammation and cell transformation that is regulated by NF-kB. Overall design: Exponentially growing U87-MG cells were stably transfected with an empty vector (V0) or vectors coding for Myc-KPC1 or Flag-p50. Cells were dissociated with trypsin, washed with PBS, and brought to a concentration of 50×10^6 cells/ml. Cell suspension (5×10^6/0.1 ml) was inoculated subcutaneously at the right flank of 7-weeks old male Balb/C nude mice (n=7). Following 21 days, mRNA from U87-MG xenografts was isolated using an RNA purification kit, and analyzed using the Illumina HiSeq 2500 sequencer. The RNASeq analysis experiment was repeated twice independently. Run1 included a total of 7 samples. Samples 1-3 were isolated from V0 – control tumors (3 different tumors), samples 4-5 were isolated from KPC1-expressing  tumors (2 different pools of 3 tumors each due to small tumor size), and samples 6-7 were isolated from p50-expressing tumors for (2 different pools of 2-3 tumors each, due to very small tumor size). Run2 included a total of 5 samples. Samples 8-10 were isolated from V0 (control) tumors (3 different tumors), samples 11-12 were isolated from KPC1 tumors (2 different pools of 3 tumors each due to small tumor size). Several repeated attempts to extract RNA from the p50-expressing tumors did not yield any results, as the tumors were miniscule.","project":"SRP045611"}
{"number_samples":6,"species":"human","abstract":"VHL inactivation is a major component of renal cancer. In this study we re-engineered RCC4 cells to re-express VHL and performed mRNA-sequencing on triplicates to analyse gene and transcript expression changes specific to VHL inactivation.","project":"SRP045624"}
{"number_samples":12,"species":"human","abstract":"Advances in cellular reprogramming and stem cell differentiation now enable ex vivo studies of human neuronal differentiation. However, it remains challenging to elucidate the underlying regulatory programs because differentiation protocols are laborious and often result in low neuron yields. Here, we overexpressed two murine Neurogenin transcription factors in human induced pluripotent stem cells, and obtained neurons with bipolar morphology in four days at greater than 90% purity. The high purity enabled mRNA and microRNA expression profiling during neurogenesis, thus revealing the genetic programs involved in the transition from stem cell to neuron. These profiles were then analyzed to identify the regulatory networks underlying the differentiation of the neurons. Overall design: Paired end RNA sequencing of iPS cells (PGP1) at 0, 1, 3, and 4 days post- doxycycline induction of murine NGN1 and NGN2. This was done using an Illumina HiSeq, and reads were aligned to hg19","project":"SRP045632"}
{"number_samples":72,"species":"human","abstract":"RNAseq data of 36 samples across human brain development by age group from LIBD","project":"SRP045638"}
{"number_samples":12,"species":"human","abstract":"An unbalanced karyotype, a condition known as aneuploidy, has a profound impact on cellular physiology and is a hallmark of cancer. Determining how aneuploidy affects cells is thus critical to understanding tumorigenesis. Here we show that aneuploidy interferes with the degradation of autophagosomes within lysosomes. Mis-folded proteins that accumulate in aneuploid cells due to aneuploidy-induced proteomic changes overwhelm the lysosome with cargo, leading to the observed lysosomal degradation defects. Importantly, aneuploid cells respond to lysosomal saturation. They activate a lysosomal stress pathway that specifically increases the expression of genes needed for autophagy-mediated protein degradation. Our results reveal lysosomal saturation as a universal feature of the aneuploid state that must be overcome during tumorigenesis. Overall design: RPE-1 cells either untreated or treated with one of Reversine, Bafilomycin A1 or MG132, each condition was done in triplicate. D14-*_Control: untreated control D14-*_Rev: cells treated with 0.5uM Reversine for 24hrs and harvested 48hrs later D14-*_Baf: cells treated with 0.1uM BafA1 for 6hrs D14-*_Mg: cells treated with 1uM MG132 for 24 hrs","project":"SRP045639"}
{"number_samples":6,"species":"human","abstract":"Characterized mRNA and miRNA transcriptomes by RNA sequencing in EEC to investigate potential molecular mechanisms underlying the pathogenesis.","project":"SRP045645"}
{"number_samples":4,"species":"human","abstract":"We used 2', 3'-cyclic phosphate cDNA synthesis and Illumina sequencing to identify and quantify ribonuclease L cleavage sites in host and viral RNAs. Overall design: To make cDNA libraries, we exploited the 2’, 3’-cyclic phosphate at the end of RNA fragments produced by RNase L and other metal-ion-independent endoribonucleases. Using Arabidopsis thaliana tRNA ligase, RNA fragments with 2’, 3’-cyclic phosphates were covalently attached to defined RNA linkers containing an 8-base long unique molecular identifier (UMI) sequence. Libraries prepared in this manner contain cDNA derived exclusively from RNA fragments with 2', 3'-cyclic phosphates. The UMI sequence allows for detailed quantitation. We applied these methods to total RNA from A549 cells infected by influenza A virus unable to express NS1. NS1 inhibits the activation of RNase L.","project":"SRP045659"}
{"number_samples":18,"species":"human","abstract":"RNAPII pausing/termination shortly after initiation is a hallmark of gene regulation. However, the molecular mechanisms involved are still to be uncovered. Here, we show that NELF interacts with Integrator complex subunits (INTScom) forming a stable complex with RNPII and Spt5. The interaction between NELF and INTScom subunits is RNA and DNA independent. Using both HIV-1 promoter and genome wide analyses, we demonstrate that Integrator subunits specifically control NELF-mediated RNAPII pause/release at coding genes. The strength of RNAPII pausing is determined by the nature of the NELF-associated complex. Interestingly, in addition to controlling RNAPII pause release INTS11 catalytic subunit of the INTScom is required for the synthesis of full length mRNA. Finally, INTScom-target genes are enriched in HIV-1 TAR/ NELF-binding element and in a 3’box sequence required for snRNA biogenesis. Revealing these unexpected functions of INTScom in regulating RNAPII pausing/release and completion of mRNA synthesis of NELF-target genes will contribute to our understanding of the gene expression cycle. Overall design: Genome-wide expression in HeLa cells in the absence of Integrator 11, or NELF or mock (control) depleted by strand-specific RNASeq (Illumina)","project":"SRP045663"}
{"number_samples":40,"species":"human","abstract":"A total of 23 participants (data available in present submission and in GSE58608) completed three months of supervised aerobic exercise training of one leg. Skeletal muscle biopsies have been collected before and after the training period. We have investigated differences between trained and untrained leg and before and after training by studying the gene and isoform expression. Additional samples present in this study has been previously published (GEO accession number GSE58608). Overall design: Analysis of transcriptome in skeletal muscle biopsy samples in response to exercise training in 22 participants (of the total 23 participants). One biopsy is collected from each leg before and after training period.","project":"SRP045666"}
{"number_samples":4,"species":"human","abstract":"Here we show that by simple modulation of extrinsic signaling pathways, a new class of pluripotent stem cells, referred to as region selective epiblast stem cells (rsEpiSCs), could be efficiently derived from different stages of the early embryo. rsEpiSCs share features of primed pluripotency yet are distinct from EpiSCs in their molecular characteristics and ability to colonize post-implantation embryos. We performed RNA-sequencing experiments and examined the global gene expression profiles of EpiSCs, rsEpiSCs, in vivo isolated four regions of E6.5 mouse epiblasts: AP (anterior-proximal), AD (anterior-distal), PP (posterior-proximal) and PD (posterior-distal), human H1 ESCs, H1 rsESCs, H9 ESCs, H9 rsESCs, rhesus monkey ORMES23 rsESCs, and chimpanzee rsiPSCs. Overall design: Examination of global gene expression profiles in 2 pluripotent stem cell types across multiple species.","project":"SRP045673"}
{"number_samples":12,"species":"human","abstract":"G quadruplex motifs are frequently located near TSS and in the first introns of transcribed genes. We investigated the role that G4 structures play in transcription by stabilizing G4 DNA with the selective G4 ligand PhenDC3 in the HT-1080 human fibrosarcoma cell line. After treatment of triplicate samples with PhenDC3 or a negative control (DMSO carrier only), we performed RNA-Seq after poly-dT selection to assess changes in gene and isoform expression, as well as changes to splicing patterns. Overall design: 6 samples total; 2 gene expression conditions (treated and untreated) in triplicate; 4 raw sequencing files per sample","project":"SRP045695"}
{"number_samples":17,"species":"human","abstract":"Understanding the molecular processes underlying intra-tumor heterogeneity is of critical importance to improve the efficiency of therapy and overcome drug resistance. In malignant melanoma, heterogeneity is though to arise -at least partly- through epigenetic rather than genetic reprogramming of proliferating cells, leading to the appearance within the primary tumors of a phenotypically distinct invasive cell subpopulation. The invasive melanoma cells are at the origin of metastatic spread and are thought to provide a reservoir of therapeutically resistant cells. To decipher the gene regulatory networks that underlie the proliferative and invasive cellular states we integrated publicly available gene expression and DNA methylation data from tumor biopsies with a newly generated compendium of open chromatin and histone modification profiling of melanoma cultures that are dominant in either of the two subpopulations. Extensive in silico analyses identified SOX10 and MITF, and AP-1 and TEADs as key upstream regulators of the proliferative and invasive transcriptomes, respectively. Knock-down experiments confirmed the implication of TEADs in the invasive gene network and, more importantly, established a causative link between activation of these transcription factors and melanoma cell invasion and sensitivity to MAPK inhibitors. Overall design: Investigation of the transcriptomes of 11 different melanoma cultures & Investigation of the effects of the Knock down of all 4 TEADs in an invasive melanoma culture (MM047).","project":"SRP045711"}
{"number_samples":81,"species":"human","abstract":"In the present study 23 participants completed three months of supervised aerobic exercise training of one leg (training period 1) followed by 9 months of rest before 12 of the participants completed a second exercise training period (training period 2) of three months of both legs. Skeletal muscle biopsies have been collected before and after the training periods. We have compared trained leg with untrained leg and studied gene and isoform expression. Additional samples included in this study has been previously submitted (GEO accession number GSE58387 and GSE60590). Overall design: Analyze of transcriptome in skeletal muscle biopsy samples in response to exercise training in 23 participants in total (in addition to data previously submitted GEO accession number GSE58387 and GSE60590). Biopsy is collected from skeletal muscle before and after training period.","project":"SRP045823"}
{"number_samples":9,"species":"human","abstract":"To understand epigenic dysregulation of host and viral genes upon EBV infection in human gastric cancer, genome wide transcripts by RNAseq were undertaken for total RNAs of 3 EBVnGC, their isogenic cell lines converted by in vitro EBV-infection and 3 EBV-naturally infected GC cell lines. Overall design: Three independent gastric cancer (GC) cell lines (NUGC3, SNU-484, SNU-638) were experimentally infected with AKATA strain of EBV to derive EBV-infected isogenic GC cell lines (NUGC3EBV, SNU-484EBV, SNU-638EBV). Three indepedent GC cell lines with natural EBV-infection (SNU-719, NCC24, YCCEL1)) were also included in this study.","project":"SRP045859"}
{"number_samples":6,"species":"human","abstract":"Quiescent MRC-5 fibroblasts were compared to young fibroblasts Overall design: 6 samples: 3 biological replicates for each age group: young and quiescent MRC-5 cells. 50bp, single-end reads, no strand-specific reads","project":"SRP045867"}
{"number_samples":6,"species":"human","abstract":"Ewing sarcoma is a highly aggressive tumor characterized by a translocation between members of the FET family of RNA binding proteins and one of several ETS transcription factors, with the most common translocation being EWS-FLI1. EWS-FLI1 leads to changes in gene expression through mechanisms that are not completely understood. We performed RNA sequencing analysis on primary pediatric human mesenchymal progenitor cells (pMPCs) expressing EWS-FLI1 in order to identify novel target genes. This analysis identified lnc277 as a previously uncharacterized long non-coding RNA upregulated by EWS-FLI1 in pMPCs. Inhibiting the expression of lnc277 diminished the ability of Ewing sarcoma cell lines to proliferate and form colonies in soft agar whereas inhibiting lnc277 had no effect on other cell types tested. By analyzing gene expression after shRNA knockdown, we found that both EWS-FLI1 and lnc277 repressed many more genes that they induced and that a significant fraction of EWS-FLI1 repressed targets were also repressed by lnc277. Analysis of primary human Ewing sarcoma RNA sequencing data further supports a role for lnc277 in mediating gene repression. We identified hnRNPK as an RNA binding protein that interacts directly with lnc277. We found a significant overlap in the genes repressed by hnRNPK and those repressed by both EWS-FLI1 and lnc277, suggesting that hnRNPK participates in lnc277 mediated gene repression. Thus, lnc277 is a previously uncharacterized long non-coding RNA downstream of EWS-FLI1 that facilitates the development of Ewing sarcoma via the repression of target genes. Our studies identify a novel mechanism of oncogenesis downstream of a chromosomal translocation and underscore the importance of lncRNA-mediated gene repression as a mechanism of EWS-FLI1 transcriptional regulation. Overall design: RNA from primary human bone marrow derived mesenchymal cells either control or with inducible expression of  EWS-FLI1 for 13 days was used to prepare PolyA selected cDNA libraries.","project":"SRP045869"}
{"number_samples":4,"species":"human","abstract":"Ewing sarcoma is a highly aggressive tumor characterized by a translocation between members of the FET family of RNA binding proteins and one of several ETS transcription factors, with the most common translocation being EWS-FLI1. EWS-FLI1 leads to changes in gene expression through mechanisms that are not completely understood. We performed RNA sequencing analysis on primary pediatric human mesenchymal progenitor cells (pMPCs) expressing EWS-FLI1 in order to identify novel target genes. This analysis identified lnc277 as a previously uncharacterized long non-coding RNA upregulated by EWS-FLI1 in pMPCs. Inhibiting the expression of lnc277 diminished the ability of Ewing sarcoma cell lines to proliferate and form colonies in soft agar whereas inhibiting lnc277 had no effect on other cell types tested. By analyzing gene expression after shRNA knockdown, we found that both EWS-FLI1 and lnc277 repressed many more genes that they induced and that a significant fraction of EWS-FLI1 repressed targets were also repressed by lnc277. Analysis of primary human Ewing sarcoma RNA sequencing data further supports a role for lnc277 in mediating gene repression. We identified hnRNPK as an RNA binding protein that interacts directly with lnc277. We found a significant overlap in the genes repressed by hnRNPK and those repressed by both EWS-FLI1 and lnc277, suggesting that hnRNPK participates in lnc277 mediated gene repression. Thus, lnc277 is a previously uncharacterized long non-coding RNA downstream of EWS-FLI1 that facilitates the development of Ewing sarcoma via the repression of target genes. Our studies identify a novel mechanism of oncogenesis downstream of a chromosomal translocation and underscore the importance of lncRNA-mediated gene repression as a mechanism of EWS-FLI1 transcriptional regulation. Overall design: A673 Ewing cells expressing an shRNA targeting hnRNPK or control were subjected to paired end RNA sequencing and compared to shGFP control.","project":"SRP045870"}
{"number_samples":6,"species":"human","abstract":"To understand how haploinsufficiency of progranulin (PGRN) protein causes frontotemporal dementia (FTD), we created induced pluripotent stem cells (iPSC) from patients carrying the GRNIVS1+5G>C mutation (FTD-iPSCs). FTD-iPSCs were fated to cortical neurons, the cells most affected in FTD and known to express PGRN. Although generation of neuroprogenitors was unaffected, their further differentiation into neurons, especially CTIP2-, FOXP2- or TBR1-TUJ1 double positive cortical neurons, was significantly decreased in FTD-neural progeny. Zinc finger nuclease-mediated introduction of PGRN cDNA into the AAVS1 locus corrected defects in cortical neurogenesis, demonstrating that PGRN haploinsufficiency causes inefficient cortical neuron generation. RNAseq analysis confirmed reversal of altered gene expression profile following genetic correction. Wnt signaling pathway, one of the top defective pathways in FTD-iPSC-derived neurons coupled with its reversal following genetic correction, makes it an important candidate. Therefore, we demonstrate for the first time that PGRN haploinsufficiency hampers corticogenesis in vitro. Overall design: We profiled 6 samples: two biological replicates for 3 conditions. Condition 1 consists of neuronal progeny derived from human Embryonic Stem Cells. Condition 2 consists of neuronal progeny derived from induced pluripotent stem cells generated from patients carrying PGRN mutation. Condition 3 consists of neuronal progeny derived from induced pluripotent stem cells generated from patients carrying PGRN mutation, genetically modified to correct the PGRN defect.","project":"SRP045876"}
{"number_samples":6,"species":"human","abstract":"Anti-angiogenic therapy is commonly used for the treatment of CRC. Although patients derive some clinical benefit, treatment resistance inevitably occurs. The MET signaling pathway has been proposed to be a major contributor of resistance to anti-angiogenic therapy. MET is upregulated in response to VEGF pathway inhibition and plays an essential role in tumorigenesis and progression of tumors. In this study we set out to determine the efficacy of cabozantinib in a preclinical CRC PDTX model. We demonstrate potent inhibitory effects on tumor growth in 80% of tumors treated. The greatest antitumor effects were observed in tumors that possess a mutation in the PIK3CA gene. The underlying antitumor mechanisms of cabozantinib consisted of inhibition of angiogenesis and Akt activation and significantly decreased expression of genes involved in the PI3K pathway. These findings support further evaluation of cabozantinib in patients with CRC. PIK3CA mutation as a predictive biomarker of sensitivity is intriguing and warrants further elucidation. A clinical trial of cabozantinib in refractory metastatic CRC is being activated. Overall design: CRC PDTX Model treated with cabozantinib","project":"SRP045898"}
{"number_samples":3,"species":"human","abstract":"Multiple sclerosis is a chronic inflammatory demyelinating disease of the central nervous system with marked heterogeneity in several aspects including pathological processes. Four histopathological patterns of MS have been described. Pattern II is characterized by infiltrating macrophages and T-cells and by antibody and complement deposition. Transcriptome analysis of three patern II demyelinating brain lesions from a multiple sclerosis patient using RNA sequencing demonstrated the presence of mRNA transcripts for genes specific of activated macrophages, T and B cells as well as genes coding for immunoglobulins, complement proteins and some pattern II associated proteins, providing additional evidence supporting pattern II demyelination. Overall design: Examination of 3 different demyelinating lesions identified by Immunohistopathology.","project":"SRP045900"}
{"number_samples":15,"species":"human","abstract":"RNA from A673 cells with shRNA-mediated knockdown of GFP (4 libraries),  EWS-FLI1 (4 libraries), or lnc277 (7 libraries) was isolated with TRIzol (Invitrogen).  Each sample was DNase treated and further purified on an RNeasy  Mini column (Qiagen) before quality analysis on an Agilent 2100 Bioanalyzer.  For each sample, 100-150ng of RNA was synthesized into cDNA, sheared on  a Covaris ultrasonicator, and amplified using the NuGen Encore Complete kit  (NuGen) to produce strand-specific and rRNA-depleted libraries. Samples were  multiplexed (4/lane) for 2x100bp paired-end sequencing on an Illumina HiSeq  2000 Overall design: RNA from A673 cells with shRNA-mediated knockdown of GFP (4 libraries), EWS-FLI1 (4 libraries), or lnc277 (7 libraries) was isolated with TRIzol (Invitrogen).","project":"SRP045905"}
{"number_samples":7,"species":"human","abstract":"We performed mRNA-seq and small RNA-seq on the newly derived ELF human naïve pluripotent stem cell line and compared with the existing H1 human primed line. Expression analysis revealed that mitochondria oxidative phosphorylation and fatty acid beta-oxidaton is up-regulated in naive state while fatty acid synthesis is up-regulated in primed state. Small RNA-seq revealed consistent expression changes in microRNAs that target key fatty acid beta-oxidation and synthesis genes. Integration with metabolomics data revealed consistent changes in the expression of NNMT (higher in naive state) and IDO1 (higher in primed state) and in the concentration of corresponding metabolic substrates and products. As a regulator of S-Adenosyl methionine (SAM) level, NNMT is proposed as a candidate regulator of epigenetic states. ChIP-seq analysis releaved that naive lines have lower H3K27me3 marks in developmental genes. Inhibition of STAT3, a known regulator of NNMT, reduces NNMT expression level and decreases overall H3K27me3 marks around transcriptional start sites. In particular, STAT3 inhibitor treatment increased H3K27me3 marks in 313 genes that also have higher H3K27me3 marks in primed state than naive state. These 313 genes are enriched with developmental functions, and include several WNT pathway genes. In summary, integrative analysis of RNA-seq, ChIP-seq and metabolomics data revealed key metabolic differences between naive and primed pluripotency and identified NNMT as a key regulator of epigeneitc state. Overall design: 2 biological replicates for ELF RNA-seq; 2 biological replicates for ELF small RNA-seq; one sample for ELF H3K27me3 ChIP-seq and one sample as input; one sample for ELF treated with STAT3 inhibitor for 6 hours and one sample as input. UPDATE: We performed RNA-seq on two additional human naïve embronic stem cell lines (H1 4iLIF and Lis1), as well as mouse in vivo preimplantation inner cell mass and postimplantation epiblast. These RNA-seq samples are included in the naïve vs. primed comparisons in the study; 2 biological replicates of H1 4iLIF; 2 biological replicates of Lis1; 3 biological replicates of mouse preimplantation inner cell mass; 2 biological replicates of mouse postimplantation epiblast.","project":"SRP045911"}
{"number_samples":6,"species":"human","abstract":"Control of NCCIT differentiation","project":"SRP045949"}
{"number_samples":8,"species":"human","abstract":"We describe a chemical method to label and purify 4-thiouridine (s4U) -containing RNA. We demonstrate that methanethiolsulfonate (MTS) reagents form disulfide bonds with s4U more efficiently than the commonly used HPDP-biotin, leading to higher yields and less biased enrichment. This increase in efficiency allowed us to use s4U-labeling to study global microRNA (miRNA) turnover in proliferating cultured human cells without perturbing global miRNA levels or the miRNA processing machinery. This improved chemistry will enhance methods that depend on tracking different populations of RNA such as 4-thiouridine-tagging to study tissue-specific transcription and dynamic transcriptome analysis (DTA) to study RNA turnover. Overall design: s4U metabolic labeling of RNA in 293T cells, followed by biochemical enrichment of labeled RNA with two biotinylation reagents, RNAs >200nt and miRNAs in separate experiments","project":"SRP045983"}
{"number_samples":6,"species":"human","abstract":"Coxiella and Leishmania both inhabit the lysozome. Not much is currently known about the differences in host response to these microbial pathogens inhabiting the same niche. Understanding differences in host response may elucidate new treatments to these difficult to treat pathogens.","project":"SRP045986"}
{"number_samples":64,"species":"human","abstract":"Comparative studies in primates are extremely restricted because we only have access to a few types of cell lines from non-human apes and to a limited collection of frozen tissues. In order to gain better insight into regulatory processes that underlie variation in complex phenotypes, we must have access to faithful model systems for a wide range of tissues and cell types. To facilitate this, we have generated a panel of 7 fully characterized chimpanzee (Pan troglodytes) induced pluripotent stem cell (iPSC) lines derived from fibroblasts of healthy donors. All lines are free of integration from exogenous reprogramming vectors, can be maintained using standard iPSC culture techniques, and have proliferative and differentiation potential similar to human and mouse lines. To begin demonstrating the utility of comparative iPSC panels, we collected RNA-seq data and methylation profiles from the chimpanzee iPSCs and their corresponding fibroblast precursors, as well as from 7 human iPSCs and their precursors, which were of multiple cell type and population origins. Overall, we observed much less regulatory variation within species in the iPSCs than in the somatic precursors, indicating that the reprogramming process has erased many of the differences observed between somatic cells of different origins. We identified 4,918 differentially expressed genes and 1,986 differentially methylated regions between iPSCs of the two species, many of which are novel inter-species differences and not observed between the somatic cells of the two species. Our panel will help realize the potential of iPSCs, and in combination with genomic technologies, transform studies of comparative evolution in primates. Overall design: We obtained RNA sequencing and methylation profiles from 7 chimpanzee iPSCs and the fibroblasts used to generate them, as well as 7 human iPSCs and the LCLs and fibroblasts used to generate them.","project":"SRP045999"}
{"number_samples":4,"species":"human","abstract":"How stroma communicates with cancer to influence treatment response is poorly understood. We show that stromal fibroblasts protect breast cancer (BrCa) against radiation and chemotherapy through an exosome-mediated anti-viral pathway and NOTCH3. Stroma increases RAB27B and transfers exosomes to BrCa. RNA within exosomes, comprised largely of non-coding transcripts and transposable elements, stimulates the pattern recognition receptor RIG-I through a 5’-triphosphate motif to activate STAT1. BrCa NOTCH3 is activated in parallel by stromal JAG1 and cooperates with STAT1 to enhance transcriptional responses of NOTCH target genes and to expand therapy resistant tumor-initiating cells. Computational modeling using primary human and mouse BrCa supports the interaction of anti-viral/NOTCH3 pathways in controlling NOTCH target genes and treatment resistance, particularly in basal subtype tumors. Gamma secretase inhibitors reverse stromal protection and abrogate radiation resistance in vivo. Thus, stroma orchestrates an intricate cross-talk with BrCa by utilizing exosomes to coax anti-viral signaling that expands therapy resistant cells through druggable pathways. Overall design: RNA profile of ceullar RNA and exosome of co-culture of breast caner cell line 1833 and stroma cell line MRC5.","project":"SRP046000"}
{"number_samples":8,"species":"human","abstract":"We devised a novel normalization approach employing chimpanzee RNA as a spike-in standard in NGS-based human gene expression analysis. In this approach, human transcripts were normalized read-by-read by spiked-in chimpanzee transcripts that were readily discriminated by natural single nucleotide variations (SNVs) between the two species. The SNV-normalized analysis enabled more accurate measurement of differential gene expression than an ordinary read-based approach.","project":"SRP046066"}
{"number_samples":24,"species":"human","abstract":"We report the application of RNA sequencing technology for high-throughput profiling of gene expression responses to human rhinovirus infection at 24 hours in air-liquid interface human airway epithelial cell cultures derived from 6 asthmatic and 6 non-asthmatic donors.  RNA-seq analysis identified sets of genes associated with asthma specific viral responses.  These genes are related to inflammatory pathways, epithelial remodeling and cilium assembly and function, including those described previously (e.g. CCL5, CXCL10 and CX3CL1), and novel ones that were identified for the first time in this study (e.g. CCRL1, CDHR3).   We concluded that air liquid interface cultured human airway epithelial cells challenged with live HRV are a useful in vitro model for the study of rhinovirus induced asthma exacerbation, given that our findings are consistent with clinical data sets.  Furthermore, our data suggest that abnormal airway epithelial structure and inflammatory signaling are important contributors to viral induced asthma exacerbation. Overall design: Differentiated air-liquid interface cultured human airway epithelial cell mRNA profiles from 6 asthmatic and 6 non-asthmatic donors after 24 hour treatment with either HRV or vehicle control were generated by deep sequencing, using Illumina HiSeq 2000.","project":"SRP046226"}
{"number_samples":14,"species":"human","abstract":"Myelodysplastic syndromes and chronic myelomonocytic leukemia (CMML) are characterized by mutations in epigenetic modifiers and aberrant DNA methylation. DNA methyltransferase inhibitors (DMTis) are used to treat these disorders, but response is highly variable with few means to predict which patients will benefit. To develop a molecular means of predicting response at diagnosis, we examined baseline differences in mutations, DNA methylation, and gene expression in 40 CMML patients responsive and resistant to decitabine (DAC). While somatic mutations did not differentiate responders and non-responders, we were able to identify for the first time 158 differentially methylated regions (DMRs) at baseline between responders and non-responders using next-generation sequencing. These DMRs were primarily localized to non-promoter regions and overlapped with distal regulatory enhancers. Using the methylation profiles, we developed an epigenetic classifier that accurately predicted DAC response at the time of diagnosis. We also found 53 differentially expressed genes between responders and non-responders. Genes up-regulated in responders were enriched in the cell cycle, potentially contributing to effective DAC incorporation. Two chemokines overexpressed in non-responders -- CXCL4 and CXCL7 -- were able to block the effect of DAC on normal CD34+ and primary CMML cells in vitro, suggesting their up-regulation contributes to primary DAC resistance. Overall design: mRNA profiling in bone marrow mononuclear cells (BM MNC) from 14 CMML patients (8 decitabine responders vs. 6 non-responders).","project":"SRP046233"}
{"number_samples":12,"species":"human","abstract":"We expressed either a wt or a phosphomutant version of ZFP36L1 in IMR90 ER:RAS cells. 7 days upon RAS induction (when the cells reach a fully senescent phenotype) we collected the RNA. ZFP36L1 is a RNA binding protein that binds to AU-rich elements in the 3’UTR of mRNAs and triggers their degradation. Our previous experiments showed that the activity of ZFP36L1 was key in the regulation of the senescent phenotype.By performing RNAseq we have uncovered the effect of expressing a constitutively active mutant of ZFP36L1 within the senescent transcriptome. Overall design: 4 samples examined: Non-senescent cells (EV - 4OHT), Senescent cells (EV + 4OHT), Senescent cells expressing ZFP36L1wt and Senescent cells expressing ZFP36L1mut","project":"SRP046254"}
{"number_samples":9,"species":"human","abstract":"Rescue of KLHL9 (WT) expression in KLHL9 null cell line, SF210, causes significant reprogramming away from a mesenchymal lineage. Overall design: An inducible expression vector carrying KLHL9 or GFP was stably introduced into SF210 cells via lentivirus transduction. Individual lines were isolated via clonal dilution and induced using doxycycline.","project":"SRP046266"}
{"number_samples":8,"species":"human","abstract":"We identified which mRNAs are dependent on the splicing factors SNW1 or PRPF8. These factors were depleted in HeLa cells by RNAi, and the levels of intronic reads in mRNAs was compared to control RNAi. Overall design: Intronic reads from polyA containing RNAs from HeLa cells and mSnw1-LAP 'rescued' cells, both depleted for endogenous SNW1, were compared. Furthermore, intronic reads from polyA containing RNAs from HeLa cells depleted for SNW1 or PRPF8 were compared to control depleted HeLa cells.","project":"SRP046271"}
{"number_samples":3,"species":"human","abstract":"Genome-wide methylome analyses reveal novel epigenetic regulation patterns in schizophrenia and bipolar disorder ","project":"SRP046293"}
{"number_samples":18,"species":"human","abstract":"The airway epithelial cell plays a central role in coordinating pulmonary response to injury and inflammation.  Here, transforming growth factor-b (TGFb) activates gene expression programs to induce stem cell-like properties, inhibit expression of differentiated epithelial adhesion proteins and express mesenchymal contractile proteins.  This process is known as epithelial mesenchymal transition (EMT); although much is known about the role of EMT in cellular metastasis in an oncogene-transformed cell, less is known about Type II EMT, that occurring in normal epithelial cells.  In this study, we applied next generation sequencing (RNA-seq) in primary human airway epithelial cells to understand the gene program controlling Type II EMT and how cytokine-induced inflammation modifies it. Generalized linear modeling was performed on a two-factor RNA-seq experiment of 6 treatments of telomerase immortalized human small airway epithelial cells (3 replicates).  Using a stringent cut-off, we identified 3,478 differentially expressed genes (DEGs) in response to EMT.  Unbiased transcription factor enrichment analysis identified three clusters of EMT regulators, one including SMADs/TP63 and another NF-kB/RelA.  Surprisingly, we also observed 527 of the EMT DEGs were also regulated by the TNF-NF-kB/RelA pathway.  This Type II EMT program was compared to Type III EMT in TGFb stimulated A549 alveolar lung cancer cells, revealing significant functional differences. Moreover, we observe that Type II EMT modifies the outcome of the TNF program, reducing IFN signaling and enhancing integrin signaling.  We confirmed experimentally that TGFb-induced the NF-kB/RelA pathway by observing a 2-fold change in NF-kB/RelA nuclear translocation.  A small molecule IKK inhibitor blocked TGFb-induced core transcription factor (SNAIL1, ZEB1 and Twist1) and mesenchymal gene (FN1 and VIM) expression.  These data indicate that NF-kB/RelA controls a SMAD-independent gene network whose regulation is required for initiation of Type II EMT.  Type II EMT dramatically affects the induction and kinetics of TNF-dependent gene networks. Overall design: A human small airway epithelial cell line was treated with TGF-Beta to induce the epithelial to mesenchymal transition. TGF-Beta treated and untreated cells were further treated with TNF-alpha for 1 and 12 hours. Three replicates for each treatment and untreated controls were performed for a total of 18 samples.","project":"SRP046376"}
{"number_samples":6,"species":"human","abstract":"We characterized expression profilings of SMYD2 knockdown and control cells treated by BIX-01294 and Rapamycin. Overall design: We used siRNAs to knockdown SMYD2 in HCT116 cells, and NC as control. We treated those cell lines with BIX-01294 and Rapamycin, and the expression profilings were perfomed by Illumina high throughput sequencing.","project":"SRP046741"}
{"number_samples":2,"species":"human","abstract":"Genome-wide mapping of transcriptional regulatory elements are essential tools for the understanding of the molecular events orchestrating self-renewal, commitment and differentiation of stem cells. We combined high-throughput identification of nascent, Pol-II-transcribed RNAs by Cap Analysis of Gene Expression (CAGE-Seq) with genome-wide profiling of histones modifications by chromatin immunoprecipitation (ChIP-seq) to map active promoters and enhancers in a model of human neural commitment, represented by embryonic stem cells (ESCs) induced to differentiate into self-renewing neuroepithelial-like stem cells (NESC). We integrated CAGE-seq, ChIP-seq and gene expression profiles to discover shared or cell-specific regulatory elements, transcription start sites and transcripts associated to the transition from pluripotent to neural-restricted stem cell. Our analysis showed that >90% of the promoters are in common between the two cell types, while approximately half of the enhancers are cell-specific and account for most of the epigenetic changes occurring during neural induction, and most likely for the modulation of the promoters to generate cell-specific gene expression programs. Interestingly, the majority of the promoters activated or up-regulated during neural induction have a “bivalent” histone modification signature in ESCs, suggesting that developmentally-regulated promoters are already poised for transcription in ESCs, which are apparently pre-committed to neuroectodermal differentiation. Overall, our study provide a collection of differentially used enhancers, promoters, transcription starts sites, protein-coding and non-coding RNAs in human ESCs and ESC-derived NESCs, and a broad, genome-wide description of promoter and enhancer usage and gene expression programs occurring in the transition from a pluripotent to a neural-restricted cell fate. Investiagtion of promoters usage changes during ESCs neural induction Overall design: ESCs and NESCs promoter usage profiling by CAGE-seq","project":"SRP046749"}
{"number_samples":13,"species":"human","abstract":"Ribosome profiling of MDA-MB-231 cells treated with Silvestrol to monitor transcriptome wide, eIF4A-dependent changes in translation efficiency Overall design: Translation efficiency (TE) of mRNAs dervied from ribosome footprints was monitored in the presence or absence of 25 nM Silvestrol, an inhibitor of eukaryotic translation initiation factor 4A (eIF4A).  Transcripts with reduced TE in the presence of Silvestrol were compare to transcripts with reduced TE in the presence of INK128, a catalytic mTOR inhbitor.","project":"SRP047065"}
{"number_samples":43,"species":"human","abstract":"Mitochondria are vital due to their principal role in energy production via oxidative phosphorylation (OXPHOS)1. Mitochondria carry their own genome (mtDNA) encoding  critical genes involved in OXPHOS, therefore, mtDNA mutations cause fatal or severely debilitating disorders with limited treatment options. 2. Clinical manifestations of mtDNA disease vary based on mutation type and heteroplasmy levels i.e. presence of mutant and normal mtDNA within each cell. 3,4. We evaluated therapeutic concepts of generating genetically corrected pluripotent stem cells for patients with mtDNA mutations. We initially generated multiple iPS cell lines from a patient with mitochondrial encephalomyopathy and stroke-like episodes (MELAS) caused by a heteroplasmic 3243A>G mutation and a patient with Leigh disease carrying a homoplasmic 8993T>G mutation (Leigh-iPS). Due to spontaneous mtDNA segregation in proliferating fibroblasts, isogenic MELAS iPS cell lines were recovered containing exclusively wild type (wt) mtDNA with normal metabolic function. As expected, all iPS cells from the patient with Leigh disease were affected. Using somatic cell nuclear transfer (SCNT; Leigh-NT1), we then simultaneously replaced mutated mtDNA and generated pluripotent stem cells from the Leigh patient fibroblasts. In addition to reversing to a normal 8993G>T, oocyte derived donor mtDNA (human haplotype D4a) in Leigh-NT1 differed from the original haplotype (F1a) at a additional 47 nucleotide sites. Leigh-NT1 cells displayed normal metabolic function compared to impaired oxygen consumption and ATP production in Leigh-iPS cells or parental fibroblasts (Leigh-fib). We conclude that natural segregation of heteroplasmic mtDNA allows the generation of iPS cells with exclusively wild type mtDNA. Moreover, SCNT offers mitochondrial gene replacement strategy for patients with homoplasmic mtDNA disease. Overall design: Duplicate cDNA libraries of fibroblasts from a Leigh patient and a MELAS patient, two sendai produced iPSC lines from the Leigh patient and three sendai produced iPSC lines from the MELAS patient, three fibroblasts lines produced by differentiating three iPS Leigh patient iPSC lines to fibroblasts, two somatic cell nuclear transfer produced NT-ESC lines from the Leigh patient, two fibroblast lines produced by differentiating two Leigh patient NT-ESC lines, four fibroblasts lines produced by differentiating four MELAS patient iPSC lines with the mutation to fibroblasts, four fibroblast lines produced by differentiating two IVF-ESC lines without mutated mtDNA genomes, four fibroblast lines produced by differentiating two somatic cell nuclear transfer NT-ESC lines without mutated mtDNA genomes, and four fibroblasts lines produced by differentiating two MELAS patient iPSC lines without the mutation to fibroblasts. The sequence reads were mapped to hg19 reference genome and hits that passed quality filters were analyzed for differential expression.","project":"SRP047082"}
{"number_samples":6,"species":"human","abstract":"In metazoans, the endoribonuclease SMG6 is thought to cleave many endogenous mRNAs targeted for nonsense-mediated mRNA decay (NMD). However, most evidence as to the identity of endogenous SMG6 substrates is indirect, and little is known about their cleavage sites. Here, we report the efficacy of an RNA degradome approach called parallel analysis of RNA ends (PARE) for identifying NMD intermediates in human cells. By specifically sequencing the 5’ ends of intermediates dependent on SMG6 and the critical NMD factor UPF1, hundreds of endogenous transcripts that are direct targets of SMG6 have been revealed. A preferred sequence motif spanning most SMG6 cleavage sites has been identified and validated by mutational analysis. For many SMG6 substrates, depletion of SMG6 leads to decapping of the RNAs. These findings provide key insights into NMD and targeting by SMG6 while also demonstrating the potential of PARE for analyzing other ribonucleases with diverse endogenous substrates. Overall design: Genome-wide degradome sequencing and mRNA profiling was done following the knockdown of SMG6 and UPF1.  SPARE libraries targeting a sequence motif surrounding the identified cleavage site were also deeply sequenced on an Illumina platform. PARE is a method of degradome analysis that captures and deeply sequences the 5' monophosphorylated ends of polyadenylated RNAs (see.German 2009 Nat Protoc 4, 356-362).  C-PARE libraries capture capped, polyadenlyated RNAs by first dephosphorylating the RNA, then decapping using tobacco acid pyrophosphatase and constructing PARE libraries, and will be described in the forthcoming paper.  SPARE captures the 5' monophosphorylated ends of specific RNAs using transcript-specific primers (see Schapire 3013 Methods 64:283-291).","project":"SRP047097"}
{"number_samples":1,"species":"human","abstract":"Recently a genome of Russian individual (somatic DNA from blood) was sequenced (Skryabin et al. 2009). That study was continued to find a linkage between genetic differences in parental alleles and bias in biallelic expression of genes.","project":"SRP047124"}
{"number_samples":4,"species":"human","abstract":"We performed whole-genome transcriptomic profiling of RNA from mononuclear cells from bone marrow aspirates taken from healthy individuals. This study complements GSE58335: transcriptomic profiling of peripheral blood mononuclear cells from healthy individuals. Overall design: High-throughput sequencing was done using the Illumina GA IIx. The RNA is from previously published samples (Stirewalt et al., Genes Chromosomes Cancer, 2008, PMID:17910043)","project":"SRP047126"}
{"number_samples":5,"species":"human","abstract":"Major features of transcription by human RNA polymerase II (Pol II) remain poorly defined due to a lack of quantitative approaches for visualizing Pol II progress at nucleotide resolution. We developed a simple and powerful approach for performing native elongating transcript sequencing (NET-seq) in human cells that globally maps strand-specific Pol II density at nucleotide resolution. NET-seq exposes a mode of antisense transcription that originates downstream and converges on transcription from the canonical promoter. Convergent transcription is associated with a distinctive chromatin configuration and is characteristic of lower-expressed genes. Integration of NET-seq with genomic footprinting data reveals stereotypic Pol II pausing coincident with transcription factor occupancy. Finally, exons retained in mature transcripts display Pol II pausing signatures that differ markedly from skipped exons, indicating an intrinsic capacity for Pol II to recognize exons with different processing fates. Overall design: 10 samples were analyzed. NET-seq data for HeLa S3 cells consisted in 2 WT biological replicates (HeLaS3_Rep1 and HeLaS3_Rep2), 1 treatment condition (HeLaS3_FP) and 1 control for the treatment condition (HeLaS3_DMSO). NET-seq data for HEK293T cells consisted in 2 WT biological replicates (HEK293T_Rep1 and HEK293T_Rep2). RNA-seq data for HeLa S3 cells consisted in the 3 cellular fractions in WT condition (HeLaS3_chromatin_RNAseq, HeLaS3_nucleoplasm_RNAseq and HeLaS3_cytoplasm_RNAseq). RNA-seq data for HEK 293T cells consisted in the cytoplasmic fraction in WT condition (HEK293T_cytoplasm_RNAseq)","project":"SRP047174"}
{"number_samples":60,"species":"human","abstract":"We compared the performance of conventional RNAseq with RNA Capture Sequencing (CaptureSeq) to assemble and quantify known RNA spike-Ins and human transcripts. We find CaptureSeq to be superior for the detection and quantification of the 37% lowest expressed genes, and comparable for the next 45% of moderately expressed genes. CaptureSeq contributes only minor technical variation and measures differential gene expression accurately. We demonstrate these advantages by the targeted sequencing of long noncoding RNAs across 20 human tissues, expanding previous annotations two-fold and simultaneously generating a quantitative atlas of expression. This analysis confirms the use of CaptureSeq as an important method for transcriptional profiling. Overall design: Long noncoding RNA assembly and expression is analysed by targeted RNA sequencing for 20 human tissues and 4 human cell lines","project":"SRP047192"}
{"number_samples":48,"species":"human","abstract":"Autism spectrum disorder (ASD) is a disorder of brain development believed, in most cases, to be of genetic origin. We use induced pluripotent stem cells (iPSCs)-derived 3-dimensional neural cultures (organoids) in patients with ASD and macrocephaly to investigate neurodevelopmental alterations that cause this form of ASD. By using transcriptome analyses, we identified modules of co-expressed genes significantly upregulated in ASD patients compared to non-ASD first-degree family members. Overall design: Total RNA was prepared from terminal differentiation day 0, 11 and 31 of iPSCs-derived neural cultures from ASD patients and non-ASD first-degree family members. A total of 4 patients and 8 controls (unaffected family members) were analyzed in replicates (two to three iPSC clones per person).","project":"SRP047194"}
{"number_samples":54,"species":"human","abstract":"Truncating mutations of CHD8, encoding a chromodomain helicase, and of many other genes with diverse functions, are strong-effect risk factors for autism spectrum disorder (ASD), suggesting multiple mechanisms of pathogenesis. We explored the transcriptional networks that CHD8 regulates in neural progenitor cells (NPCs) by reducing its expression and then integrating transcriptome sequencing (RNA-seq) with genome-wide CHD8 binding (ChIP-seq). Suppressing CHD8 to levels comparable with loss of a single allele caused altered expression of 1,756 genes, 64.9% of which were up-regulated.  CHD8 showed widespread binding to chromatin, with 7,324 replicated sites that marked 5,658 genes. Integration of these data suggests that a limited array of direct regulatory effects of CHD8 produced a much larger network of secondary expression changes. Genes indirectly down-regulated (i.e., without CHD8 binding sites) reflect pathways involved in brain development, including synapse formation, neuron differentiation, cell adhesion, and axon guidance, whereas CHD8-bound genes are strongly associated with chromatin modification and transcriptional regulation. Genes associated with ASD were strongly enriched among indirectly down-regulated loci (p = 1.01x10-9) and CHD8-bound genes (p = 4.34x10-3), which align with previously identified co-expression modules during fetal development. We also find an intriguing enrichment of cancer related gene-sets among CHD8-bound genes (p < 1.9x10-11). In vivo suppression of chd8 in zebrafish produced macrocephaly comparable to that of humans with inactivating mutations. These data indicate that heterozygous disruption of CHD8 precipitates a network of gene expression changes involved in neurodevelopmental pathways in which many ASD-associated genes may converge on shared mechanisms of pathogenesis. Overall design: RNA-seq in NPCs treated with shRNAs targeting CHD8. For controls, NPCs were treated with shRNAs targeting GFP and LacZ. Infection and sequencing was carried out in two separate batches, with one GFP and one LacZ sample in each batch. All samples were sequenced in two technical replicates.","project":"SRP047233"}
{"number_samples":1,"species":"human","abstract":"To investigated a detailed analysis of the transcriptional response between epithelial Caco-2 cells with different Bifidobacterium animals subsp. lactis KLDS 2.0603 cells ( Control and cells treated by digestive tract fluids simulated ).","project":"SRP047266"}
{"number_samples":9,"species":"human","abstract":"Diabetes is one of the leading causes of blindness in the western world. TNFa is a known regulator of diabetic retinopathy pathology. This study was performed to determine the effect of TNFa on retinal microvascular endothelial cells and whether NFAT signaling has a role in mediating this effect.","project":"SRP047271"}
{"number_samples":14,"species":"human","abstract":"Increased MITF expression contributes to melanoma progression and resistance to BRAF pathway inhibition. We show that, unexpectedly, lack of MITF is associated with more severe resistance to a range of inhibitors. Indeed, the presence of endogenous MITF was essential for robust drug responses. Both in primary and acquired resistance, MITF levels inversely correlated with expression of several activated receptor tyrosine kinases, most commonly AXL. The MITF-low/AXL-high/drug resistance phenotype was seen in roughly half of BRAF mutant and the majority of NRAS mutant melanoma cell lines. The dichotomous behavior of MITF in drug response was corroborated in vemurafenib-resistant biopsies, including MITF high and low clones in a relapsed patient. Drug cocktails containing AXL inhibitor enhanced melanoma cell elimination by BRAF or ERK inhibition. Our results demonstrate that a low MITF/AXL ratio predicts early resistance to multiple targeted drugs, and warrant clinical validation of AXL inhibitors to combat resistance of BRAF and NRAS mutant MITF-low melanomas. Overall design: Experssion analysis by RNAseq of 14 melanoma cell lines.","project":"SRP047299"}
{"number_samples":3,"species":"human","abstract":"RNA-Seq study from Caco2 cells following transduction with control shRNA and shRNA against CHD6","project":"SRP047307"}
{"number_samples":9,"species":"human","abstract":"Burst-forming unit-erythroid (BFU-E) and colony-forming unit-erythroid (CFU-E) cells are erythroid progenitors traditionally defined by colony assays. We developed a flow cytometry-based strategy for isolating human BFU-E and CFU-E cells based on the changes in expression of cell surface markers during in vitro erythroid cell culture. BFU-E and CFU-E are characterized by CD45+GPA-IL-3R-CD34+CD36-CD71low and CD45+GPA-IL-3R-CD34-CD36+CD71high phenotypes, respectively. Colony assays validated phenotypic assignment giving rise to BFU-E and CFU-E colonies, both at a purity ~90%. The BFU-E colony forming ability of CD45+GPA-IL-3R-CD34+CD36-CD71low cells required SCF and erythropoietin, while the CFU-E colony forming ability of CD45+GPA-IL-3R-CD34-CD36+CD71high cells required only erythropoietin. Bioinformatic analysis of the RNA-seq data revealed unique transcriptomes in each differentiation stage. The sorting strategy was validated in uncultured primary cells isolated from bone marrow and peripheral blood, indicating that marker expression is not an artifact of in vitro cell culture, but represents an in vivo characteristic of erythroid progenitor populations. The ability to isolate highly pure human BFU-E and CFU-E progenitors will enable detailed cellular and molecular characterization of these distinct progenitor populations and define their contribution to disordered erythropoiesis in inherited and acquired hematological disease. Our data provide important resource for future studies. Overall design: Transcription profiles of Human erythroid progenitors at distinct developmental stages were generated by deep sequencing, in triplicate, using IlluminaHiSeq 2000. The complete dataset comprises 4 sample types: CD34, BFU, CFU, and Pro (reanalysis of GSM1304777-GSM1304779).","project":"SRP047323"}
{"number_samples":6,"species":"human","abstract":"Recent studies have identified a Lys 27-to-methionine (K27M) mutation at one allele of H3F3A, one of the two genes encoding histone H3 variant H3.3, in 60% of high-grade pediatric glioma cases. The median survival of this group of patients after diagnosis is ~1 yr. Here we show that the levels of H3K27 di- and trimethylation (H3K27me2 and H3K27me3) are reduced globally in H3.3K27M patient samples due to the expression of the H3.3K27M mutant allele. Remarkably, we also observed that H3K27me3 and Ezh2 (the catalytic subunit of H3K27 methyltransferase) at chromatin are dramatically increased locally at hundreds of gene loci in H3.3K27M patient cells. Moreover, the gain of H3K27me3 and Ezh2 at gene promoters alters the expression of genes that are associated with various cancer pathways. These results indicate that H3.3K27M mutation reprograms epigenetic landscape and gene expression, which may drive tumorigenesis. Overall design: We performed chromatin-immunoprecipitation of H3K27me3, H3K4me3, and EZH2 in SF7761 and NSC cell lines. And do RNA-seq in SF7761, SF8828 and NSC cell lines. SF7761 and SF8628 cell lines from patients harboring the histone H3.3 K27M mutation were obtained from Hashizume et al. (2012). NSCs (N7800-100) were purchased from Invitrogen and cultured and maintained in NSC medium (A10509-01, StemPro NSC SFM, Invitrogen).","project":"SRP047339"}
{"number_samples":9,"species":"human","abstract":"Whole transcriptome study on lung cancer","project":"SRP047358"}
{"number_samples":4,"species":"human","abstract":"Fibroblasts from PRDM12 patients and unaffected wildtype relatives were cultured until near confluency. The transcriptional profile of those cells was determined by mRNA sequencing and uncovered differential expression in several known pain and neurodevelopmental genes. Overall design: Transcriptome comparison of human PRDM12 mutant and wildtype fibroblasts","project":"SRP047363"}
{"number_samples":2,"species":"human","abstract":"The U2AF heterodimer has been well studied for its role in defining functional 3’ splice sites in pre-mRNA splicing, but many fundamental questions still remain unaddressed regarding the function of U2AF in mammalian genomes. Through genome-wide analysis of U2AF-RNA interactions, we report that U2AF has the capacity to directly define ~88% of functional 3’ splice sites in the human genome, but numerous U2AF binding events also occur in intronic locations. Mechanistic dissection reveals that upstream intronic binding events interfere with the immediate downstream 3’ splice site associated with either the alternative exon to cause exon skipping or with the competing constitutive exon to induce exon inclusion. We further demonstrate partial functional impairment with mutations in U2AF35, but not U2AF65, in regulated splicing. These findings reveal the genomic function and regulatory mechanism of U2AF in both normal and disease states. Overall design: Examination of U2AF heterodimer regulated splicing in Hela cells with CLIP-seq (U2AF65), paired-end RNA-seq (si-NC and si-U2AF65) and RASL-seq (respective three biological replicates of WT, si-NC, si-U2AF65, si-U2AF35, si-NC + pcDNA3.0, si-U2AF65 + pcDNA3.0, and si-U2AF65 + Flag-U2AF35)","project":"SRP047366"}
{"number_samples":4,"species":"human","abstract":"Primary human bronchial epithelial cells were transduced with control or hYAP(S127A) lentivirus in sphere forming conditions. Bronchospheres were harvested on day 18-20 for RNAseq analysis Overall design: Passage 1 Primary HBECs from 2 independent donors were transduced with control or hYAP lentivirus. 48 hours post infection, cells were plated on transwell inserts in a 50-50 mixture of ALI medium-Cultrex BME reduced growth factor (RGF) to form spheres. Well differentiated bronchospheres were harvested for RNA-seq analysis on day 18-20 by combining 3 wells of each group for each donor.","project":"SRP047383"}
{"number_samples":9,"species":"human","abstract":"Whole transcriptome sequencing of lung cancer specimens","project":"SRP047399"}
{"number_samples":15,"species":"human","abstract":"Human pluripotent stem cells can be derived from somatic cells by forced expression of defined factors, and more recently by nuclear-transfer into human oocytes, revitalizing a debate on whether one reprogramming approach might be advantageous over the other. Here we compared the genetic and epigenetic stability of human nuclear-transfer embryonic stem cell (NT-ESC) lines and isogenic induced pluripotent stem cell (iPSC) lines, derived from the same somatic cell cultures of fetal, neonatal and adult origin. Both cell types shared similar genome-wide gene expression and DNA methylation profiles. Importantly, NT-ESCs and iPSCs have comparable numbers of de novo coding mutations but significantly higher than parthenogenetic ESCs. Similar to iPSCs NT-ESCs displayed clone- and gene-specific aberrations in DNA methylation and allele-specific expression of imprinted genes, similarly to iPSCs. The occurrence of these genetic and epigenetic defects in both NT-ESCs and iPSCs suggests that they are inherent to reprogramming, regardless of the underlying technique. Overall design: RNA sequencing analysis was performed on a total of 12 human cell lines, including: an isogenic set of 3 nuclear-transfer embryonic stem cell (NT-ESC) lines, 2 RNA-reprogrammed induced pluripotent stem cell (iPSC) lines and their parental neonatal fibroblast cell line; an isogenic set of 1 NT-ESC line, 3 iPSC lines and their parental adult fibroblast cell line (derived from a type 1 diabetic subject); as well as 1 control embryonic stem cell (ESC) line.","project":"SRP047407"}
{"number_samples":6,"species":"human","abstract":"Human embryonic stem cells were differentiated into peripheral sensory neurons via the intermediate generation of neural crest like cell (NCC). Using various markers we identified these cells as LTMR. We then analyzed there complete transcriptional profile in comparison to the intermediate neural crest like cells. Overall design: mRNA expression data of human ESC-derived sensory neuron clusters (10-20 cells) and human ESC-derived neural crest like cells (~100 cells) was generated by illumina deep sequencing","project":"SRP047422"}
{"number_samples":10,"species":"human","abstract":"The suitability of small RNA sequencing on the Illumina® HiSeq™ platform for quantification of small RNAs in formalin-fixed and paraffin embedded tissue was assessed in this project.","project":"SRP047429"}
{"number_samples":8,"species":"human","abstract":"Breast cancer is genetically heterogeneous, and recent studies have underlined a prominent contribution of epigenetics to the   development of this disease. To uncover new synthetic lethalities with known breast cancer oncogenes, we screened an  epigenome-focused short hairpin RNA library on a panel of engineered breast epithelial cell lines. Here we report a selective  interaction between the NOTCH1 signaling pathway and the SUMOylation cascade. Knockdown of the E2-conjugating enzyme  UBC9 (UBE2I) as well as inhibition of the E1-activating complex SAE1/UBA2 using ginkgolic acid impairs the growth of NOTCH1-activated breast epithelial cells. We show that upon inhibition of SUMOylation NOTCH1-activated cells proceed slower through the  cell cycle and ultimately enter apoptosis. Mechanistically, activation of NOTCH1 signaling depletes the pool of unconjugated small  ubiquitin-like modifier 1 (SUMO1) and SUMO2/3 leading to increased sensitivity to perturbation of the SUMOylation cascade.  Depletion of unconjugated SUMO correlates with sensitivity to inhibition of SUMOylation also in patient-derived breast cancer cell  lines with constitutive NOTCH pathway activation. Our investigation suggests that SUMOylation cascade inhibitors should be  further explored as targeted treatment for NOTCH-driven breast cancer. Overall design: We treated MCF10A and NOTCH1 cells with either DMSO or ginkgolic acid 30 uM for 3 days. Two replicates have been analysed for each condition.","project":"SRP047459"}
{"number_samples":72,"species":"human","abstract":"Genetic variants that impact gene regulation are important contributors to human phenotypic variation. For this reason, considerable efforts have been made to identify genetic associations with differences in mRNA levels of nearby genes, namely, cis expression quantitative trait loci (eQTLs). The phenotypic consequences of eQTLs are presumably due, in most cases, to their ultimate effects on protein expression levels. Yet, only few studies have quantified the impact of genetic variation on proteins levels directly. It remains unclear how faithfully eQTLs are reflected at the protein level, and whether there is a significant layer of cis regulatory variation acting primarily on translation or steady state protein levels. To address these questions, we measured ribosome occupancy by high-throughput sequencing, and relative protein levels by high-resolution quantitative mass spectrometry, in a panel of lymphoblastoid cell lines (LCLs) in which we had previously measured transcript expression using RNA sequencing. We then mapped genetic variants that are associated with changes in transcript expression (eQTLs), ribosome occupancy (rQTLs), or protein abundance (pQTLs). Most of the QTLs we detected are associated with transcript expression levels, with consequent effects on ribosome and protein levels. However, we found that eQTLs tend to have significantly reduced effect sizes on protein levels, suggesting that their potential impact on downstream phenotypes is often attenuated or buffered. Additionally, we confirmed the presence of a class of cis QTLs that specifically affect protein abundance with little or no effect on mRNA levels; most of these QTLs have little effect on ribosome occupancy, and hence may arise from differences in post-translational regulation. Overall design: We measured level of translation transcriptome-wide in lymphoblastoid cell lines derived from 72 HapMap Yoruba individuals using ribosome profiling assay, for which we have transcript level, protein level (62 out of 72) and genotype information collected.","project":"SRP047476"}
{"number_samples":1,"species":"human","abstract":"The human HEK293 / 293T and rat cardiomyoblast H9c2 cell lines are commonly employed for microRNA-mRNA interaction studies. Here, I provide microRNA sequencing data obtained from each of these lines to better document which microRNAs are endogenously expressed at high or low levels. Overall design: Small RNA sequencing profiles were generated from cultured HEK293 and H9c2 cells on Illumina HiSeq 2000 instruments.","project":"SRP047494"}
{"number_samples":8,"species":"human","abstract":"Our data throw light upon the effect of PTEN deficiency on gene expression, epigenomic modifications(H3K4me3, H3K27me3, 5-Hydroxymethylcytosine and 5-methylcytosin). Overall design: RNA-seq of NSCs-PTEN+/+ and NSCs-PTEN-/-; RRBS and TA-RRBS of NSCs-PTEN+/+, NSCs-PTEN-/-; H3K4me3 ChIP-seq of NSCs-PTEN+/+ and NSCs-PTEN-/-; H3K27me3 ChIP-seq of NSCs-PTEN+/+ and NSCs-PTEN-/-.","project":"SRP047516"}
{"number_samples":2,"species":"human","abstract":"We have found aerobic glycolysis is increased in cancer-associated fibroblasts (CAF). To explore the mechanism by which glucolysis was switched from oxidative phosphorylation, two in vitro CAF models were successfully established. To screen genes potentially regulate aerobic glysolysis, we analyzed gene expression profiling of CAF, and compared with control fibroblasts. Overall design: Primary fibroblasts separately treated with PDGF or TGF-B for 96 hours to induce CAF formation, control fibroblast were treated with 1% BSA. mRNA profiling were analyzed by deep sequencing.","project":"SRP047519"}
{"number_samples":8,"species":"human","abstract":"DNase I hypersensitive sites (DHSs) provide important information on the presence of transcriptional regulatory elements and the state of chromatin in mammalian cells1, 2, 3. Conventional DNase sequencing (DNase-seq) for genome-wide DHSs profiling is limited by the requirement of millions of cells4, 5. Here we report an ultrasensitive strategy, called single-cell DNase sequencing (scDNase-seq) for detection of genome-wide DHSs in single cells. We show that DHS patterns at the single-cell level are highly reproducible among individual cells. Among different single cells, highly expressed gene promoters and enhancers associated with multiple active histone modifications display constitutive DHS whereas chromatin regions with fewer histone modifications exhibit high variation of DHS. Furthermore, the single-cell DHSs predict enhancers that regulate cell-specific gene expression programs and the cell-to-cell variations of DHS are predictive of gene expression. Finally, we apply scDNase-seq to pools of tumour cells and pools of normal cells, dissected from formalin-fixed paraffin-embedded tissue slides from patients with thyroid cancer, and detect thousands of tumour-specific DHSs. Many of these DHSs are associated with promoters and enhancers critically involved in cancer development. Analysis of the DHS sequences uncovers one mutation (chr18: 52417839G>C) in the tumour cells of a patient with follicular thyroid carcinoma, which affects the binding of the tumour suppressor protein p53 and correlates with decreased expression of its target gene TXNL1. In conclusion, scDNase-seq can reliably detect DHSs in single cells, greatly extending the range of applications of DHS analysis both for basic and for translational research, and may provide critical information for personalized medicine. Overall design: Exploring the landscape of chromatin accessibility in single cells and clinical samples","project":"SRP048222"}
{"number_samples":20,"species":"human","abstract":"RNA sequencing of lung adenocarcinoma, lung squamous carcinoma and lung large cell cancer samples","project":"SRP048484"}
{"number_samples":4,"species":"human","abstract":"CD24 is the one of cell surface protein that anchored via glycosyl phosphatidylinositol that links to cell surface and modulates growth and differentiation signal many of cell types. A small population of cells, namely ‘Cancer Stem Cells (CSCs)’, charges highly aggressive and metastatic character of cancer cells and shows conformational change of ‘epithelial to mesenchymal transition (EMT). To understand the role of CD24 in CSC and EMT, CD24 was knocked down using siRNA in the MCF7 cell line (MCF-7 hCD24 KD) and analyzed transcriptome profiles by mRNA-sequencing. Corresponding author: Chul Geun Kim, Department of Life Science, Hanyang University, Seoul 133-791, Korea (e-mail, cgkim@hanyang.ac.kr). Overall design: mRNA profiles of MCF-7 hCD24 KD cells [‘monolayer (m) cultured MCF7 wild type (WT) and mMCF7 CD24KD’ and ‘sphere-forming (s) MCF7 WT and sMCF7 CD24KD’] were generated by deep sequencing using Illumina GAIIx.","project":"SRP048536"}
{"number_samples":2,"species":"human","abstract":"To characterize the transcriptional programs that underlie pancreas differentiation and identity, we have generated genome-scale expression profiles by RNA-seq from human embryonic stem cell derived liver progenitors and human fetal pancreatic tissue (days 54-57 post conception).  These samples were compared to those already published transcriptomes (Xie et al., 2013).  Together, these samples were used to perform principles compotent analysis.  Once this was performed, we were able to identify transcription factors that were important in the identity of each cell type. Overall design: To generate genome-scale expression profiles by RNA-seq from human embryonic stem cell derived liver progenitors and human fetal pancreatic tissue (days 54-57 post conception), total RNA was isolated from human embryonic stem cell derived liver progenitors and frozen human fetal pancreas.  Libraries were sequenced and mapped to the hg19 version of the human genome.  Gene expression was determined using Sailfish. These samples were compared to those already published transcriptomes (Xie et al., 2013).  Together, these samples were used to perform principles compotent analysis.","project":"SRP048556"}
{"number_samples":2,"species":"human","abstract":"To characterize the transcriptional programs that underlie pancreas differentiation and identity, we have generated genome-scale expression profiles by RNA-seq from human embryonic stem cell derived liver progenitors and human fetal pancreatic tissue (days 54-57 post conception).  These samples were compared to those already published transcriptomes (Xie et al., 2013).  Together, these samples were used to perform principles compotent analysis.  Once this was performed, we were able to identify transcription factors that were important in the identity of each cell type. Overall design: To generate genome-scale expression profiles by RNA-seq from human embryonic stem cell derived liver progenitors, total RNA was isolated from human embryonic stem cell derived liver progenitors.  Libraries were sequenced and mapped to the hg19 version of the human genome.   Gene expression was determined using Sailfish. These samples were compared to those already published transcriptomes (Xie et al., 2013).  Together, these samples were used to perform principles compotent analysis.","project":"SRP048557"}
{"number_samples":6,"species":"human","abstract":"We show that EWS-FLI1, an aberrant transcription factor responsible for the pathogenesis of Ewing sarcoma, reprograms gene regulatory circuits by directly inducing or directly repressing enhancers. At GGAA repeats, which lack regulatory potential in other cell types and are not evolutionarily conserved, EWS- FLI1 multimers potently induce chromatin opening, recruit p300 and WDR5, and create de novo enhancers. GGAA repeat enhancers can loop to physically interact with target promoters, as demonstrated by chromosome conformation capture assays. Conversely, EWS-FLI1 inactivates conserved enhancers containing canonical ETS motifs by displacing wild-type ETS transcription factors and abrogating p300 recruitment. Overall design: Ewing sarcoma cell lines (A673 and SKNMC) were analyzed by RNA-seq. EWS-FLI1 was depleted by infection with lentiviral shRNAs (shFLI1 and shGFP control).","project":"SRP048562"}
{"number_samples":19,"species":"human","abstract":"Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. By tandem affinity purification and mass spectrometry, we present a comprehensive characterisation of the MITF interactome comprising multiple novel cofactors involved in transcription, DNA replication and repair and chromatin organisation, including a BRG1 chromatin remodelling complex comprising CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. MITF, SOX10 and YY1 bind between two BRG1-occupied nucleosomes thus defining both a combinatorial signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of MAREs. Nevertheless, BRG1 silencing enhances MITF occupancy at MAREs showing that BRG1 acts to promote dynamic MITF interactions with chromatin. Overall design: 19 samples corresponding to mRNA profiles of 501Mel and Hermes3A after MITF, BRG1 or control shRNA-mediated knockdown were generated by deep sequencing in triplicate (in duplicate for 501_shMITF and corresponding control 501_shSCR2), using HiSeq2500.","project":"SRP048577"}
{"number_samples":12,"species":"human","abstract":"Genetic studies in T-cell acute lymphoblastic leukemia have uncovered a remarkable complexity of oncogenic and loss-of-function mutations. Amongst this plethora of genetic changes, NOTCH1 activating mutations stand out as the most frequently occurring genetic defect, identified in more than 50% of T-cell acute lymphoblastic leukemias, supporting an essential driver role for this gene in T-cell acute lymphoblastic leukemia oncogenesis. In this study, we aimed to establish a comprehensive compendium of the long non-coding RNA transcriptome under control of Notch signaling. For this purpose, we measured the transcriptional response of all protein coding genes and long non-coding RNAs upon pharmacological Notch inhibition in the human T-cell acute lymphoblastic leukemia cell line CUTLL1 using RNA-sequencing. Similar Notch dependent profiles were established for normal human CD34+ thymic T-cell progenitors exposed to Notch signaling activity in vivo. In addition, we generated long non-coding RNA expression profiles (array data) from GSI treated T-ALL cell lines, ex vivo isolated Notch active CD34+ and Notch inactive CD4+CD8+ thymocytes and from a primary cohort of 15 T-cell acute lymphoblastic leukemia patients with known NOTCH1 mutation status. Integration of these expression datasets with publically available Notch1 ChIP-sequencing data resulted in the identification of long non-coding RNAs directly regulated by Notch activity in normal and malignant T-cell context. Given the central role of Notch in T-cell acute lymphoblastic leukemia oncogenesis, these data pave the way towards development of novel therapeutic strategies that target hyperactive Notch1 signaling in human T-cell acute lymphoblastic leukemia. Overall design: CUTLL1 cell lines were treated with Compound E (GSI) or DMSO (solvent control). Cells were collected 12 h and 48 h after treatment. This was performed for 3 replicates. RNA-sequencing was performed on these samples.","project":"SRP048603"}
{"number_samples":4,"species":"human","abstract":"Genetic studies in T-cell acute lymphoblastic leukemia have uncovered a remarkable complexity of oncogenic and loss-of-function mutations. Amongst this plethora of genetic changes, NOTCH1 activating mutations stand out as the most frequently occurring genetic defect, identified in more than 50% of T-cell acute lymphoblastic leukemias, supporting an essential driver role for this gene in T-cell acute lymphoblastic leukemia oncogenesis. In this study, we aimed to establish a comprehensive compendium of the long non-coding RNA transcriptome under control of Notch signaling. For this purpose, we measured the transcriptional response of all protein coding genes and long non-coding RNAs upon pharmacological Notch inhibition in the human T-cell acute lymphoblastic leukemia cell line CUTLL1 using RNA-sequencing. Similar Notch dependent profiles were established for normal human CD34+ thymic T-cell progenitors exposed to Notch signaling activity in vivo. In addition, we generated long non-coding RNA expression profiles (array data) from GSI treated T-ALL cell lines, ex vivo isolated Notch active CD34+ and Notch inactive CD4+CD8+ thymocytes and from a primary cohort of 15 T-cell acute lymphoblastic leukemia patients with known NOTCH1 mutation status. Integration of these expression datasets with publically available Notch1 ChIP-sequencing data resulted in the identification of long non-coding RNAs directly regulated by Notch activity in normal and malignant T-cell context. Given the central role of Notch in T-cell acute lymphoblastic leukemia oncogenesis, these data pave the way towards development of novel therapeutic strategies that target hyperactive Notch1 signaling in human T-cell acute lymphoblastic leukemia. Overall design: CD34+ cells of 2 healthy donors are cultured on a OP9-GFP or OP9-DLL1 feeder layer.","project":"SRP048604"}
{"number_samples":8,"species":"human","abstract":"Here we report the discovery of highly potent and selective EZH2 small molecule inhibitors, their validation by a cellular thermal shift assay, their application across a large lymphoma cell panel and their efficacy in GCBDLBCL xenograft models. Overall design: RNA-seq of KARPAS-422 cell line RNA, in duplicate, treated with DMSO as control, and EZH2 inhibitors CPI360, EPZ-6438 and GSK126. Eight samples in total.","project":"SRP048640"}
{"number_samples":3,"species":"human","abstract":"The antibody gene mutator AID promiscuously damages oncogenes and B cell identity genes leading to chromosomal translocations and tumorigenesis. Why non-immunoglobulin loci are susceptible to AID activity is unknown. Here we study AID-mediated lesions in the context of nuclear architecture and the B cell regulome. We show that AID targets are not randomly distributed across the genome, but are predominantly clustered within super-enhancers. Unexpectedly, in these domains AID deaminates highly active promoters and eRNA+ enhancers interconnected in some instances over megabases of linear chromatin. Using genome editing we demonstrate that 3D-linked targets cooperate to recruit AID-mediated breaks. Furthermore, a comparison of hypermutation in mouse B cells, AID-induced kataegis in human lymphomas, and translocations in MEFs reveals that AID damages different genes in different cell types. Yet, in all cases, the targets are predominantly associated with topological complex, highly transcribed super-enhancers, demonstrating that these compartments are key mediators of AID recruitment. Overall design: Examination of AID activity in human cells via Single Nucleotide Variant discovery in H3K4me3 ChIP-seq data from 26 MSH2-/-; AIDtg; UGItg, 18 AICDA-/- and 2 unmodified RAMOS clonal populations.  Examination of PolII mediated long-range interactions via Chia-PET of RAMOS cells (2 sample).  Identification of super-enhancers from H3K27Ac ChIP-Seq data from activated B cells (3 replicates and 1 input control) and RAMOS cells (1 sample and 1 input control), 2 preparations of naive and 2 of germinal center (GC) B cells from human tonsilectomy samples.  Mapping of regulatory elements in RAMOS based on H3K4me1 (1 sample) and Nipbl (2 replicates) ChIP-Seq.  RNA expression analyses of activated B cells from 3 WT and 3 Il4raU/U mice and RAMOS cells (3 replicates).  Mapping of long-range interactions by 4C in activated B cells from a WT and an Il4raU/U mouse with the IL4ra and Il21r locus, respectively, as a viewpoint.  Mapping of Super-Enhancers in activated B cells from Il4raU/U and WT control mice (2 samples).","project":"SRP048660"}
{"number_samples":7,"species":"human","abstract":"We compared untreated HCC1419 cells with Lapatinib resistant HCC1419 cells that were either treated with Lapatinib for only 9 days before harvesting (drug tolerant persisters, DTPs) or were growing in the presence of Lapatinib (>70 days) (drug tolerant expanded persisters, DTEPs). We show that the Notch pathway is significantly over-expressed in DTEPs when compared to untreated cells. Overall design: RNA-Seq of HCC1419 cells either untreated, treated with 1µM Lapatinib for 9 days or treated with 1µM Lapatinib for greater than 70 days","project":"SRP048664"}
{"number_samples":6,"species":"human","abstract":"The activation of cellular quality control pathways to maintain metabolic homeostasis and mitigate diverse cellular stresses is emerging as a critical growth and survival mechanism in many cancers. Autophagy, a highly conserved cellular self-degradative process, is a key player in the initiation and maintenance of pancreatic ductal adenocarcinoma (PDA). However, the regulatory circuits that activate autophagy, and how they enable reprogramming of PDA cell metabolism are unknown. We now show that autophagy regulation in PDA occurs as part of a broader program that coordinates activation of lysosome biogenesis, function and nutrient scavenging, through constitutive activation of the MiT/TFE family of bHLH transcription factors. In PDA cells, the MiT/TFE proteins - MITF, TFE3 and TFEB - override a regulatory mechanism that controls their nuclear translocation, resulting in their constitutive activation. By orchestrating the expression of a coherent network of genes that induce high levels of lysosomal catabolic function, the MiT/TFE factors are required for proliferation and tumorigenicity of PDA cells. Importantly, unbiased global metabolite profiling reveals that MiT/TFE-dependent autophagy-lysosomal activation is specifically required to maintain intracellular AA pools in PDA.  This AA flux is part of a program that is essential for metabolic homeostasis and bioenergetics of PDA but not for their non-transformed counterparts. These results identify the MiT/TFE transcription factors as master regulators of the autophagy-lysosomal system in PDA and demonstrate a central role of the autophagosome-lysosome compartment in maintaining tumor cell metabolism through alternative amino acid acquisition and utilization. Overall design: Examination of mRNA levels in pancreatic ductal adenocarcinoma (PDA) cell line 8988T after treatment with siRNA for control or TFE3","project":"SRP048669"}
{"number_samples":36,"species":"human","abstract":"We used mRNA-seq to profile transcriptomes of purified human ovarian granulosa cells Overall design: 12 female individuals","project":"SRP048674"}
{"number_samples":6,"species":"human","abstract":"The non-small cell lung cancer (NSCLC) cell line HCC827 harbors an activating EGFR mutation (exon 19 deletion) that confers sensitivity to the FDA-approved EGFR inhibitor erlotinib. By applying the ClonTracer barcoding system, we were able to show the presence of pre-existing sub-populations in HCC827 that contribute to erlotinib resistance. Prior studies implicated that MET amplification confers resistance to erlotinib in this cell line. Therefore we examined the effects of the c-Met inhibitor crizotinib on the barcoded HCC827 population when treated either sequentially or simultaneously with both inhibitors. Despite the significant reduction in barcode complexity, the erlotinib/crizotinib combination treatment failed to eradicate all of the resistant clones implying the presence of an erlotinib/crizotinib dual resistant subpopulation. We performed transcriptome profiling (RNA-seq) to elucidate the potential resistance mechanisms of the dual resistant subpopulation in comparison to vehicle-treated or single agent erlotinib-resistant HCC827 cell populations as controls. Overall design: mRNA profiling of the subpopulations of human NSCLC cell line HCC827 that contribute to EGFR inhibitor erlotinib and MET inhibitor crizotinib resistance","project":"SRP048700"}
{"number_samples":16,"species":"human","abstract":"KCL-22 is a chronic myeloid leukemia (CML) cell line derived from a patient in blast crisis phase and harbors the BCR-ABL translocation. The catalytic (ATP-competitive) BCR-ABL inhibitors imatinib and nilotinib have dramatically improved CML patient outcome, but the development of resistance remains a clinical challenge. The recent identification of allosteric BCR-ABL inhibitors, such as GNF-2, which target the enzyme by binding to the myristoyl pocket rather than catalytic site of ABL1, may provide a strategy to broadly overcome resistance to the class of ABL1 ATP competitive inhibitors. We therefore wanted to use the ClonTracer barcoding system to compare the clonal responses of KCL-22 to imatinib, nilotinib and GNF-2. RNA-seq was employed to characterize genetic alterations and gene expression signatures in the pooled cell populations resistant to BCR-ABL inhibitors as well as single clones showing differential response to the three inhibitors. Overall design: mRNA profiling of the subpopulations and single clones of human CML cell line KCL-22 that contribute to BCR-ABL inhibitor resistance","project":"SRP048701"}
{"number_samples":18,"species":"human","abstract":"To study the transcriptomic changes in progression of cervical pre-cancer (from normal cells to LGSIL to HGSIL) from co-existing tissues in a single FFPE block","project":"SRP048735"}
{"number_samples":4,"species":"human","abstract":"Release of promoter-proximal paused RNA polymerase II (Pol II) during early elongation is a critical step in transcriptional regulation in metazoan cells. Paused Pol II release is thought to require the kinase activity of cyclin-dependent kinase 9 (CDK9) for the phosphorylation of DRB sensitivity-inducing factor, negative elongation factor, and C-terminal domain (CTD) serine-2 of Pol II. We found that Pol II-associated factor 1 (PAF1) is a critical regulator of paused Pol II release, that positive transcription elongation factor b (P-TEFb) directly regulates the initial recruitment of PAF1 complex (PAF1C) to genes, and that the subsequent recruitment of CDK12 is dependent on PAF1C. These findings reveal cooperativity among P-TEFb, PAF1C, and CDK12 in pausing release and Pol II CTD phosphorylation. Overall design: Comparison of the chromatin occupancy of [1] PAF1, CDC73, LEO1, CTR9, total Pol II, and CTD serine 2-phosphorylated Pol II by ChIP-seq in THP1 cells; [2] PAF1, Pol II, Pol II (ser-5p), CDK12, and CDK9 by ChIP-seq in control and PAF1 knockdown cells; [3] LEO1 and Pol II by ChIP-seq in control and flavopiridol treated THP1 cells.","project":"SRP048744"}
{"number_samples":434,"species":"human","abstract":"RNA sequencing of human leukemia Overall design: The goals of this project are to obtain a comprehensive study of mutations and gene expression in human acute myeloid leukemia (AML). Methods: AML cells were thawed. DNA and RNA (polyA) was extracted and sequences were obtained with an illumina HiSeq 2000 sequencer. Results are pending.","project":"SRP048759"}
{"number_samples":310,"species":"human","abstract":"We report the global pattern of ileal gene expression in a cohort of 310 treatment-naïve pediatric Crohn Disease patients and controls.  We focus on genes with consistent altered expression in the ileum of younger (Paris age A1a) vs older (Paris age A1b) patients. Overall design: Ileal biopsies were obtained during diagnostic colonoscopies of children and adolescents aged less than 17 years, who presented with IBD-like symptoms.  All patients underwent baseline colonoscopy and histological characterization; non-IBD controls were those with suspected IBD, but with normal radiographic, endoscopic, and histologic findings.  Biopsies were stored at -80 degrees.","project":"SRP048801"}
{"number_samples":4,"species":"human","abstract":"Kruppel-like factor-9 (KLF9), a member of the large KLF transcription factor family, has emerged as a regulator of oncogenesis, cell differentiation and neural development; however, the molecular basis for KLF9’s diverse contextual functions remains unclear. This study focuses on the functions of KLF9 in human glioblastoma stem-like cells. We establish for the first time a genome-wide map of KLF9-regulated targets in human glioblastoma stem-like cells, and show that KLF9 functions as a transcriptional repressor and thereby regulates multiple signaling pathways involved in oncogenesis and stem cell regulation. A detailed analysis of one such pathway, integrin signaling, shows that the capacity of KLF9 to inhibit glioblastoma cell stemness and tumorigenicity requires ITGA6 repression. These findings enhance our understanding of the transcriptional networks underlying cancer cell stemness and differentiation, and identify KLF9-regulated molecular targets applicable to cancer therapeutics. Overall design: Two cell lines were used as biological replicates. Each cell line has one KLF9 induction sample and one control sample.","project":"SRP048804"}
{"number_samples":8,"species":"human","abstract":"An emerging theme of gene regulation is the involvement of architectural chromosomal molecules in transcription control. Condensins are critical regulators of mitotic chromosomes, but their interphase chromatin localization and functions remain poorly understood. Here we report that both the condensin I and condensin II complexes exhibit an unexpected, dramatic 17-?-estradiol-induced preferential recruitment to oestrogen receptor ? (ER-?)-bound active enhancers in interphase breast cancer cells, exhibiting non-canonical interaction with ER-? distinct from classic cofactors. Condensins prove to positively regulate ligand-dependent gene and eRNA transcription by modulating a binding equilibrium of enhancer-associated coactivators/corepressors, including p300 and RIP140. This activity was achieved by the condensin-dependent recruitment of an E3 ubiquitin ligase, HECTD1, to active enhancers, where it polyubiquitinates and dismisses corepressor RIP140 to stimulate eRNA transcription. Collectively, our results reveal an important, unanticipated transcriptional role of interphase condensins in modulating enhancer activation, providing new insights into enhancer function in the regulated transcriptional programs Overall design: The GRO-seq measures the trancription of nascent RNAs in the genome. From MCF7 cells treated with veichle or estrodial, we could identify estrogen-regulated eRNAs and subsequently could study their functions.","project":"SRP048811"}
{"number_samples":14,"species":"human","abstract":"Most B cell lymphomas arise in the germinal center (GC), where humoral immune responses evolve from potentially oncogenic cycles of mutation, proliferation, and clonal selection. Although lymphoma gene expression diverges significantly from GC-B cells, underlying mechanisms that alter the activities of corresponding regulatory elements (REs) remain elusive. Here we define the complete pathogenic circuitry of human follicular lymphoma (FL), which activates or decommissions transcriptional circuits from normal GC-B cells and commandeers enhancers from other lineages. Moreover, independent sets of transcription factors, whose expression is deregulated in FL, target commandeered versus decommissioned REs. Our approach reveals two distinct subtypes of low-grade FL, whose pathogenic circuitries resemble GC-B or activated B cells. Remarkably, FL-altered enhancers also are enriched for sequence variants, including somatic mutations, which disrupt transcription factor binding and expression of circuit-linked genes. Thus, the pathogenic regulatory circuitry of FL reveals distinct genetic and epigenetic etiologies for GC-B transformation. Overall design: Expression profiles of follicular lymphoma, centrocyte and peripheral blood B cells were generated by deep sequencing, using Illumina Hi-Seq 2000.","project":"SRP048820"}
{"number_samples":30,"species":"human","abstract":"Metazoan development depends on accurate execution of differentiation programs that allow pluripotent stem cells to adopt specific fates. Differentiation is brought about by global changes to chromatin architecture and transcriptional networks, yet whether other regulatory events support cell fate determination is less well understood. Using human embryonic stem cell and Xenopus models, we identified the vertebrate-specific ubiquitin ligase Cul3KBTBD8 as an essential regulator of neural crest specification. Cul3KBTBD8 monoubiquitylates NOLC1 and its paralog TCOF1, whose mutation underlies the craniofacial disorder Treacher Collins Syndrome that is characterized by a loss of cranial neural crest cells. Ubiquitylation of NOLC1 and TCOF1 drives formation of a platform that connects RNA polymerase I with ribosome modification enzymes, thereby altering the translational program of differentiating cells to support the generation of neural crest cells. We conclude that the dynamic regulation of ribosome function is an important feature of cell fate determination. Overall design: Ribosome profiling and mRNA-Seq","project":"SRP048825"}
{"number_samples":2,"species":"human","abstract":"Unraveling complexity of DNA methylome is essential to decipher DNA methylation mechanism in life. However, this has been subjected to technological constraints to balance between cost and accurate measurement of the DNA methylation level. In this study, by innovatively introducing C-hydroxylmethylated adapters, we have developed MeDIP-Bisulfite sequencing (MB-seq), which could obtain DNA methylome of repertoire CpGs at single-base resolution. We found MB-seq only costs 10% of MethylC-seq, but covers 85% of total CpGs in human genome. Unlike absolute methylation levels determined by MethylC-seq and RRBS, MB-seq presented relative methylation levels that are linearly inflated. This has enlightened us to develop a MB-seq corresponding correction method for methylation level based on ridge regression, which integrates the data of MB-seq and RRBS to predict the methylation level of total 28.2 million CpGs on human genome with high accuracy (Pearson correlation coefficient, PCC=0.90). Moreover, by employing MB-seq, we generated the DNA methylome of an ovarian epithelial cell line (T29) and its oncogenic counterpart (T29H), respectively. After ridge regression, we identified 131,790 differential methylation regions (DMRs) with high accuracy between T29 and T29H, far more than 7,567 obtained from RRBS. Taken together, our result demonstrated that the MB-seq combined with ridge regression is a wide applicable approach for profiling of DNA methylome. Overall design: Total RNAs were extracted from T29 and T29H with RNeasy Mini Kit (QIAGEN, Germany). RNA quality was quality-controlled by Bioanalyser 2100 (RNA nano kits, Agilent). mRNA-Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 (Illumina, San Diego, CA), and then subjected to transcriptome sequencing on the Illumina Hiseq 2000","project":"SRP048842"}
{"number_samples":8,"species":"human","abstract":"We describe a novel population of human adult cardiac resident stem cells (CRSCs), which are positive for W8B2 antigen, originating from human adult atrial appendages. W8B2+ CRSCs exhibit a spindle-shaped morphology, are clonogenic and able to self-renew. W8B2+ CRSCs show high expression of mesenchymal but not hematopoietic nor endothelial markers. W8B2+ CRSCs expressed GATA4, HAND2, and TBX5, but not C-KIT, SCA-1, NKX2.5, PDGFRa, ISL1 or Wilm’s tumor gene-1 (WT1). W8B2+ CRSCs can differentiate into the cardiovascular lineages and secrete various cytokines. Overall design: Comparative RNA sequencing was performed using W8B2+ cell from human atrial appendages (passage 2 from 3 different patients), c-Kit+ cell from human atrial appendages (passage 2 from 3 different patients) and W8B2+ cell from bone marrow (passage 3 from 2 different patients, from PromoCell, Heidelberg, Germany).","project":"SRP048889"}
{"number_samples":12,"species":"human","abstract":"Whole transcriptome sequencing of K-562 cells expressing WT or mutant species of ETV6.","project":"SRP048957"}
{"number_samples":7,"species":"human","abstract":"Background: Although genome-wide association studies (GWAS) have identified hundreds of variants associated with risk of autoimmune and immune-related disorders (AID), our understanding of the diseases mechanisms is limited. In particular, more than 90% of the risk variants lie in non-coding regions, and almost 10% of these map to long non-coding RNA transcripts (lncRNAs). LncRNAs are known to show more cell-type specificity than protein-coding genes. Methods: In this study, we aimed to characterize lncRNAs and protein-coding genes located in loci associated with nine AID which have been well-defined by Immunochip analysis, by transcriptome analysis across seven peripheral blood leukocyte populations (granulocytes, monocytes, NK cells, B-cells, memory-T cells, naive CD4+ and naive CD8+ T-cells) and four cord blood derived T-helper cell populations (precursor, primary, polarized (Th1, Th2) T-helper cells). Results: We show that lncRNAs mapping to loci shared between AIDs are significantly enriched in immune cell types when compared to lncRNAs from the whole genome (a<0.005). We were not able to prioritize single cell types relevant for specific diseases, but we observed five cell types enriched (a<0.005) in five AID (NK cells for inflammatory bowel disease, juvenile idiopathic arthritis, primary biliary cirrhosis, psoriasis; memory-T and CD8+ T-cells in juvenile idiopathic arthritis, primary biliary cirrhosis, psoriasis, rheumatoid arthritis; Th0 and Th2 cells for inflammatory bowel disease, juvenile idiopathic arthritis, primary biliary cirrhosis, psoriasis, rheumatoid arthritis). Furthermore we show that co-expression analyses of lncRNAs and protein-coding genes can predict the signaling pathways in which these AID-associated lncRNAs are involved. Conclusions: The observed enrichment of lncRNA transcripts in AID loci implies an important role for lncRNAs in AID etiology and suggests that lncRNA genes should be studied in more detail to correctly interpret GWAS findings. The co-expression results strongly support a model in which the lncRNA and protein-coding genes function together in the same pathways. Overall design: 7 immune cell types","project":"SRP048971"}
{"number_samples":12,"species":"human","abstract":"Stem cell differentiation timecourse, six time points through induction from induced pluripotency (day0) towards beating cardiomyocytes, mature at day14. Accompanying study investigates careful differentiation protocols.","project":"SRP048993"}
{"number_samples":6,"species":"human","abstract":"Synthetic transcription factors can be applied to many areas of biotechnology, medicine, and basic research.  Currently, the most common method for engineering synthetic transcription factors has been based on programmable DNA-binding domains of zinc finger proteins, Transcription Activator-Like Effectors (TALEs), and most recently the CRISPR/Cas9 system. These transcription factor platforms consist of the DNA-binding domain fused to potent transcriptional activation domains, most commonly the tetramer of the minimal transactivation domain of the VP16 protein from herpes simplex virus, referred to as VP64. Although many applications are well-suited for the targeted activation of a single gene, genetic reprogramming requires the coordinated regulation of many nodes of natural gene networks as is typically performed by naturally occurring reprogramming factors. Thus we sought to combine principles from each of these approaches by attaching potent transcriptional activation domains to a natural reprogramming factor to increase the efficiency and/or rate of cell fate conversion. In this study, we evaluated the effects of fusing potent activation domains to the transcription factor MyoD, the master regulator of the skeletal myoblast lineage. In certain non-myogenic lineages, MyoD overexpression causes upregulation of the myogenic gene network and conversion to a myoblast phenotype including cell fusion into multinucleated myotubes. Compared to wild-type MyoD, the VP64-MyoD fusion protein induced greater overall reprogramming of global gene expression. This simple approach for increasing the potency of natural reprogramming factors circumvents the need for screening engineered proteins and leads to a more robust cellular reprogramming compared to treatment with the wild type transcription factor. Overall design: Human dermal fibroblasts were transduced with a single tet inducible lentivirus that expresses either WT-MyoD or VP64-MyoD in response to treatment with doxycycline. Untreated human dermal fibroblast served as the negative control. Gene expression was measured using mRNA-seq, and differential expression was calculated using DESeq. All experiments were performed in biological duplicates.","project":"SRP049004"}
{"number_samples":3,"species":"human","abstract":"Generating human serotonergic neurons from fibroblasts","project":"SRP049028"}
{"number_samples":18,"species":"human","abstract":"Tumor epithelial cells develop within a microenvironment consisting of extracellular matrix, growth factors, and cytokines produced by non-epithelial stromal cells. In response to paracrine signals from tumor epithelia, stromal cells modify the microenvironment to promote tumor growth and metastasis. Here, we identify interleukin (IL)-33 as an epithelial cell-derived regulator of stromal cell activation and mediator of intestinal polyposis. IL-33 expression was elevated in the tumors and serum of colorectal cancer patients and induced in the adenomatous polyps of ApcMin/+ mutant mice. Genetic and antibody suppression of IL-33 signaling in ApcMin/+ mice inhibited proliferation, induced apoptosis, and suppressed angiogenesis in polyps, which reduced both tumor number and size. In ApcMin/+ polyps, IL-33 expression localized to tumor epithelial cells and expression of the IL-33 receptor, IL1RL1, associated with two stromal cell types, namely subepithelial myofibroblasts (SEMFs) and mast cells, whose activation was previously associated with polyposis. In vitro IL-33 stimulation of human SEMFs induced the expression of extracellular matrix components and growth factors associated with intestinal tumor progression. IL-33 deficiency reduced mast cell accumulation in ApcMin/+ polyps and expression of mast cell-derived proteases and cytokines known to promote polyposis. Together, our results suggest that IL-33 is a tumor epithelial cell-derived paracrine signal that promotes polyposis through the coordinated activation of stromal cells and the formation of a reactive stroma microenvironment. Overall design: Six T-75 flasks of CCD-18Co cells were grown to 80% confluency; three were treated with rhIL-33, three were given vehicle control; cells were trypsinized and split in two--half of each flask used for sequencing and half for qPCR validation post-sequencing","project":"SRP049061"}
{"number_samples":16,"species":"human","abstract":"BACKGROUND & AIMS- More frequent interaction of bacteria with the colonic epithelium is associated with ulcerative colitis (UC).  The identities of all proteins which promote bacterial clearance in colonic epithelial cells are unknown.  Previously, we discovered that dCAP-D3 (Chromosome Associated Protein-D3), regulates responses to bacterial infection.  We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium. METHODS- Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing CAP-D3 shRNA.  CAP-D3 levels in colonic epithelial cells from healthy and UC patient tissues were analyzed by immunoblot.  RNA-sequencing identified bacterially-induced CAP-D3 target genes.  The role of CAP-D3 target genes in bacterial clearance was analyzed by gentamycin protection assays, immunofluorescent staining, and by using pharmacologic inhibitors. RESULTS- CAP-D3 expression was reduced in colonic epithelial cells from UC patients with active disease. Reduction of CAP-D3 expression inhibited autophagy and decreased intracellular bacterial clearance.  The components of the heterodimeric SLC7A5/SLC3A2 amino acid transporter were identified as CAP-D3 target genes; their levels increased in infected, CAP-D3 deficient cell lines and in cells from UC patients.  In HT-29 cells, this resulted in earlier SLC7A5 recruitment to Salmonella-containing vacuoles, increased mTOR activity, and enhanced bacterial survival.  Inhibition of SLC7A5/SLC3A2 or mTOR activity rescued the bacterial clearance defect in CAP-D3 deficient cells. CONCLUSIONS- CAP-D3 attenuates amino acid transporter transcription to promote bacterial autophagy in colon epithelial cells.  CAP-D3 protein levels are decreased in patients with active UC, suggesting that CAP-D3 is a potential therapeutic target to restore mucosal homeostasis in UC patients. Overall design: Three RNA samples from 3 independent experiments including timepoints taken at 0, 0.5 and 7 hours post-infection were analyzed on a bioanalyzer for quality; one of the 0.5 hour post-infection samples was excluded at this time due to poor RNA purity.  Directional, cDNA libraries made from cellular mRNAs were generated from the other 8 samples and sequenced (paired-end sequencing of 100 bp reads) in the Genomics Core at the University of Chicago on an Illumina HiSeq2000.","project":"SRP049063"}
{"number_samples":29,"species":"human","abstract":"A panel of 29 melanoma cell lines were gene expression profiled by RNA-Seq. Overall design: mRNA profiles of 29 melanoma cell lines","project":"SRP049068"}
{"number_samples":12,"species":"human","abstract":"Characterization of the selectivity of SMN splicing modifiers in SMA type I fibroblasts by RNASeq Overall design: In total 12 samples were analyzed, divided into four distinct groups (treated with SMN-C3 @ 500 nM; controls for SMN-C3; treated with SMN-C1 @ 100 nM; controls for SMN-C1) containing 3 replicates each.","project":"SRP049090"}
{"number_samples":54,"species":"human","abstract":"Leiomyosarcoma (LMS) is a malignant neoplasm with smooth muscle differentiation, and there are three molecular subtypes of LMS which have been defined previously by our lab. To further validate these subtypes and identify potential therapeutic targets in each subtype, we profiled the LMS cases from each subtype with RNA-Seq technology. Overall design: 8 LMS cases from subtype I, 6 cases from subtype II, 3 cases from subtype III and 7 cases of normal tissues, were selected for RNA-Seq. Then the expression levels and potential mutations were analyzed on these cases.","project":"SRP049097"}
{"number_samples":5,"species":"human","abstract":"DNA methylation can be abnormally regulated in human disease and associated with effects on gene transcription that appear to be causally related to pathogenesis.  The potential to use pharmacological agents that reverse this dysregulation is therefore an attractive possibility.  To test how 5-aza-2’-deoxycytidine (5-aza-CdR) influences the genome therapeutically, we exposed non-malignant cells in culture to the agent and used genome-wide assays to assess the cellular response.  We found that cells allowed to recover from 5-aza-CdR treatment only partially recover DNA methylation levels, retaining an epigenetic “imprint” of drug exposure.  We show very limited transcriptional responses to demethylation of not only protein-coding genes but also loci encoding non-coding RNAs, with a limited proportion of the induced genes acquiring new promoter activation within gene bodies.  The data revealed an uncoupling of DNA methylation effects at promoters, with demethylation mostly unaccompanied by transcriptional changes.  The limited panel of genes induced by 5-aza-CdR resembles those activated in other human cell types exposed to the drug, and represents loci targeted for Polycomb-mediated silencing in stem cells, suggesting a model for the therapeutic effects of the drug.  Our results do not support the hypothesis of DNA methylation having a predominant role to regulate transcriptional noise in the genome, and indicate that DNA methylation acts only as part of a larger complex system of transcriptional regulation.  The targeting of 5-aza-CdR effects with its clastogenic consequences to euchromatin raises concerns that the use of 5-aza-CdR has innate tumorigenic consequences, requiring its cautious use in diseases involving epigenetic dysregulation. Overall design: mRNA profile of 5 different samples of HEK 293T cells treated with 5-aza-CdR","project":"SRP049156"}
{"number_samples":3,"species":"human","abstract":"Quantitative profiling of initiation ribosomes in vivo","project":"SRP049168"}
{"number_samples":14,"species":"human","abstract":"We characterized the gene expression differences in mDA neurons from all PD (Parkinson's disease) cases (6 independent samples) and controls (8 independent samples), identifying 1,028 differentially expressed genes making up the PD expression signature. Strikingly, MAOB gene was identified as significantly differentially expressed (p = 0.046). The heat map clearly differentiates cases from controls, where interestingly most differentially expressed genes had lower expression in PD cases compared to controls. In the clustering, the RNA expression pattern of the control (C2) with a family history of PD located close to the PD expression signature suggested a susceptibility to PD. Overall design: RNA was isolated from FAC-sorted cells of 14 samples (biological duplicates for each cell line, 7 cell lines in total) using RNeasy Micro Kit (QIAGEN). Quality control of the RNA was carried out with the Agilent Bio-analyzer, Qubit 2.0 at the MPSR of Columbia University. 100 ng of RNA with RIN = 9 were used for generating mRNA-focused libraries using TruSeq RNA Sample Preparation Kit v2 and sequencing on an Illumina 2000/2500 V3 Instrument offered by the Columbia Genome Center.","project":"SRP049203"}
{"number_samples":6,"species":"human","abstract":"Identifying the interaction partners of non-coding RNAs is essential for elucidating their functions. We have developed an approach, termed microRNA-cross-linking and immunoprecipitation (miR-CLIP), using pre-miRNAs modified with psoralen and biotin to capture their targets in cells. Photo-cross-linking and Argonaute 2-immunopurification followed by streptavidin affinity-purification of probe-linked RNAs provided selectivity in the capture of targets, identified by deep-sequencing. MiR-CLIP with pre-miR-106a, a miR-17-5p family member, identified hundreds of putative targets in HeLa cells, many carrying conserved sequences complementary to the miRNA seed but also many that were not predicted computationally. MiR-106a overexpression experiments confirmed that miR-CLIP captured functional targets, including H19, a long-non-coding RNA that is expressed during skeletal muscle cell differentiation. We showed that miR-17-5p family members bind H19 in HeLa cells and myoblasts. During myoblast differentiation levels of H19, miR-17-5p family members and mRNA targets changed in a manner suggesting that H19 acts as a sponge for these miRNAs. Overall design: Two replicates of three cDNA libraries were submitted to deep sequencing: a sample from RNA-7-transfected cells; a sample from pre-miR-106a transfected cells; and a control sample.","project":"SRP049237"}
{"number_samples":4,"species":"human","abstract":"Identifying the interaction partners of non-coding RNAs is essential for elucidating their functions. We have developed an approach, termed microRNA-cross-linking and immunoprecipitation (miR-CLIP), using pre-miRNAs modified with psoralen and biotin to capture their targets in cells. Photo-cross-linking and Argonaute 2-immunopurification followed by streptavidin affinity-purification of probe-linked RNAs provided selectivity in the capture of targets, identified by deep-sequencing. MiR-CLIP with pre-miR-106a, a miR-17-5p family member, identified hundreds of putative targets in HeLa cells, many carrying conserved sequences complementary to the miRNA seed but also many that were not predicted computationally. MiR-106a overexpression experiments confirmed that miR-CLIP captured functional targets, including H19, a long-non-coding RNA that is expressed during skeletal muscle cell differentiation. We showed that miR-17-5p family members bind H19 in HeLa cells and myoblasts. During myoblast differentiation levels of H19, miR-17-5p family members and mRNA targets changed in a manner suggesting that H19 acts as a sponge for these miRNAs. Overall design: Two replicates of two cDNA libraries were submitted to deep sequencing: a sample from siH19-transfected cells and a control sample.","project":"SRP049238"}
{"number_samples":8,"species":"human","abstract":"Mutations in the gene encoding the transcription factor forkhead box P1 or FOXP1 occur in patients with neurodevelopmental disorders, including autism. However, the function of FOXP1 in the brain remains mostly unknown. Here, we identify the gene expression program regulated by FoxP1 in both human neural cells and mouse brain and demonstrate a conserved role for FOXP1 transcriptional regulation of autism and Fragile X Mental Retardation Protein (FMRP) mediated pathways. Coexpression networks support a role for Foxp1 in neuronal activity, and we show that Foxp1 is necessary for neuronal excitability. Using a Foxp1 mouse model, we observe defects in ultrasonic vocalizations. This behavioral phenotype is reflected at the genomic level as striatal Foxp1-regulated overlap with genes known to be important in rodent vocalizations. These data support an integral role for FOXP1 in regulating signaling pathways vulnerable in developmental disorders and the specific regulation of pathways important for vocal communication. Overall design: We carried out RNA-sequencing (RNA-seq) and ChIP-sequencing of human neural progenitors cells. We carried out RNA-sequencing (RNA-seq) of mouse striatal tissue, mouse hippocampal tissue and mouse cortical tissue. For the RNA-seq, four indipendent replicates were used for the neural progenitor cells and mouse tissues. For the Chip-seq, a single neural progenitor cell line was used.","project":"SRP049264"}
{"number_samples":4,"species":"human","abstract":"Patient specific therapy is emerging as an important possibility for many cancer patients.  However, to identify such therapies it is essential to determine the genomic and transcriptional alterations present in one tumor relative to control samples. This presents a challenge since use of a single sample precludes many standard statistical analysis techniques.  We reasoned that one means of addressing this issue is by comparing transcriptional changes in one tumor with those observed in a large cohort of patients analyzed by The Cancer Genome Atlas (TCGA).  To test this directly, we devised a bioinformatics pipeline to identify differentially expressed genes in tumors resected from patients suffering from the most common malignant adult brain tumor, glioblastoma (GBM).  We performed RNA sequencing on tumors from individual GBM patients and filtered the results through the TCGA database in order to identify possible gene networks that are overrepresented in GBM samples relative to controls.  Importantly, we demonstrate that hypergeometric-based analysis of gene pairs identifies gene networks that validate experimentally.  These studies identify a putative workflow for uncovering differentially expressed patient specific genes and gene networks for GBM and other cancers. Overall design: RNAseq on 2 gliosblastoma samples and 2 epileptic samples","project":"SRP049333"}
{"number_samples":166,"species":"human","abstract":"Induced pluripotency is a promising avenue for disease modeling and therapy, but the molecular principles underlying this process, particularly in human cells, remain poorly understood due to donor-to-donor variability and intercellular heterogeneity. Here we constructed and characterized a clonal, inducible human reprogramming system that provides a reliable source of cells at any stage of the process. This system enabled integrative transcriptional and epigenomic analysis across the human reprogramming timeline at high resolution. We observed distinct waves of gene network activation, including the ordered re-activation of broad developmental regulators followed by early embryonic patterning genes and culminating in the emergence of a signature reminiscent of pre-implantation stages. Moreover, complementary functional analyses allowed us to identify and validate novel regulators of the reprogramming process. Altogether, this study sheds light on the molecular underpinnings of induced pluripotency in human cells and provides a robust cell platform for further studies. Overall design: mRNA sequencing of primary and secondary fibroblasts with reference BJ (supplementary file fibroblasts), reprogramming intermendiates from untreated hiF-T reprogramming (supplementary file reprogramming), or selective time points upon LSD1 inhibitor treatment (supplementary file LSD1i). RNA samples used for mRNA sequencing are the same used for smallRNA sequencing.","project":"SRP049340"}
{"number_samples":10,"species":"human","abstract":"We descrive a joint model of transcriptional activation and mRNA accumulation, using estrogen receptor ERa activation in MCF-7 breast cancer cell line, which can be used for inference of transcription rate, RNA processing delay and degradation rate given data from high-throughput sequencing time course experiments. Overall design: MCF-7 cells were mock treated or with 10nM 17b-E2 to nine time points (5', 10', 20', 40', 80', 160', 320', 640' and 1280'). Genome-wide identification of RNA polymerase II (RNAPII) occupancy and transcriptome profiling (RNA-seq) following E2 induction of MCF-7 cells Please note that the information in the wig.txt files is in gene-specific coordinates, not chromosomic coordinates, as this is the most sensible format for the associated project/paper.","project":"SRP049355"}
{"number_samples":17,"species":"human","abstract":"Purpose: The goal was to capture the transcriptional activity due to over-expression of HER2 protein. We profiled this transcriptional activity using two different RNA-Seq alignment and quantification pipelines. We also used these samples to generate a gene expression signature of HER2 pathway activity. Over-expression was validated using Western blots. Illumina RNA-Seq technology was used to capture the downstream transcriptional activity. Reads were 101 base pairs long and single ended. An R open source package “Rsubread” was used to align and quantify the read using UCSC hg19 annotation. The integer-based gene counts were later normalized in FPKM and TPM . Overall design: A profile of gene expression, downstream of ERBB2/HER2 over-expression, was generated in cells derived from breast and used to generate a gene-expression signature reflective of HER2 pathway activation status.","project":"SRP049377"}
{"number_samples":6,"species":"human","abstract":"This SuperSeries is composed of the SubSeries listed below. Purpose: Kaposi’s sarcoma (KS)-associated herpesvirus (KSHV) causes several lymphoproliferative disorders, including KS, a common AIDS-associated malignancy. Cellular and viral microRNAs (miRNAs) have been shown to play important roles in regulating the expression of genes in oncogenesis. Herpesviruses, including KSHV, encode for miRNAs that are involved in angiogenesis, inflammation and apoptosis. A better knowledge of the miRNA-mediated pathways that regulate KSHV infection is therefore essential for an improved understanding of viral infection and pathogenesis. Methods: In this study, we used deep sequencing to analyze miRNA, both viral and human, and mRNA expression in KS tumor-derived human cells. Results: This approach revealed 153 differentially expressed human miRNAs between KSHV-positive and -negative cells. Differential expression of eight miRNAs was independently confirmed by qRT-PCR. We additionally showed that a majority (~73%) of KSHV-regulated miRNAs are down-regulated, including most members of the 14q32 miRNA cluster. Specifically, human miR-409-3p, which is known to target the pro-angiogenic growth factor angiogenin and the inflammation marker fibrinogen-beta, was significantly down-regulated in KSHV-infected cells based on deep sequencing and qRT-PCR. Despite this substantial down-regulation of cellular miRNAs, hsa-miR-708-5p was significantly up-regulated by KSHV and has been shown to directly inhibit pro-apoptotic protease Caspase-2. Finally, we evaluated to what extent there was an inverse correlation between miRNA and mRNA expression levels. Using filtered datasets, we identified relevant canonical pathways that were significantly enriched. Conclusion: Taken together, our data demonstrate that most human miRNAs affected by KSHV are repressed and our findings highlight the relevance of studying the post-transcriptional gene regulation of miRNAs for KSHV-associated malignancies. Overall design: Refer to individual Series. 6 samples analyzed (one cell type). Two experimental conditions: uninfected vs. chronically KSHV-infected cells (n=3). Two sequencing platforms: microRNA-Seq and mRNA-Seq.","project":"SRP049391"}
{"number_samples":6,"species":"human","abstract":"Implantation is dependent on synchronized interactions between the conceptus and surrounding decidual cells but the involvement of clock genes in this process is not well understood. Circadian oscillations are predicated on transcriptional-translational feedback loops, which balance the activities of the transcriptional activators CLOCK and BMAL1 and repressors encoded by PER and CRY genes. Here we show that loss of PER2 expression silences circadian oscillations in decidualizing human endometrial stromal cells (HESCs). Downregulation was preceded by reduced CLOCK binding to a noncanonical E-box enhancer in the PER2 promoter and occurred between 12 - 24 h after exposure to a deciduogenic stimulus. RNA sequencing revealed that premature inhibition of PER2 by siRNA knockdown leads to a grossly disorganised decidual response. Gene ontology analysis highlighted a preponderance of cell cycle regulators amongst the 1,121 genes perturbed upon PER2 knockdown. Congruently, PER2 inhibition abrogated mitotic expansion of differentiating HESCs by inducing cell cycle block at G2/M. Analysis of mid-luteal endometrial biopsies revealed an inverse correlation between PER2 transcript levels and the number of miscarriages in women suffering reproductive failure. Thus, PER2 synchronizes mitotic expansion of HESCs with a periodic decidual gene expression; uncoupling of these events may cause persistent pregnancy failure. Overall design: Endometrial mRNA profiles of paired control (siRNA-NT) and siRNA-PER2 were generated by deep sequencing, in triplicate using Illumina","project":"SRP049409"}
{"number_samples":4,"species":"human","abstract":"We sequenced polyadenylated RNA isolated from Jurkat cells infected or not with HIV-1 to investigate in detail alterations in the transcriptome of the CD4+ T cells upon viral infection.","project":"SRP049410"}
{"number_samples":4,"species":"human","abstract":"RNA seq experiment","project":"SRP049426"}
{"number_samples":21,"species":"human","abstract":"Purpose: Aim of the study is to identify functional differences between the P1 and P2-HNF4a isoforms. To do this, we generated Tet-On inducible lines that express either the human (P1) HNF4a2 or (P2) HNF4a8 under control of DOX in the HCT116 human colon cancer cells. Methods: HNF4a2 and Parental lines were induced with 0.3 µg/mL DOX, while HNF4a8 line was induced with either 0.1 or 0.3 µg/mL DOX for 24 hours. Samples were generated by deep sequencing, using the Illumina TruSeq RNA. Result: There were common and unique dysregulated genes identified in the HNF4a2 and HNF4a8 lines (+DOX); more upregulated genes than downregulated genes in both the lines. Conclusion: The functional difference between HNF4a2 and HNF4a8 is that the latter tends to upregulate genes involved in proliferation and anti-apoptosis while HNF4a2 upregulates genes involved in growth suppression and cell death. Overall design: Tet-On inducible HCT116 cell (Parental, HNF4a2, and HNF4a8) lines, treated with (0.0, 0.1, or 0.3 µg/mL) DOX for 24 hours, were 50bp pair-ended sequenced in triplicate using Illumina TruSeq RNA Sample Prep v2 Kit.","project":"SRP049436"}
{"number_samples":18,"species":"human","abstract":"We report the identification of miRNAs and transcripts that are regulated during human cardiac maturation. Overall design: Examination of  hESC-derived cardiomyocytes , human fetal  tissue and human adult heart tissue for cardiomyocyte maturation.","project":"SRP049449"}
{"number_samples":2,"species":"human","abstract":"Human Gastric Cancer Cells with Tanshinone IIA treatment","project":"SRP049450"}
{"number_samples":4,"species":"human","abstract":"Sequence data of HeLa cells were generated based on ribominus RNA sequencing with or without RNase R treatment (RNaseR+/-), respectively. These data sets were used for prediction of circRNAs.","project":"SRP049453"}
{"number_samples":4,"species":"human","abstract":"We identified spatially restricted transcription factors and found SOX15 expression confined to stratified esophageal epithelium, with attenuation in Barrett's esophagus. SOX15 binds esophagus-specific loci and its loss in human esophageal cells affected esophagus-specific transcripts Overall design: [RNA-Seq] Total RNA isolated from CPA control cells and CPA cells following SOX15 depletion, samples were prepared for sequencing using the TruSeq RNA Sample Preparation Kit (Illumina) according to the manufacturer's instructions. 75 base pair single-end reads were sequenced on an Illumina NextSeq 500 instrument. The data include 2 independent biological replicates per genotype. [ChIP-Seq] Examine SOX15-chromatin binding in CPA cells.","project":"SRP049465"}
{"number_samples":13,"species":"human","abstract":"We characterized the gene expression by Hierarchical Clustering and one-matrix clustering in hESC, day 12 progenitors, day 25-day 27, day82 differentiated hypothalamic neurons from hESCs and day 45 neurons derived from iPSCs generated from controls (2 independent) and BBS (Bardet-Biedl Syndrome, 3 independent) subjects. Overall design: RNA was isolated from cells of 13 samples (1 hESC, triplicate for day 12 progenitors, 1 day 25 neuron sample, duplicate for day 27 neuron samples, 1 day 82 neuron sample, five day 45 neuron samples made from 5 independent iPSC lines ) using RNeasy Micro Kit (QIAGEN). Quality control of the RNA was carried out with the Agilent Bio-analyzer, Qubit 2.0 at the MPSR of Columbia University. 100 ng of RNA with RIN = 9 were used for generating mRNA-focused libraries using TruSeq RNA Sample Preparation Kit v2 and sequencing on an Illumina 2000/2500 V3 Instrument offered by the Columbia Genome Center.","project":"SRP049475"}
{"number_samples":14,"species":"human","abstract":"Although recent evidence suggests that overlapping sense/antisense transcription is a common feature in higher eukaryotes, the possibility that overlapping transcripts could interact to each other and bear a specific biological function has not been explored. Here we show that a plethora of sense/antisense transcript pairs are co-expressed from 8q24.21 within the same cell and acquire a stable double-stranded RNA conformation. Interestingly, these molecules display predominantly nuclear localization and establish specific interactions with nuclear components. A detailed characterization of a particular sense/antisense pair (ndsRNA-2a) revealed that this molecule displays differential localization throughout the cell cycle, interacts with RCC1 and RAN and through the latter with the mitotic RANGAP1-SUMO1/RANBP2 complex. Notably, an increased number of bi/multi-nucleated cells and chromatin bridges were observed upon ndsRNA-2a overexpression, whereas strand-specific ndsRNA-2a knockdown leads to mitotic catastrophe and cell death. This suggests a functional role of ndsRNA-2a in cell cycle progression that critically requires its double stranded nature. Finally, the identification of hundreds of sense/antisense transcripts pairs harboring ndsRNA profile signatures and their regulation by cellular cues suggests that ndsRNAs constitute a novel class of regulatory molecules that are likely to be involved in a plethora of biological processes. Overall design: PLB985 long (3x datasets) and small (3x datasets) strand specific RNA-Seq for captured RNAs. Global PLB985 for long (2x datasets) and small RNAs (2x datasets). Global libraries for EtOH (vehicle) treated (1x dataset) or retinoic acid induced differentiated PLB985 cells (1x dataset).","project":"SRP049479"}
{"number_samples":6,"species":"human","abstract":"Investigation of bile acid signaling and liver size in a mouse model wherein the mouse liver is repopulated with human liver cells","project":"SRP049484"}
{"number_samples":2,"species":"human","abstract":"Single-cell genomics and single-cell transcriptomics have recently emerged as powerful tools to study the biology of single cells at a genome-wide scale. Here we describe a method that allows the integration of genomic DNA and mRNA sequencing from the same cell. We use this method to correlate DNA copy number variation to transcriptome variability among individual cells. Overall design: First, hand-picked single cells are lysed and reverse transcribed using a poly-A primer including cell-specific barcodes, a 5' Illumina adapter and a T7 promoter overhang to convert mRNA to single stranded cDNA (ss cDNA). The gDNA and single stranded cDNA are then subjected to quasilinear whole genome amplification, as previously described, using an adapter with a defined 27 nucleotide sequence at the 5’ end followed by 8 random nucleotides. After 7 rounds of amplification, the gDNA and cDNA are copied to generate a variety of different short amplicon (0.5–2.5 kb) species, with a majority of amplicons containing adapter Ad-2 at both ends and a small fraction of cDNA derived amplicons containing Ad-2 at one end and Ad-1x at the other. Next, the sample is split into two tubes to further amplify gDNA and cDNA. The tube used to sequence gDNA is amplified using PCR. Following sonication, adapter Ad-2 removal, and cell-specific indexed Illumina library preparation, this half is used to sequence gDNA. The tube used to sequence cDNA is converted to double-stranded cDNA and amplified using in vitro transcription such that the amplified RNA (aRNA) is uniquely produced from cDNA but not gDNA. 3’ Illumina adapters are then ligated to the aRNA followed by reverse transcription and PCR, allowing quantification of mRNA.","project":"SRP049500"}
{"number_samples":5,"species":"human","abstract":"Disappearance of the Barr body has long been considered a hallmark of cancer, although whether this corresponds to epigenetic instability and transcriptional reactivation, or to genetic loss, has remained unclear. Here we show that in breast cancer cell lines as well as primary breast tumors, the inactive X chromosome frequently displays a highly abnormal 3D nuclear organization, with global perturbations in its characteristic heterochromatic state, including apparent gain of euchromatic marks and lessening of repressive marks such as H3K27me3 and promoter DNA methylation. Genome-wide profiling of chromatin and transcription reveal modified epigenomic landscapes in cancer cells accompanied by a significant degree of X-linked gene reactivation, affecting genes previously implicated in cancer, including the histone deacetylase, HDAC8 and transducin (Beta)-Like 1X-Linked, TBL1X. We provide proof of principle that epigenetic deregulation can indeed perturb the dosage of some X-linked factors and demonstrate that many of these genes are reactivated in primary breast tumors. Our study establishes that the inactive X is subject to epigenetic erosion in a cancer context and sets the stage for the use of chromatin marks and X- chromosome genes as potential biomarkers to assess epigenetic changes in cancer. Overall design: Examination of allele specific expression of genes in chrX using histone marks, transcriptome, exome and SNP data on two normal cell-line and three cancer cell-line.","project":"SRP049507"}
{"number_samples":4,"species":"human","abstract":"siRNAs with dN are a promising and easy approach to characterize essential genes by minimizing the off-target effect in cancer cells. Overall design: mRNA profiles of HCT 116 cells after the transfection of siRNAs or blank control by deep sequencing, using HiSeq 2000","project":"SRP049510"}
{"number_samples":6,"species":"human","abstract":"Purpose: By integrating DNA methylation and gene expression of COPD lung tissues, we identified EPAS1 as a key regulator whose downstream genes significantly overlapped with multiple genes sets associated with COPD disease severity. EPAS1 is distinct in comparison with other key regulators in terms of methylation profiles and downstream target genes. Genes predicted to be regulated by EPAS1 were enriched for biological processes including signaling, cell communications, and system developement. As EPAS1 downstream genes were significantly enriched for hypoxia responsive genes in endothelial cells, we tested EPAS1 function in human and mouse endothelial cells HUVEC and C166. EPAS1 knockdown by siRNA in endothelial cells impacted genes that significantly overlapped with EPAS1 downstream genes in lung tissue including hypoxia responsive genes. Methods: The cell lines of HUVEC (Lonza, MD, USA) and C166 (American Type Culture Collection, VA, USA) were cultured in the appropriate media at 37°C with 5% CO2. The cells were transfected with EPAS1 siRNA and non-targeting negative control siRNA (Life Technologies, CA, USA) using Lipofectamine RNAiMAX as recommended transfection protocols by the manufacturer. After the treatments with 5nM Silencer® Select siRNA (s4700 for EPAS1, s65525 for Epas1; Life Technologies, USA) for 48 hours, the total RNA was purified with RNeasy Mini Kit (QIAGEN, Germany). The efficiencies of knocked down the EPAS1 expression were assessed by qPCR with 1.4% for HUVEC, 3.2% for C166. Approximately 250 ng of total RNA per sample were used for library construction by the TruSeq RNA Sample Prep Kit (Illumina) and sequenced using the Illumina HiSeq 2500 instrument with 100nt single read setting according to the manufacturer's instructions. Sequence reads were aligned to human genome assembly hg19 and mouse genome assembly mm10, respectively, using Tophat [96]. Total 23,228 human and 22,609 mouse genes were quantified using Cufflinks [96]. siRNA signatures were derived by comparing expression profiles of EPAS1 or Epas1 siRNAs with non-targeting siRNAs at paired t-test p-value cutoff 0.05 with resulting signature sizes of 2,796 and 3,730, and corresponding q-values [97] 0.11 and 0.07 for HUVEC and C166, respectively. Results: When comparing endothelial cells treated with EPAS1 siRNAs and scrambled siRNAs, we identified an EPAS1 siRNA signature consisting of 2796 and 3730 genes in human and mouse endothelial cell lines, respectively, whose expression levels significantly changed (t-test p-value<0.05), including EPAS1 itself (p-value = 0.002 and 0.02) and the EPAS1 downstream target gene VEGFA (p-value = 0.03 and 0.01). The EPAS1 siRNA signatures derived from human and mouse cell lines were highly consistent, with 695 genes in common to both signatures (p-value = 7.2x10-65). Both signatures not only significantly overlapped with EPAS1 downstream genes (p-value = 7.3x10-7 and 1.5x10-12), but also with hypoxia response genes in endothelial cells (Fisher’s Exact Test p-value = 5.8x10-8 and 1.2x10-12 in the human and mouse signatures, respectively). Moreover, the EPAS1 siRNA signatures consistently overlapped genes associated with the COPD severity phenotypes. These results together validate that EPAS1 causally regulates the downstream target genes we predicted, and that these genes in turn affect COPD development and progression. Overall design: For each of human and mouse cell lines,  3 siRNA control cells, and 3 siRNA EPAS1 knockdown cells were used and analyzed. To identify EPAS1 signatures, paired t-test was performed between control siRNA and EPAS1 siRNA samples.","project":"SRP049514"}
{"number_samples":3,"species":"human","abstract":"Technical evaluation of MALBAC transcriptome amplification","project":"SRP049515"}
{"number_samples":12,"species":"human","abstract":"During cerebellar development, the main portion of the cerebellar plate neuroepithelium (NE) gives birth to Purkinje cells and interneurons, while the germinal zone at its dorsal edge, called the rhombic lip (RL), generates granule cells and cerebellar nuclei neurons. However, it remains elusive how these components work together to generate the intricate structure of the cerebellar anlage. In this study, we found that a polarized cerebellar anlage structure self-organizes in three-dimensional (3D) human ES cell (hESC) culture. This NE is capable of differentiating into electrophysiologically functional Purkinje cells. The addition of FGF19 promotes spontaneous generation of dorsoventrally polarized NE structures containing cerebellar and basal plates. Furthermore, further addition of SDF1 promoted the generation of stratified cerebellar plate NE with RL-like germinal zones self-forming at the edge. Thus, hESC-derived cerebellar progenitors exhibit substantial self-organizing potential for generating a polarized structure reminiscent of the early human cerebellar anlage at the first trimester. Overall design: Examination of mRNA profile in two different treated human ES cells .","project":"SRP049553"}
{"number_samples":12,"species":"human","abstract":"Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this work is comprehensive analysis of target genes co-regulated by CREB1 and FoxA1 in prostate cancer cell models, tissues, and circulating tumor cells (CTCs). Expression of CREB1/FoxA1 target genes corresponds with disease recurrence and aggressive clinical features. Methods: LNCaP cells between passage number 32-34 and abl between 62-64 were used for assay. For RNA-seq, cells are transfected with FoxA1,CREB1 specific or nonspecific siRNA and total RNA is extracted with Qiangen RNeasy Mini kit(cat 74104) and library is generated with  TruSeq RNA Sample Preparation Kit v2 from Illumina(cat  RS-122-2001). For Chip-seq,  LNCaP cells between passage number 32-34 and abl between 62-64 were used for assay. After crosslinking with 1% formaldehyde, ChIP was performed. the ChIP producted was further used to generate library with illumina ChIP-seq kit. Hi-seq 2500 was used for sequencing and the data was analyzed by MACs for peaks. Overall design: LNCaP cells and LNCaP cells were used as cell model. Library was sequenced using Illumina HISeq 2500.","project":"SRP049591"}
{"number_samples":406,"species":"human","abstract":"We apply the cellular reprogramming experimental paradigm to two disorders caused by symmetrical copy number variations (CNV) of 7q11.23 and displaying a striking combination of shared as well as symmetrically opposite phenotypes: Williams Beuren syndrome (WBS) and 7q microduplication syndrome (7dupASD). Through a uniquely large and informative cohort of transgene-free patient-derived induced pluripotent stem cells (iPSC), along with their differentiated derivatives, we find that 7q11.23 CNV disrupt transcriptional circuits in disease-relevant pathways already at the pluripotent state. These alterations are then selectively amplified upon differentiation into disease-relevant lineages, thereby establishing the value of large iPSC cohorts in the elucidation of disease-relevant developmental pathways. In addition, we functionally define the quota of transcriptional dysregulation specifically caused by dosage imbalances in GTF2I (also known as TFII-I), a transcription factor in 7q11.23 thought to play a critical role in the two conditions, which we found associated to key repressive chromatin modifiers. Finally, we created an open-access web-based platform (accessible at http://bio.ieo.eu/wbs/ ) to make accessible our multi-layered datasets and integrate contributions by the entire community working on the molecular dissection of the 7q11.23 syndromes. Overall design: We reprogrammed skin fibroblasts from patients harbouring a 7q11.23 hemi-deletion (WBS, 4 patients; +1 atypical deletion, AtWBS) or microduplication (7dupASD; 2 patients), as well as from one unaffected relative and two unrelated controls, using integration-free mRNA-reprogramming, leading to the establishment of a total of 27 characterized iPSC clones. We profiled these by RNAseq (either polyA or ribo-zero). To isolate the contribution of GTF2I to the transcriptional dysregulation, we created stable lines expressing a short hairpin against GTF2I from a representative subset of these iPSC clones, and profiled by RNAseq 7 such lines along with their respective scramble controls. Finally, we also profiled by RNAseq mesenchymal stem cells (MSC) derived from a representative subset of the lines.","project":"SRP049593"}
{"number_samples":73,"species":"human","abstract":"To determne JunB target gene in human keratincoytes Mice with epidermal deletion of JunB transcription factor displayed a psoriasis-like inflammation. The relevance of these findings to humans and the mechanisms mediating JunB function are not fully understood. Here, we demonstrate that impaired JunB function via gene silencing or overexpression of a dominant negative mutant increased human keratinocyte cell proliferation but decreased cell barrier function. RNA-seq revealed over 500 genes affected by JunB loss-of-function which included an upregulation of an array of proinflammatory molecules relevant to psoriasis. Among these were TNFa, CCL2, CXCL10, IL6R and SQSTM1, an adaptor protein involved in NF-kB activation. ChIP-Seq and gene reporter analyses showed that JunB directly suppressed SQSTM1 through binding to a consensus AP-1 cis-element located around 2 Kb upstream of SQSTM1-trasncription start site. Similar to JunB loss-of-function, SQSTM1-overexpression induced TNFa, CCL2 and CXCL10. Conversely, NF-kB-inhibition genetically with a mutant IkBa or pharmacologically with PDTC prevented cytokine, but not IL6R, induction by JunB-deficiency.  Taken together, our findings indicate that JunB controls epidermal growth, barrier formation and proinflammatory responses through direct and indirect mechanisms, pinpointing SQSTM1 as a key mediator of JunB-suppression of NF-kB-dependent inflammation. Overall design: 3 indepdent set of priamry human keratinocytes isolated from foreskin skin samples were transfected with nonsilencing control or siRNA oligonucleotides targeting JunB. mRNA was then isolated and used for cDNA library construction followed by RNA-sequencing.","project":"SRP049599"}
{"number_samples":10,"species":"human","abstract":"We report that knockdown of EJC core proteins, eIF4A3, Y14, Magoh, causes a transcript-wide changes in alternative splicing, as well as some transcriptional changes. These changes are specific to EJC core proteins, and KD of UPF1 protein caused different sets of alterantive splicing changes. These changes are linked to the rate of transcription. Overall design: Examination of 4 different knockdown, as well as GFP knockdown in HeLa cells, 2 replicates each condition.","project":"SRP049611"}
{"number_samples":6,"species":"human","abstract":"We have found that thyroid hormones (THs), acting as soluble integrin avß3 ligands, activate growth-related signaling pathways in T-cell lymphomas (TCL). Specifically, TH-activated avß3 integrin signaling promotes TCL proliferation and angiogenesis, in part, via the up-regulation of VEGF. Overall design: CUTLL1 cells were treated with T3- and T4-bound agarose or agarose alone for 24hrs. Total RNA was harvested from cells and used for expression profiling via RNA-seq.","project":"SRP049648"}
{"number_samples":22,"species":"human","abstract":"Extranodal NK/T-cell lymphomas (NKTCLs) and gamma-delta peripheral T-cell lymphomas (?d-PTCLs) are rare lymphomas with poor prognosis. There is little insight into the role of oncogenic mutations in these allied cancers. Here, we analyzed the genome-wide mutation profile on 17 NKTCL cases and relevant NK/T-Cell lines by performing RNA sequencing and observed frequent oncogenic mutations in STAT3, STAT5B and other JAK-STAT pathway genes.","project":"SRP049695"}
{"number_samples":2,"species":"human","abstract":"Fibrolamellar hepatocellular carcinoma (FL-HCC) is a very rare and distinct subtype of hepatocellular carcinoma that occurs in the previously healthy livers of adolescents and young adults. The low incidence rate has kept FL-HCC an understudied cancer, thus the list of FL-HCC markers is limited and the genetic alterations driving its growth have remained unknown. In order to gain a better understanding of the molecular mechanisms underlying FL-HCC initiation and progression, we performed an in-depth genomic analyses of one tumor followed by immunohistochemistry validation on seven other tumors. We showed the expression of neuroendocrine markers in FL-HCC. In addition, DNA/RNA sequencing revealed that common cancer pathways are not visibly altered in FL-HCC but identified two novel structural variants, both resulting in fusion transcripts. Through in vitro studies, we demonstrated the oncogenic properties of these fusion transcripts. These experiments further... (for more see dbGaP study page.)","project":"SRP049705"}
{"number_samples":25,"species":"human","abstract":"In this study, we have integrated RNA-seq data from subcellular fractionated RNA (i.e., cytoplasm, nucleoplasm, and chromatin-associated) with GRO-seq data using a novel bioinformatics pipeline. This has yielded a comprehensive catalog of polyadenylated lncRNAs in MCF-7 cells, about half of which have not been annotated previously and about a quarter of which are estrogen-regulated. Knockdown of selected lncRNAs, such as lncRNA152 and lncRNA67 followed by RNA-seq suggest that these lncRNAs regulate the expression of cell cycle genes. Overall design: characterization of long noncoding RNAs","project":"SRP049713"}
{"number_samples":7,"species":"human","abstract":"We report molecular characterization of human brown and white adipocytes. We showed that PAZ6 and SW872 cells exhibit classical molecular and phenotypic markers of brown and white adipocytes, respectively. However, SGBS cells presented a versatile phenotype of adipocyte Overall design: Sequencing of three human adipocytes cell lines (SGBS, SW872 and PAZ6) in undifferentiated and differentiated stages.","project":"SRP049714"}
{"number_samples":16,"species":"human","abstract":"DNA methylation is thought to induce a transcriptional silencing through the combination of two mechanisms: the repulsion of transcriptional activators that do not recognize their binding sites when methylated, and the recruitment of transcriptional repressors that specifically bind methylated DNA. Methyl CpG Binding Domain proteins MeCP2, MBD1 and MBD2 belong to the latter category. However, the exact contribution of each protein in the DNA methylation dependent transcriptional repression occurring during development and diseases remains elusive. Here we present MBD2 ChIPseq data generated from the endogenous protein in an isogenic cellular model of human mammary oncogenic transformation. In immortalized or transformed cells, MBD2 was found in one fourth of methylated regions and associated with transcriptional silencing. Depletion of MBD2 induces upregulations of genes bound by MBD2 and methylated in their transcriptional start site regions. MBD2 was partially redistributed on methylated DNA during oncogenic transformation, independently of DNA methylation changes. Genes downregulated during this transformation preferentially gained MBD2 binding sites on their promoter. Depletion of MBD2 in transformed cells induced the upregulation of some of these repressed genes, independently of the strategy used for the abrogation of oncosuppressive barriers. Our data confirm that MBD2 is a major interpret of DNA methylation, and show an unreported dynamic in this interpretation during oncogenic transformation. Overall design: RNAseq of untreated HMEC-hTERT cells, siCtrl, siMBD2 or DAC treated HMLER cells, siCtrl or siMBD2 treated HME-ZEB1-RAS and HME-shP53-RAS cells, in duplicates.","project":"SRP049756"}
{"number_samples":4,"species":"human","abstract":"Aggressive double and triple hit (DH/TH) DLBCL feature activation of Hsp90 stress pathways.  Herein, we show that Hsp90 controls post-transcriptional dynamics of key mRNA species including those encoding BCL6, MYC and BCL2. Using a proteomics approach, we found that Hsp90 binds to and maintains activity of eIF4E (eukaryotic translation initiation factor 4E). EIF4E drives nuclear export and translation of BCL6, MYC and BCL2 mRNA. eIF4E RIP-sequencing in DLBCL suggests that nuclear eIF4E controls an extended program that includes BCR signaling, cellular metabolism and epigenetic regulation. Accordingly, eIF4E was required for survival of DLBCL including the most aggressive subtypes DH/TH lymphomas. Indeed, eIF4E inhibition induces tumor regression in cell line and patient-derived tumorgrafts of TH-DLBCL, even in the presence of elevated Hsp90 activity. Targeting Hsp90 is typically limited by counter-regulatory elevation of Hsp70B, which induces resistance to Hsp90 inhibitors. Surprisingly, we identify Hsp70 mRNA as an eIF4E target. In this way, eIF4E inhibition can overcome drug resistance to Hsp90 inhibitors. Accordingly, rational combinatorial inhibition of eIF4E and Hsp90 inhibitors resulted in cooperative anti-lymphoma activity in DH/TH DLBCL in vitro and in vivo. Overall design: We found that eIF4E activity regulates the nuclear export of BCL6, MYC, and BCL2 in DH/TH DLBCLs. To determine the extent of nuclear eIF4E activity in DH/TH DLBCLs and how these programs can support the oncogenic activity of BCL6, MYC and/or BCL2 transcripts, we conducted eIF4E-RIP of nuclear RNA followed by RNA-sequencing in OCI-Ly1 cells in triplicates. To understand the changes in gene expression after ribavarin in a clinically relevant sample, we generated a patient-derived xenograft (PDX) in NSG mice from a de-identified specimen isolated from a patient prior to treatment harboring a triple-hit ABC-type DLBCL. PDX cells from passage four (PDX-4) were implanted into NSG mice. When tumors were palpable, mice were randomized to receive vehicle or 80 mg/kg/b.i.d. ribavarin intraperitoneally for 10 days.  We isolated RNA from tumors treated with vehicle (n=2) or ribavarin (n=2) and performed mRNA-seq.","project":"SRP049769"}
{"number_samples":2,"species":"human","abstract":"A cell supsension containing an equal mix of HEK and 3T3 cells was used in Drop-Seq, a novel technology for high-throughput single cell mRNAseq.  The cells were mixed at a final concentration of 50 cells per microliter. Overall design: An estimated 1000 STAMPs and 100 STAMPs were separately selected for sequencing to obtain both high coverage and broad coverage data","project":"SRP049775"}
{"number_samples":11,"species":"human","abstract":"Homo sapiens Transcriptome or Gene expression from lifeTech total brain RNA (AM6050)","project":"SRP049776"}
{"number_samples":8,"species":"human","abstract":"Rationale: Slit2 is a possible modulator of vascular endothelial growth factor (VEGF) - induced angiogenesis, but its effects have not been tested in large animal models. Objective: We studied the effect of Slit2 on therapeutic angiogenesis induced by VEGF receptor 2 (VEGFR2) ligands Vammin and VEGF-D?N?C in vivo in rabbit skeletal muscles. The Slit2 target genes were also studied by RNA sequencing (RNA-Seq) in endothelial cells. Methods and Results: Adenoviral intramuscular gene transfers were performed into rabbit hindlimbs. Confocal and multiphoton microscopy were used for blood vessel imaging. Signaling experiments and gene expression analyses were performed to study mechanisms of Slit2 action. Slit2 decreased VEGFR2-mediated vascular permeability. It also reduced VEGFR2-mediated increase in blood perfusion and capillary enlargement, whereas sprouting of the capillaries was increased. Slit2 gene transfer alone did not have any effects on vascular functions or morphology. VEGFR2 activation was not affected by Slit2, but eNOS phosphorylation was diminished. The transcriptome profiling showed Slit2 downregulating angiogenesis-related genes such as nuclear receptor subfamily 4 group A member 1 (NR4A1) and Stanniocalcin-1 (STC-1) as well as genes related to endothelial cell migration and vascular permeability. Conclusions: Combining Slit2 with VEGFs adjusts VEGFR2-mediated angiogenic effects into a more physiological direction. This possibly allows the use of higher VEGF vector doses to achieve a more widespread vector and VEGF distribution in the target tissues leading to a better therapeutic outcome while reducing excess vascular permeability. Overall design: HUVEC mRNA profiles after adenoviral vector gene transfers in duplicate.","project":"SRP049805"}
{"number_samples":83,"species":"human","abstract":"Background: Sepsis involves aberrant immune responses to infection, but the exact nature of this immune dysfunction remains poorly defined. Bacterial endotoxins like lipopolysaccharide (LPS) are potent inducers of inflammation, which has been associated with the pathophysiology of sepsis, but repeated exposure can also induce a suppressive effect known as endotoxin tolerance or cellular reprogramming. It has been proposed that endotoxin tolerance might be associated with the immunosuppressive state that was primarily observed during late-stage sepsis. However, this relationship remains poorly characterised. Here we clarify the underlying mechanisms and timing of immune dysfunction in sepsis. Methods: We defined a gene expression signature characteristic of endotoxin tolerance. Gene-set test approaches were used to correlate this signature with early sepsis, both newly and retrospectively analysing microarrays from 593 patients in 11 cohorts. Then we recruited a unique cohort of possible sepsis patients at first clinical presentation in an independent blinded controlled observational study to determine whether this signature was associated with the development of confirmed sepsis and organ dysfunction. Findings: All sepsis patients presented an expression profile strongly associated with the endotoxin tolerance signature (p < 0.01; AUC 96.1%). Importantly, this signature further differentiated between suspected sepsis patients who did, or did not, go on to develop confirmed sepsis, and predicted the development of organ dysfunction. Interpretation: Our data support an updated model of sepsis pathogenesis in which endotoxin tolerance-mediated immune dysfunction (cellular reprogramming) is present throughout the clinical course of disease and related to disease severity. Thus endotoxin tolerance might offer new insights guiding the development of new therapies and diagnostics for early sepsis. Overall design: For the RNA-Seq study reported here, 73 patients were recruited with deferred consent at the time of first examination in an emergency ward based on the opinion of physicians that there was a potential for the patient's condition to develop into sepsis. These were retrospectively divided into groups based on clinical features and compared to 11 non-urgent surgical controls.","project":"SRP049820"}
{"number_samples":6,"species":"human","abstract":"Transcription by RNA Polymerase II (Pol II) in metazoan is regulated in several steps, including  preinitiation complex (PIC) formation, initiation, Pol II escape, productive elongation, cotranscriptional  RNA-processing and termination. Genome-wide studies have demonstrated that  the phenomenon of promoter-bound Pol II pausing is widespread, especially for genes involved  in developmental and stimulus-responsive pathways. However, a mechanistic understanding of  the paused Pol II states at promoters is limited. For example, at a global level, it’s unclear to  what extent the engaged paused Pol II is stably tethered to the promoter or undergoes rapid  cycles of initiation and termination. Here we used the small molecule Triptolide (TPL), an  XPB/TFIIH inhibitor, to block transcriptional initiation followed by measuring Pol II occupancy  by ChIP-seq. This inhibition of initiation enables us to investigate different states of paused Pol  II. Specifically, our global analysis reveals that most genes with paused Pol II, as defined by  pausing index, show significant clearance of Pol II during the period of TPL treatment. Our  study further identifies a group of genes with unexpectedly stably-paused Pol II, with unchanged  Pol II occupancy even after one hour of inhibition of initiation. This group of genes constitutes a  small portion of all paused genes defined by the conventional criterion of pausing index. These  findings could pave the way for evaluating the contribution of different elongation/pausing  factors on different states of Pol II pausing in developmental and other stimulus-responsive  pathways. Overall design: ChIP-Seq of total/Ser5P Pol II in HCT116 cells treated with DMSO or TPL in serum starvation/activation or normal conditions. Nascent RNA-seq in HCT116 cells treated with DMSO or TPL in starved condition.","project":"SRP049823"}
{"number_samples":4,"species":"human","abstract":"Dynamic interactions of nuclear lamins with chromatin through so-called lamin-associated domains (LADs) contribute to spatial arrangements of the genome. Here, we provide evidence for pre-patterning of differentiation-driven formation of lamin A/C LADs by domains of histone H2B modified by the nutrient sensor O-linked N-acetylglucosamine (H2BGlcNAc), which we term GADs. We demonstrate a two-step process of lamin A/C LAD formation during in vitro adipogenesis, involving (i) a spreading of lamin A/C-chromatin interactions during the transition from progenitor cell proliferation to cell cycle arrest, and (ii) a genome-scale redistribution these interactions through a process of LAD ‘exchange’ within hours of adipogenic induction. Chromatin state modeling reveals that lamin A/C LADs can be found both in active and repressive chromatin contexts which can be influenced by cell differentiation status. De novo formation of adipogenic lamin A/C LADs occurs non-randomly on GADs, consisting of megabase-size intergenic and repressive chromatin domains. Accordingly, while pre-differentiation lamin A/C LADs are gene-rich, post-differentiation LADs harbor repressive features reminiscent of lamin B1 LADs. Moreover, release of lamin A/C from genes directly involved in glycolysis concurs with their transcriptional upregulation after adipogenic induction, and with concordant downstream elevations in H2BGlcNAc levels and O-GlcNAc cycling. Our results unveil an epigenetic pre-patterning of adipogenic LADs by GADs, suggesting a coupling of developmentally regulated lamin A/C-genome interactions to a metabolically sensitive chromatin modification. Overall design: Examination of LMNA and H2BGlcNAc binding in ASCs across differentiation","project":"SRP049946"}
{"number_samples":4,"species":"human","abstract":"Nasopharyngeal carcinoma (NPC) has extremely skewed ethnic and geographic distributions, is poorly understood at the genetic level and is in need of effective therapeutic approaches. Here we determined the mutational landscape of 128 cases with NPC using whole-exome and targeted deep sequencing, as well as SNP array analysis. These approaches revealed a distinct mutational signature and nine significantly mutated genes, many of which have not been implicated previously in NPC. Notably, integrated analysis showed enrichment of genetic lesions affecting several important cellular processes and pathways, including chromatin modification, ERBB-PI3K signaling and autophagy machinery. Further functional studies suggested the biological relevance of these lesions to the NPC malignant phenotype. In addition, we uncovered a number of new druggable candidates because of their genomic alterations. Together our study provides a molecular basis for a comprehensive understanding of, and exploring new therapies for, NPC Overall design: Examination of mRNA expression levels in 4 individuals dignosed for Nasopharyngeal Carcinoma using RNA sequencing.","project":"SRP049967"}
{"number_samples":19,"species":"human","abstract":"We performed RNA-Seq analyses on 15 human fetal samples at 53-137 days of development, 9 female and 5 male, and identified the transcriptional changes during the transition of human cKIT+ primordial germ cells (PGCs), the precursors of gametes, to the generation of Advanced Germline Cells. Comparing the transcriptional profile of PGCs to that of H1 and UCLA1 hESCs identifies differences between the two cell types and pinpoints molecules that can be used in the development of in vitro germ cell differentiation protocols starting from human pluripotent stem cells. Overall design: RNA-Seq of cKIT+ cells analyzed from 6 biological samples for testes and 9 samples for ovaries from 53-137 days. 2 biological replicates of TRA-1-81+ cells sorted from H1 and UCLA1 hESCs. WGBS of cKIT+ cells analyzed from 4 biological samples of ovaries and 1 biological sample of testes at 57-137 days of development.","project":"SRP049981"}
{"number_samples":56,"species":"human","abstract":"Ets homologous factor (EHF) is an Ets family transcription factor expressed in many epithelial cell types including those lining the respiratory system. Disruption of the airway epithelium is central to many lung diseases, and a network of transcription factors coordinates its normal function. EHF can act as a transcriptional activator or a repressor, though its targets in lung epithelial cells are largely uncharacterized. RNA-seq after EHF depletion or overexpression showed significant alterations in the expression of genes involved in response to wounding. EHF knockdown also targeted genes in pathways of epithelial development and differentiation and locomotory behavior. Overall design: mRNA profiles from Calu-3 cells treated with negative control (NC) or EHF siRNA, in quintuplicate. mRNA profiles from 3 pcDNA (empty vector control) clones and 3 pcDNA-EHF overexpression A549 clones, 3-4 replicates each.","project":"SRP049988"}
{"number_samples":129,"species":"human","abstract":"Background: Sepsis, a leading cause of morbidity and mortality, is not a homogeneous disease but rather a syndrome encompassing many heterogeneous pathophysiologies. Patient factors including genetics predispose to poor outcomes, though current clinical characterizations fail to identify those at greatest risk of progression and mortality. Results: The Community Acquired Pneumonia and Sepsis Outcome Diagnostic study enrolled 1,152 subjects with suspected sepsis. We sequenced peripheral blood RNA of 129 representative subjects with systemic inflammatory response syndrome (SIRS) or sepsis (infection with SIRS), including 78 sepsis survivors and 28 sepsis nonsurvivors, who had previously undergone plasma proteomic and metabolomic profiling. The expression of 338 genes differed between subjects with SIRS and those with sepsis, primarily reflective of immune activation in sepsis. The expression of 1,238 genes differed with sepsis outcome: Nonsurvivors had lower expression of many immune function-related genes. Functional genetic variants associated with sepsis mortality were sought based on a common disease – rare variant hypothesis. VPS9D1, whose expression was increased in sepsis survivors, had a higher burden of missense variants in sepsis survivors, and these were associated with altered expression of 3,799 genes, primarily reflecting Golgi and endosome biology. Conclusions: Host response in sepsis survivors – activation of immune response-related genes – was muted in sepsis nonsurvivors. The association of sepsis survival with robust immune response and presence of missense variants in VPS9D1 warrants replication and further functional studies. Overall design: We sequenced peripheral blood RNA of 129 representative subjects with systemic inflammatory response syndrome (SIRS, n=23) or sepsis (infection with SIRS), including 78 sepsis survivors and 28 sepsis nonsurvivors, who had previously undergone plasma proteomic and metabolomic profiling.","project":"SRP050000"}
{"number_samples":14,"species":"human","abstract":"Intrahepatic cholangiocarcinoma (iCCA) is a fatal bile duct cancer with dismal prognosis and limited therapeutic options. By performing RNA- and exome sequencing analyses we have discovered a novel fusion event, FGFR2-PPHLN1 (16%), and damaging mutations in the ARAF oncogene (11%). Overall design: Methods: mRNA and gDNA were exctracted from fresh frozen tumor tissues and corresponding normal tissue (n=8 pairs) from patients with iCCA who underwent surgical resection. RNA-seq was performed using Illumina HiSeq 2500 System with 100 nucleotide single-end reads. One sample and its paired non-tumoral tissue were eliminated from the subsequent analysis because of bad RNa quality. The same 8 paired tumors were also analyzed by whole-exome seq. *** Submitter confirms there are no patient privacy concerns with these data. ***","project":"SRP050003"}
{"number_samples":25,"species":"human","abstract":"We have compared the proteome, transcriptome and metabolome of two isogenic cell lines: MCF-10A, derived from human breast epithelium, and the mutant MCF-10A-H1047R. These cell lines differ by a single amino acid substitution (H1047R) caused by single nucleotide change in one allele of the PIK3CA gene which encodes the catalytic subunit p110a of phosphatidylinositol 3-kinase (PI3K). The H1047R mutation of PIK3CA is one of the most frequently encountered somatic cancer-specific mutations. In MCF-10A, this mutation induces an extensive cellular reorganization that far exceeds the known signaling activities of PI3K. The changes are highly diverse; with examples in structural protein levels, the DNA repair machinery and sterol synthesis. Gene set enrichment analysis reveals a highly significant concordance of the genes differentially expressed in MCF-10A-H1047R cells and the established protein and RNA signatures of basal breast cancer. No such concordance was found with the specific gene signatures of other histological types of breast cancer. Our data document the power of a single base mutation, inducing an extensive remodeling of the cell toward the phenotype of a specific cancer. Overall design: 2 cell lines (H1047R and WT), 4 time points (0, 6, 12, 24 hours), 3 replicates","project":"SRP050036"}
{"number_samples":5,"species":"human","abstract":"The endoribonuclease Dicer is known for its central role in the biogenesis of eukaryotic small RNAs/microRNAs. Despite its importance, Dicer target transcripts have not been directly mapped. Here, we apply biochemical methods to human cells and C. elegans and identify thousands of Dicer binding sites. We find known and hundreds of novel miRNAs with high sensitivity and specificity. We also report structural RNAs, promoter RNAs, and mitochondrial transcripts as Dicer targets. Interestingly, most Dicer binding sites reside on mRNAs/lncRNAs and are not significantly processed into small RNAs. These passive sites typically harbor small, Dicer-bound hairpins within intact transcripts and generally stabilize target expression. We show that passive sites can sequester Dicer and reduce microRNA expression. mRNAs with passive sites were in human and worm significantly associated with processing-body/granule function. Together, we provide the first transcriptome-wide map of Dicer targets and suggest conserved binding modes and functions outside the miRNA pathway. Overall design: Regulatory impact of Dicer binding was assessed by knock down experiments in human HEK293 cells. Drosha knockdown and mock transfections were used as controls. Knockdown was performed with two independent siRNAs each. In total 5 samples.","project":"SRP050055"}
{"number_samples":6,"species":"human","abstract":"The AML1-ETO fusion protein, a transcription factor generated by the t(8;21) translocation in acute myeloid leukaemia (AML), dictates a leukemic program by increasing self-renewal and inhibiting differentiation.  Here we demonstrate that the histone demethylase JMJD1C functions as a co-activator for AML1-ETO and is required for its transcriptional program. JMJD1C is directly recruited by AML1-ETO to its target genes and regulates their expression by maintaining low H3K9me2 levels. Analyses in JMJD1C knockout mice also establish a JMJD1C requirement for AML1-ETO’s ability to increase proliferation. We also show a critical role for JMJD1C in the survival of multiple human AML cell lines, suggesting that it is required for leukemic programs in different AML cell types through its association with key transcription factors. Overall design: Examination of RNA expression when Kasumi-1 cells are treated with control shRNA or two different JMJD1C shRNAs; in duplicate. Please note that the 'shAML1_ETO_vs_shControl.all_gene_exp.tb.txtl' was generated comparing control and shRNA treated RNA abundance-using previously published data [GSE43834; GSM1071857 and GSM1071852].","project":"SRP050058"}
{"number_samples":6,"species":"human","abstract":"Total RNA extracted from prostate cancer LNCaP cells transfected with siRNA against CTCF(siCTCF), or negative control siRNA (si-)were processed, and sequenced by two different companies using Illumina Hi-seq 2000 platform to generate RNA sequencing with two output sequences: paired-end 50bp and 101bp in read length. Nearly 100 million and 50 million raw reads were yielded from each sample respectively. We used FastQC to confirm the quality of raw fastq sequencing data, and SOAPfuse software to detect fusion transcripts. Overall design: Discovering fusion genes from siCTCF and si- in LNCaP cells.","project":"SRP050061"}
{"number_samples":3,"species":"human","abstract":"To assay the effect of depletion of the RNA exosome on RNAs shorter than the standard length captured by RNA-seq (>200 nt), we created RNA-seq libraries using fragmented RNA and a linker-ligation-based protocol that does not deplete RNAs shorter than 200 nt. Note: these data relate to Figure 6E in Lubas, Andersen et al., Cell Reports 2014 (accepted) Overall design: These samples constitute RNA-seq libraries prepared to enrich for short RNA fragments such as snRNA and snoRNAs. Three different HeLa cell RNAi experiments were used to generate the RNA samples applied in the library construction: control transfected, hRRP40-depleted and triple-depleted of the known RNA exosome-associated ribonucleases (DIS3, DIS3L and hRRP6 knock-down). By these means the data offers reveal RNA exosome substrates via their up-regulation in the respective knock-downs NOTE: The 'Figure6E_RNAseq_DataTable_PlottedValues.txt' was generated from total 5 samples, with two additional published samples [SRP031620] and provided to better allow readers to fully replicate the analyses presented in the publication.","project":"SRP050074"}
{"number_samples":16,"species":"human","abstract":"To comprehensively characterize microRNAs (miRNA) expression and their target genes in thyroid cancer, we performed next-generation sequencing expression analysis of this disease. Recent studies have found that only the most abundant microRNAs mediate significant target suppression. We sequenced small RNA from 8 papillary thyroid carcinomas (PTC) with paired samples of normal thyroid tissue. We found that only a small set of abundant miRNAs are differentially expressed after pair-wise comparison (12 upregulated and 8 downregulated) reaching the minimum threshold amount to repress target mRNAs. We integrated computational prediction of potential targets and mRNA sequencing from the paired normal and tumor thyroid tissues from the same eight patients with PTC. The integrated analyses identified a master microRNA regulatory network in PTC that is involved in essential biological processes such as thyroid differentiation. As both mature products of miR-146b (miR-146b-5p and -3p) were among the most abundant upregulated in tumors, we unveil their target genes and found that miR-146b-3p specifically binds to the 3`UTR of PAX8 and NIS, leading to an impaired translation of the proteins and subsequently decreasing the iodide uptake of the cells.  Furthermore, we show that mir-146b and PAX8 regulate each other, describing a novel regulatory circuit that determines the differentiated phenotype of PTC. In conclusion, our integrative genomic analysis uncovers the target genes of two of the most upregulated miRNAs and highlights the importance of a miR-146b3p-PAX8-NIS regulatory circuit that determines thyroid differentiation in thyroid cancer. Overall design: Samples from Papillary Thyroid Carcinoma tumors (n=8) and contralateral normal thyroid tissue from the same patient (n=8) were collected at the Biobank of the Hospital Universitario La Paz (Madrid, Spain). The clinical characteristics of patients are summarized in Table S1. Surgically removed tissues were quickly frozen in liquid nitrogen until analysis. The samples were snap frozen on dry ice and stored at -80°C.","project":"SRP050087"}
{"number_samples":10,"species":"human","abstract":"We constructed a genome wide target profile of hsa-miR-191 by sequencing RNA isolated from Ago2 immunoprecipitations and total RNA samples following miR-191 transfection Overall design: Examination of mRNA levels in HeLa cells and Ago2 immunoprecipitations from HeLa cells following miR-191 mature duplex or control siRNA transfection","project":"SRP050138"}
{"number_samples":17,"species":"human","abstract":"The splicing factor SF3B1 is the most commonly mutated gene in the myelodysplastic syndromes (MDS), particularly in patients with refractory anemia with ring sideroblasts (RARS). MDS is a disorder of the hematopoietic stem cell and we thus studied the transcriptome of CD34+ cells from MDS patients with SF3B1 mutations using RNA-sequencing. Genes significantly differentially expressed at the transcript and/or exon level in SF3B1 mutant compared to wildtype cases include genes involved in MDS pathogenesis (ASXL1, CBL), iron homeostasis and mitochondrial metabolism (ALAS2, ABCB7, SLC25A37) and RNA splicing/processing (PRPF8, HNRNPD). Many genes regulated by a DNA damage-induced BRCA1-BCLAF1-SF3B1 protein complex showed differential expression/splicing in SF3B1 mutant cases. Our data indicate that SF3B1 plays a critical role in MDS by affecting the expression and splicing of genes involved in specific cellular processes/pathways, many of which are relevant to the known RARS pathophysiology, suggesting a causal link. Overall design: RNA-Seq was performed to compare the transcriptome of bone marrow CD34+ cells from eight MDS patients with SF3B1 mutation, four MDS patients with no known splicing mutation and five healthy controls.","project":"SRP050146"}
{"number_samples":24,"species":"human","abstract":"We report HERV-K rec iCLIP-seq binding data, ribosome profiling data, and RNA-seq from ELF1 naïve hESC and RNA-seq from NCCIT cells. Overall design: HERV-K Rec iCLIP-seq: 2 replicates in NCCIT. Ribosome profiling: 4 replicates each of Rec-overexpressing NCCIT vs. control NCCIT; RNAseq: 3 replicates each of HERV-K Rec siRNA vs. control siRNA in NCCIT; RNA-seq: 3 replicates each of ELF1 naïve hESC vs. primed hESC.","project":"SRP050147"}
{"number_samples":12,"species":"human","abstract":"The identification of RNA editing has proven to be a challenging task. Mapping and alignment artifacts hamper robust detection of RNA editing sites. RNA editing sites are characterized by RNA-DNA-differences (RDDs) that are mediated by two well known classes of RNA editing enzymes. The most prominent class of ADARs (Adeosine deaminases that act on RNA) catalyzes the hydrolytic deamination of adenosine to inosine which is recognized as guanosine by the translational and transcriptional machinery. RNA editing sites are identified by means of A->G substitution in RNS-seq assays. The less frequent C-to-U editing is mediated by the family of APOBECs (apolipoprotein B mRNA editing enzymes, catalytic polypeptide-like) – APOBEC3 has been shown to edit single stranded DNA as a means of defending against foreign DNA. In this study, we have conducted knockdown experiments in HEK293 of editing enzymes to identify edited sites by means of comparing RNA-seq samples of total RNA. In one assay ADAR1 and ADAR2 have been knockdown by using known siRNAs from the literature. In the second assay a set of siRNAs has been applied to knockdown APOBEC3 (B, C, F).","project":"SRP050149"}
{"number_samples":45,"species":"human","abstract":"Senescent human fibroblasts were compared to young proliferating fibroblasts. Five different cell lines were compared. Illumina sequencing (HiSeq2000) was applied to generate 50bp single-end reads. Overall design: 48 samples: 3 biological replicates for each group: young proliferating and senescent BJ cells;  young proliferating and senescent Wi-38 cells;  young proliferating and senescent IMR-90 cells; 5 population doubling from young proliferating to senescent cell for HFF and MRC-5 cells","project":"SRP050179"}
{"number_samples":2,"species":"human","abstract":"Comparison between mesenchymal and epithelial population within the CD44+/CD24- population of the SRC transformed MCF10A cells.","project":"SRP050191"}
{"number_samples":28,"species":"human","abstract":"Triple negative breast cancer (TNBC) is a heterogeneous and clinically aggressive disease for which there is no targeted therapy. Here we report the preferential and high sensitivity of TNBCs to BET bromodomain inhibitors such as JQ1 manifested by cell cycle arrest in early G1, apoptosis, and induction of markers of luminal epithelial differentiation in vitro and in vivo. The sensitivity of TNBC and other tumor types to BET inhibition establishes a rationale for clinical investigation, and a motivation to understand mechanisms of resistance. After engendering acquired resistance to BET inhibition in previously sensitive TNBCs, we utilized integrative approaches to identify a unique mechanism of epigenomic resistance to this epigenetic therapy. Resistant cells remain dependent on BRD4, confirmed by RNA interference. However, TNBC cells adapt to BET bromodomain inhibition by re-recruitment of unmutated BRD4 to super-enhancers, now in a bromodomain-independent manner. Proteomic studies of resistant TNBC identify hyper-phosphorylation of BRD4 and strong association with MED1. Together, these studies provide a rationale for BET inhibition in TNBC and argue for combination strategies to anticipate clinical drug resistance. Overall design: RNA-Seq in parental and JQ1 resistant triple negative breast cancer (TNBC) in response to DMSO or JQ1 treatment over time","project":"SRP050193"}
{"number_samples":12,"species":"human","abstract":"Transcription and pre-mRNA alternative splicing was analyzed by  in isogenic human HEK293 cell lines that inducibly express a-amanitin resistant mutants of the RNA polymerase II large subunit with slow and fast elongation rates. Overall design: Alternative splicing was analyzed by, RNA-seq and RASL-seq of polyA+ RNA from a-amanitin treated cells; transcription elongation rate was analyzed by BrUTP-labelled GRO-seq of a-amanitin treated cells at time points after release from a DRB (5,6-dichloro-1-bold beta-D-ribofuranosylbenzimidazole) block.","project":"SRP050215"}
{"number_samples":402,"species":"human","abstract":"Purpose: The purpose of this study is to identify functionally inter-connected group of miRNAs whose reduced expression promotes leukemia development in vivo. We searched for relevant target genes of these miRNAs that are upregulated in T-ALL relative to controls. Methods: In order to examine the global gene expression, we generated 9 T-ALL patients and 4 normal controls by deep sequencing using Illumina Hi-Seq sequencer. The sequence reads that passed quality filters were analyzed using Spliced Transcripts Alignment to a Reference aligner (STAR) followed by differential gene expression analysis using DESeq. Results: Using an optimized data analysis workflow, we mapped reads per sample to the human genome (build hg19) and identified transcripts in both patient and controls with STAR workflow. We applied a machine learning approach to eliminate targets with redundant miRNA-mediated control. This strategy finds a convergence on the Myb oncogene and less prominent effects on the Hpb1 transcription factor. The abundance of both genes is increased in T-ALL and each can promote T-ALL in vivo. Conclusion: Our study reveals a Myc regulated network of tumor suppressor miRNAs in T-ALL. We identified a small number of functionally validated tumor suppressor miRNAs.  These miRNAs are repressed upon Myc activation and this links their expression directly to Myb a key oncogenic driver in T-ALL. Overall design: Examination of global gene expression in 9 T-ALL patients and 4 normal controls using total RNA sequencing. BaseMeanA in DESeq_results.xlsx is the control.","project":"SRP050223"}
{"number_samples":12,"species":"human","abstract":"Using xenograft-based experimental evolution, we characterize the full life history from initiation to metastasis of a tumor at the genomic and transcriptomic levels.","project":"SRP050242"}
{"number_samples":6,"species":"human","abstract":"We report a negative correlation of invasiveness with REST expression. In addition, one alternatively spliced product (ASP) of REST, REST-003, shows a positive correlation with invasiveness. REST has a well-established role in regulating transcription of genes important for neuronal development.  Its role in cancer, though significant, is less well understood.  We would like to investigate the effect of REST on invasive phenotype.  In order to do so, we downregulate REST by siRNA treatment in weakly invasive MCF-7 breast cancer cells in which REST is expressed highly: 1) si-GAPDH (control), two si-RESTs (2)si-REST_1 and 3) si-REST_2).  Conversely, we  overexpress REST by transfection of wt-REST cDNA in highly invasive MDA-MB-231 cells in which REST is expressed at the low level:  4) EGFP (control), 5) mt-REST (another control) and 6) wt-REST. REST (repressor element-1 (RE-1) silencing transcription factor) contains a DNA-binding domain that is localized within eight zinc fingers and two repressor domains located at the N-terminal and C-terminal, respectively.   REST suppresses expression of neural-specific genes.  mt-REST lacks two repressor domains, so it can be used as a control for wt-REST.  In contrast, REST-003 is one of alternatively spliced products (ASPs) of REST. Overall design: Downregulation of REST using two siRNAs in MCF-7 cells or overexpression of REST with wt- or mt-REST cDNA in MDA-MB-231 cells REST_ISOFORM_FIG1.pdf was provided to explain *rawRestIsoformCnts.txt files described in the README1.txt file.","project":"SRP050245"}
{"number_samples":4,"species":"human","abstract":"Fifty six genes from DESeq were differentially expressed in the treated versus control samples.  More than 20% were related to immune, defense, wounding and inflammatory responses Overall design: Downregulation of  REST-003 using siRNAs in MDA-MB-231 cells; we used siRNA against REST-003, as REST-003 may control invasiveness.  We transfected si-Control (scramble) or si-REST-003 in MDA-MB-231:  duplicate of both (total 4 samples).","project":"SRP050246"}
{"number_samples":24,"species":"human","abstract":"Gene expression study of MDA-MB-231 and in HBL-100 cell lines exposed to the antimetastatic drug NAMI-A","project":"SRP050249"}
{"number_samples":6,"species":"human","abstract":"Here we present the whole genome ChIP-Seq analyses of a wide variety of histone marks, H3K27ac, H3K4me1, H3K4me3, and H3K27me3 in the brain, heart, and liver, along with the RNA-seq data of these organs of early human embryos 12 weeks after gestation. Overall design: In total, brain, heart, and liver of early human post-implantation embryos were used, and four histone modifications were detected, including H3K27ac, H3K4me1, H3K4me3 and H3K27me3. Also, the transcriptomes of these three organs were analyzed.","project":"SRP050260"}
{"number_samples":83,"species":"human","abstract":"Long noncoding RNAs (lncRNAs) are transcripts longer than 200 nucleotides located within the intergenic stretches or overlapping antisense transcripts of protein coding genes. LncRNAs are involved in numerous biological roles including imprinting, epigenetic regulation, apoptosis and cell-cycle. To determine whether lncRNAs are associated with clinical features and recurrent mutations in older patients (aged =60 years) with cytogenetically normal (CN) acute myeloid leukemia (AML), we evaluated lncRNA expression in 148 untreated older CN-AML cases using a custom microarray platform. Overall design: In this study, we analyzed a large set of older CN-AML patients using custom lncRNA microarrays to investigate whether lncRNA expression is associated with clinical features, molecular abnormalities and outcome and to build a prognostic lncRNA signature that was subsequently validated using RNA sequencing. This submission represents RNA-Seq component of study.","project":"SRP050272"}
{"number_samples":48,"species":"human","abstract":"Analysis of C4-2 Prostate cancer cell line after 72 hours of knockdown. CHKA is overexpressed in a number of solid tumours, including prostate cancer. Results provide insight into the molecular mechanisms of CHKA in prostate carcinogenesis. Overall design: This experiment was designed to understand the regulation of transcriptome by Choline kinase alpha (CHKA) which is an important enzyme in Kennedy pathway. In order to achieve this, the endogenous protein was knocked down using siRNA pool that targets the CHKA mRNA.","project":"SRP050331"}
{"number_samples":8,"species":"human","abstract":"We employed high-throughput sequencing of both short (~18-24nt) and long (>200nt) RNAs in human erythrocytes. We obtained blood from five healthy individuals for the short (small) RNA-seq library preparations and blood from three individuals for the long RNA-seq library preparations. We identified an abundant, diverse population of RNAs. Both polyadenylated and nonpolyadenlated long RNAs were identified. Additionally, known and novel microRNAs were identified in the short RNA dataset using the probabalistic modeling algorithm miRDeep. These RNAs lend insight into erythrocyte biology and provide utility as potential biomarkers. To determine both shared and unique aspects of the erythrocyte long RNA transcriptome, we compared this transcriptome with that of the PBMC and CD34+ erythroid progenitor transcriptomes. Overall design: Sequencing of RNAs from mature erythrocytes of healthy individuals","project":"SRP050333"}
{"number_samples":4,"species":"human","abstract":"This study's goal is to perform gene expression profiling using RNA-seq on FFPE tissue, to compare with that from fresh frozen tissue, and to evaluate the feasibility of RNA-seq for discovery and validation of biomarkers.","project":"SRP050335"}
{"number_samples":10,"species":"human","abstract":"A basal (MDAMB468) and luminal (ZR75-1) cell line were treated with DMSO or PKC412 for 6h Overall design: 2 DMSO and 3 PKC412 treated samples for each cell line","project":"SRP050365"}
{"number_samples":17,"species":"human","abstract":"We sequenced mRNA from a total of 12 samples (6 different cell types, each with two biological replicates) to infer the relationship among those cell types Overall design: Examination of mRNA levels in six different human cell types grown in culture with two biological replicates for each cell type","project":"SRP050374"}
{"number_samples":15,"species":"human","abstract":"Constitutive low level DNA damage in RNASEH2 deficiency is linked to innate immune activation. Hierarchical clustering of over 16000 transcripts revealed remarkably similar profiles in patients with lupus erythematosus and patients with AGS with up-regulation of genes involved in DNA damage signaling and type I-IFN signaling. Overall design: Comparison of transcriptional profiles of native RNASEH2-deficient patient fibroblasts with wild type cells.","project":"SRP050397"}
{"number_samples":62,"species":"human","abstract":"Following the discovery of BRD4 as a non-oncogene addiction target in acute myeloid leukemia (AML), BET inhibitors are being explored as promising therapeutic avenue in numerous cancers. While clinical trials have reported single-agent activity in advanced hematologic malignancies, mechanisms determining the response to BET inhibition remain poorly understood. To identify factors involved in primary and acquired BET resistance in leukemia, we performed a chromatin-focused shRNAmir screen in a sensitive MLL/AF9; NrasG12D-driven AML model, and investigated dynamic transcriptional profiles in sensitive and resistant murine and human leukemias. Our screen reveals that suppression of the PRC2 complex, contrary to effects in other contexts, promotes BET resistance in AML. PRC2 suppression does not directly affect the regulation of Brd4-dependent transcripts, but facilitates the remodeling of regulatory pathways that restore the transcription of key targets such as Myc. Similarly, while BET inhibition triggers acute MYC repression in human leukemias regardless of their sensitivity, resistant leukemias are uniformly characterized by their ability to rapidly restore MYC transcription. This process involves the activation and recruitment of WNT signaling components, which compensate for the loss of BRD4 and drive resistance in various cancer models. Dynamic ChIP- and STARR-seq enhancer profiles reveal that BET-resistant states are characterized by remodeled regulatory landscapes, involving the activation of a focal MYC enhancer that recruits WNT machinery in response to BET inhibition. Together, our results identify and validate WNT signaling as a driver and candidate biomarker of primary and acquired BET resistance in leukemia, and implicate the rewiring of transcriptional programs as an important mechanism promoting resistance to BET inhibitors and, potentially, other chromatin-targeted therapies. Overall design: RNA-Seq of DMSO- or JQ1-treated cancer cell lines; ChIP-seq for H3K36me3 and H3K27me3 in a leukemia cell line treated either with DMSO or JQ1, ChIP-seq for H3K27ac in resistant and sensitive mouse and human leukemia. Functional enhancer mapping (STARR-seq) in K-562 treated either with DMSO or JQ1.","project":"SRP050440"}
{"number_samples":8,"species":"human","abstract":"To elucidate miRNA-mediated temporal crosstalk during productive infection, we identified genome-wide miRNA target sites using Argonaute-crosslinking and immunoprecipitation followed by high-throughput sequencing (AGO-CLIPseq) in human cytomegalovirus (HCMV)-infected cells and evaluated the targeting efficacy by applying our new AGO-CLIPseq enrichment (ACE)-scoring algorithm. Overall design: To uncover the miRNA targetome in uninfected or infected human foreskin fibroblasts with HCMV (24, 48 and 72 post-infection hour) were subjected to take AGO-CLIPseq as well as mRNAseq/smallRNAseq.","project":"SRP050468"}
{"number_samples":12,"species":"human","abstract":"Stabilin-1/CLEVER-1 is a multidomain protein present in lymphatic and vascular endothelial cells and in M2 immunosuppressive macrophages. Stabilin-1 functions in scavenging, endocytosis and leukocyte adhesion to and transmigration through the endothelial cells. We have analyzed the putative functions of Stabilin-1 in blood monocytes and found that in healthy individuals 60-80% of both CD14+CD16- and CD14+C16+ monocytes, but not CD14dimCD16+ monocytes, expressed Stabilin-1 on the surface. Microarray and RNAseq analysis was performed to get more insight into the effect of Stabilin-1 expression on human monocytes transcriptome. Overall design: The transcriptome of human monocytes transfected with Stabilin-1 siRNA was compared to that of control siRNA transfected monocytes","project":"SRP050490"}
{"number_samples":22,"species":"human","abstract":"Somatic mutations in the spliceosome gene ZRSR2— located on the X chromosome — are associated with myelodysplastic syndrome (MDS). ZRSR2 is involved in the recognition of 3? splice site during the early stages of spliceosome assembly; however, its precise role in RNA splicing has remained unclear. Here, we characterize ZRSR2 as an essential component of the minor spliceosome (U12-dependent) assembly. shRNA mediated knockdown of ZRSR2 leads to impaired splicing of the U12-type introns, and RNA-Sequencing of MDS bone marrow reveals that loss of ZRSR2 activity causes increased mis-splicing. These splicing defects involve retention of the U12-type introns while splicing of the U2-type introns remain mostly unaffected. ZRSR2 deficient cells also exhibit reduced proliferation potential and distinct alterations in myeloid and erythroid differentiation in vitro. These data identify a specific role for ZRSR2 in RNA splicing and highlight dysregulated splicing of U12-type introns as a characteristic feature of ZRSR2 mutations in MDS. Overall design: RNA sequencing was performed on 16 bone marrow samples (MDS and normal)  and six samples of control or ZRSR2 shRNA transduced TF-1 cells and data was analysed for aberrant splicing caused by ZRSR2 mutations/deficiency.","project":"SRP050493"}
{"number_samples":6,"species":"human","abstract":"We have compared the genome-wide effects on the transcriptome after treatment with ICG-001 (the specific CBP inhibitor) versus C646, a compound that competes with acetyl-coA for the Lys-coA binding pocket of both CBP and p300. We found that both drugs cause large-scale changes in the transcriptome of HCT116 colon cancer cells and PANC1 pancreatic cancer cells, and reverse some tumor-specific changes in gene expression. Interestingly, although the epigenetic inhibitors affect cell cycle pathways in both the colon and pancreatic cancer cell lines, the WNT signaling pathway was affected only in the colon cancer cells. Notably, WNT target genes were similarly down-regulated after treatment of HCT116 with C646 as with ICG-001. Overall design: To identify genes affected by direct targeting of a component of the transcriptional complex implicated in WNT regulation, we used siRNAs to knockdown TCF7L2 in PANC1 cells. Cells were treated with control siRNAs or siRNAs specific for TCF7L2 and RNA was analyzed by RNA-seq.","project":"SRP050497"}
{"number_samples":328,"species":"human","abstract":"Germ cells are vital for transmitting genetic information from one generation to the next and for maintaining the continuation of species. Here, we analyze the transcriptome of human primordial germ cells (PGCs) from the migrating stage to the gonadal stage at single-cell and single-base resolutions. Human PGCs show unique transcription patterns involving the simultaneous expression of both pluripotency genes and germline-specific genes, with a subset of them displaying developmental stage-specific features. Furthermore, we analyze the DNA methylome of human PGCs and find global demethylation of their genomes. Approximately 10-11 weeks after gestation, the PGCs are nearly devoid of any DNA methylation; with only 7.8% and 6.0% of the median methylation levels in male and female PGCs, respectively. Our work paves the way toward deciphering the complex epigenetic reprogramming of the germline with the aim of restoring totipotency in fertilized oocytes. Overall design: In total, we analyzed the transcriptomes of 233 individual male and female human PGCs from 15 embryos at between 4 and 19 weeks of gestation as well as 86 neighboring somatic cells from 13 of these embryos using a single-cell RNA-Seq method that we developed. Furthermore, we analyzed the DNA methylomes of both male and female human gonadal PGCs as well as the neighboring somatic cells of eleven of these embryos at between 7 and 19 weeks of gestation using whole-genome bisulfite sequencing (WGBS). In total, we generated 1.3 billion 100 bp paired-end reads for transcriptome analyses and 4.1 billion 100 bp or 150 bp paired-end reads for DNA methylome analyses.","project":"SRP050499"}
{"number_samples":4,"species":"human","abstract":"Analysis of gene expression of Pdx-EGFP1+ pancreatic progenitors before or after co-culture at mRNA level. The hypothesis tested in the study was that the overall gene expression in Pdx1-EGFP+ does not alter after co-culture with endothelial cells. The result supported our hypothesis. Overall design: Total RNA isolated from Pdx1-EGFP+ progenitors from the Pdx1-EGFP HUES8 cell-derived pancreatic progenitor population before (none) and after co-culture (AKT-HUVEC, MPEC, or BJ) Fig 2d in publication.","project":"SRP050533"}
{"number_samples":2,"species":"human","abstract":"Analysis of overall gene expression in endothelial cells, MPEC and AKT-HUVEC, comparing to a human fibroblast line, BJ. We are looking for the highly abundant genes in endothelial cells comparing to fibroblasts. Overall design: Total RNA isolated from MPEC, AKT-HUVEC and BJ Fig 3b in publication.","project":"SRP050534"}
{"number_samples":4,"species":"human","abstract":"Divergent transcription, in which reverse-oriented transcripts occur upstream of eukaryotic promoters in regions devoid of annotated genes, has been suggested to be a general property of active promoters. Here we show that the human basal RNA polymerase II transcriptional machinery and core promoter are inherently unidirectional, and that reverse-oriented transcripts originate from their own cognate reverse-directed core promoters. In vitro transcription analysis and mapping of nascent transcripts in cells revealed that core promoters are unidirectional and that sequences at reverse start sites are similar to those of their forward counterparts. The use of DNase I accessibility to define proximal promoter borders revealed that about half of promoters are unidirectional and that these unidirectional promoters are depleted at their upstream edges of reverse core promoter sequences and their associated chromatin features. Divergent transcription is thus not an inherent property of the transcription process, but rather the consequence of the presence of both forward- and reverse-directed core promoters. Overall design: Using 5'-GRO-seq and GRO-seq to determine mechanisms of divergent transcription initiation","project":"SRP050563"}
{"number_samples":7,"species":"human","abstract":"Germline Gain-of-Function Mutations in AFF4 Cause a Novel Syndrome and Functionally Link the Super Elongation Complex and Cohesin","project":"SRP050576"}
{"number_samples":7,"species":"human","abstract":"Skeletal myocytes are metabolically active and susceptible to insulin resistance, thus implicated in type 2 diabetes (T2D). This complex disease involves systemic metabolic changes and their elucidation at the systems level requires genome-wide data and biological networks. Genome-scale metabolic models (GEMs) provide a network-context to integrate high-throughput data. We generated myocyte-specific RNA-seq data and investigated their correlation with proteome data. These data were then used to reconstruct a comprehensive myocyte GEM. Next, we performed a meta-analysis of six studies comparing muscle transcription in T2D versus healthy subjects. Transcriptional changes were mapped on the myocyte GEM, revealing extensive transcriptional regulation in T2D, particularly around pyruvate oxidation, branched-chain amino acid catabolism, and tetrahydrofolate metabolism, connected through the down-regulated dihydrolipoamide dehydrogenase. Strikingly, the gene signature underlying this metabolic regulation successfully classifies the disease state of individual samples, suggesting that regulation of these pathways is a ubiquitous feature of myocytes in response to T2D. Overall design: Isolated skeletal muscle precursor cells from six normal glucose tolerant and non-obese males and females were differentiated in vitro. RNA from fully differentiated myotubes was sequenced using RNA-seq.","project":"SRP050596"}
{"number_samples":303,"species":"human","abstract":"Human pluripotent stem cell-based in vitro models that reflect human physiology have the potential to reduce the number of drug failures in clinical trials, and offer a cost effective approach for assessing chemical safety.  Here, human embryonic stem (ES) cell-derived neural progenitor cells, endothelial cells, mesenchymal stem cells, and microglia/macrophage precursors were combined on chemically-defined poly(ethylene glycol) (PEG) hydrogels and cultured in serum-free media to model cellular interactions of the developing brain.  The precursors self-assembled into 3-dimensional (3D) neural constructs with cortically organized neuronal and glial cells, interconnected vascular networks, and ramified microglia. Replicate constructs were highly reproducible by RNA sequencing (Spearman’s correlation coefficients, ? = 0.97) and robustly expressed neurogenesis, vasculature development, and microglia genes.  Finally, linear support vector machines were used to construct a predictive model from RNA sequencing data for 240 neural constructs treated with 60 toxic and non-toxic chemicals, which then correctly classified 9/10 blinded compounds. Overall design: Note that all cell types were derived from the H1 human embryonic stem cell line. 11 samples for initial quality control (triplicate day 13 neural progenitor cells; quadruplicate day 21 neural progenitor cells cocultured with mesenchymal stem cells and endothelial cells; quadruplicate day 21 neural progenitor cells cocultured with mesenchymal stem cells and endothelial cells and migroglia/macrophage precursor cells), quadruplicate samples of H1 ES cells as a control for comparing to untreated toxicity study samples, and 288 samples associated with toxicity screening (all samples formed using neural progenitor cells, endothelial cells, mesenchymal stem cells, and microglia/macrophage precursors).","project":"SRP050892"}
{"number_samples":4,"species":"human","abstract":"The human genome encodes a family of nine protein arginine methyltransferases (PRMT1-9). Different members of this enzyme family catalyze different types of protein methylation at the terminal nitrogen atoms of arginine residues, forming monomethylated arginine (MMA), asymmetrically dimethylated arginine (ADMA) and symmetrically dimethylated arginine (SDMA). The last member of this family, PRMT9, is characterized in detail here. We identify two spliceosome-associated proteins, SAP145 (SF3B2) and SAP49 (SF3B4), as PRMT9 binding partners, linking PRMT9 to U2snRNP maturation.  We show that SAP145 is methylated by PRMT9 at arginine 508 (R508). Amino acid analysis and a methyl-specific antibody revealed the formation of MMA and SDMA, and PRMT9 thus joins PRMT5 as the only mammalian enzymes that can deposit the SDMA mark. Methylation of the SAP145R508 generates a binding site for the Tudor domain of SMN, and RNA-seq analysis reveals gross splicing changes when PRMT9 levels are attenuated. These studies identify PRMT9 as a non-histone methyltransferase that primes the U2snRNP for interaction with SMN. Overall design: RNA sequencing was carried out using RNA samples from two biological replicates of control and PRMT9 knockdown HeLa cells.","project":"SRP050943"}
{"number_samples":16,"species":"human","abstract":"We hypothesize that the observed differences in incidences of pleural and peritoneal malignant mesothelioma (MM) are the result of differences in the direct response of these cell types to asbestos rather than to differences mediated by the in vivo microenvironment. To test this hypothesis, we characterized cellular responses to asbestos in a controlled environment using high-throughput RNA sequence and other assays. Overall design: Examination of asbestos-treated versus untreated mesothelial cells from four cell lines representing two tissue types in culture.","project":"SRP050954"}
{"number_samples":45,"species":"human","abstract":"Together with the GSE54456 data, we used in total RNA-seq data from 99 lesional psoriatic, 27 uninvolved psoriatic, and 90 normal skin biopsies and applied computational approaches to identify and characterize expressed lncRNAs. Overall design: 7 psoriatic, 27 uninvolved, and 8 normal skin samples","project":"SRP050971"}
{"number_samples":460,"species":"human","abstract":"Oscillatory gene expression is fundamental to mammalian development, but technologies to monitor expression oscillations are limited. We have developed a statistical approach called Oscope to identify and characterize the transcriptional dynamics of oscillating genes in single-cell RNA-seq data from an unsynchronized cell population. Applications to a number of data sets, include a single-cell RNA-seq data set of human embroyonic stem cells (hESCs), demonstrate advantages of the approach and also identify a potential artifact in the Fluidigm C1 platform. Overall design: Total 213 H1 single cells and 247 H1-Fucci single cells were sequenced. The 213 H1 cells were used to evaluate Oscope in identifying oscillatory genes. The H1-Fucci cells were used to confirm the cell cycle gene cluster identified by Oscope in the H1 hESCs. Normalized expected counts are provided in GSE64016_H1andFUCCI_normalized_EC.csv.gz","project":"SRP050992"}
{"number_samples":4,"species":"human","abstract":"To identify which genes were regulated by CIC in human prostate cancer cell, the effect of CIC knockdown on PC3 cell transcriptome was determined Overall design: The transcriptome profiles were obtained from control PC3 and shCIC treated PC3","project":"SRP051000"}
{"number_samples":2,"species":"human","abstract":"The project is part of an effort at the benchmarking of available tools for the estimation of isoform-level transcript abundances from RNA-seq data, a question of high current interest in high-throughput mRNA expression profiling. Data associated with this project include those from RNA and RNA 3' end sequencing experiments with human (Jurkat) and murine (NIH-3T3) cell lines as well as in silico-generated read sequences generated with Flux Simulator (doi:10.1093/nar/gks666; available at http://sammeth.net/confluence/display/SIM/Home). In particular, the following datasets are provided: raw short read data in FASTQ format and processed genome alignments in BAM format (used as input to benchmarked tools).","project":"SRP051039"}
{"number_samples":160,"species":"human","abstract":"Purpose: The aim of this study is to compare different RNA extraction methods using a mixture design that allows the relative changes of the majority of genes profiled to be estimated. A number of samples were degraded to allow us to compare methods for dealing with more variable samples. Methods - Cell Culture: Lung adenocarcinoma cell lines NCI-H1975 and HCC827 from a range of passages (2-4) were grown on 3 separate occasions in RPMI media (Gibco) supplemented with Glutamax and 10% fetal calf serum to a 70% confluence. To replicate common experimental conditions cell lines were treated with 0.01% Dimethyl sulfoxide (Sigma), which is commonly used as a vehicle in drug treatment experiments. After 6 hours of treatment, cells were collected, snap-frozen on dry ice and stored at -80 degrees C until required. Methods - RNA preparation: Total RNA was extracted from between half a million and million cells using Total RNA Purification Kit (Norgen Biotek) with on-column DNAse treatment accorting to the kit instructions. RNA concentration for each pair of samples to be mixed was equalised to ~100 ng/µl using Qubit RNA BR Assay Kit (Life Technologies). Replicates were pooled in known proportions to obtain mixtures ranging from pure NCI-H1975 (100:0) to pure HCC827 (0:100) and intermediate mixtures ranging from 75:25 to 50:50 to 25:75  NCI-H1975:HCC827. All mixtures corresponding to the second replicate were split into two equal aliquots. One aliquot was left intact (we refer to this as the 'good' replicate), while the second aliquot was degraded to produce known outlier samples by incubation at 37 degrees C for 7 days in a thermal cycler with a heated lid. 10 µl from each replicated mixture (both good and degraded) were used for Next Generation Sequencing library preparation using two kits: Illumina TruSeq Total Stranded RNA with Ribozero (TotalRNA) and Illumina TruSeq RNA v2 (mRNA) according to the manufacturer's instructions. Completed libraries were sequenced on HiSeq 2500 with TruSeq SBS Kit v4- HS reagents (Illumina) as 100 bp single-end reads at the Australian Genome Research Facility (AGRF), Melbourne. Approximately 30 million 100 bp single-end reads were obtained for each sample. Reads were aligned to the human reference genome hg19 and mapped to known genomic features at the gene level using the Rsubread package (version 1.16.1) (Liao et al. 2013). Single reads were then summarized into gene-level counts using FeatureCounts (Liao et al. 2014). Overall design: Total RNA was extracted from lung adenocarcinoma cell lines NCI-H1975 and HCC827 (3 independent samples for each cell line) and mixed in known ratios. Both mRNA and Total RNA transcriptomes from these mixtures were profiled by RNA-Seq.","project":"SRP051083"}
{"number_samples":40,"species":"human","abstract":"We compare the performance of two library preparation protocols (poly(A) and exome capture) in in vitro degraded RNA samples Overall design: VcaP cell were grown, and treated with MDV3100 (enzalutamide) or DHT (dihydrotestosterone), intact RNA was isolated and samples were prepared in technical triplicates using two library preparation protocol. Also cells were subject to in vitro degradation through incubation of the whole cell lysate in 37C for increasing amounts of time. Following incbation paired capture and poly(A) libraries were prepared.","project":"SRP051102"}
{"number_samples":5,"species":"human","abstract":"We used targeted RNA sequencing to identify gene fusions in various cell types","project":"SRP051118"}
{"number_samples":2,"species":"human","abstract":"Pol II RIP sequencing","project":"SRP051138"}
{"number_samples":2,"species":"human","abstract":"The inflammatory gene response requires activation of the protein kinase TAK1, but it is currently unknown how TAK1-derived signals coordinate transcriptional programs in the genome. We determined the genome-wide binding of the TAK1-controlled NF-?B subunit p65 in relation to active enhancers and promoters of transcribed genes by ChIP-seq experiments. Out of 35,000 active enhancer regions, 410 H3K4me1-positive enhancers show interleukin (IL)-1-induced H3K27ac and p65 binding. Inhibition of TAK1, IKK2 or depletion of p65 blocked inducible enhancer activation and gene expression. As exemplified by the CXC chemokine cluster located on chromosome 4, the TAK1-p65 pathway also regulates the recruitment kinetics of the histone acetyltransferase CBP, of NF-?B p50 and of AP-1 transcription factors to both, promoters and enhancers. This study provides a high resolution view of epigenetic changes occurring during the IL-1 response and allows the first genome-wide identification of a novel class of inducible p65 NF-?B-dependent enhancers in epithelial cells. Overall design: RNA-seq of KB cells either untreated or treated with IL-1 alpha","project":"SRP051170"}
{"number_samples":21,"species":"human","abstract":"Characterization of host-pathogen interactions is critical for the development of next-generation therapies and vaccines. Classical approaches involve the use of transformed cell lines and/or animal models which may not reflect the complexity and response of the human host. We reconstituted the ciliated human bronchial epithelium in vitro using primary bronchial epithelial cells to simultaneously monitor the infection-linked global changes in nontypeable Haemophilus influenzae (NTHi) and infected host epithelia gene expression by dual RNA-seq. Acquisition of a total of nearly 2,5 billion sequences allowed construction of high-resolution strand-specific transcriptome maps of NTHi during infection of host mucosal surface and monitoring of metabolic as well as stress-induced host-adaptation strategies of this pathogen. As a part of our screening, we identified a global profile of noncoding transcripts that are candidate small RNAs regulated during human host infection in Haemophilus species. Temporal analysis of host mRNA signatures revealed significant dysregulation of target cell cytoskeleton elicited by bacterial infection, with a profound effect on intermediate filament network of bronchial epithelium. Our data provide a robust and comprehensive catalogue of regulatory responses that drive NTHi pathogenesis and gives novel insights into complex crosstalk between the host and the invading pathogen. Overall design: Primary human bronchial epithelium was infected with NTHi at a multiplicity of 100:1. Total RNA was isolated at 1, 6, 24 and 72 h post-infection in three biologically-independent experiments and cDNA libraries were prepared and sequenced with Illumina HiSeq 2500 sequencer. At each time point, between 60 and 180 million total reads per sample were obtained of which approximately one-third could be aligned to non-rRNA regions of the bacterial and human genomes","project":"SRP051182"}
{"number_samples":22,"species":"human","abstract":"The proinflammatory cytokine tumor necrosis factor (TNF) plays a central role in low-grade adipose tissue inflammation and development of insulin resistance during obesity. In this context, nuclear factor kappa-light-chain-enhancer of activated B cells (NFkB), is directly involved and required for the acute activation of the inflammatory gene program. Here we show that the major transactivating subunit of NF?B, v-rel avian reticuloendotheliosis viral oncogene homolog A (RELA), is also required for acute TNF-induced suppression of adipocyte genes. Notably, this repression does not involve RELA binding to the associated enhancers but rather loss of cofactors and enhancer RNA (eRNA) selectively from high occupancy sites within super-enhancers. Based on these data we have developed models that with high accuracy predict which enhancers and genes are repressed by TNF in adipocytes. We show that these models are applicable to other cell types where TNF represses genes associated with super-enhancers in a highly cell type-specific manner. Our results propose a novel paradigm for NF?B-mediated repression, whereby NF?B selectively redistributes cofactors from high occupancy enhancers, thereby specifically repressing super-enhancer-associated cell identity genes. Overall design: Genome-wide assesment of the acute transcriptional response to TNF in human SGBS adipocytes using RNA- ChIP- and DHS-seq. Total RNA-seq and RNAPII-ChIP seq for vehicle and TNF treated adipocytes are available under GSE60462","project":"SRP051211"}
{"number_samples":35,"species":"human","abstract":"The pervasive expression of circular RNA from protein coding loci is a recently discovered feature of many eukaryotic gene expression programs.  Computational methods to discover and quantify circular RNA are essential to the study of the mechanisms of circular RNA biogenesis and potential functional roles they may play.  In this paper, we present a new statistical algorithm that increases the sensitivity and specificity of circular RNA detection.by discovering and quantifying circular and linear RNA splicing events at both annotated exon boundaries and in un-annotated regions of the genome  Unlike previous approaches which rely on heuristics like read count and homology between exons predicted to be circularized to determine confidence in  prediction of circular RNA expression, our algorithm is a statistical approach. We have used this algorithm to discover general induction of circular RNAs in many tissues during human fetal development.  We find that some regions of the brain show marked enrichment for genes where circular RNA is the dominant isoform. Beyond this global trend, specific circular RNAs are tissue specifically induced during fetal development, including a circular isoform of NCX1 in the developing fetal heart that, by 20 weeks, is more highly expressed than the linear isoform as well as beta-actin. In addition, while the vast majority of circular RNA production occurs at canonical U2 (major spliceosome) splice sites, we find the first examples of developmentally induced circular RNAs processed by the U12 (minor) spliceosome, and an enriched propensity of U12 donors to splice into circular RNA at un-annotated, rather than annotated, exons.  Together, our algorithm and its results suggest a potentially significant role for circular RNA in human development. Overall design: 35 human fetal samples from 6 tissues (3 - 7 replicates per tissue) collected between 10 and 20 weeks gestational time were sequenced using Illumina TruSeq Stranded Total RNA with Ribo-Zero Gold sample prep kit.","project":"SRP051249"}
{"number_samples":2,"species":"human","abstract":"Transcriptome of A549 cells after interaction with Aspergillus fumigatus","project":"SRP051309"}
{"number_samples":28,"species":"human","abstract":"Pharmacological inhibition of chromatin co-regulatory factors represents a clinically validated strategy to modulate oncogenic signaling through selective attenuation of gene expression. Here, we demonstrate that CBP/EP300 bromodomain inhibition preferentially abrogates the viability of multiple myeloma cell lines. Phenotypic effects are preceded by the direct transcriptional suppression of the lymphocyte-specific transcription factor IRF4 and the subsequent down-regulation of the IRF4 transcriptional program. Ectopic expression of IRF4 antagonizes the phenotypic effects of CBP/EP300 bromodomain inhibition and prevents the suppression of the IRF4 target c-MYC. These findings suggest that CBP/EP300 bromodomain inhibition represents a viable therapeutic strategy for targeting multiple myeloma and other lymphoid malignancies dependent on the IRF4 network. Overall design: Through the use of CBP/EP300 bromodomain inhibitors (CBP/EP300i), we demonstrate that MYC expression in BETi-resistant cells is dependent on CBP/EP300 bromodomains and that treatment with CBP/EP300i restores phenotypic sensitivity.","project":"SRP051320"}
{"number_samples":30,"species":"human","abstract":"Human DIS3 is a nuclear, catalytic subunit of the exosome complex containing exonucleolytic and endonucleolytic active domains. To identify DIS3 targets genome-wide we conducted comprehensive transcriptomic analysis of HEK293 cells producing mutated DIS3 versions and Photoactivatable Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation (PAR-CLIP) experiments. Pervasive transcription products like Promoter Upstream Transcripts (PROMPTs) accumulated robustly in catalytic DIS3 mutants, representing ~8% of PAR-CLIP reads. Importantly, RNAs originating from unannotated genomic regions increased ~2.5 times in double DIS3 mutants, covering ~70% of genome and allowing for discovery of thousands of  novel transcripts. The first intron of many pre-mRNAs accumulated in DIS3 mutants indicating a widespread premature RNA polymerase II termination. The short form of NEAT1 lincRNA was overexpressed in DIS3 mutants, leading to increased number of paraspeckles. Moreover, there was a global deregulation of mRNAs in DIS3 double mutant. Finally, snoRNA precursors accumulated, which correlated with a strong PAR-CLIP signal indicating that DIS3 but not RRP6 is a main snoRNA processing enzyme. In aggregate, we demonstrate that DIS3 is a major nucleoplasmic activity responsible for shaping the human transcriptome. Overall design: RNA-seq experiments were performed in triplicates for DIS3 wild type (control), DIS3 PIN, DIS3 RNB domain mutants and DIS3 PIN RNB double mutant. RNA-seq samples from DIS3 wild type and DIS3 double mutant were additionally sequenced in deeper resolution, also in triplicates. DIS3 PAR-CLIP experiment was performed in duplicate. Pol II ChIP-seq experiment in WT and DIS3 PIN RNB double-mutants cells was performed in triplicates.","project":"SRP051331"}
{"number_samples":18,"species":"human","abstract":"investigation of lncRNAs deregulated in oncogenic induced senescence.","project":"SRP051359"}
{"number_samples":48,"species":"human","abstract":"DNA methylation is an epigenetic mark thought to be robust to environmental perturbations on a short time scale. Here, we challenge that view by demonstrating that the infection of human dendritic cells with a live pathogenic bacteria is associated with rapid changes in methylation levels at thousands of loci. We performed an integrated analysis of data on genome-wide DNA methylation, histone mark patterns, chromatin accessibility, and gene expression, before and after infection. We found that infection-induced changes in methylation rarely occur at promoter regions and instead localize to distal enhancer elements. Active demethylation is associated with extensive epigenetic remodeling, including the gain of histone activation marks and the induction of enhancer RNAs, and is strongly predictive of changes in the expression levels of nearby genes. Collectively, our observations show that active, rapid changes in DNA methylation in enhancers play a previously unappreciated role in regulating the transcriptional response of immune cells to infection. Overall design: Transcriptional profiles (polyA+) of 6 non-infected and 6 MTB-infected dendritic cell samples.","project":"SRP051368"}
{"number_samples":6,"species":"human","abstract":"DNA methylation is an epigenetic mark thought to be robust to environmental perturbations on a short time scale. Here, we challenge that view by demonstrating that the infection of human dendritic cells with a live pathogenic bacteria is associated with rapid changes in methylation levels at thousands of loci. We performed an integrated analysis of data on genome-wide DNA methylation, histone mark patterns, chromatin accessibility, and gene expression, before and after infection. We found that infection-induced changes in methylation rarely occur at promoter regions and instead localize to distal enhancer elements. Active demethylation is associated with extensive epigenetic remodeling, including the gain of histone activation marks and the induction of enhancer RNAs, and is strongly predictive of changes in the expression levels of nearby genes. Collectively, our observations show that active, rapid changes in DNA methylation in enhancers play a previously unappreciated role in regulating the transcriptional response of immune cells to infection. Overall design: Transcriptional profiles (ribo-minus) of non-infected and MTB-infected dendritic cell samples, in triplicate.","project":"SRP051369"}
{"number_samples":12,"species":"human","abstract":"There is some emerging evidence that members of the Schlafen (SLFN) family of  proteins mediate antineoplastic responses, but the mechanisms accounting for these  effects are not known. We provide evidence that human SLFN5, an interferon (IFN)-  inducible member of the family, exhibits key roles in controlling motility and  invasiveness of renal cell carcinoma (RCC) cells. Our studies define the mechanism by  which this occurs, demonstrating that SLFN5 negatively controls expression of matrix  metalloproteinases (MMP)-1 and -13 and several other genes involved in the control of  malignant cell motility. Importantly, our data establish that SLFN5 expression correlates  with a better overall survival in a large cohort of patients with RCC. The inverse  relationship between SLFN5 expression and RCC aggressiveness raises the possibility of  developing unique therapeutic approaches in the treatment of RCC, by modulating  SLFN5 expression. Overall design: Examination of 2 SLFN5 knockdown cells and 2 controls, in triplicate.","project":"SRP051401"}
{"number_samples":6,"species":"human","abstract":"The Kaposi's sarcoma associated herpesvirus (KSHV) is an oncogenic virus that causes Kaposi's sarcoma, primary effusion lymphoma (PEL), and some forms of multicentric Castleman's disease. The KSHV ORF57 protein is a conserved posttranscriptional regulator of gene expression that is essential for virus replication. ORF57 is multifunctional, but most of its activities are directly linked to its ability to bind RNA. We globally identified virus and host RNAs bound by ORF57 during lytic reactivation in PEL cells using high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP). As expected, ORF57-bound RNA fragments mapped throughout the KSHV genome, including the known ORF57 ligand PAN RNA. In agreement with previously published ChIP results, we observed that ORF57 bound RNAs near the oriLyt regions of the genome. Examination of the host RNA fragments revealed that a subset of the ORF57-bound RNAs was derived from transcript 5´ ends. The position of these 5´-bound fragments correlated closely with the 5´-most exonintron junction of the pre-mRNA. We selected four candidates (BTG1, EGR1, ZFP36, and TNFSF9) and analyzed their pre-mRNA and mRNA levels during lytic phase. Analysis of both steady-state and newly made RNAs revealed that these candidate ORF57-bound pre-mRNAs persisted for longer periods of time throughout infection than control RNAs, consistent with a role for ORF57 in pre-mRNA metabolism. In addition, exogenous expression of ORF57 was sufficient to increase the pre-mRNA levels and, in one case, the mRNA levels of the putative ORF57 targets. These results demonstrate that ORF57 interacts with specific host pre-mRNAs during lytic reactivation and alters their processing, likely by stabilizing pre-mRNAs. These data suggest that ORF57 is involved in modulating host gene expression in addition to KSHV gene expression during lytic reactivation. Overall design: HITS-CLIP was performed on TREx BCBL-Rta cells 20 hpi using antibodies against ORF57. Three biological replicates were performed.","project":"SRP051468"}
{"number_samples":71,"species":"human","abstract":"We discovered induction of circular RNA in human fetal tissues, including the heart. In this study, we were able to recapitulate this induction by in vitro directed differentiation of hESCs to cardiomyocytes, paving the way for future studies into circular RNA regulation. Overall design: We harvested hESCs at sequential stages of differentiation: undifferentiated (day 0), mesoderm (day 2), cardiac progenitor (day 5) and definitive cardiomyocyte (day 14). We performed RNA sequencing in biological triplicate, with 3-8 technical replicates each.","project":"SRP051472"}
{"number_samples":21,"species":"human","abstract":"Aberrant expression of cancer genes and non-canonical RNA species is a hallmark of cancer. However, the mechanisms driving such atypical gene expression programs are incompletely understood. Here, our transcriptional profiling of a cohort of 50 primary clear cell renal cell carcinoma (ccRCC) samples from The Cancer Genome Atlas (TCGA) reveals that transcription read-through beyond the termination site is a source of transcriptome diversity in cancer cells. Amongst the genes most frequently mutated in ccRCC, we identified SETD2 inactivation as a potent enhancer of transcription read-through. We further show that invasion of neighbouring genes and generation of RNA chimeras are functional outcomes of transcription read-through. We identified the BCL2 oncogene as one of such invaded genes and detected a novel chimera, the CTSC-RAB38, in 20% of ccRCC samples. Collectively, our data highlight a novel link between transcription read-through and aberrant expression of oncogenes and chimeric transcripts that is prevalent in cancer. Overall design: RNA-seq of SETD2 mutant and wild-type ccRCC cell lines.","project":"SRP051485"}
{"number_samples":6,"species":"human","abstract":"The transcription factor forkhead box D3 (FoxD3) is a transcriptional factor which belongs to forkhead box (Fox) transcription factor family. The functions of FOXD3 in embryogenesis and in the development of neural crest cells have been clearly defined. Its tumor suppressor function hasbeen found in many types of cancer in recent years. However, the study about its roles in lung cancerdevelopment is still lacking. Our study found that deficiency of FoxD3 in lung cancer enhanced cell growth and cell invasion. RNA-sequence analysis demonstrated that loss of FoxD3 mainly affected cell cycle progression related gene expression. Knockdown of FoxD3 led to G2-M cell accumulation with up-regulation of DNA replication licensing factor MCM5 and MCM4 as well as cell cycle regulator polo-like kinase-1 (PLK1) and CDC6. Over-expression of those genesin lung cancer was associated with poor clinical outcomes of lung cancer patients. We also identified high mobility group box-1(HMGB1), Hras and Ephrin B1 gene expression increase after FoxD3 silencing which may participate in enhanced lung cancer cell invasion. Our study identifiedtumor suppressor function of FoxD3 in lung cancer andwe did the first comprehensive analysis of the genes regulated by FoxD3 for cell proliferation and invasion in lung cancer. The identified genes regulated by FoxD3 through our analysis will provide valuable information to uncover the mechanism of FoxD3 tumor suppressor function. Overall design: Three control and three FOXD3 knockdown A549 cell lines were subjected to RNA sequencing.","project":"SRP051544"}
{"number_samples":6,"species":"human","abstract":"The aim of the study is to identify AR target gens in LNCaP cells Overall design: 6 samples correponding to 2 times 3 replicates were used for the study","project":"SRP051583"}
{"number_samples":18,"species":"human","abstract":"Through a genetic screen in BRAF mutant tumor cells, we show that the Hippo pathway effector YAP acts as a parallel survival input to promote resistance to RAF-MEK inhibitor therapy. Our data uncover YAP as a novel mechanism of resistance to RAF-MEK targeted therapy. The findings unveil the synthetic lethality of YAP and RAF-MEK co-suppression as a promising strategy to enhance response and patient survival. Overall design: RNAseq analysis of HCC364 (lung adenocarcinoma) cells in the context of drug treatment with PLX4720 (vemurafenib, a BRAF inhibitor) or Trametinib (a MEK inhibitor) alongside shRNA knockdown of the gene YAP1","project":"SRP051595"}
{"number_samples":60,"species":"human","abstract":"Human fibroblasts at different population doublings were treated with low amounts of rotenone (mild stress) and compared to untreated fibroblasts. Two different cell lines were used (MRC-5, HFF). Illumina sequencing (HiSeq2000) was applied to generate 50bp single-end reads. Overall design: 60 samples: 3 biological replicates for each group: MRC-5 cells at 4 different population doublings (PD) with and without rotenone; HFF cells at 6 different population doublings with and without rotenone","project":"SRP051599"}
{"number_samples":29,"species":"human","abstract":"The histological grade of carcinomas describes the ability of tumor cells to organize differentiated epithelial structures and has prognostic impact. Molecular control of differentiation in normal and cancer cells relies on lineage-determining transcription factors (TFs) that activate the repertoire of cis-regulatory elements controlling cell type-specific transcriptional outputs. TF recruitment to cognate genomic DNA binding sites results in the deposition of histone marks characteristic of enhancers and other cis-regulatory elements. Here we integrated transcriptomics and genome-wide analysis of chromatin marks in human pancreatic ductal adenocarcinoma (PDAC) cells of different grade to identify first, and then experimentally validate the sequence-specific TFs controlling grade-specific gene expression. We identified a core set of TFs with a pervasive binding to the enhancer repertoire characteristic of differentiated PDACs and controlling different modules of the epithelial gene expression program. Defining the regulatory networks that control the maintenance of epithelial differentiation of PDAC cells will help determine the molecular basis of PDAC heterogeneity and progression. Overall design: Poly(A) fraction of the total RNA from human pancreatic ductal adenocarcinoma cell lines was extracted and subjected to by multiparallel sequencing. Experiments were carried out in unmodified cells in duplicate, genome edited clonal CFPAC1 cells (2 KLF5-deleted CRISPR-Cas9 clones, 3 ELF3-deleted CRISPR-Cas9 clones and 2 wt clones) and CFPAC1 cells ectopically expressing ZEB1 or empty vector control (in duplicate).","project":"SRP051606"}
{"number_samples":18,"species":"human","abstract":"Estrogens play an important role in breast cancer (BC) development and progression, where the two isoforms of the estrogen receptor (ERa and ERß) are generally co-expressed and mediate the effects of these hormones in cancer cells. ERß has been suggested to exert an antagonist role toward the oncogenic activities of ERa, and for this reason it is considered an oncosuppressor. As clinical evidence regarding a prognostic role for this receptor subtype in hormone-responsive BC is still limited and conflicting, more knowledge is required on the biological functions of ERß in cancer cells. We described previously the ERß and ERa interactomes of BC cells, identifying specific and distinct patterns of protein interactions for the two receptors. In particular, we identified factors involved in mRNA splicing and maturation as important components of both ERa and ERß pathways. Guided by these findings, we investigated here in depth the differences in the early transcriptional events and RNA splicing patterns induced in ERa vs ERa+ERß cells, by expressing ERß in ERa+ human BC MCF-7 cells. High-throughput mRNA sequencing was then performed in both cell lines after stimulation with 17b-estradiol, and the results obtained were compared. Overall design: We investigated here in depth the differences in the early transcriptional events and RNA splicing patterns induced in ERa vs ERa+ERß cells, by expressing ERß in ERa+ human BC MCF-7 cells. High-throughput mRNA sequencing was then performed in both cell lines after stimulation with 17b-estradiol, and the results obtained were compared.","project":"SRP051628"}
{"number_samples":25,"species":"human","abstract":"We constructed a genome wide target profile of hsa-miR-503, hsa-miR-103, and hsa-miR-494 by sequencing RNA isolated from Ago2 immunoprecipitations and total RNA samples following transfection of the respective miRNA in mature duplex form Overall design: Examination of mRNA levels in HeLa cells and Ago2 immunoprecipitations from HeLa cells following miR-503, miR-103, or miR-494 mature duplex or control siRNA transfection","project":"SRP051644"}
{"number_samples":8,"species":"human","abstract":"We investigated the effect of SIRT6-knockout on gene expression and H3K4me3 modification profile in human mesenchymal stem cells. Overall design: RNAs isolated from SIRT6+/+ and SIRT6-/- hMSCs at early and late passages were sequenced, respectively. And, H3K4me3 ChIP-seq was performed upon the SIRT6 deleted hMSC and WT at early stage, respectively.","project":"SRP051674"}
{"number_samples":1,"species":"human","abstract":"Pluripotent stem cells (PSC) represent an alternative source of hematopoietic stem cells (HSCs). Clinical translation is impeded by limited engraftment of human (h)PSC-multipotent progenitor cells (MPP). This barrier suggests that additional cues are required for definitive hematopoiesis. We hypothesized that vascular niche producing Notch ligands Jagged-1 (JAG1) and Delta-like ligand-4 (DLL4) would drive definitive hematopoiesis. To test our hypothesis, hes2 human embryonic stem cells (hESC) 2 and Macaca nemestrina (Mn) iPSC line-7 were differentiated with cytokines ± endothelial cells (EC), which express JAG1 and DLL4. EC co-culture supported emergence of 8-fold more CD34+CD45+ cells compared to co-culture with cytokines ± ECs with JAG1 or DLL4 knockdown. EC-induced cells exhibit Notch activation and express HSC-specific targets of Notch signaling RUNX1 and GATA2. EC-induced PSC-MPP engraft at a higher level in NSG mice compared to cytokine-induced cells (10% >5 months), and selection increased engraftment (30%). Long-term engraftment and the myeloid-to-lymphoid ratio achieved with vascular niche induction is similar to levels achieved for cord blood MPP and up to 20-fold higher than hPSC-MPP engraftment. Our findings identify a previously underappreciated role for endothelial Notch ligands in PSC definitive hematopoiesis and production of long-term engrafting CD34+ cells and suggest they are critical for HSC emergence. Overall design: Transcriptome sequencing of Macaca nemestrina (Mn) iPSCs","project":"SRP051675"}
{"number_samples":56,"species":"human","abstract":"Systems biology is an approach to comprehensively study complex interactions within a biological system. Most published systems vaccinology studies have utilized whole blood or peripheral blood mononuclear cells (PBMC) to monitor the immune response after vaccination.  Because human blood is comprised of multiple hematopoietic cell types, the potential for masking responses of under-represented cell populations is increased when analyzing whole blood or PBMC. To investigate the contribution of individual cell types to the immune response after vaccination, we established a rapid and efficient method to purify human T and B cells, natural killer (NK) cells, myeloid dendritic cells (mDC), monocytes, and neutrophils from fresh venous blood. Purified cells were fractionated and processed in a single day. RNA-Seq and quantitative shotgun proteomics were performed to determine expression profiles for each cell type prior to and after inactivated seasonal influenza vaccination. Our results show that transcriptomic and proteomic profiles generated from purified immune cells differ significantly from PBMC. Differential expression analysis for each immune cell type also shows unique transcriptomic and proteomic expression profiles as well as changing biological networks at early time points after vaccination. This cell type-specific information provides a more comprehensive approach to monitor vaccine responses. Overall design: PBMC and six purified cell types from two vaccinated donors were isolated prior to (d0) and at days 1, 3, and 7 post-TIV vaccination for RNA-seq analysis","project":"SRP051688"}
{"number_samples":8,"species":"human","abstract":"Hepatitis C virus uniquely requires the liver specific microRNA-122 for replication, yet global effects on endogenous miRNA targets during infection are unexplored. Here, high-throughput sequencing and crosslinking immunoprecipitation (HITS-CLIP) experiments of human Argonaute (Ago) during HCV infection showed robust Ago binding on the HCV 5’UTR, at known and predicted miR-122 sites. On the human transcriptome, we observed reduced Ago binding and functional mRNA de-repression of miR-122 targets during virus infection. This miR-122 “sponge” effect could be relieved and redirected to miR-15 targets by swapping the miRNA tropism of the virus. Single-cell expression data from reporters containing miR-122 sites showed significant de-repression during HCV infection depending on expression level and number of sites. We describe a quantitative mathematical model of HCV induced miR-122 sequestration and propose that such miR-122 inhibition by HCV RNA may result in global de-repression of host miR-122 targets, providing an environment fertile for the long-term oncogenic potential of HCV. Overall design: mRNA-seq libraries were generated from mock or J6/JFH1 Clone2 infected Huh7.5 cells. Cells were infected at an MOI of 1-2 and harvested at 72 hours and 96 hours post-infection for CLIP. Libraries were generated using Illumina Truseq technology.","project":"SRP051705"}
{"number_samples":10,"species":"human","abstract":"Interleukin-2 (IL-2) is a pleiotropic cytokine that regulates lymphocyte function by signaling through heterodimerization of the IL-2Rß and ?c receptor subunits. Previously, we engineered an IL-2 “superkine” (H9) with enhanced affinity for IL-2Rß. Here, we describe next-generation IL-2 variants that function as “receptor signaling clamps.” They retain high-affinity for IL-2Rß, thereby inhibiting binding of endogenous IL-2, but their engagement of ?c is weakened, thereby attenuating IL-2Rß-?c heterodimerization. These IL-2 analogues act as partial agonists and can differentially affect lymphocytes poised at distinct activation thresholds. Moreover, one of these variants potently antagonized IL-2 and IL-15 signaling and function better than blocking antibodies against IL-2Ra or IL-2Rß. Furthermore, this mutein prolonged survival in a model of graft versus host disease and blocked spontaneous proliferation of smoldering adult T-cell leukemia (ATL) T cells ex vivo. This receptor-clamping approach may be a general mechanism-based strategy for engineering cytokine partial agonists for therapeutic immunomodulation. Overall design: Genome-wide transcription factors binding of STAT5 and mRNA-Sequencing of gene expression profiles in human pre-activated CD8+ T cells.","project":"SRP051736"}
{"number_samples":7,"species":"human","abstract":"During activation, T cells integrate multiple signals from APCs and cytokine milieu. The blockade of these signals can have clinical benefits as exemplified by CTLA4-Ig, which blocks interaction of B7 co-stimulatory molecules on APCs with CD28 on T cells. Variants of CTLA4-Ig, abatacept and belatacept are FDA approved as immunosuppressive agents in arthritis and transplantation whereas murine studies suggested that CTLA4-Ig can be beneficial in a number of other diseases.  However, detailed analysis of human CD4 cell hyporesponsivness induced by CTLA4-Ig has not been performed. Herein, we established a model to study effect of CTLA4-Ig on the activation of human naïve T cells in a human mixed lymphocytes system. Comparison of human CD4 cells activated in the presence or absence of CTLA4-Ig, showed that co-stimulation blockade during TCR activation does not affect NFAT signaling but results in decreased activation of NF-kB and AP-1 transcription factors followed by profound decrease in proliferation and cytokine production. The resulting T cells become hyporesponsive to secondary activation and, although capable of receiving TCR signals, fail to proliferate or produce cytokines, demonstrating properties of anergic cells. However, unlike some models of T cell anergy, these cells did not possess increased levels of TCR signaling inhibitor CBLB. Rather, the CTLA4-Ig induced hyporesponsiveness was associated with an elevated level of p27kip1 cyclin-dependent kinase inhibitor. Overall design: Time series. Human resting and activated T cell dUTP mRNA-Seq profiles were generated on Illumina HiSeq2500","project":"SRP051737"}
{"number_samples":41,"species":"human","abstract":"Drug resistance invariably limits the clinical efficacy of targeted therapy with kinase inhibitors against cancer. We found that targeted therapy with BRAF, ALK, or EGFR inhibitors induces a complex network of secreted signals in drug-stressed melanoma and lung adenocarcinoma cells. This therapy-induced secretome (TIS) stimulates the outgrowth, infiltration and metastasis of drug-resistant cancer clones in the tumour. Additionally, the TIS supports the survival of drug-sensitive cells, contributing to incomplete tumour regression. We used transcriptomic analysis of sensitive tumour cells and xenograft tumours treated with vehicle, vemurafenib, or crizotinib to identify the transcriptional drivers and to dissect the TIS in melanoma (A375, Colo800, UACC62) and lung adenocarcinoma (H3122). In addition, we utilize cell type–specific mRNA purification by translating ribosome affinity purification (TRAP) to identify pathways that are up-regulated in resistant cells (A375R) in response to the regressing tumour microenvironment. Overall design: Analysis of the response of drug sensitive melanoma and lung adenocarcinoma cells to pharmacological inhibition of their driver oncogene and gene expression analysis of drug resistant cancer cells responding to different tumor microenvironments.","project":"SRP051765"}
{"number_samples":9,"species":"human","abstract":"Purpose: We have succeeded in the generation and long-term expansion of SOX9-expressing CD271+PDGFRa+CD73+ chondrogenic ectomesenchymal cells from the PAX3/SOX10/FOXD3-expressing MIXL1-CD271hiPDGFRaloCD73- neural crest-like progeny of human pluripotent stem cells in a chemically defined medium supplemented with Nodal/Activin/TGFb inhibitor (SB) and FGF2 (hereafter called FSB). When “primed” with TGFb, such cells efficiently formed translucent cartilage particles, which were completely mineralized in 12 weeks in immunocompromized mice. Under the FSB condition, ectomesenchymal cells were expandable without loss of chondrogenic potential for at least 16 passages, maintained normal karyotype for at least 10 passages, which any conditions deviated from it (e.g. FGF2 alone or SB alone) failed to support. In order to address the molecular basis of such effects of FSB, a series of RNA-seq experiments were carried out. Methods: We generated and compared the transcriptome profiles of human ectomesenchymal cells expanded under FSB  with those cultured under FSB first then under FGF2 alone (F).  As a control, we also generated transcriptome of ectomesenchymal cells expanded from the begining under F conditions. RNA-sequencing libraries were prepared using a SureSelect Strand Specific RNA Library Preparation kit (Agilent technologies, Santa Clara, CA). Sequencing was performed on an Illumina HiSeq 1500 using a TruSeq Rapid SBS kit (Illumina, San Diego, CA) in a 50-base single-end mode. Sequenced reads were mapped against the human reference genome (GRCh37), using TopHat v2.0.12 (http://ccb.jhu.edu/software/tophat/index.shtml). Expression levels were calculated as fragments per kilobase of exon per million mapped fragments (FPKMs) using Cufflinks v2.1.1 (http://cole-trapnell-lab.github.io/cufflinks). Results: Ectomesenchymal cells maintained under FSB conditions preferentially expressed genes representing embryonic progenitors (SOX4/12, LIN28A/B, MYCN), cranial mesenchymes (ALX1/3/4) and chondroprogenitors (SOX9, COL2a1) of the neural crest origin (SOX8/9, NGFR, NES). In contrast, those cultured under FSB then F, still expressed SOX4/11/12, but lost LIN28, MYCN, ALX1/3/4, NGFR, COL2a1 expression. Interestingl it enhances expresion ofTGFß-inducible genes such as THBS1/2 and DCN  and osteogenesis-related genes such as  COL1a1/2 and RUNX1/2. Conclusions: The CD271+CD73+ ectomesenchymal cells accumulated under FSB conditions possess an mRNA profile of proliferating primitive neural crest/ectomesenchymal cells, although they lacked SOX10 expression, which is critical for neural and melanocytic lineage commitment. Transition from FSB to F conditions supressed the proliferation-related genes expression and enhanced the ossification/mineralization, vasculogenesis/angiogenesis, and cardiac myogenesis-related gene expression. Thus, suppression of TGFß signaling by SB does not seem to freeze the developmental stage of the hPSC-derived neural crest during expansion. Such suppression may instead simply support the high proliferative potential of the cells as well as the expression of SOX9 (and COL2a1), and thereby maintain chondrogenic activity. SOX9 expression initiated at the specification and pre-migratory stages is transient in trunk neural crest but persists in cranial neural crest. The chondrogenic CD271+CD73+ ectomesenchymal cells that maintain SOX9 transcription and translation may therefore represent proliferating cranial neural crest, with a slight commitment to non-neural lineages. Overall design: Examination of human ES-derived neural crest-like progenies expanded in 3 different culture media. Each group contains three biological replicates.","project":"SRP051772"}
{"number_samples":31,"species":"human","abstract":"Multiple Myeloma (MM) is a fatal proliferation of plasma cells that primarily affects elderly individuals, afflicting over 21,000 patients and accounting for over 10,000 deaths in the US each year. Most patients with MM relapse and ultimately become refractory to chemotherapy. Molecular and cytogenetic stratification of patients can identify patients at high risk of relapse, who have a particularly poor survival. Even though gene expression profiling (GEP) has been shown to be better than standard criteria and leads to better treatment stratification, a notable proportion of patients with high-risk gene expression signature can also achieve very good long-term survival, whereas some patients with low-risk gene expression signature myeloma (MM) can experience early relapse. Thus, newer molecular markers are needed for better risk stratification and most importantly newer therapeutic targets are desperately needed for patients with high-risk and relapsed disease. We hypothesize that integrative genomic analysis can define biologically and clinically distinct forms of myeloma beyond what can be gleaned by gene expression profiling alone, and by bringing together multiple types of molecular measures and understanding their relationships, we can reveal a more complete picture of multiple myeloma pathology and therapeutic opportunities. We are performing high-resolution genome-wide genomic, exome and transcriptome and epigenomic characterization of primary MM using next-generation sequencing and correlating these data with survival outcomes in a well-annotated cohort of patients treated at Mt. Sinai. We expect our results to yield novel insights into pathogenesis of relapsed MM, improved prognostic models and new therapeutic targets not identified by current technologies, and to identify approved therapies that could be repurposed for improving the treatment of relapsed MM.","project":"SRP051819"}
{"number_samples":2,"species":"human","abstract":"Interferons (IFNs) are key players in the antiviral response. IFN sensing by the cell activates transcription of IFN-stimulated genes (ISGs) able to induce an antiviral state by affecting viral replication and release. IFN also induces the expression of ISGs that function as negative regulators to limit the strength and duration of IFN response. The ISGs identified so far belong to coding genes. However, only a small proportion of the transcriptome corresponds to coding transcripts and it has been estimated that there could be as many coding as long non-coding RNAs (lncRNAs). To address whether IFN can also regulate the expression of lncRNAs, we analyzed the transcriptome of HuH7 cells treated or not with IFNa2 by expression arrays. Analysis of the arrays showed increased levels of several well-characterized coding genes that respond to IFN both at early or late times. Furthermore, we identified several IFN-stimulated or -downregulated lncRNAs (ISRs and IDRs). Further validation showed that ISR2, 8, and 12 expression mimics that of their neighboring genes GBP1, IRF1, and IL6, respectively, all related to the IFN response. These genes are induced in response to different doses of IFNa2 in different cell lines at early (ISR2 or 8) or later (ISR12) time points. IFNß also induced the expression of these lncRNAs. ISR2 and 8 were also induced by an influenza virus unable to block the IFN response but not by other wild-type lytic viruses tested. Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV. Increased levels of ISR2 were also detected in patients chronically infected with HIV. This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response. Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response. Overall design: HuH7 cells were treated with 10000 units/ml of IFN a2 and RNA was isolated 3 days post-treatment","project":"SRP051822"}
{"number_samples":14,"species":"human","abstract":"Necrotizing enterocolitis (NEC) is the most frequent life-threatening gastrointestinal disease experienced by premature infant occuring in neonatal intensive care units. NEC is associated with severe intestinal inflammation, intestinal perforation leading to mortality. The challenge for neonatologists is to detect early clinical manifestations of NEC. Therefore, one of the strategies to prevent or treat NEC would be to develop an early diagnostic tool allowing identification of preterm infants either at risk of developing NEC or at the onset of the disease. Illumina’s deep sequencing technology (RNA-seq) was used to establish the gene expression profile between resected ileal healthy preterm (control, n=5) and NEC diagnosed preterm infant (NEC, n=9) and analyzed by IPA Core analysis system. IPA analysis indicated that the most significant functional pathways overrepresented in NEC neonates were associated with innate immune functions, such as altered T and B cell signaling, B cell development, and the role of pattern recognition receptors in recognition of bacteria and viruses. Among genes that were strongly modulated in NEC neonates, we observed a high degree of similarity with those linked to the development of IBD. By comparing gene expression patterns between NEC and Crohn’s disease, we identified several new potential protein targets for helping to predict and/or diagnose NEC in preterm infant. Gene expression profile revealed an uncontrolled innate immune response in the intestine of NEC neonates. Moreover, comparative analysis between NEC and Crohn’s disease evidenced high degree of similarity between these two inflammatory diseases and allowed us to identify several new potential NEC biomarkers. Overall design: Illumina’s deep sequencing technology (RNA-seq) was used to establish the gene expression profile between resected ileal healthy preterm (control, n=5) and NEC diagnosed preterm infant (NEC, n=9)","project":"SRP051825"}
{"number_samples":69,"species":"human","abstract":"Huntington’s Disease (HD) is a devastating neurodegenerative disorder that is caused by an expanded CAG trinucleotide repeat in the Huntingtin (HTT) gene. Transcriptional dysregulation in the human HD brain has been documented but is incompletely understood. Here we present a genome-wide analysis of mRNA expression in human prefrontal cortex from 20 HD and 49 neuropathologically normal controls using next generation high-throughput sequencing. Surprisingly, 19% (5,480) of the 28,087 confidently detected genes are differentially expressed (FDR<0.05) and are predominantly up-regulated. A novel hypothesis-free geneset enrichment method that dissects large gene lists into functionally and transcriptionally related groups discovers that the differentially expressed genes are enriched for immune response, neuroinflammation, and developmental genes. Markers for all major brain cell types are observed, suggesting that HD invokes a systemic response in the brain area studied. Unexpectedly, the most strongly differentially expressed genes are a homeotic gene set (represented by Hox and other homeobox genes), that are almost exclusively expressed in HD, a profile not widely implicated in HD pathogenesis. The significance of transcriptional changes of developmental processes in the HD brain is poorly understood and warrants further investigation. The role of inflammation and the significance of non-neuronal involvement in HD pathogenesis suggest anti-inflammatory therapeutics may offer important opportunities in treating HD. Overall design: 20 Huntington's Disease and 49 neurologically normal control samples from post-mortem human subjects","project":"SRP051844"}
{"number_samples":188,"species":"human","abstract":"The molecular factors involved in the development of Post-Traumatic Stress Disorder (PTSD) remain poorly understood. Previous transcriptomic studies investigating the mechanisms of PTSD apply targeted approaches to identify individual genes under a cross-sectional framework lack a holistic view of the behaviours and properties of these genes at the system-level. Here we sought to apply an unsupervised gene-network based approach to a prospective experimental design using whole-transcriptome RNA-Seq gene expression from peripheral blood leukocytes of U.S. Marines (N=188), obtained both pre- and post-deployment to conflict zones. We identified discrete groups of co-regulated genes (i.e., co-expression modules) and tested them for association to PTSD. We identified one module at both pre- and post-deployment containing putative causal signatures for PTSD development displaying an over-expression of genes enriched for functions of innate-immune response and interferon signalling (Type-I and Type-II). Importantly, these results were replicated in a second non-overlapping independent dataset of U.S. Marines (N=96), further outlining the role of innate immune and interferon signalling genes within co-expression modules to explain at least part of the causal pathophysiology for PTSD development. A second module, consequential of trauma exposure, contained PTSD resiliency signatures and an over-expression of genes involved in hemostasis and wound responsiveness suggesting that chronic levels of stress impair proper wound healing during/after exposure to the battlefield while highlighting the role of the hemostatic system as a clinical indicator of chronic-based stress. These findings provide novel insights for early preventative measures and advanced PTSD detection, which may lead to interventions that delay or perhaps abrogate the development of PTSD. We used RNA-Sequencing gene expression to characterize both prognostic and diagnostic molecular signatures associated to PTSD risk and PTSD status compared to control subjects. Overall design: Peripheral blood luekocytes gene expression was subject to transcriptional analysis for 94 service members both prior-to and following-deployment to conflict zones. Half of the subjects returned with Post-traumatic stress disorder (PTSD), while the other half did not.","project":"SRP051848"}
{"number_samples":1,"species":"human","abstract":"Recent advances in transcriptome sequencing have enabled the discovery of thousands of long non-coding RNAs (lncRNAs) across many species. Though several lncRNAs have been shown to play important roles in diverse biological processes, the functions and mechanisms of most lncRNAs remain unknown. Two significant obstacles lie between transcriptome sequencing and functional characterization of lncRNAs: identifying truly non-coding genes from de novo reconstructed transcriptomes, and prioritizing the hundreds of resulting putative lncRNAs for downstream experimental interrogation. We present slncky, a computational lncRNA discovery tool that produces a high-quality set of lncRNAs from RNA-sequencing data and further uses evolutionary constraint to prioritize lncRNAs that are likely to be functionally important. Our automated filtering pipeline is comparable to manual curation efforts and more sensitive than previously published computational approaches. Furthermore, we develop a sensitive alignment pipeline for aligning lncRNA loci and propose new evolutionary metrics relevant for analyzing sequence and transcript evolution. Our analysis reveals that evolutionary selection acts in several distinct patterns, and uncovers two notable classes of intergenic lncRNAs: one showing strong purifying selection on RNA sequence and another where constraint is restricted to the regulation but not the sequence of the transcript. Overall design: To study a comprehensive and comparable set of lncRNAs expressed in the pluripotent state, we analyzed RNA-Seq data from pluripotent cells derived from several strains and species, and grown in two physiological conditions. First we derived “naïve” ES cells (ESCs) from each of three different mice strains: 129SvEv, NOD, and Mus musculus castaneus (cast) mouse, a wild mouse subspecies originally from Thailand, as well as naïve induced pluripotent stem (iPS) cells from rat and human. Next, to facilitate comparisons across pluripotent cells from different species, we also cultured mouse and rat cells in “primed” epiblast stem cell (EpiSC) culture conditions, since human iPS cells in culture are much more similar molecularly and functionally to mouse primed EpiSCs rather than to a ground state naïve ESCs. We polyA selected RNA from each cell type and deeply sequenced on HiSeq2500","project":"SRP051853"}
{"number_samples":22,"species":"human","abstract":"RNA-Sequencing analysis of 18 papillary thyroid carcinoma biopsies and of 4 healthy donors' thyroids. In this analysis we assessed differential gene expression and investigated the mutational landscape in this tumor type. Analysis of gene fusion was also performed, leading to the identification of a novel chimeric transcript, potential driver in tumor initiation. Overall design: Total RNA isolated from 18 papillary thyroid carcinoma biopsies and 4 healthy donors' thyroids.","project":"SRP052056"}
{"number_samples":6,"species":"human","abstract":"Expression from CD133+ cells isolated from adult human exocrine tissue was compared to a CD133-depleted cell population Overall design: Islet-depleted exocrine tissue from three independent adult human cadaveric pancreata were cultured for four days in Miami media 1A. Following trypsinization, cells were isolated using anti-CD133 immunomagnetic beads to >95% CD133+.  CD133-negative cells were further depleted of CD133+ cells to <1% CD133+.","project":"SRP052057"}
{"number_samples":9,"species":"human","abstract":"A growing body of genomic data on human cancers poses the critical question of how genomic variations translate to cancer phenotypes. We used standardized shotgun proteomics and targeted protein quantitation platforms to analyze a panel of 10 colon cancer cell lines differing by mutations in DNA mismatch repair (MMR) genes. In addition, we performed transcriptome sequencing (RNA-seq) to enable detection of protein sequence variants from the proteomic data. In addition, RNA-seq data also provide transcript expression information, which can be combined with protein expression levels to identify regulatory changes in biologic systems","project":"SRP052201"}
{"number_samples":42,"species":"human","abstract":"Roberts syndrome (RBS) is a human developmental disorder caused by mutations in the cohesin acetyltransferase ESCO2. We previously reported that mTORC1 was inhibited and overall translation was reduced in RBS cells. Treatment of RBS cells with L-leucine partially rescued mTOR function and protein synthesis, correlating with increased cell division. In this study, we use RBS as a model for mTOR inhibition and analyze transcription and translation with ribosome profiling to determine genome-wide effects of L-leucine. The translational efficiency of many genes is increased with Lleucine in RBS cells including genes involved in ribosome biogenesis, translation, and mitochondrial function. snoRNAs are strongly upregulated in RBS cells, but decreased with L-leucine. Imprinted genes, including H19 and GTL2, are differentially expressed in RBS cells consistent with contribution to mTORC1 control. This study reveals dramatic effects of L-leucine stimulation of mTORC1 and supports that ESCO2 function is required for normal gene expression and translation. Overall design: 42 samples of human fibroblast cell lines with various genotypes (wt, corrected, and esco2 mutants) are treated with l-leucine or d-leucine (control) for 3 or 24 hours. Biological replicates are present.","project":"SRP052229"}
{"number_samples":10,"species":"human","abstract":"The evolutionary expansion of the human neocortex reflects increased amplification of basal progenitors in the subventricular zone, producing more neurons during fetal corticogenesis. Here, we analyze the transcriptomes of distinct progenitor subpopulations isolated by a novel approach from developing mouse and human neocortex. We identify 56 genes preferentially expressed in human apical and basal radial glia that lack mouse orthologs. Among these, ARHGAP11B has the highest degree of radial glia-specific expression. ARHGAP11B arose from partial duplication of the Rho GTPase-activating-protein–encoding ARHGAP11A on the human lineage after separation from the chimpanzee lineage. Expression of ARHGAP11B in embryonic mouse neocortex promotes basal progenitor generation and self-renewal, and can increase cortical plate area and induce gyrification. Hence, ARHGAP11B may have contributed to evolutionary expansion of human neocortex. Overall design: Gene expression profiles of mouse and human purified neocortical progenitor types and neurons were generated by RNA-seq and analyzed including inter- and intra-species comparison.","project":"SRP052294"}
{"number_samples":2,"species":"human","abstract":"Purpose: The goal of this study is to investigate the alteration of gene expression pattern of alveolar macrophages by allergen challenge in human asthmatics. Method: By using subsegmental bronchial provocation with allergen (SBP-AG) protocol, we obtained BAL fluids, before and 48 hours after allergen challenge in the subjects enrolled in the protocol. Alveolar macrophages were purified from the BAL fluids and total RNA was isolated. Next-generation sequencing data were generated by using the Illumina system. Results: Using an optimized data analysis workflow, we mapped about 75 million sequence reads per sample to the human genome and identified 29,691 transcripts in the macrophage mRNAs. Among them, the change in the expression profiles of 37 transcripts were statistically significant. Conclusions: It has been well accepted that Th2 cytokine enriched environment transforms the phenotype of macrophages into alternatively activated form. However, the details of a genome-wide gene expression profiles of macrophages were not well investigated. Using RNA-seq technology, we provided comprehensive data of macrophage gene expression profiles in allergic lung inflammation. Our data could offer a framework to study biologic functions of alternatively activated macrophage in chronic inflammatory diseases. Overall design: mRNA profiles of alveolar macrophages obtained from asthmatics, before and after allergen challenge.","project":"SRP052491"}
{"number_samples":20,"species":"human","abstract":"In order to try and identify characteristics of gene expression in the endometrium of women suffering infertility or recurrenty miscarriage, we performed RNAseq on endometrial pipelle biopsies from 20 women.  The endometrial transcriptome in the mid-luteal phase of the cycle (window of implantation) is highly divergent in women suffering infertility or miscarriages. Overall design: 20 mid-luteal endometrial biopsies were analysed from infertile women and patients suffering recurrent pregnancy loss. ","project":"SRP052612"}
{"number_samples":8,"species":"human","abstract":"Background: Lymphocytic colitis (LC) causes watery diarrhea. This study aimed to identify the inherent pathomechanisms of epithelial transport and barrier dysfunction and regulatory inputs. Methods: 8 (4 LC samples and 4 unrelated control samples) biopsies of fresh sigmoid colon were analysed by paired end sequencing (Illumina High Seq 2500) Conclusions: ENaC-mediated Na+ malabsorption via ERK1/2 and epithelial barrier dysfunction from tight junction downregulation through claudin-4, -5, and -8 are mechanisms causing malabsorptive and leak flux diarrhea in LC resulting from the key effector cytokines TNFa and INFg. Overall design: Sigmoid colon mRNA profiles from 8 unrelated individuals (4 with LC, 4 as control) were generated using paired end sequencing with Illumina High Seq 2500","project":"SRP052615"}
{"number_samples":12,"species":"human","abstract":"To determine the global effects of ASCT2 inhibition, we used next generation sequencing to determine mRNA expression changes in PC-3 cells treated with BenSer or GPNA for 48 h. Overall design: Examination of two different ASCT2 inhibitors BenSer and GPNA in prostate cancer cell line PC-3.","project":"SRP052620"}
{"number_samples":6,"species":"human","abstract":"We report the effects of Rapamycin treatment on the transcriptome of normal human dermal fibroblasts isolated from foreskin (designated 2DD). We sequenced mRNA from 2 replicates of proliferative (PRO) quiescent (QUI, serum starved) or treated with 500nM Rapamycin for 5 days (RAP). Comparative analyses with PRO transcripts a baseline indicate that genes that changed expression from Rapamycin treated fibroblasts are significantly different from those of quiescence cells. Rapamycin treated cells showed a significant enrichment for cytokines from the Il-6 cascade. Overall design: Examination of mRNAs from proliferative, quiescent (serum starvation) and Rapamycin (5oonM, 5days) treated 2DD normal human dermal/foreskin fibroblasts.","project":"SRP052706"}
{"number_samples":6,"species":"human","abstract":"We characterized the marine natural product cortistatin A (CA) as an inhibitor of CDK8 to determine whether pharmacologic inhibition of CDK8 regulates super-enhancer function and inhibits AML proliferation. In this series, we examine the transcriptional effect on insensitive HCT116 cells of 3hrs exposure to CA. Overall design: HCT116 cells were treated in triplicate with either DMSO or CA for 3hrs after which RNA was harvested and prepared for RNA sequencing to assess transcriptional changes.","project":"SRP052713"}
{"number_samples":4,"species":"human","abstract":"Quantitative and qualitative analysis of small RNAs in human endothelial cells and exosomes provides insights into localized RNA processing, degradation and sorting","project":"SRP052732"}
{"number_samples":169,"species":"human","abstract":"Melanoma resistance to MAPK- or T cell checkpoint-targeted therapies represents a major clinical challenge, and treatment failures of MAPK-targeted therapies due to acquired resistance often require salvage immunotherapies. We show that genomic analysis of acquired resistance to MAPK inhibitors revealed key driver genes but failedto adequately account for clinical resistance. From a large-scale comparative analysis of temporal transcriptomes from patient-matched tumor biopsies, we discovered highly recurrent differential expression and signature outputs of c-MET, LEF1 and YAP1 as drivers of acquired MAPK inhibitor resistance. Moreover, integration of gene- and signature-based transcriptomic analysis revealed profound CD8 T cell deficiency detected in half of resistant melanomas in association with downregulation of dendritic cells and antigen presentation. We also propose a major methylomic basis to transcriptomic evolution under MAPK inhibitor selection. Thus, this database provides a rich informational resource, and the current landscape represents a benchmark to understanding melanoma therapeutic resistance. Overall design: Melanoma biopsies pre and post MAPKi treatment were sent for RNAseq analysis","project":"SRP052740"}
{"number_samples":5,"species":"human","abstract":"The goal of this project is to find out the different genes regulated by LSD2","project":"SRP052760"}
{"number_samples":4,"species":"human","abstract":"To study the transcription and translation of tumor-mutated alleles in HCC patients","project":"SRP052763"}
{"number_samples":6,"species":"human","abstract":"To investigate the effect of CEBPA and its mutant isoform P30 on the expression of mRNAs and long non-coding RNAs (lncRNAs), we utilized the K562 AML cell line carrying a stable and Tet-on inducible CEBPA  or P30 allele. Overall design: Based on the expression of known CEBPA transcriptional targets, we selected RNA extracted from 48 hours of induction (CEBPA or P30)  together with RNA extracted from control-induced cells (CTR).  2 biological replicates for each sample have been utilized.","project":"SRP052781"}
{"number_samples":4,"species":"human","abstract":"Characterization of circular RNAs in Ovarian cancer","project":"SRP052817"}
{"number_samples":9,"species":"human","abstract":"We used RNA sequencing to identify differentially expressed genes during esophageal epithelial differentiation and in the presence of interleukin 13 using an air-liquid interface culture system. Overall design: RNA sequencing was performed on a human esophageal epithelial cell line (EPC2-hTERT) grown submerged (day 8) or at the air-liquid interface (ALI) (day 14, untreated or treated with interleukin 13 [100 ng/mL])","project":"SRP052856"}
{"number_samples":7,"species":"human","abstract":"Mutations within genes encoding spliceosomal proteins are the most common class of mutations in patients with myelodysplastic syndromes, yet it is currently not well understood how these mutations impact hematopoiesis or RNA splicing. Here we report that mutations affecting the splicing factor SRSF2 alter its normal RNA recognition activity, resulting in impaired hematopoietic differentiation and myelodysplasia. Commonly occurring SRSF2 mutations impaired wildtype SRSF2’s normal RNA-binding avidity and preference for specific exonic splicing enhancer RNA motifs. Integration of murine and human transcriptome data identified recurrent mis-splicing of key transcriptional regulators in the presence of mutant SRSF2, including promotion of a highly conserved “poison” exon of EZH2 that results in nonsense-mediated decay and contributes to impaired hematopoiesis. These data provide a mechanistic basis for the enrichment of specific mutations in spliceosomal proteins in myelodysplasia, and suggest that altered RNA recognition activity is a novel mechanism of leukemogenesis. Overall design: mRNA profiles of murine model and K562 cells expressing SRSF2 WT, mutants and knockdown of SRSF2 in TF-1 cells generated by deep sequencing.","project":"SRP052874"}
{"number_samples":4,"species":"human","abstract":"Pro-inflammatory response of VSMCs is triggered by endothelial damage and a causative step for thrombosis and neointimal thickening in the arterial vessels. Therefore, we investigate a role of cytosolic Hsp60 as a novel pro-inflammatory mediator in VSMCs.  Hsp60 was detected in the cytosol of VSMCs. The selective depletion of cytosolic Hsp60 in VSMCs reduced the IKK activation, repressed the induction of NF-?B-dependent pro-survival genes (MnSOD and Bfl-1/A1), and enhanced apoptotic death in response to TNF-a. Moreover, a quantitative RNA sequencing revealed that the expression of 75 genes among the 774 TNF-a-inducible genes was significantly reduced by the depletion of cytosolic Hsp60. In particular, the expression of pro-inflammatory cytokines/chemokines, such as CCL2, CCL20, and IL-6, was regulated by the cytosolic Hsp60 in VSMCs. Finally, the depletion of cytosolic Hsp60 markedly inhibited the neointimal thickening in the balloon-injured arterial vessels by inducing apoptotic cell death and inhibiting chemokine production.  This study provides the first evidence that cytosolic Hsp60 could be a therapeutic target for preventing inflammation-driven VSMC hyperplasia in the injured vessels. Overall design: Hsp60 normal vs knockout with TNF-alpha treatment","project":"SRP052879"}
{"number_samples":32,"species":"human","abstract":"Through the study of EGFR-mutant lung adenocarcinoma we show that NFkB signaling is rapidly engaged by EGFR oncogene inhibition to promote tumor cell persistence and therapy resistance. Unexpectedly, we found that EGFR oncogene inhibition induced an EGFR-TRAF2-RIP1-IKK complex that stimulated an NFkB-mediated transcriptional survival program. We identified a direct pharmacologic NFkB inhibitor, PBS-1086, that suppressed this adaptive survival program and increased both the magnitude and duration of initial EGFR TKI response in cellular and in vivo tumor models, including a novel patient-derived NSCLC xenograft. These findings unveil NFkB as a critical adaptive survival mechanism engaged in response to EGFR oncogene inhibition and identify PBS-1086 as a promising NFkB inhibitor to eliminate disease persistence and potentially prevent the emergence of resistance in patients. Overall design: RNAseq analysis of 11-18 (EGFR-mutant lung adenocarcinoma) cells in the context of drug treatment with erlotinib and/or genetic or pharmacological inactivation of NFkB","project":"SRP052950"}
{"number_samples":10,"species":"human","abstract":"To explore the primary cause of Dilated Cardiomyopathy in heart samples from DCM-diagnosed patients who had undergone heart transplant (hDCM), we set out to identify differentially expressed genes by massively parallel sequencing of heart samples. Overall design: Methods: Heart mRNA profiles from DCM-diagnosed patients who had undergone heart transplant (hDCM) were generated by deep sequencing, in triplicate, using Illumina GAIIx.","project":"SRP052978"}
{"number_samples":6,"species":"human","abstract":"To find the expression of transfected genes","project":"SRP052983"}
{"number_samples":8,"species":"human","abstract":"Injury to anterior cruciate ligament (ACL) is common in young individuals and a frequent cause of functional instability and early onset of osteoarthritis. The healing potential of an injured ACL is known to decay over time. The molecular origin of this healing deficiency largely remains elusive but plausibly involves gene transcripts associated with tissue healing. To explore this possibility, we set out to identify transcript expression differences in injured ACL remnants recovered at the time of surgical reconstruction, via microarray (n=24) and RNA-seq (n=8) technologies in transcriptome profiling. We found that time-from-injury was an important determinant of changes in gene expression signatures predominately resulting in repression of several biological processes as identified by gene ontology. The most interesting observation was a time-dependent decline in the gene transcripts as well as the biological processes common to both microarray and RNA-seq analyses. Compared to acute tears, in chronic several important biological processes were namely extracellular matrix organization, angiogenesis, cell adhesion, wound healing, mesenchyme transition, and response to hypoxia. Furthermore, the cross-platform concordance in terms of differentially expressed transcripts or enriched pathways was linearly correlated (r=0.64). Microfluidic digital PCR confirmed the expression of selected differentially expressed transcripts. These intriguing findings suggest an initial attempt of the injured ACL to repair, which drops with time. These findings have implications for efforts to repair the ACL and may be relevant for its reconstruction. These findings also emphasize the utility of differentially expressed transcripts as prognostic biomarkers in patients with ACL injury. Overall design: Examination of transcript expression differences by time-from-injury in anterior cruciate ligament","project":"SRP052991"}
{"number_samples":20,"species":"human","abstract":"Senescence of stromal fibroblasts has been linked to establishment of cancer associated fibroblasts (CAF) and aging-associated increase of tumors. However, in clinically occurring carcinomas, density and proliferation of CAFs are frequently increased rather than decreased. We previously showed that genetic deletion or down-modulation of the canonical Notch effector CSL/RBP-J? in skin dermal fibroblasts is sufficient for CAF activation with consequent development of multifocal keratinocyte tumors. We now show that CSL deletion or knockdown induces senescence of primary fibroblasts derived from dermis, oral mucosa, breast and lung. CSL functions in these cells as a constitutive direct repressor of multiple senescence- and CAF-effector genes. At the same time, it physically interacts with p53, repressing its activity, and p53 activation provides a failsafe mechanism against compromised CSL function. Concomitant loss of CSL and p53 overcomes fibroblast senescence, enhances expression of CAF effector genes and, in vivo, promotes tumour and stromal cell expansion. Together, the findings support a CAF activation/stromal evolution model under convergent CSL/p53 control. Overall design: Human Dermal Fibroblasts were transfected with two different siRNA against CSL in parallel with a control siRNA. Total RNA was extracted 3 days post-transfection, followed by RNA-Seq analysis.","project":"SRP052998"}
{"number_samples":22,"species":"human","abstract":"We sorted 2PN stage human embryos by predicted viability using a noninvasive measurement we previously developed. We then performed RNA-seq on each embryo (which was a single cell) and looked for differences in expression between viable and nonviable embryos. Overall design: We sequenced the transcriptomes of 22 human zygotes (2PN stage) embryos, of which 11 were predicted nonviable and 11 were predicted viable. We excluded 5 which had no siblings in this data set. From the remaining 17 embryos, we analyzed the set of genes that was differentially expressed between those predicted viable and those predicted nonviable.","project":"SRP053004"}
{"number_samples":6,"species":"human","abstract":"In response to DNA damage tissue homoeostasis is ensured by elaborate protein networks promoting DNA repair, cell cycle arrest or apoptosis. DNA damage-response signaling pathways coordinate these processes, in part, by propagating gene expression-modulating signals. DNA damage does not only influence abundance of mRNAs, but also their coding information through alternative splicing. This level of regulation is thus far poorly understood. Here we show that transcription-blocking DNA lesions promote displacement of activated spliceosomes and initiate a positive feedback loop centered on the DNA damage signaling kinase ATM. Pausing of elongating RNA polymerase at DNA lesions induces spliceosome displacement and R-loop formation. This in turn activates ATM, which signals to further impede splicing activity. These findings define the R-loop-dependent ATM activation by transcription-blocking lesions as an important event in the DNA damage response of non-replicating cells and highlight a key role for spliceosome displacement in this process.","project":"SRP053034"}
{"number_samples":6,"species":"human","abstract":"Fanconi anemia (FA) is a disorder of genomic instability characterized by progressive bone marrow failure (BMF), developmental abnormalities and an increased susceptibility to cancer. Although various consequences in hematopoietic stem/progenitor cells have been attributed to FA-BMF, the quest to identify the initial pathological event is still ongoing. To address this issue, we established induced pluripotent stem cells (iPSCs) from fibroblasts of six FA patients with FANCA mutations. An improved reprogramming method yielded iPSC-like colonies from all patients, and iPSC clones were propagated from two patients. Quantitative evaluation of the differentiation ability demonstrated that the differentiation propensity toward the hematopoietic and endothelial lineages is already defective in early hemoangiogenic progenitors. The expression levels of critical transcription factors were significantly downregulated in these progenitors. These data indicate that the hematopoietic consequences in FA patients originate from the early hematopoietic stage, and highlight the potential usefulness of iPSC technology for elucidating the pathogenesis of FA-BMF. Overall design: The investigation of the RNA-seq analysis of iPSC-derived HAPCs.","project":"SRP053042"}
{"number_samples":4,"species":"human","abstract":"Spinal muscular atrophy (SMA) is a neuromuscular disorder caused by mutations of the survival of motor neuron 1 (SMN1) gene. In the pathogenesis of SMA, pathological changes of the neuromuscular junction (NMJ) precede the motor neuronal loss. Therefore, it is critical to evaluate the NMJ formed by SMA patients’ motor neurons (MNs), and to identify drugs that can restore the normal condition. We generated NMJ-like structures using motor neurons (MNs) derived from SMA patient-specific induced pluripotent stem cells (iPSCs), and found that the clustering of the acetylcholine receptor (AChR) is significantly impaired. Valproic acid and  antisense oligonucleotide treatment ameliorated the AChR clustering defects, leading to an increase in the level of full-length SMN transcripts. Thus, the current in vitro model of AChR clustering using SMA patient-derived iPSCs is useful to dissect the pathophysiological mechanisms underlying the development of SMA, and to evaluate the efficacy of new therapeutic approaches.  Overall design: to evaluate the effects of VPA on the expression profiles of the MNs","project":"SRP053043"}
{"number_samples":9,"species":"human","abstract":"To reveal dynamic changes in networks of gene expression and epigenetic regulation during healthy human T cell aging. Overall design: Peripheral blood was obtained from 90 healthy newborn (0 y), 90 healthy middle-aged (age range from 40 to 55 years) and 90 long-lived subjects (age range from 90 to 110 years). The NB samples came from umbilical cord blood. 90 individuals from each age stage were distributed equally into 3 pools with identical male/female ratios. Peripheral blood mononuclear cells (PBMCs) were isolated from the pooled blood samples using density gradient centrifugation, and CD4+ T cells were isolated by positive selection using CD4 beads.","project":"SRP053046"}
{"number_samples":2,"species":"human","abstract":"Recently, RNA sequencing has achieved single cell resolution, but what is limiting is an effective way to routinely isolate and process large numbers of individual cells for in-depth sequencing, and to do so quantitatively. We have developed a droplet-microfluidic approach for parallel barcoding thousands of individual cells for subsequent RNA profiling by next-generation sequencing. This high-throughput method shows a surprisingly low noise profile and is readily adaptable to other sequencing-based assays. Using this technique, we analyzed mouse embryonic stem cells, revealing in detail the population structure and the heterogeneous onset of differentiation after LIF withdrawal. The reproducibility and low noise of this high-throughput single cell data allowed us to deconstruct cell populations and infer gene expression relationships. Overall design: A total of 8 single cell data sets are submitted: 3 for mouse embryonic stem (ES) cells (1 biological replicate, 2 technical replicates); 3 samples following LIF withdrawal (days 2,4, 7); one pure RNA data set (from human lymphoblast K562 cells); and one sample of single K562 cells.","project":"SRP053052"}
{"number_samples":4,"species":"human","abstract":"The c-MYC oncogene is a key transcription factor deregulated in most human tumors. Histone marks associated with transcriptionally active genes in euchromatic islands define the set of high-affinity c-MYC targets. The mechanisms involved in their recognition by c-MYC are not known but likely involve chromatin-remodelling and chromatin-modifying complexes. Here, we show that c-MYC interacts with BPTF, a core subunit of the NURF complex that binds active chromatin. BPTF is required for the activation of the full c-MYC transcriptional programme in fibroblasts. BPTF knockdown leads to a decrease in c-MYC recruitment to DNA and to changes in chromatin accessibility. Using BPTF-null MEFs we show that BPTF is necessary for c-MYC-driven proliferation, G1-S progression, and replication stress, but not for c-MYC-driven apoptosis. Consistently, BPTF is required for the proliferation of cells driven by c-MYC, such as Burkitt lymphoma, and its expression in human cancer lines correlates with the activation of c-MYC gene signatures. Our findings point to the c-MYC-BPTF axis as a potential therapeutic target in cancer. Overall design: To assess whether BPTF is required for the transcriptional activity of c-MYC, human foreskin fibroblasts (HFF) were stably transduced with the chimeric MYC-ER cDNA (HFF MYC-ER) and infected with lentiviruses coding for either control (shNt) or BPTF-targeting shRNAs. Cells were serum-starved for 2 days to achieve quiescence and then treated with 4-hydroxytamoxifen (4-OHT)","project":"SRP053098"}
{"number_samples":88,"species":"human","abstract":"Bariatric surgery is the most effective therapy of severe human obesity. It is associated with improvements in metabolic and non metabolic co-morbidities which are thought to be mediated by a decrease of adipose tissue inflammation. However, the molecular mechanisms behind these beneficial effects are poorly understood. We analyzed expression profiles in subcutaneous adipose tissue from 22 obese women before and 3 months after surgery using the RNA-seq technology. Of 15,972 detected genes, 1214 were differentially expressed after surgery. Upregulated genes were mostly involved in the basal cellular machinery. Downregulated genes were enriched in metabolic functions of adipose tissue. At baseline, we identified 26 modules of coexpressed genes. The four most stable modules reflected the innate and adaptive immune responses of adipose tissue, including a general signature of innate immune cells, an adaptive immune response elicited by T lymphocytes, a neutrophil-mediated inflammatory signature and an interferon-signaling pathway, respectively. After surgery, a few crucial molecules involved in chemotaxis and activation of immune cells were disconnected from their respective networks. These molecules may represent therapeutic targets against adipose inflammation. Overall design: mRNA sequencing of subcutaneous adipose tissue (SAT) samples from 22 obese women before and 3 months after bariatric surgery","project":"SRP053101"}
{"number_samples":9,"species":"human","abstract":"TNFa-induced inflammation is important in diabetic retinopathy. This study was performed to determine the effect of TNFa on human microvascular endothelial cells as well as the role of the PPARbeta/delta antagonist GSK0660 on TNFa-stimulated cells.","project":"SRP053124"}
{"number_samples":27,"species":"human","abstract":"IL-6 and IL-27 have antagonistic and overlapping functions, signal through a shared receptor subunit and employ the same downstream STAT proteins. To evaluate the degree of specificity and redundancy for these cytokines, we quantified global transcriptomic changes induced by the two cytokines and determined the relative contributions of STAT1 and STAT3 using genetic models and ChIP-seq. We found a high degree of overlap of the transcriptomes induced by IL-6 and IL-27 and extremely few examples in which the cytokines acted in opposition. Using STAT deficient cells and T cells from patients with gain-of-function STAT1 mutations, we show that STAT3 was responsible for the overall transcriptional output driven by both cytokines, whereas STAT1 was the driver of cytokine specificity. STAT1 did not compensate for the lack STAT3; on the contrary, much of STAT1 binding to chromatin was STAT3 dependent. Thus, STAT1 shapes the specific cytokine signature superimposed upon STAT3’s action. Overall design: Integrated analysis of transcriptome and transcription factor binding data from cytokine treated CD4+T cells","project":"SRP053186"}
{"number_samples":32,"species":"human","abstract":"Background: Transcription control of mitochondrial metabolism is essential for cellular function. A better understanding of this process will aid the elucidation of mitochondrial disorders, in particular of the many genetically unsolved cases of oxidative phosphorylation (OXPHOS) deficiency. Yet, to date only few studies have investigated nuclear gene regulation in the context of OXPHOS deficiency. In this study, we combined RNA sequencing of human complex I-deficient patient cells across 32 conditions of perturbed mitochondrial metabolism, with a comprehensive analysis of gene expression patterns, co-expression calculations and transcription factor binding sites. Results: Our analysis shows that OXPHOS genes have a significantly higher co-expression with each other than with other genes, including mitochondrial genes. We found no evidence for complex-specific mRNA expression regulation in the tested cell types and conditions: subunits of different OXPHOS complexes are similarly (co-)expressed and regulated by a common set of transcription factors. However, we did observe significant differences between the expression of OXPHOS complex subunits compared to assembly factors, suggesting divergent transcription programs. Furthermore, complex I co-expression calculations identified 684 genes with a likely role in OXPHOS biogenesis and function. Analysis of evolutionarily conserved transcription factor binding sites in the promoters of these genes revealed almost all known OXPHOS regulators (including GABP, NRF1/2, SP1, YY1, E-box factors) and a set of six yet uncharacterized candidate transcription factors (ELK1, KLF7, SP4, EHF, ZNF143, and EL2). Conclusions: OXPHOS genes share an expression program distinct from other mitochondrial genes, indicative of targeted regulation of this mitochondrial sub-process. Within the subset of OXPHOS genes we established a difference in expression between subunits and assembly factors. Most transcription regulators of genes that co-express with complex I are well-established factors for OXPHOS biogenesis. For the remaining six factors we here suggest for the first time a link with transcription regulation in OXPHOS deficiency. Overall design: RNA-SEQ of whole cell RNA in 2 control and 2 complex I deficient patient fibroblast cell lines treated with 4 compounds in duplicate, resulting in a total of 2x2x4x2=32 samples","project":"SRP053190"}
{"number_samples":4,"species":"human","abstract":"Much remains unknown concerning the mechanism by which the splicing machinery pinpoints short exons within intronic sequences and how splicing factors are directed to their pre-mRNA targets. Part of the explanation probably lies in differences in chromatin organization between exons and introns. Proteomic, co-immunoprecipitation, and sedimentation analyses described here indicated that SF3B1, an essential splicing component of the U2 snRNP complex, is strongly associated with nucleosomes. ChIP-seq and RNA-seq analyses revealed that SF3B1 is specifically bound to nucleosomes located at exonic positions. SF3B1 binding is enriched at nucleosomes positioned over short exons flanked by long introns that are also characterized by differential GC content between exons and introns. Disruption of SF3B1 binding to such nucleosomes affected the splicing of these exons similarly to inhibition of SF3B1 expression. Our findings suggest that the association of SF3B1 with nucleosomes is functionally important for splice site recognition and that SF3B1 conveys splicing-relevant information embedded in chromatin structure. Overall design: MNase-seq on Input and SF3B1 pull-down, mRNA-seq on control and SF3B1 si-RNA treated cells as well as on TSA (Trichostatin A) treated and untreated cells.","project":"SRP053195"}
{"number_samples":72,"species":"human","abstract":"We examine how NGS sequencing of sperm can provide a window as to how particular perturbations of the sperm RNA profile from baseline may be indicative of male factor infertility, and may thus provide direction as to proper course of infertility treatment for couple. Overall design: NGS RNA-seq of 72 sperm samples from male partner of couples undergoing fertility treatment","project":"SRP053246"}
{"number_samples":34,"species":"human","abstract":"Recent technological advances have made transcriptome sequencing (RNA-seq) possible in cells with low RNA copy number including platelets. Resulting studies have used RNA-seq in platelets isolated from healthy individuals to characterize the platelet transcriptome. However, platelets, possibly through gene expression changes, contribute to the etiology of and response to cardiovascular disease and events. To address this, we performed the largest human platelet RNA-seq analysis to date in 34 platelet samples: 16 ST-segment elevation myocardial infarction (STEMI), 16 non-STEMI (NSTEMI), and 2 controls. Overall design: RNA-seq of platelet samples from 34 individuals: 16 with ST-elevation myocardial infarction (STEMI), 16 with non-STEMI, and 2 non-myocardial infarction controls","project":"SRP053296"}
{"number_samples":3,"species":"human","abstract":"Phospholamban R14del mutazion (PLN-R14del) has been identified in a large family pedigree in which heterozygous carriers exhibited inherited dilated cardiomyopathy (DCM) and death by middle age. To better understand the causal link between the mutations in PLN and DCM pathology, we derived induced pluripotent stem cells from a DCM patient carrying the PLN R14del mutation. We showed that iPSC-derived cardiomyocytes recapitulated the DCM-specific phenotype and demonstrated that either TALEN-mediated genetic correction or combinatorial gene therapy resulted in phenotypic rescue. Our findings offer novel insights into the pathogenesis caused by mutant PLN and point to the development of potential new therapeutics of pathogenic genetic variants associated with inherited cardiomyopathies. Overall design: iPSCs were derived from a female patient carrying a heterozygous mutation (R14del) in the PLN gene. Tree samples were analyzed: Cardiomyocytes derived from PLN-R41del iPSC cells (R14del-CM); R14del-CMs infected with AAV6-EGFP-miR-PLN and R14del-CMs infected with AAV6-EGFP-miR-luc used as a negative control","project":"SRP053366"}
{"number_samples":32,"species":"human","abstract":"We recently identified ISRIB as a potent inhibitor of the integrated stress response (ISR). ISRIB renders cells resistant to the effects of eIF2a phosphorylation and enhances long-term memory in rodents (10.7554/eLife.00498).  Here we show by genome-wide in vivo ribosome profiling that translation of a restricted subset of mRNAs is induced upon ISR activation. ISRIB substantially reversed the translational effects elicited by phosphorylation of eIF2a and induced no major changes in translation or mRNA levels in unstressed cells. eIF2a phosphorylation-induced stress granule (SG) formation was blocked by ISRIB. Strikingly, ISRIB addition to stressed cells with pre-formed SGs induced their rapid disassembly, liberating mRNAs into the actively translating pool. Restoration of mRNA translation and modulation of SG dynamics may be an effective treatment of neurodegenerative diseases characterized by eIF2a phosphorylation, SG formation and cognitive loss. Overall design: Ribosome profiling with paired RNA-seq","project":"SRP053402"}
{"number_samples":40,"species":"human","abstract":"Purpose: provide evidence that RNA-seq can add information to transcriptome profiling already discovered by other technologies for atopic dermatitis Methods: mRNA profiles of 20 atopic dermatitis were analyzed to compare lesional and non-lesional skin, then transcriptomes found by reads were compared to Microarray and RT-PCR Results:RNA-seq provided complementary genes to AD transcriptome IL-36 and TREM-1 Conclusions: Our study represents the first analysis of lesional AD tissue by RNA-seq and comparison to microarray and RT-PCR Overall design: paired biopsies from lesional and non-lesional tissue of 20 patients sequenced by RNA-seq","project":"SRP053794"}
{"number_samples":10,"species":"human","abstract":"We applied ribosome profiling and RNA sequencing to examine gene expression regulation during oncogenic cell transformation. One model involves normal mammary epithelial cells (MCF10A) containing ER-Src. Treatment of such cells with tamoxifen rapidly induces Src, thereby making it possible to kinetically follow the transition between normal and transformed cells. The other model consists of three isogenic cell lines derived from primary fibroblasts in a serial manner (Hahn et al., 1999). EH cell is immortalized by overexpression of telomerase (hTERT), and exhibits normal fibroblast morphology. EL cell expresses hTERT along with both large and small T antigens of Simian virus 40, and it displays an altered morphology but is not transformed. ELR cell expresses hTERT, T antigens, and an oncogenic derivative of Ras (H-RasV12). Overall design: Ribosome profiling and RNA sequencing in two cancer cell models","project":"SRP054971"}
{"number_samples":8,"species":"human","abstract":"RNA-seq of HEK293T over-expressing WWTR-CAMTA1 fusion","project":"SRP055005"}
{"number_samples":18,"species":"human","abstract":"Elucidating the extent and consequences of genetic differences between humans is essential for understanding phenotypic diversity and personalized medicine. Although variation in RNA levels, transcription factor binding and chromatin have been explored, little is known about global variation in translation and its genetic determinants among humans. We used ribosome profiling, RNA sequencing, and mass spectrometry to perform an integrated analysis in lymphoblastoid cell lines from a diverse group of individuals. We find significant differences in RNA levels, translation, and protein abundance suggesting diverse mechanisms of personalized gene expression control. Combined analysis of RNA expression and ribosome occupancy improves the identification of individual protein level differences.  Finally, we identify genetic differences that specifically modulate ribosome occupancy - many of these differences lie close to start codons and upstream ORFs. Our results reveal a new level of gene expression variation among humans and indicate that genetic variants can cause changes in protein levels through effects on translation. Overall design: Ribosome profiling and RNA sequencing experiments from human lymphoblastoid cells","project":"SRP055009"}
{"number_samples":6,"species":"human","abstract":"Circular RNAs (circRNAs) are an endogenous class of animal RNAs. Despite their abundance, their function and expression in the nervous system are unknown. Therefore, we sequenced RNA from different brain regions, primary neurons, isolated synapses, as well as during neuronal differentiation. Using these and other available data, we discovered and analyzed thousands of neuronal human and mouse circRNAs. circRNAs were extraordinarily enriched in the mammalian brain, well conserved in sequence, often expressed as circRNAs in both human and mouse, and sometimes even detected in Drosophila brains. circRNAs were overall upregulated during neuronal differentiation, highly enriched in synapses, and often differentially expressed compared to their mRNA isoforms. circRNA expression correlated negatively with expression of the RNA-editing enzyme ADAR1. Knockdown of ADAR1 induced elevated circRNA expression. Together, we provide a circRNA brain expression atlas and evidence for important circRNA functions and values as biomarkers. Overall design: To assess circRNA expression in mammalian brain, we sequenced and analyzed mouse brain regions (hippocampus, cerebellum, prefrontal cortex and olfactory bulb), various neuronal differentiation (mouse P19 and human SH-SY5Y cells) and maturation (mouse cortical neurons) stages, and subcellular compartments in mouse (synaptoneurosomal fraction, cytoplasmic fraction, whole brain lysate).","project":"SRP055027"}
{"number_samples":4,"species":"human","abstract":"Purpose: The oxidation resistance gene 1 (OXR1) is crucial for protecting against oxidative stress, limiting neurodegeneration and maintaining normal lifespan in eukaryotes. However, the molecular function of OXR1 is still unknown. Previously we showed that human OXR1 regulates expression of antioxidant genes GPX2 and HO-1 via the p21 signaling pathway.  To examine the role of hOXR1 in global transcription regulation during cellular stress, we employed RNA sequencing to investigate the transcription profile in hOXR1 depleted HeLa cells. Methods: Control siRNA (siCon) and human OXR1 siRNA (siOXR1) transfected HeLa cells were either harvested directly (siCon_NT, siOXR1_NT) or exposed to 0.5 mM H2O2 for 1 h and then harvested immediately without recovery (siCon_R0h, siOXR1_R0h). Total RNA pooled from duplicate samples was used for RNA sequencing on an Illumina HiSeq2000 platform. The sequence reads that passed quality filters were analyzed at gene level. The Blast2GO program was used to generate gene ontology (GO) annotation of differentially expressed genes (DEGs).The WEGO software was used to further perform GO functional classification and to predict pathways affected. qRT–PCR validation was performed using SYBR Green assays. Results: In total, in non-treated and hydrogen peroxide exposed cells, hOXR1 depletion results in the down-regulation of 554 genes and the up-regulation of 253 genes. These differentially expressed genes include transcription factors (i.e. HIF1A, SP6, E2F8 and TCF3) and numerous genes of the p53 signaling pathway involved in cell-cycle arrest (i.e. cyclin D, CDK6 and p21) and apoptosis (i.e. CytC and CASP9). Western blot analysis reveals that hOXR1 suppresses CASP9 protein expression and reduces post-translational cleavage into its active form. After exposure to hydrogen peroxide (1 h), 56 early response genes were up-regulated in hOXR1 depleted cells, in which 38 of these genes were not induced in control cells. In addition, a subset of the commonly up-regulated early response genes showed a stronger induction in hOXR1 depleted cells (i.e.  FOS, JUN and DUSP1). Out of a total of 52 differentially expressed transcription factors in hOXR1 depleted cells under normal physiology and oxidative stress condition, 14 genes (including HIF1A, STAT5A, E2F8 and TCF3) are differentially regulated under H2O2 treatment in hOXR1 silenced cells as compared to control cells.  Finally, we demonstrate that hOXR1 depleted cells undergo cell cycle arrest in G2/M phase during oxidative stress. Conclusions: Human OXR1 is important for regulation of the early stress response to oxidative stress in HeLa cells. HOXR1 modulates the p53 signaling pathway via regulation of genes involved in cell cycle arrest, apoptosis and anti-oxidation. Further, hOXR1 regulates numerous transcription factors of importance for cellular stress responses. In summary, hOXR1 may act as a sensor of cellular oxidative stress to regulate the transcriptional networks required to detoxify reactive oxygen species (ROS) and modulate cell cycle arrest and cell death (apoptosis). Overall design: The mRNA profiles of hOXR1 depleted and control Hela cells with or without H2O2 treatment 1 h were generated by RNA sequencing using Illumina Hiseq 2000.","project":"SRP055048"}
{"number_samples":15,"species":"human","abstract":"The corneal endothelium plays a primary role in maintaining corneal homeostasis and clarity, and must be surgically replaced with allogenic donor corneal endothelium in the event of visually significant dysfunction. However, a worldwide shortage of donor corneal tissue has led to a search for alternative sources of transplantable tissue. Cultured human corneal endothelial cells (HCEnC) have been shown to restore corneal clarity in experimental models of corneal endothelial dysfunction in animal models, but characterization of cultured HCEnC remains incomplete. To this end, we utilized next-generation RNA sequencing technology to compare the transcriptomic profile of ex vivo human corneal endothelium (evHCEnC) with that of primary HCEnC and HCEnC lines, and to determine the utility of cultured and immortalized corneal endothelial cells as models of in vivo corneal endothelium. Multidimensional analyses of the transcriptome datasets demonstrated that primary HCEnC have a closer relationship to evHCEnC than do immortalized HCEnC. Subsequent analyses showed that the majority of the genes specifically expressed in HCEnC (not expressed in ex vivo corneal epithelium or fibroblasts) demonstrated a marked variability of expression in cultured cells compared with evHCEnC. In addition, genes associated with either corneal endothelial cell function or corneal endothelial dystrophies were investigated. Significant differences in gene expression and protein levels were observed in the cultured cells compared with evHCEnC for each of the genes tested except for AGBL1 and LOXHD1, which were not detected by RNA-seq or qPCR. Our transcriptomic analysis suggests that at a molecular level primary HCEnC most closely resemble evHCEC and thus represent a viable therapeutic option for managing corneal endothelial dysfunction. Our findings also suggest that investigators should perform an assessment of the entire transcriptome of cultured HCEnC prior to determination of the potential clinical utility of the cultured HCEnC for the management of corneal endothelial cell failure. Overall design: Transcriptomes from ex vivo corneal endothelium, primary cultures and three cell lines were compared. Three samples of each endothelial cell group were submitted for RNA sequencing for a total of 15 samples. The transcriptome for the ex vivo corneal endothelium was used as the reference (i.e., proxy for in vivo corneal endothelium). Transcript abundances for a subset of genes associated with corneal endothelial cell function or disease were validated with qPCR and western blot. Samples of ex vivo endothelium used for validation were independent replicates not used for RNA-sequencing.","project":"SRP055101"}
{"number_samples":6,"species":"human","abstract":"To clarify the lineage relationship between IL3Rahigh- and IL3Ralow precursor cells and to find potential molecules involved in their differentiation, we compared the IL3Rahigh- and IL3Ralow precursor populations from three independent donors by mRNA deep sequencing and used the Ingenuity Pathway Analysis (IPA)- and Multi-experimental Viewer (MeV) to analyze the differentially expressed genes (p<0.001). Analysis of the protein coding genes showed that the samples from IL3Rahigh precursor cells clustered together, as did the IL3Ralow samples. This indicated that the gene expression patterns of these cells are likely to be conserved. Further analysis revealed a list of (649) differentially expressed molecules between the two populations.  Among these, most notably, genes involved in the differentiation of cell in general, amongst which differentiation of MF, OC and antigen presenting cells appeared to be activation increased. Overall design: Examination of two hematopoietic precursor populations in human BM","project":"SRP055103"}
{"number_samples":18,"species":"human","abstract":"Alu elements are major contributors to lineage-specific new exons in primate and human genomes. Recent studies indicate that some Alu exons have high transcript inclusion levels or tissue-specific splicing profiles, and may play important regulatory roles in modulating mRNA degradation or translational efficiency. However, the contribution of Alu exons to the human proteome remains unclear and controversial. The prevailing view is that exons derived from young repetitive elements, such as Alu elements, are restricted to regulatory functions and have not had adequate evolutionary time to be incorporated into stable, functional proteins. We adopt a proteotranscriptomics approach to systematically assess the contribution of Alu exons to the human proteome. Using RNA sequencing, ribosome profiling and proteomics data from human tissues and cell lines, we provide evidence for the translational activities of Alu exons and the presence of Alu exon derived peptides in human proteins. These Alu exon peptides represent species-specific protein differences between primates and other mammals, and in certain instances between humans and closely related primates. In the case of the RNA editing enzyme ADARB1, which contains an Alu exon peptide in its catalytic domain, RNA sequencing analyses of A-to-I editing demonstrate that both the Alu exon skipping and inclusion isoforms encode active enzymes. The Alu exon derived peptide may fine tune the overall editing activity and, in limited cases, the site selectivity of ADARB1 protein products. Our data indicate that Alu elements have contributed to the acquisition of novel protein sequences during primate and human evolution. Overall design: Comparing the A-I RNA editing levels of HEK293 cells transfected with empty vector (CV) control, Alu exon inclusion (long) isoform of ADARB1, and Alu exon skipping (short) isoform of ADARB1. Each group has 3 replicates.","project":"SRP055105"}
{"number_samples":6,"species":"human","abstract":"We identify perhexiline, a small molecule inhibitor of mitochondrial carnitine palmitoyltransferase-1, as a HES1-signature antagonist drug with robust antileukemic activity against NOTCH1 induced leukemias in vitro and in vivo. Overall design: RNA-Seq from CUTLL1 cell lines treated with Perhexiline or vehicle for 3 days","project":"SRP055108"}
{"number_samples":98,"species":"human","abstract":"We performed single-cell and bulk transcriptome profiling in two different human cell lines. We performed single-cell RNA sequencing in live and fixed cells. Overall design: Single cell RNA sequencing of live and fixed cells, bulk RNA sequencing in two cell lines.","project":"SRP055153"}
{"number_samples":15,"species":"human","abstract":"Effect of various synthetic ligands called diabodies on primary human megakaryocyte-erythrocyte progenitors (MEPs) isolated from bone marrow of a normal donor. The diabodies used in our studies act as either agonists or antagonists for erythropoietin receptor (EPOR) expressed on the surface of MEPs.","project":"SRP055180"}
{"number_samples":52,"species":"human","abstract":"Chronic lymphocytic leukemia (CLL) is a biologically and clinically heterogeneous disease. The somatic hypermutation status of the immunoglobulin heavy chain variable (IGHV) genes has been identified as one of the most robust prognostic markers in CLL. Patients with unmutated IGHV status (U-CLL) typically experience an inferior outcome compared to those whose clones express mutated IGHV genes (M-CLL). We conducted a genome-wide DNA methylation analysis in CD19+ B-cells from a group of 43 CLL patients using reduced representation bisulfite sequencing (RRBS). Using base-pair resolution methylation sequencing, 2323 differentially methylated regions between CLL and normal B-cells (CLL-specific DMRs) and 569 between M-CLL and U-CLL samples (IGHV-specific DMRs) were identified in the CLL genomes. The IGHV-specific DMRs are mostly unique when compared to the CLL-specific DMRs. Less than 10% of the IGHV-specific DMRs are located in promoter regions; however, more than half of these overlap with known DNase I hypersensitive sites, enhancer regions marked by histone modification (H3K4Me1 and H3K27Ac), and transcription factor binding sites in the ENCODE datasets, which indicates that these DMRs contain regulatory sequences. Distinctive DNA methylation patterns were observed in M-CLL and U-CLL samples. Overall, U-CLL was found to contain 50% more hypermethylated regions than M-CLL samples. The hypermethylated loci observed in the U-CLL samples also appear to be hypermethylated in normal naïve B-cells as compared memory B-cells, suggesting that M-CLL and U-CLL differ in differentiation status corresponding to normal B-cell differentiation stages. RNA-seq analysis performed using matched samples (n=34), in which both DNA methylation and gene expression data were available, demonstrated excellent correlation between DNA methylation and gene expression. Several genes whose expression status was previously shown to be associated with CLL prognosis such as ZAP70, CRY1, LDOC1, SEPT10, LAG3, and LPL were differentially methylated in the promoter regions between M-CLL and U-CLL samples indicating that DNA methylation plays an important role in defining the gene expression patterns of these prognostic genes. We further validated 9 genes with IGHV-specific DMRs in the promoter regions using bisulfite pyrosequencing, and the results demonstrated excellent correlation between differential methylation and IGHV mutation status. These novel differentially methylated genes could be developed into biomarkers for CLL prognosis. In addition, DNA hypomethylation was observed in a significant number of genes involved in lymphocyte activation such as PDCD1, NFAT1, and CD5. DNA hypomethylation was observed in the proximal promoter and far up-stream enhancer regions of CD5, an important cell surface marker that uniquely identifies CLL. Overall, the DNA methylation landscape in CLL patients indicates that CLL B cells possess an active B-cell phenotype; at the same time, U-CLL and M-CLL are faithfully committed to their lineage resembling either naïve or memory B-cells.  In summary, this comprehensive DNA methylation analysis has identified a large number of novel epigenetic changes in CLL patients. The results from this study will further advance our understanding of the epigenetic contribution to molecular subtypes in CLL. Overall design: To perform a transcriptome analysis in CLL, we generated sequencing libraries from total RNA isolated from purified B-cells of CLL patients and healthy donnors. The RNA-seq libraries were sequenced using Illumina HiSeq2000 sequencer with a read length of 100bp.  11 CLL B-cell samples, 3 normal control samples including one each of normal CD19+ B cells were studied. We generated 20-30 million Illumina sequencing reads for each sample.","project":"SRP055390"}
{"number_samples":6,"species":"human","abstract":"We performed RNA sequencing in isogenic models of COX-1 proficient (OV3/SCR) and COX-1 deficient (OV3/COX1KD) OVCAR-3 ovarian cancer cells. COX-1 knockdown was associated with a coordinated anti-oncogenic phenotype, with growth, angiogenesis, migration/invasion, and epithelial-mesenchymal transition among the pathways down-regulated. Overall design: RNA sequencing was performed at Vanderbilt Technologies for Advanced Genomics (VANTAGE) using Illumina HiSeq 2500.","project":"SRP055392"}
{"number_samples":11,"species":"human","abstract":"c-MYC (MYC) overexpression or hyperactivation is one of the most common drivers of human cancer. Despite intensive study, the MYC oncogene remains recalcitrant to therapeutic inhibition. Like other classic oncogenes, hyperactivation of MYC leads to collateral stresses onto cancer cells, suggesting that tumors harbor unique vulnerabilities arising from oncogenic activation of MYC. Herein, we discover the spliceosome as a new target of oncogenic stress in MYC-driven cancers. We identify BUD31 as a MYC-synthetic lethal gene, and demonstrate that BUD31 is a splicing factor required for the assembly and catalytic activity of the spliceosome. Core spliceosomal factors (SF3B1, U2AF1, and others) associate with BUD31 and are also required to tolerate oncogenic MYC. Notably, MYC hyperactivation induces an increase in total pre-mRNA synthesis, suggesting an increased burden on the core spliceosome to process pre-mRNA. In contrast to normal cells, partial inhibition of the spliceosome in MYC-hyperactivated cells leads to global intron retention, widespread defects in pre-mRNA maturation, and deregulation of many essential cell processes. Importantly, genetic or pharmacologic inhibition of the spliceosome in vivo impairs survival, tumorigenicity, and metastatic proclivity of MYC-dependent breast cancers. Collectively, these data suggest that oncogenic MYC confers a collateral stress on splicing and that components of the spliceosome may be therapeutic entry points for aggressive MYC-driven cancers. Overall design: Examination of intron rentention in MYC-ER HMECs, in 4 conditions","project":"SRP055411"}
{"number_samples":33,"species":"human","abstract":"There is a need for reliable prognostic markers that can guide therapeutic intervention in Crohn’s disease (CD). We examined whether different behavioral phenotypes in CD can be classified based on colonic miRNA or mRNA expression and if miRNAs have prognostic utility for CD. We perform high-throughput sequencing of small RNA and mRNA isolated from colon tissue from CD patients and non-IBD (NIBD) controls. To identify miRNA and genes associated with specific behavioral phenotypes of CD, patients were stratified according to disease behavior (non-stricturing, non-penetrating; stricturing; penetrating) and compared miRNA profiles in each class with those of the NIBD group. Using a novel statistical simulation approach applied to colonic RNA-seq data for CD patients and NIBD controls, we identify at drivers of gene expression profiles associated with CD. Overall design: Macroscopically non-inflamed colon tissue from well-characterized Crohn's disease patients and normal controls were obtained. Small RNA-seq and RNA-seq were performed on these samples. Additionally, we investigated the effect of inflammation on miRNA expression by performing small RNA-seq on matched colon samples obtained from macroscopically inflamed regions from a subset (six) of these patients with Crohn's Disease.","project":"SRP055438"}
{"number_samples":9,"species":"human","abstract":"The human cerebral cortex depends for its normal development and size on a precisely controlled balance between self-renewal and differentiation of diverse neural progenitor cells. Specialized progenitors that are common in humans, but virtually absent in rodents, called ‘outer radial glia’ (ORG), have been suggested to be crucial to the evolutionary expansion of the human cortex. We combined cell type-specific sorting with transcriptome-wide RNA-sequencing to identify genes enriched in human ORG, including targets of the transcription factor Neurogenin, and previously uncharacterized, evolutionarily dynamic, long noncoding RNAs. Single-cell transcriptional profiling of human, ferret, and mouse progenitors showed that more human RGC co-express proneural Neurogenin targets than in ferret or mouse, suggesting greater self-renewal of neuronal lineage-committed progenitors in humans. Finally, we show that activating the Neurogenin pathway in ferret RGC promotes delamination and outward migration. Thus, we find that the abundance of human ORG is paralleled by increased transcriptional heterogeneity of cortical progenitors. Overall design: Three biological replicates of human late mid-fetal cortex (18 to 19 weeks  of gestation) were dissociated and immunolabeled. Apical and outer radial glial cells were purified by FACS and compared to an immunonegative population, predominantly neurons.","project":"SRP055440"}
{"number_samples":17,"species":"human","abstract":"IGHV mutation status is a well-established prognostic factor in chronic lymphocytic leukemia, and also provides crucial insights into tumor cell biology and function. Currently, determination of IGHV transcript sequence, from which mutation status is calculated, requires a specialized laboratory procedure. RNA sequencing is a method that provides high resolution, high dynamic range transcriptome data that can be used for differential expression, isoform discovery, and variant determination. In this paper, we demonstrate that unselected next-generation RNA sequencing can accurately determine the IGH@ sequence, including the complete sequence of the complementarity-determining region 3 (CDR3), and mutation status of CLL cells, potentially replacing the current method which is a specialized, single-purpose Sanger-sequencing based test. Overall design: CLL cells were sequenced by mRNA-seq on the Illumina platform then subjected to the costom bioinformatic pipeline Ig-ID which yields IGH data","project":"SRP055444"}
{"number_samples":15,"species":"human","abstract":"Most RNAs within polarized cells such as neurons are sorted subcellularly in a coordinated manner. Despite advances in the development of methods for profiling polyadenylated RNAs from small amounts of input RNA, techniques for profiling coding and non-coding RNAs simultaneously are not well established. Here, we optimized a transcriptome profiling method based on double-random priming and applied it to serially diluted total RNA down to 10 pg. Read counts of expressed genes were robustly correlated between replicates, indicating that the method is both reproducible and scalable. Our transcriptome profiling method detected both coding and long non-coding RNAs sized >300 bases. Compared to total RNA-seq using a conventional approach, our protocol detected 70% more genes due to reduced capture of ribosomal RNAs. We used our method to analyze the RNA composition of compartmentalized motoneurons. The somatodendritic compartment was enriched for transcripts with post-synaptic functions as well as for certain nuclear non-coding RNAs such as 7SK. In axons, transcripts related to translation were enriched including the cytoplasmic non-coding RNA 7SL. Our profiling method can be applied to a wide range of investigations including perturbations of subcellular transcriptomes in neurodegenerative diseases, and investigations of microdissected tissue samples such as anatomically defined fiber tracts. Overall design: Total number of samples is 43","project":"SRP055447"}
{"number_samples":9,"species":"human","abstract":"Gene expression profiling of normal human fibroblasts treated with Novartis compound NVS-SM1, Novartis compound NVS-SM3 (NVS-SM1 inactive analog), and DMSO for 24 hours. This study allowed to elucidate the mechanism of action of NVS-SM1, a potent modulator of SMN2 splicing.","project":"SRP055454"}
{"number_samples":12,"species":"human","abstract":"To improve our understanding of lncRNA expression in T cells, we used whole genome sequencing (RNA-seq) to identify lncRNAs expressed in human T cells and those selectively expressed in T cells differentiated under TH1, TH2, or TH17 polarizing conditions.  The majority of these lineage-specific lncRNAs are co-expressed with lineage-specific protein-coding genes.  These lncRNAs are predominantly intragenic with co-expressed protein-coding genes and are transcribed in sense and antisense orientations with approximately equal frequencies.  Further, genes encoding TH lineage specific mRNAs are not randomly distributed across the genome but are highly enriched in the genome in genomic regions also containing genes encoding TH lineage-specific lncRNAs.  Our analyses also identify a cluster of antisense lncRNAs transcribed from the RAD50 locus that are selectively expressed under TH2 polarizing conditions and co-expressed with IL4, IL5 and IL13 genes.  Depletion of these lncRNAs via selective siRNA treatment demonstrates the critical requirement of these lncRNAs for expression of the TH2 cytokines, IL-4, IL-5 and IL-13.  Collectively, our analyses identify new lncRNAs expressed in a TH lineage specific manner and identify a critical role for a cluster of lncRNAs for expression of genes encoding TH2 cytokines. Overall design: Human peripheral blood mononuclear cells (PBMC) were cultured under TH1, TH2, and TH17 polarizing conditions. TH1, TH2, and TH17 primary and effector cultures were isolated and poly(A)+ and total RNA sequencing performed.","project":"SRP055474"}
{"number_samples":12,"species":"human","abstract":"In several developmental lineages, an increase in expression of the MYC proto-oncogene drives the transition from quiescent stem cells to transit amplifying cells. The mechanism by which MYC restricts self-renewal of adult stem cells is unknown. Here, we show that MYC activates a stereotypic transcriptional program of genes involved in protein translation and mitochondrial biogenesis in mammary epithelial cells and indirectly inhibits the YAP/TAZ co-activators that are essential for mammary stem cell self-renewal. We identify a phospholipase of the mitochondrial outer membrane, PLD6, as the mediator of MYC activity. PLD6 mediates a change in the mitochondrial fusion/fission balance that promotes nuclear export of YAP/TAZ in a LATS- and RHO-independent manner. Mouse models and human pathological data confirm that MYC suppresses YAP/TAZ activity in mammary tumors. PLD6 is also required for glutaminolysis, arguing that MYC-dependent changes in mitochondrial dynamics balance cellular energy metabolism with the self-renewal potential of adult stem cells. Overall design: RNA-Seq Experiments in 2 different primary breast epithelial cell lines (HMLE, which were sorted according to CD44/CD24 surface markers & unsorted IMEC). Both cell lines expressed a doxycycline-inducible version of MYC. For the HMLE cell line DGE analysis was performed for the uninduced (EtOH) situation, comparing CD44high vs CD44 low and for the induced situation Dox vs. EtOH for the CD44high population. For the IMEC cell line DGE was performed by comparing Dox-treated populations expressing either Dox-inducible MYC or a vector control which allows to filter out potential effects due to doxycycline treatment.","project":"SRP055475"}
{"number_samples":12,"species":"human","abstract":"Tandem DCIS/IDC are defined as ductal carcicnoma in situ (DCIS) lesions that have concurrent invasive ductal carcinoma (IDC) within the same breast. These are identified radiologically by an area of clustered microcalcifications adjacent to (contiguous with) an invasive mass. Our radiologist (Dr. William P. Smith) has provided us with biopsy cores from each region. One core from each region (DCIS and IDC) has bas been collected and subjected to RNA sequencing for our studies to compare changes from DCIS to IDC in each individual patient. Overall design: 6 pairs of DCIS-IDC samples were collected, and analysed by RNA sequencing","project":"SRP055512"}
{"number_samples":111,"species":"human","abstract":"Gene expression is highly dynamic during fetal development and determines tissue specification and function. In humans, the transcriptional profile of different organs during development has not been systematically studied. However, understanding how a particular tissue acquires its tissue identity will give insight into the development and maturation of tissues from a developmental biology perspective. Overall design: Next-generation sequencing (DeepSAGE) dataset of 111 RNA samples representing 21 different human fetal organs and the maternal endometrium at three timepoints (gestational ages) during first and second trimester development (W9, W16-18, W22)","project":"SRP055513"}
{"number_samples":76,"species":"human","abstract":"Genome-wide expression profiles in peripheral monocytes (PM) from 19 obese women before and 3 months after bariatric surgery using the RNA-seq technology. This dataset is linked to the dataset GSE65540 providing expression profiles in subcutaneous adipose tissue (SAT) in the same population. Due to exclusion of some individuals for technical reasons, the overlap between the 2 datasets is of 18 women. Overall design: mRNA sequencing of peripheral monocyte (PM) samples from 19 obese women before and 3 months after bariatric surgery","project":"SRP055514"}
{"number_samples":643,"species":"human","abstract":"Single cell transcriptomics has emerged as a powerful approach to dissecting phenotypic heterogeneity in complex, unsynchronized cellular populations.  However, many important biological questions demand quantitative analysis of large numbers of individual cells. Hence, new tools are urgently needed for efficient, inexpensive, and parallel manipulation of RNA from individual cells. We report a simple microfluidic platform for trapping single cell lysates in sealed, picoliter microwells capable of “printing” RNA on glass or capturing RNA on polymer beads. To demonstrate the utility of our system for single cell transcriptomics, we developed a highly scalable technology for genome-wide, single cell RNA-Seq. The current implementation of our device is pipette-operated, profiles hundreds of individual cells in parallel with library preparation costs of ~$0.10-$0.20/cell, and includes five lanes for simultaneous experiments. We anticipate that this system will ultimately serve as a general platform for large-scale single cell transcriptomics, compatible with both imaging and sequencing readouts.!Series_type = Expression profiling by high throughput sequencing Overall design: A microfluidic device that pairs sequence-barcoded mRNA capture beads with individual cells was used to barcode cDNA from individual cells which was then pre-amplified by in vitro transcription in a pool and converted into an Illumina RNA-Seq library. Libraries were generated from ~600 individual cells in parallel and extensive analysis was done on 396 cells from the U87 and MCF10a cell lines and from ~500 individual cells with extensive analysis on 247 cells from the U87 and WI-38 cell lines. Sequencing was done on the 3'-end of the transcript molecules.  The first read contains cell-identifying barcodes that were present on the capture bead and the second read contains a unique molecular identifier (UMI) barcode, a lane-identifying barcode, and then the sequence of the transcript.","project":"SRP055569"}
{"number_samples":11,"species":"human","abstract":"Introduction: Expansion of antigen (Ag)-specific natural occurring regulatory T cells (nTregs) is required to obtain sufficient numbers of cells for cellular immunotherapy. In this study, different allogeneic stimuli were studied for their capacity to generate functional alloAg-specific nTregs. Methods: A highly enriched nTreg-fraction (CD4+CD25brightCD127- T cells) was alloAg-specific expanded using HLA-mismatched immature, mature monocyte-derived dendritic cells (moDC) or peripheral blood mononuclear cells (PBMC). The allogeneic mature moDC-expanded nTregs were fully characterized by analysis of the demethylation status within the TSDR of the FOXP3 gene and the expression of both protein and mRNA of FOXP3, HELIOS, CTLA4 and cytokines. In addition, the antigen-specific suppressive capacity of these expanded nTregs was tested. Results: Allogeneic mature moDC and skin-derived DC were superior in inducing nTreg-expansion compared to immature moDC or PBMC in an HLA-DR and CD80/CD86-dependent way. Remarkably, the presence of exogenous IL-15 without IL-2, could facilitate optimal mature moDC-induced nTreg-expansion. Allogeneic mature moDC-expanded nTregs were at low ratios (<1:320), potent suppressors of alloAg-induced proliferation without significant suppression of completely HLA-mismatched-Ag-induced proliferation. Mature moDC-expanded nTregs were highly demethylated at the TSDR within the FOXP3 gene and highly expressed of FOXP3, HELIOS and CTLA4.  A minority of the expanded nTregs produced IL-10, IL-2, IFN-g and TNF-a but very few IL-17 producing nTregs were found. Next generation sequencing of mRNA of moDC-expanded nTregs revealed a strong induction of Treg-associated mRNAs. Conclusions: Human allogeneic mature moDC are highly efficient stimulator cells, in presence of exogenous IL-15, for expansion of stable alloAg-specific nTregs with superior suppressive function. Overall design: Four different batches of highly pure regulatory T cells (all from the same donor) were expanded in two different ways, and compared to non-expanded samples.","project":"SRP055675"}
{"number_samples":4,"species":"human","abstract":"Genomic mechanism responsible for malignant transformation has been a major concern in glioma research, but differing conclusions have been drawn based on diverse study conditions. Therefore, it is indispensable to secure the direct evidence using longitudinal samples in the same patient. We have analyzed genomic data using next generation sequencing technology for the serial samples from 3 patients with IDH1 mutated gliomas who had undergone progression of the disease from histologically benign to malignant phenotype. Comprehensive analysis of genomic dynamics in clonal evolution during the malignant transformation and validation with public database revealed alterations in regulating machinery of gene expression involving spliceosomal complex, transcriptional factor, or chromatin remodeler. And, consequential expression changes implied activation of genes associated with restoration of stemness of the cancer cells. Despite being limited to small number of cases, this analysis provides direct example of genomic changes responsible for malignant transformation in gliomas.","project":"SRP055730"}
{"number_samples":16,"species":"human","abstract":"Glucocorticoids have major effects on adipose tissue metabolism. To study tissue mRNA expression changes induced by chronic elevated endogenous glucocorticoids, we performed RNA sequencing on subcutaneous adipose tissue from patients with Cushing’s disease (n=5) compared to patients with non-functioning pituitary adenomas (n=11). We found higher expression of transcripts involved in several metabolic pathways, including lipogenesis, proteolysis and glucose oxidation as well as decreased expression of transcripts involved in inflammation and protein synthesis. To further study this in a model system, we subjected mice to dexamethasone treatment for 12 weeks and analyzed their inguinal (subcutaneous) fat pads, which led to similar findings.  Additionally, mice treated with dexamethasone showed drastic decreases in lean body mass as well as increased fat mass, further supporting the human transcriptomic data. These data provide insight to transcriptional changes that may be responsible for the co-morbidities associated with chronic elevations of glucocorticoids Overall design: DESIGN: Patients with cushing's (n=5) or non-functioning pituitary adenoma (n=11) were prospectively observed from March 2011 to June 2012.","project":"SRP055749"}
{"number_samples":10,"species":"human","abstract":"Biochemical fractionation of HEK293 nuclei and RNA-seq of chromatin-associated and soluble-nuclear RNA. Overall design: Nuclei from three biological replicates were isolated by detergent lysis, fractionated, then three chromatin and three soluble RNA samples were converted to cDNA using Illumina TruSeq stranded protocol, and sequenced on Illumina HiSeq2000","project":"SRP055770"}
{"number_samples":6,"species":"human","abstract":"In recent years, several studies have shed light into the processes that regulate epidermal specification and homeostasis. We previously showed that a broad-spectrum ?–secretase inhibitor DAPT promoted early keratinocyte specification in human embryonic stem cells triggered to undergo ectoderm specification. Here, we show that DAPT accelerates human embryonic stem cell differentiation and induces expression of the ectoderm protein AP2. Furthermore, we utilize RNA sequencing to identify several candidate regulators of ectoderm specification including those involved in epithelial and epidermal development in human embryonic stem cells. Genes associated with transcriptional regulation and growth factor activity are significantly enriched upon DAPT treatment during specification of human embryonic stem cells to the ectoderm lineage. The human ectoderm cell signature identified in this study contains several genes expressed in ectodermal and epithelial tissues. Importantly, these genes are also associated with skin disorders and ectodermal defects, providing a platform for understanding the biology of human epidermal keratinocyte development under diseased and homeostatic conditions. Overall design: 6 samples were analyzed, 3 replicates of ETOH treated H1 HESCs and 3 replicates of DAPT treated H1 HESCs","project":"SRP055809"}
{"number_samples":86,"species":"human","abstract":"Here we provide fundamental insights into early human development by single-cell RNA-sequencing of human and mouse preimplantation embryos. We elucidate conserved transcriptional programs along with those that are human-specific. Importantly, we validate our RNA-sequencing findings at the protein level, which further reveals differences in human and mouse embryo gene expression. For example, we identify several genes exclusively expressed in the human pluripotent epiblast including the transcription factor KLF17. Key components of the TGF-ß signaling pathway including NODAL, GDF3, TGFBR1/ALK5, LEFTY1, SMAD2, SMAD4 and TDGF1 are also enriched in the human epiblast. Intriguingly, inhibition of TGF-ß signaling abrogates NANOG expression in human epiblast cells, consistent with a requirement for this pathway in pluripotency. Although key trophectoderm factors Id2, Elf5, and Eomes are exclusively localized to this lineage in the mouse, the human orthologues are either absent or expressed in alternative lineages. Importantly, we also identify genes with conserved expression dynamics including Foxa2/FOXA2, which we show is restricted to the primitive endoderm in both human and mouse embryos. Comparisons of the human epiblast to existing embryonic stem cells (hESCs) reveals conservation of pluripotency but also additional pathways more enriched in hESCs. Our analysis highlights significant differences in human preimplantation development compared to mouse and provides a molecular blueprint to understand human embryogenesis and its relationship to stem cells. Overall design: Single-Cell RNA-seq","project":"SRP055810"}
{"number_samples":8,"species":"human","abstract":"Despite its biological importance, transfer RNA (tRNA) could not be adequately sequenced due to the abundant presence of post-transcriptional modifications and extensive structure that interfere with cDNA synthesis and adapter ligation. We achieve efficient and quantitative tRNA sequencing by removing base methylations using engineered demethylases and using a highly processive thermo-stable reverse transcriptase without the need for adapter ligation (DMTRT-tRNA-seq). Our method should be applicable for biological investigations of tRNA in all organisms. Overall design: Development of tRNA-Seq method","project":"SRP055858"}
{"number_samples":14,"species":"human","abstract":"Identification of differentially spliced genes by wild type or S34F mutation of U2AF1 Overall design: Examination of effects on splicing events by overexpressing wipdtype or S34F mutation of two U2AF1 isoforms in A549 cells. All experimental conditions are performed in duplicate.","project":"SRP055860"}
{"number_samples":6,"species":"human","abstract":"Cancer cells must adapt metabolically to survive and proliferate when nutrients are limiting. Here we used combined transcriptional-metabolomic network analysis to identify metabolic pathways that support glucose-independent tumor cell growth. We found that glucose deprivation stimulated the re-wiring of the tricarboxylic acid (TCA) cycle and co-opting early steps of gluconeogenesis to promote cell proliferation under low glucose. Glucose limitation promoted the production of phosphoenolpyruvate (PEP) from glutamine, which was used to fuel biosynthetic pathways normally sustained by glucose, including serine and purine biosynthesis. Mitochondrial PEP-carboxykinase (PCK2) was required for this glutamine-dependent metabolic reprogramming. PCK2 expression was dependent on the hypoxia-inducible factors (HIFs), and required to maintain tumor cell proliferation under limiting glucose availability. Elevated PCK2 expression is observed in several human tumor types, and enriched in tumor tissue from non-small cell lung cancer (NSCLC) patients. Our results define a novel role for PCK2 in cancer cell metabolic reprogramming that promotes glucose-independent cell growth and metabolic stress resistance in human tumors. Overall design: A549 cell line was obtained from ATCC (Manassas, VA, USA). A549 cells were cultured in the presence (25 mM) or absence of glucose for 48 hours prior to RNA extraction.","project":"SRP055863"}
{"number_samples":14,"species":"human","abstract":"It is fundamentally unknown how normal cellular processes or responses to extracellular stimuli may invoke polyadenylation and degradation of ncRNA substrates or if human disease processes exhibit defects in polyadenylation of ncRNA substrates as part of their pathogenesis. Our results demonstrate that mononuclear cells from subjects with relapsing-remitting multiple sclerosis (RRMS) exhibit pervasive increases in levels of polyadenylated ncRNAs including Y1 RNA, 18S and 28S rRNA, and U1, U2, and U4 snRNAs and these defects are unique to RRMS. Defects in expression of both Ro60 and La proteins in RRMS appear to contribute to increased polyadenylation of ncRNAs. Further, IFN-ß1b, a common RRMS therapy, restores both Ro60 and La levels to normal as well as levels of polyadenylated Y1 RNA and U1 snRNA suggesting that aberrant polyadenylation of ncRNA substrates may have pathogenic consequences. Overall design: We extracted RNA from peripheral whole blood in healthy control subjects and patients with established relapsing-remitting multiple sclerosis using PaxGene tubes.","project":"SRP055874"}
{"number_samples":5,"species":"human","abstract":"We present a more extensive and yet precise assessment to elucidate differences and similarities in performance at numerous aspects including signal range, sensitivity to fold-change, and fidelity with TaqMan qRT-PCR. There were three levels of data examined: entire data sets, data derived from gene name annotation oriented subset of 15442 RefSeq genes, and data derived from transcript pattern defined subset of 7034 RefSeq genes.  Our results showed a fair degree of overall correlation between all 6 platforms evaluated; but, to varying degrees, two RNA-seq protocols outperformed three of the microarray platforms in most categories.  Notably, a fourth microarray platform, Agilent, was comparable, or marginally superior, to the RNA-seq protocols within these same assessments.  Furthermore, 3 platforms (Agilent and two RNA-seq methods) demonstrated over 80% concordance with the gold standard TaqMan assay in terms of fold-change accuracy.  Our study suggests that the use of transcript patterns can enhance a number of the observed cross-platform correlations, indicating a potential usefulness for similar evaluations. Overall design: The study assessed differences and similarities in performance at numerous aspects including signal range, sensitivity to fold-change, and fidelity with TaqMan qRT-PCR. There were three levels of data examined: entire data sets, data derived from gene name annotation oriented subset of 15442 RefSeq genes, and data derived from transcript pattern defined subset of 7034 RefSeq genes.","project":"SRP055916"}
{"number_samples":5,"species":"human","abstract":"We present a more extensive and yet precise assessment to elucidate differences and similarities in performance at numerous aspects including signal range, sensitivity to fold-change, and fidelity with TaqMan qRT-PCR. There were three levels of data examined: entire data sets, data derived from gene name annotation oriented subset of 15442 RefSeq genes, and data derived from transcript pattern defined subset of 7034 RefSeq genes.  Our results showed a fair degree of overall correlation between all 6 platforms evaluated; but, to varying degrees, two RNA-seq protocols outperformed three of the microarray platforms in most categories.  Notably, a fourth microarray platform, Agilent, was comparable, or marginally superior, to the RNA-seq protocols within these same assessments.  Furthermore, 3 platforms (Agilent and two RNA-seq methods) demonstrated over 80% concordance with the gold standard TaqMan assay in terms of fold-change accuracy.  Our study suggests that the use of transcript patterns can enhance a number of the observed cross-platform correlations, indicating a potential usefulness for similar evaluations. Overall design: The study assessed differences and similarities in performance at numerous aspects including signal range, sensitivity to fold-change, and fidelity with TaqMan qRT-PCR. There were three levels of data examined: entire data sets, data derived from gene name annotation oriented subset of 15442 RefSeq genes, and data derived from transcript pattern defined subset of 7034 RefSeq genes.","project":"SRP055917"}
{"number_samples":80,"species":"human","abstract":"Background: Gene expression variation is a phenotypic trait of particular interest as it represents the initial link between genotype and other phenotypes. Analyzing how such variation apportions among and within groups allows for the evaluation of how genetic and environmental factors influence such traits. It also provides opportunities to identify genes and pathways that may have been influenced by non-neutral processes. Here we use a population genetics framework and next generation sequencing to evaluate how gene expression variation is apportioned among four human groups in a natural biological tissue, the placenta. Results: We estimate that on average, 33.2%, 58.9% and 7.8% of the placental transcriptome is explained by variation within individuals, among individuals and among human groups, respectively.  Additionally, when technical and biological traits are included in models of gene expression they account for roughly 2% of total gene expression variation. Notably, the variation that is significantly different among groups is enriched in biological pathways associated with immune response, cell signaling and metabolism. Many biological traits demonstrated correlated changes in expression in numerous pathways of potential interest to clinicians and evolutionary biologists. Finally, we estimate that the majority of the human placental transcriptome (65% of expressed genes) exhibits expression profiles consistent with neutrality; the remainder are consistent with stabilizing selection (26%), directional selection (4.9%), or diversifying selection (4.8%). Conclusion: We apportion placental gene expression variation into individual, population and biological trait factors and identify how each influence the transcriptome. Additionally, we advance methods to associate expression profiles with different forms of selection. Overall design: Placental mRNA was sequenced on an Illumina GAIIx. Samples were derived from 4 human groups, 10 individuals per group, 2 samples per individual","project":"SRP055918"}
{"number_samples":12,"species":"human","abstract":"Pneumonia remains one of the leading causes of death in both adults and children worldwide. Although viral and fungal acute airway infections can result in pneumonia, bacteria are the most common cause of community-acquired pneumonia, with Streptococcus pneumoniae isolated in nearly 50% of cases. Pneumolysin is a cholesterol-dependent cytolysin or pore-forming toxin produced by Streptococcus pneumonia and has been shown to play a critical role in bacterial pathogenesis. Airway epithelium is the initial site of many bacterial contacts and its barrier and mucosal immunity functions are central to infectious lung diseases. We decided to assess changes in the transcriptome of human airway epithelial cells exposed to toxin, statin or both. Our current work provides the first global view in human airway epithelial cells of the transcriptome under statin exposure that results in cellular protection from pneumolysin.","project":"SRP055938"}
{"number_samples":5,"species":"human","abstract":"RNA-Seq of mRNA from human primary CD4+ T cells infected with the low passage isolate HIV89.6 and uninfected controls","project":"SRP055981"}
{"number_samples":1,"species":"human","abstract":"A cell supsension containing an equal mix of HEK and 3T3 cells was used in Drop-Seq, a novel technology for high-throughput single cell mRNAseq.  Cells were mixed at a final concentration of 12.5 cells per microliter in droplets. Overall design: An estimated 570 STAMPs were selected for amplification.","project":"SRP056000"}
{"number_samples":12,"species":"human","abstract":"Study goals for AlkB-facilitated RNA Methylation Sequencing (ARM-Seq) were to identify small RNAs containing certain nucleoside modifications that are known to interfere with reverse transcription required for RNA-seq library preparation. In order to facilitate sequencing RNA samples were treated using the Escherichia coli dealkylating enzyme AlkB to demethylate 1-methyladenosine (m1A), 3-methylcytidine (m3C) and 1-methylguanosine (m1G) residues prior to RNA-seq library preparation. These modifications are known to be common in transfer RNAs (tRNAs). Without treatment RNAs containing m1A, m3C or m1G modifications are expected to block progression of reverse transcriptase, resulting in failure of PCR amplification and sequencing, and under-representation in sequencing results. Modified RNAs revealed by ARM-Seq were identified by comparing sequencing results for AlkB-treated samples to those from untreated samples using software for differential expression analyses.","project":"SRP056032"}
{"number_samples":8,"species":"human","abstract":"Knockdown of mutant and/or wild-type SF3B1 in MEL202 cell line by Degron knock-in, followed by RNA-seq, to identify splicing events governed by mutant SF3B1. Overall design: Control: parental MEL202 cell line. Experiments: mutant-SF3B1 knockdown; wildtype-SF3B1 knockdown; mutant SF3B1 knockout. Treatments: each of these four conditions plus and minus shld.","project":"SRP056033"}
{"number_samples":12,"species":"human","abstract":"Epigenetic modifications determine the structure and regulation of eukaryotic genomes and define key signatures of cell lineage specification. Technologies that facilitate the targeted manipulation of epigenetic marks could be used to precisely control cell phenotype or interrogate the relationship between the epigenome and transcriptional control. Here we have generated a programmable acetyltransferase based on the CRISPR/Cas9 gene regulation system, consisting of the nuclease-null dCas9 protein fused to the catalytic core of the human acetyltransferase p300. This fusion protein catalyzes acetylation of histone H3 lysine 27 (H3K27) at its target sites, leading to robust transcriptional activation of target genes from promoters, proximal enhancers, and distal enhancers. In contrast to conventional dCas9-based activators, the acetyltransferase fusion effectively activated genes from enhancer regions and with individual guide RNAs. The core p300 domain was also portable to other programmable DNA-binding proteins. This technology enables the targeted perturbation of native epigenetic architecture and will be useful for reprogramming the epigenome for applications in genomics, genetics, disease modeling, and manipulating cell fate. Overall design: HEK293T cells were transfected in triplicate with plasmids expressing synthetic transcription factors. The synthetic TFs were either (a) dCas9-VP64 fusion protein and a targeting guide RNA (gRNA), or (b)dCas9-p300 fusion protein containing the catalytic domain of p300 and a targeting guide RNA (gRNA).  As a control, cells were transfected with plasmids expressing dCas9 alone and dCas9 fused with a aceryltransferase null mutatnt form of the p300 catalytic domain (D1399Y, as in text). After transfection, RNA-seq was used to identify differential expressin at on-target and off-target  sites.","project":"SRP056036"}
{"number_samples":20,"species":"human","abstract":"Decoding genome-wide effects of experimental tissue-tissue or cell-cell interactions is important, for example, to better understand tumor-stroma interactions after transplantation (xenografting). Transcriptome analysis of intermixed human and mouse cells has frequently relied on the need to separate the two cell populations prior to transcriptome analysis, which introduces confounding effects on gene expression. To circumvent this problem, we perform a bioinformatics-based genome-wide transcriptome analysis technique separating the mouse and human transcriptome part of a dataset, which allows a mixed mouse and human cell population to be sequenced without prior cell sorting. We use the new technology – which we call S3 (“S-cube” for Species-specific sequencing) technology – to provide new insights into the Notch downstream response following Notch ligand-stimulation and to explore transcriptional changes following transplantation of luminal (MCF7) and basal-type (MDA-MB-231) human breast cancer cells into mammary fat pad tissue in mice. Overall design: Analysis of Notch response to ligand in three settings: co-culture, immobilized ligand and xenografted tumors","project":"SRP056041"}
{"number_samples":16,"species":"human","abstract":"Myelin-reactive T cells have been identified in patients with multiple sclerosis (MS) and healthy subjects with comparable frequencies, but the contribution of these autoreactive T cells to disease pathology remains unknown. A total of 13,324 T cell libraries generated from blood of 23 patients and 22 healthy controls were interrogated for reactivity to myelin antigens. Libraries derived from CCR6+ myelin-reactive T cells from patients with MS exhibited significantly enhanced production of IFN-?, IL-17, and GM-CSF compared to healthy controls. Single-cell clones isolated by MHC/peptide tetramers from CCR6+ T cell libraries also secreted more pro-inflammatory cytokines while clones isolated from controls secreted more IL-10. The transcriptomes of myelin-specific CCR6+ T cells from patients with MS were distinct from those derived from healthy controls, and of note, were enriched in Th17-induced experimental autoimmune encephalitis (EAE) gene signatures and gene signatures derived from Th17 cells isolated other human autoimmune diseases. These data, although not casual, imply that functional differences between antigen specific T cells from MS and healthy controls is fundamental to disease development and support the notion that IL-10 production from myelin-reactive T cells may act to limit disease progression, or even pathogenesis. Overall design: Four conditions of purified T cells with between 3 and 5 replicates per condition","project":"SRP056049"}
{"number_samples":3,"species":"human","abstract":"Huntington’s disease (HD) is a devastating neurological disorder that is caused by an expansion of the poly-Q tract in exon 1 of the Huntingtin gene (HTT). HTT is an evolutionarily conserved and ubiquitously expressed protein that has been linked to a variety of functions including transcriptional regulation, mitochondrial function, and vesicle transport. This large protein has numerous caspase and calpain cleavage sites and can be decorated with several post-translational modifications such as phosphorylations, acetylations, sumoylations, and palmitoylations. However, the exact function of HTT and the role played by its modifications in the cell is still not well understood. Scrutiny of HTT function has been focused on a single, full length, mRNA.  In this study, we report the discovery of 5 novel HTT mRNA splice isoforms that are expressed in normal and HD-hESC lines as well as cortical neurons differentiated from hESCs. Interestingly, none of the novel isoforms generates a truncated protein. Instead, 4 of the 5 new isoforms specifically eliminate domains and modifications to generate smaller HTT proteins. The fifth novel isoform incorporates a previously unreported additional exon, dubbed 41b, which is hominid-specific and introduces a potential phosphorylation site in the protein. The discovery of this hominid-specific isoform may shed light on human-specific pathogenic mechanisms of HTT, which could not be investigated with current mouse models of the disease.  Furthermore, it provides a new human-specific target for drug screening in Huntington’s disease. Overall design: We performed RNAseq of human embryonic stem cells in pluripotency conditions to check expression of multiple HTT isoforms.","project":"SRP056066"}
{"number_samples":6,"species":"human","abstract":"Haematopoietic stem and progenitor cell (HSPC) transplant is a widely used treatment for life-threatening conditions including leukemia; however, the molecular mechanisms regulating HSPC engraftment of the recipient niche remain incompletely understood. Here, we developed a competitive HSPC transplant method in adult zebrafish, using in vivo imaging as a non-invasive readout. We used this system to conduct a chemical screen and identified epoxyeicosatrienoic acids (EET) as a family of lipids that enhance HSPC engraftment.  EETs’ pro-haematopoietic effects are conserved in the developing zebrafish, where this molecule promotes HSPC specification through activating a unique AP-1/runx1 transcription program autonomous to the haemogenic endothelium. This effect requires the activation of PI3K pathway, specifically PI3Kg. In adult HSPCs, EETs induce transcriptional programs including AP-1 activation, modulating multiple cellular processes, such as migration, to promote engraftment. Finally, we demonstrated that the EET effects on enhancing HSPC homing and engraftment are conserved in mammals. Our study established a novel method to explore the molecular mechanisms of HSPC engraftment, and discovered a previously unrecognized, evolutionarily conserved pathway regulating multiple haematopoietic generation and regeneration processes. EETs may have clinical application in marrow or cord blood transplantation. Overall design: To analyze the effect of 11,12-EET on gene expression of human blood cells, we treated human CD34+ cells (positively selected from cord blood) and the human leukemic cell line U937 with 5uM 11,12-EET for 2 hrs. Control treatment was done with DMSO.","project":"SRP056074"}
{"number_samples":5,"species":"human","abstract":"We report the differential roles of an HDAC inhibitor-TSA during hESC nerual commitment. In the initiation of hESC differentiation, TSA could inhibit the downregulation of pluripotency genes to maintain pluripotency, whereas in the neural commitment stage, TSA could promote neural gene expression to assist hESC nerual determination. Overall design: Examination of gene expression patterns in hESCs, day 4 or day 8 differentiated cells without or with TSA treatment","project":"SRP056076"}
{"number_samples":6,"species":"human","abstract":"We used RNA-seq to examine the effect of Myc activation on U2OS cells transcriptome. We also examined these effects in the presence of Torin-1, an inhibitor of mTOR Overall design: We measure gene expression profiles in U2OS cells containing an inducible Myc expression vector that were induced or mock-treated in duplicates for 36 hours. In addition, we repeated the experiments in the presence of Torin-1, an inhibitor of mTOR.","project":"SRP056084"}
{"number_samples":10,"species":"human","abstract":"Noncoding RNAs (ncRNAs) comprise an important class of natural regulators that mediate a vast array of biological processes, including the modulation of chromatin architecture. Moreover, artificial ncRNAs have revealed that the functional capabilities of RNA are extremely broad. To further investigate and harness these capabilities, we developed CRISPR-Display (\"CRISP-Disp\"), a targeted localization strategy that uses Cas9 to deploy large RNA cargos to specific DNA loci. We demonstrate that exogenous RNA domains can be functionally appended onto the CRISPR scaffold at multiple insertion points, allowing the construction of Cas9 complexes with RNAs nearing one kilobase in length, with structured RNAs, protein-binding cassettes, artificial aptamers and pools of random sequences. CRISP-Disp also allows the simultaneous multiplexing of disparate functions at multiple targets. We anticipate that this technology will provide a powerful method with which to ectopically localize functional RNAs and ribonuceloprotein complexes at specified genomic loci. Overall design: Whole cell poly(A) selected RNA seq, from HEK293FT cells bearing lentivirally-integrated Gaussia and Cypridina luciferase reporter loci.  Cells were transiently transfected with dCas9~VP64 alone, or with dCas9~VP and one of several modified sgRNAs,each targeting the Gaussia reporter.","project":"SRP056086"}
{"number_samples":8,"species":"human","abstract":"IFN-g primes macrophages for enhanced inflammatory activation by TLRs and microbial killing, but little is known about the regulation of cell metabolism or mRNA translation during priming. We found that IFN-g regulates macrophage metabolism and translation in an integrated manner by targeting mTORC1 and MNK pathways that converge on the selective regulator of translation initiation eIF4E. Physiological downregulation of the central metabolic regulator mTORC1 by IFN-g was associated with autophagy and translational suppression of repressors of inflammation such as HES1. Genome-wide ribosome profiling in TLR2-stimulated macrophages revealed that IFN-g selectively modulates the macrophage translatome to promote inflammation, further reprogram metabolic pathways, and modulate protein synthesis. These results add IFN-g-mediated metabolic reprogramming and translational regulation as key components of classical inflammatory macrophage activation. Overall design: RPF and RNAseq libraries were generated from mock or IFN-g-primed human macrophages. Cells were stimulated with Pam3Cys and harvested at 4 hours. Libraries were generated using protocol modified from Illumina Truseq technology.","project":"SRP056098"}
{"number_samples":12,"species":"human","abstract":"Pancreatic cancers (PCs) are highly metastatic with poor prognosis, mainly due to delayed detection. We hypothesized that intercellular communication is critical for metastatic progression. Here, we show that PC-derived exosomes induce liver pre-metastatic niche formation in naïve mice and consequently increase liver metastatic burden. Uptake of PC-derived exosomes by Kupffer cells caused transforming growth factor ß secretion and upregulation of fibronectin production by hepatic stellate cells. This fibrotic microenvironment enhanced recruitment of bone marrow-derived macrophages. We found that macrophage migration inhibitory factor (MIF) was highly expressed in PC-derived exosomes, and its blockade prevented liver pre-metastatic niche formation and metastasis. Compared to patients whose pancreatic tumors did not progress, MIF was markedly higher in exosomes from stage I PC patients who later developed liver metastasis. These findings suggest that exosomal MIF primes the liver for metastasis and may be a prognostic marker for the development of PC liver metastasis. Overall design: Normal pancreas and Pancreatic cancer exosomes education of human von Kupffer cells in vitro","project":"SRP056146"}
{"number_samples":2,"species":"human","abstract":"RNA sequencing of SETD2 isogenic renal cell carcinoma cell lines. Overall design: Examination of RNA expression in SETD2 isogenic cell lines","project":"SRP056153"}
{"number_samples":3,"species":"human","abstract":"We sequenced mRNA from 3 neutrophil cells taken from 3 male adult to generate the gene expression profile of human neutrophil cells Overall design: Examination of mRNA levels in human neutrophils.","project":"SRP056159"}
{"number_samples":81,"species":"human","abstract":"RNA sequencing of human leukemia Overall design: The goals of this project are to obtain a comprehensive study of mutations and gene expression in human acute myeloid leukemia (AML). Methods: AML cells were thawed. DNA and RNA (polyA) was extracted and sequences were obtained with an illumina HiSeq 2000 sequencer. Results are pending.","project":"SRP056197"}
{"number_samples":10,"species":"human","abstract":"We used Ribo-seq to examine the effect of Myc activation on protein translation in U2OS cells and correalted these changes with alterations in RNA level measured by RNA-seq on tye same conditions. We also examined these effects in the presence of Torin-1, an inhibitor of mTOR Overall design: We measure ribosome occupancy profiles in U2OS cells containing an inducible Myc expression vector that were induced or mock-treated in duplicates for 36 hours. In addition, we repeated the experiments in the presence of Torin-1, an inhibitor of mTOR.","project":"SRP056200"}
{"number_samples":4,"species":"human","abstract":"We used GRO-seq to examine the effect of Myc activation on RNA transcription in U2OS cells. Overall design: We measure in duplicates gene transcription rates in U2OS cells containing an inducible Myc expression vector that were induced or mock-treated in duplicates for 5 hours.","project":"SRP056201"}
{"number_samples":43,"species":"human","abstract":"We had previously discovered that the transcription factor OVO-like 1 (OVOL1) was highly induced during trophoblast differentiation. In this study, we used an lentiviral shRNA strategy to decrease OVOL1 expression in BeWo trophoblast cells. Control cells were transduced with shRNAs targeting no known mammalian transcript (shCont). Following stimulation of differentiation (48h exposure to 8-bromo-cyclic adenosine monophosphate), a RNA-seq approach was used to determine global transcript differences in OVOL1-knockdown cells compared to control cells. Overall design: Trophoblast cells transduced with control shRNAs were used as controls. Cells transduced with shRNAs targeting OVOL1 were used as treatment. All cells received 250 uM 8-bromo-cyclic adenosine monophosphate to stimulate differentiation. Three independent replicates of control and treatment groups were analyzed.","project":"SRP056220"}
{"number_samples":10,"species":"human","abstract":"Development of an Induced Pluripotent Stem Cell Derived Human Neuronal Model of Chemotherapeutic Neuropathy","project":"SRP056287"}
{"number_samples":24,"species":"human","abstract":"Gene expression pattern in follicular lymphomas","project":"SRP056293"}
{"number_samples":525,"species":"human","abstract":"RNA sequencing of human leukemia Overall design: The goals of this project are to obtain a comprehensive study of mutations and gene expression in human acute myeloid leukemia (AML). Methods: AML cells were thawed. DNA and RNA (polyA) was extracted and sequences were obtained with an illumina HiSeq 2000 sequencer. Results are pending.","project":"SRP056295"}
{"number_samples":3,"species":"human","abstract":"Promoter-proximal RNA polymerase II (Pol II) pausing is implicated in the regulation of gene transcription. However, the mechanisms of pausing including its dynamics during transcriptional responses remain to be fully understood. We performed global analysis of short capped RNAs and Pol II Chromatin Immunoprecipitation sequencing in MCF-7 breast cancer cells to map Pol II pausing across the genome, and used permanganate footprinting to specifically follow pausing during transcriptional activation of several genes involved in the Epithelial to Mesenchymal Transition (EMT). We find that the gene for EMT master regulator Snail (SNAI1), but not Slug (SNAI2), shows evidence of Pol II pausing before activation. Transcriptional activation of the paused SNAI1 gene is accompanied by a further increase in Pol II pausing signal whereas activation of non-paused SNAI2 gene results in the acquisition of a typical pausing signature. The increase in pausing signal reflects increased transcription initiation without changes in Pol II pausing. Activation of the heat shock HSP70 gene involves pausing release that speeds up Pol II turnover, but does not change pausing location. We suggest that Pol II pausing is retained during transcriptional activation and can further undergo regulated release in a signal-specific manner. Overall design: Untreated MCF-7 cells were analyzed for the distribution of Pol II using ChIP-sequencing with Anti-Pol II N-20 antibody (two independent biological replicates, A, B), and for the distribution of paused RNA polymerase II by sequencing of short capped RNAs (scRNAs) prepared from nuclei (three independent biological replicates, 1-3). All samples were sequenced on a MiSeq instrument in paired-end format","project":"SRP056296"}
{"number_samples":6,"species":"human","abstract":"The prostate gland mainly contains basal and luminal cells constructed as a pseudostratified epithelium. Annotation of prostate epithelial transcriptomes provides a foundation for discoveries that can impact disease understanding and treatment. Here, we describe a whole-genome transcriptome analysis of human benign prostatic basal and luminal populations by using deep RNA sequencing. Combined with comprehensive molecular and biological characterizations, we show that the differential gene expression profiles account for their distinct functional phenotypes. Strikingly, in contrast to luminal cells, basal cells preferentially express gene categories associated with stem cells, neural and neuronal development and RNA processing. Consistent with their expression profiles, basal cells functionally exhibit intrinsic stem-like and proneural properties with enhanced ribosome RNA (rRNA) transcription activity. Of clinical relevance, the treatment failed castration-resistant and anaplastic prostate cancers molecularly resemble a basal-like phenotype. Therefore, we link the cell-type specific gene signatures to subtypes of prostate cancer development, and identify genes associated with patient clinical outcome. Overall design: Human total RNA profiles of 3 pairs of benign prostatic basal and luminal populations freshly purified from prostate tissues of three prostate cancer patients by deep RNA-seq.","project":"SRP056330"}
{"number_samples":2,"species":"human","abstract":"In order to comprehensively identify RNA-expression changes after p53-activation, total RNA was isolated and subjected to next generation seqencing (RNA-Seq) after activation of a conditional p53 allele in SW480 cells. Overall design: SW480/pRTR-p53-VSV cells were subjected to RNA-Seq analysis after 48 hours doxycycline-treatment.","project":"SRP056369"}
{"number_samples":1,"species":"human","abstract":"This study assessed the transcriptional profile of SiHa cells. SiHa is a cervical cancer cell line with integrated HPV16, and was used as a model to study human gene expression in the context of integrated virus. Gene expression in SiHa, calculated by Cufflinks, was scored in windows around the locations of known viral integrations in patients or cell lines to determine if there was an association between gene expression and viral integration. We found that SiHa gene expression was higher near loci of integration for HPV18 vs. HPV16, cervical tissues vs. head and neck cancers, and cervical cancers vs. in vitro integrations. This study provides insight into the factors that may influence where viruses integrate in the human genome. Overall design: Gene Expression in untreated SiHa cells.","project":"SRP056378"}
{"number_samples":12,"species":"human","abstract":"Human embryonic stem cells (hESCs) have the unique property of immortality, ability to infinitely self-renew and survive in vitro. In contrast to tumor-deribed cells, their immortality are free from any genomic abberations. Instead, they depend on the AKAP-Lbc/Rho signaling cascade. To understand the downstream way, we performed RNA-seq analyses between normal and AKAP-Lbc-depleted hESCs using the doxycyclin-inducible gene silensing strategy. Overall design: We use the genetically modified hESCs in which AKAP-13-targeting shRNA is induced by doxycyclin(dox) treatment. To minimize cell loss during treatment, anti-apoptotic factor Bcl-XL is overexpressed. We collected RNA from dox-treated and untreated cells in biological triplicate. We measured gene expression in these 2 sample groups using RNA-seq (illumina HiSeq) .","project":"SRP056395"}
{"number_samples":4,"species":"human","abstract":"This experiment evaluated the transcriptome response of the SKBR3 HER2-positive breast cancer cell line to 48 hours treatment with 300nM lapatinib, 300nM JQ1, or the combination treatment, relative to a DMSO-only control.","project":"SRP056433"}
{"number_samples":4,"species":"human","abstract":"This experiment evaluated the transcriptome response of the SKBR3 HER2-positive breast cancer cell line to 48 hours treatment with 300nM lapatinib, 300nM JQ1, or the combination treatment, relative to a DMSO-only control.","project":"SRP056437"}
{"number_samples":4,"species":"human","abstract":"This experiment determined the transcriptome response of SKBR3 HER2-positive breast cancer cells made resistant to 300nM lapatinib, to long-term treatment (8 days) of 300nM JQ1 or 1uM I-BET151. Resistant cells were kept in the presence of lapatinib for these treatments.","project":"SRP056438"}
{"number_samples":24,"species":"human","abstract":"We report two clusters in the overall profiles of expression among the samples. At the parasitemia onset, there is a strong interferon response reflected in up-regulation of co-regulated transcripts, while unexpectedly we also see down-regulation of transcripts related to TLR signaling and innate immunity. RNASeq also suggested differential expression of reticulocytes and a subset of T cell function. No obvious difference in the transcriptomes of naïve and semi-immune volunteers was seen, however several hundred genes were up-regulated in naïve individuals. Overall design: RNA-seq analysis was performed for 12 individuals (6 each from Buenaventura and Cali) for two of the time points, namely the diagnosis day and baseline (pre-challenge day).","project":"SRP056443"}
{"number_samples":53,"species":"human","abstract":"Increasing evidence suggests that defective RNA processing contributes to the development of amyotrophic lateral sclerosis (ALS). This may be especially true for ALS caused by a repeat expansion in C9orf72 (c9ALS), in which the accumulation of RNA foci and dipeptide-repeat proteins are expected to modify RNA metabolism. We report extensive alternative splicing (AS) and alternative polyadenylation (APA) defects in the cerebellum of c9ALS cases (8,224 AS, 1,437 APA), including changes in ALS-associated genes (e.g. ATXN2 and FUS), and cases of sporadic ALS (sALS; 2,229 AS, 716 APA). Furthermore, hnRNPH and other RNA-binding proteins are predicted as potential regulators of cassette exon AS events for both c9ALS and sALS. Co-expression and gene-association network analyses of gene expression and AS data revealed divergent pathways associated with c9ALS and sALS. Overall design: Examination transcriptiome profiles in c9orf72-associated ALS, sporadic ALS and healthy control","project":"SRP056477"}
{"number_samples":9,"species":"human","abstract":"General activation of hypoxia-inducible factor (HIF) pathways is classically associated with adverse prognosis in cancer and has been proposed to contribute to oncogenic drive. In clear cell renal carcinoma (CCRC) HIF pathways are upregulated by inactivation of the von-Hippel-Lindau tumour suppressor. However HIF-1a and HIF-2a have contrasting effects on experimental tumour progression. To better understand this paradox we examined pan-genomic patterns of HIF DNA binding and associated gene expression in response to manipulation of HIF-1a and HIF-2a and related the findings to CCRC prognosis. Our findings reveal distinct pan-genomic organization of HIF isoform-specific DNA binding at thousands of sites. Overall associations were observed between HIF-1a-specific binding, and genes associated with favourable prognosis and between HIF-2a-specific binding and adverse prognosis.  However within each isoform-specific set, individual gene associations were heterogeneous in sign and magnitude, suggesting that activation of each HIF-a isoform contributes a highly complex mix of pro- and anti-tumorigenic effects Overall design: ChIP and RNASeq of HIF-1a and HIF-2a transfection in 786-O cell lines","project":"SRP056530"}
{"number_samples":8,"species":"human","abstract":"Our data provide a comprehensive list of transcriptomics alterations and warrant holistic approach including both coding and non-coding RNAs in functional studies aimed to understand the pathophysiology of LOAD Overall design: We performed directional RNA sequencing on high quality RNA samples extracted from hippocampi of 4 late onset Alzheimer's diseas (LOAD) and 4 age-matched controls.","project":"SRP056604"}
{"number_samples":20,"species":"human","abstract":"Gene expression profile in Calu-3 cells infected with MERS-CoV or SARS-CoV","project":"SRP056612"}
{"number_samples":6,"species":"human","abstract":"Somatic differentiation requires induction of lineage specific genes to fulfill specialized tissue functions yet the genomic control of this process is incompletely understood. Using the epidermis as a model, we show here that the BAF chromatin remodeling complex is essential to maintain a subset of open chromatin regions, which are strikingly enriched for the DNA binding motif of the stratified epithelial lineage-determining transcription factor p63 (p=1X10-1786). These p63 sites are intrinsically inaccessible. BAF is required to displace nucleosomes ~40 base pairs away from p63 motifs to enable p63 binding, to maintain active histone marks, and to recruit RNA polymerase II to promote gene expression. In this process, p63 was itself also essential for full BAF recruitment to p63 binding sites. Intrinsically, p63 binding sequences favor nucleosome occupancy. Our data therefore suggest a model where a lineage-specific transcription factor cooperates with the BAF complex to establish genome accessibility and engage tissue-specific gene expression. Overall design: We examined chromatin accessibility controlled by the cooperation between BAF and p63 using ATAC-seq and ChIP-seq. ATAC-seq was performed in differentiating primary human keratinocytes in control and Brg1/Brm knock down conditions, two replicates each. We further compared the ChIP-seq signal of H3K27me3, H3K4me1, PolII, H3K27Ac, and p300 in keratinocytes between Brg1/Brm loss versus control. To test whether p63 recruits BAF to its binding sites, we also performed BAF ChIP-seq in keratinocytes between p63 loss versus control. In addition, we performed RNA-seq in BAF loss, p63 loss, and control conditions.","project":"SRP056637"}
{"number_samples":12,"species":"human","abstract":"Gene expression analysis by RNA-Sequencing of SF268 glioblastoma cells. Total RNA was extracted from SF268 cells 48h after transfection with two individual siRNAs targeting YAP1 and unspecific control siRNAs.","project":"SRP056665"}
{"number_samples":55,"species":"human","abstract":"Purpose: To gain molecular insights of HBV integration that may contribute to HCC tumorigenesis, we performed whole transcriptome sequencing and whole genome copy number profiling of hepatocellular carcinoma (HCC) samples from 50 Chinese patients. Results: We identified a total of 33 HBV-human integration sites in 16 of 44 HBV-positive HCC tissues, which were enriched in HBV genotype C-infected patients. In addition, significantly recurrent HBV-MLL4 integration (18%; 8/44) was found in this cohort of patients. Conclusions: This is the first report on the molecular basis of the MLL4 integration driving MLL4 over-expression. HBV-MLL4 integration occurred frequently in Chinese HCC patients, representing a unique molecular segment for HCC with HBV infection. Overall design: mRNA profiles of 50 Chinese Hepatocellular Carcinoma patients and 5 adjacent tissues by Illumina Hiseq 2000.","project":"SRP056696"}
{"number_samples":156,"species":"human","abstract":"The innate immune system provides the first response to pathogen infection and orchestrates the activation of the adaptive immune system. Though a large component of the innate immune response is common to all infections, pathogen-specific innate immune responses have been documented as well. The innate immune response is thought to be especially critical for fighting infection with Mycobacterium tuberculosis (MTB), the causative agent of tuberculosis (TB). While TB can be a deadly disease, only 5-10% of individuals infected with MTB develop active disease, and this inter-individual variation is, at least partly, heritable. Studies of inter-individual variation in the innate immune response to MTB infection may therefore shed light on the genetic basis for variation in susceptibility to TB. Yet, to date, we still do not know which properties of the innate immune response are specific to MTB infection and which represent a general response to pathogen infection. To begin addressing this gap, we infected macrophages with eight different bacterial pathogens, including different MTB strains and related mycobacteria, and studied the transcriptional response to infection. We found that although the gene expression changes were largely consistent across the bacterial infection treatments, we were able to identify a novel subset of genes whose regulation was affected specifically by infection with mycobacteria. Genetic variants that are associated with regulatory differences in these genes should be considered candidate loci for explaining inter-individual susceptibility TB. Overall design: RNA-seq of monocyte-derived macrophages isolated from 6 healthy European males at 4, 18, and 48 hours post-infection with the following 8 bacteria:  Mycobacterium tuberculosis (MTB) H37Rv, Mycobacterium tuberculosis GC1237, MTB GC1237, bacillus Calmette-Guérin (BCG), Mycobacterium smegmatis, Yersinia pseudotuberculosis, Salmonella typhimurium, and Staphylococcus epidermidis. table-s1.txt is a tab-delimited text file that contains the batch-corrected log2 counts per million for each of the 156 samples, as well as the Ensembl gene ID and gene name. BCG = bacillus Calmette-Guérin GC =  Mycobacterium tuberculosis GC1237 Rv = Mycobacterium tuberculosis (MTB) H37Rv Rv+ = heat-inactivated MTB H37Rv Salm = Salmonella typhimurium Smeg = Mycobacterium smegmatis Staph = Staphylococcus epidermidis Yers = Yersinia pseudotuberculosis","project":"SRP056733"}
{"number_samples":10,"species":"human","abstract":"Myocyte enhancer factor 2B (MEF2B) is a transcription factor with somatic mutation hotspots at K4, Y69 and D83 in diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL). The recurrence of these mutations indicates that they may drive lymphoma development. However, inferring the mechanisms by which they may drive lymphoma development was complicated by our limited understanding of MEF2B’s normal functions. To expand our understanding of the cellular activities of wildtype (WT) and mutant MEF2B, I developed and addressed two hypotheses: (1) identifying genes regulated by WT MEF2B will allow identification of cellular phenotypes affected by MEF2B activity and (2) contrasting the DNA binding sites, effects on gene expression and effects on cellular phenotypes of mutant and WT MEF2B will help refine hypotheses about how MEF2B mutations may contribute to lymphoma development. To address these hypotheses, I first identified genome-wide WT MEF2B binding sites and transcriptome-wide gene expression changes mediated by WT MEF2B. Using these data I identified and validated novel MEF2B target genes.  I found that target genes of MEF2B included the cancer genes MYC, TGFB1, CARD11, NDRG1, RHOB, BCL2 and JUN.  Identification of target genes led to findings that WT MEF2B promotes expression of mesenchymal markers, promotes HEK293A cell migration, and inhibits DLBCL cell chemotaxis. I then investigated how K4E, Y69H and D83V mutations change MEF2B’s activity. I found that K4E, Y69H and D83V mutations decreased MEF2B DNA binding and decreased MEF2B’s capacity to promote gene expression in both HEK293A and DLBCL cells. These mutations also reduced MEF2B’s capacity to alter HEK293A and DLBCL cell movement. From these data, I hypothesize that MEF2B mutations may promote DLBCL and FL development by reducing expression of MEF2B target genes that would otherwise function to help confine germinal centre B-cells to germinal centres. Overall, my research demonstrates how observations from genome-scale data can be used to identify cellular effects of candidate driver mutations. Moreover, my work provides a unique resource for exploring the role of MEF2B in cell biology: I map for the first time the MEF2B ‘regulome’, demonstrating connections between a relatively understudied transcription factor and genes significant to oncogenesis. Overall design: RNA-seq was performed on cells expressing V5 tagged WT or mutant MEF2B and on empty vector control cells. One biological replicates was performed on cell treated with either ionomycin or a solvent-only control.","project":"SRP056742"}
{"number_samples":35,"species":"human","abstract":"Background:   Personalized medicine is predicated on the notion that individual biochemical and genomic profiles are relatively constant in times of good health and to some extent predictive of disease or therapeutic response.  We report a pilot study quantifying gene expression and methylation profile consistency over time, addressing the reasons for individual uniqueness, and its relation to N=1 phenotypes. Methods:  Whole blood samples from 4 African American women, 4 Caucasian women, and 4 Caucasian men drawn from the Atlanta Center for Health Discovery and Well Being study at three successive 6-month intervals were profiled by RNASeq, miRNASeq, and Illumina Methyl-450 arrays.  Standard regression approaches were used to evaluate the proportion of variance for each type of omic measure that is among individuals, and to quantify correlations among measures and with clinical attributes related to wellness. Results:  Longitudinal omic profiles are in general highly consistent over time, with an average of 67% of the variance in transcript abundance, 42% of CpG methylation level (but 88% for the most differentiated CpG per gene), and 50% of miRNA abundance among individuals, which are all comparable to 74% of the variance among individuals for 74 clinical traits.  One third of the variance can be attributed to differential blood cell type abundance, which is also fairly stable over time, and a lesser amount to eQTL effects, whereas seven conserved axes of covariance that capture diverse aspects of immune function explain over half of the variance.  These axes also explain a considerable proportion of individually extreme transcript abundance, namely approximately 100 genes that are significantly up- or down-regulated in each person and are in some cases enriched for relevant gene activities that plausibly associate with clinical attributes.  A similar fraction of genes have individually divergent methylation levels, but these do not overlap with the transcripts, and fewer than 20% of genes have significantly correlated methylation and gene expression. Conclusions:  People express an “omic personality” consisting of peripheral blood transcriptional and epigenetic profiles that are constant over the course of a year and reflect various types of immune activity.  Baseline genomic profiles can provide a window into the molecular basis of traits that might be useful for explaining medical conditions or guiding personalized health decisions. Overall design: Whole blood samples from 12 subjects drawn from the Atlanta Center for Health Discovery and Well Being study at three successive 6-month intervals were profiled by RNASeq, miRNASeq, and Illumina Methyl-450 arrays.","project":"SRP056784"}
{"number_samples":24,"species":"human","abstract":"Around 20-25% of childhood acute lymphoblastic leukemias carry the TEL-AML1 (TA) fusion gene. It is a fusion of two central hematopoietic transcription factors, TEL (ETV6) and AML1 (RUNX1). Despite its prevalence, the exact genomic targets of TA have remained elusive. We evaluated gene loci and enhancers targeted by TA genome-wide in precursor B acute leukemia cells  using global nuclear run-on sequencing (GRO-seq). Overall design: Nascent RNA expression profiles were generated with GRO-seq after TEL-AML1 expression in the Nalm6 pre-B-ALL cell line in four different time points (0, 4, 12 and 24 h). TEL-AML1-mut and luciferase induction cell lines were used as controls. Two replicates were included for all six samples.","project":"SRP056802"}
{"number_samples":24,"species":"human","abstract":"Understanding distinct gene expression patterns of normal adult and developing fetal human pancreatic a and b cells is crucial for developing stem cell therapies, islet regeneration strategies, and therapies designed to increase b cell function in patients with diabetes (type 1 or 2). Toward that end, we have developed methods to highly purify a, b, and d cells from human fetal and adult pancreata by intracellular staining for the cell-specific hormone content, sorting the sub-populations by flow cytometry and, using next generation RNA sequencing, we report on the detailed transcriptomes of fetal and adult a and b cells. We observed that human islet composition was not influenced by age, gender, or body mass index and transcripts for inflammatory gene products were noted in fetal b cells. In addition, within highly purified adult glucagon-expressing a cells, we observed surprisingly high insulin mRNA expression, but not insulin protein expression. This transcriptome analysis from highly purified islet a and b cell subsets from fetal and adult pancreata offers clear implications for strategies that seek to increase insulin expression in type 1 and type 2 diabetes. Overall design: RNA-sequencing of highly purified human adult and fetal islet cell subset was performed using our newly developed method. Using this data, we can study and compare the detailed transcriptome or alpha and beta cells during development.","project":"SRP056835"}
{"number_samples":133,"species":"human","abstract":"A systems biology approach was used to comprehensively examine the impact of renal disease and hemodialysis (HD) on host response during critical illness. We examined the metabolome, proteome, and transcriptome of 150 patients with critical illness, stratified by renal function. Plasma metabolite values showed greater changes as renal function declined, with the greatest derangements in patients receiving chronic HD. Specifically, 6 uremic retention molecules, 17 other protein catabolites, 7 modified nucleosides, and 7 pentose phosphate sugars increased as renal function declined, consistent with decreased excretion or increased catabolism of amino acids and ribonucleotides. Similarly, the proteome showed increased levels of low-molecular weight proteins and acute phase reactants. The transcriptome revealed a broad-based decrease in mRNA levels among HD patients. Systems integration revealed an unrecognized association between plasma RNASE1 and several RNA catabolites and modified nucleosides. Further, allantoin, N1-methyl-4-pyridone-3-carboxamide, and n-acetylaspartate showed inverse correlations with the majority of significantly down-regulated genes. In conclusion, renal function broadly affected the plasma metabolome, proteome, and peripheral blood transcriptome during critical illness. These changes were not effectively mitigated by hemodialysis. These studies suggest several novel mechanisms whereby renal dysfunction contributes to critical illness. Overall design: We sequenced peripheral blood RNA of 133 representative subjects with systemic inflammatory response syndrome that had Acute Kidney Injury (AKI) or Hemodialysis (HD). No injury (AKI0; n= 58); AKI Stage 1 (AKI1; n= 36); AKI stage 2 and 3 (AKI23; n= 17); HD (N=22).","project":"SRP056840"}
{"number_samples":15,"species":"human","abstract":"Sequencing samples from the brain bank of Emory University","project":"SRP056863"}
{"number_samples":3,"species":"human","abstract":"The hnRNP A1 and A2 proteins regulate processes such as alternative pre-mRNA splicing and mRNA stability. Here, we report that a reduction in the levels of hnRNP A1 and A2 by RNA interference or their cytoplasmic retention by osmotic stress drastically increases the transcription of a reporter gene. Based on previous work, we propose that this effect may be linked to a decrease in the activity of the transcription elongation factor P-TEFb. Consistent with this hypothesis, the transcription of the reporter gene was stimulated when the catalytic component of P-TEFb, CDK9, was inhibited with DRB. While low levels of A1/A2 stimulated the association of RNA polymerase II with the reporter gene, they also increased the association of CDK9 with the repressor 7SK RNA, and compromised the recovery of promoter-distal transcription on the Kitlg gene after the release of pausing. Transcriptome analysis revealed that more than 50% of the genes whose expression was affected by the siRNA-mediated depletion of A1/A2 were also affected by DRB. RNA polymerase II-chromatin immunoprecipitation assays on DRB-treated and A1/A2-depleted cells identified a common set of repressed genes displaying increased occupancy of polymerases at promoter-proximal locations, consistent with pausing. Overall, our results suggest that lowering the levels of hnRNP A1/A2 elicits defective transcription elongation on a fraction of P-TEFb-dependent genes, hence favoring the transcription of P-TEFb-independent genes. Overall design: two treatements and one control","project":"SRP056912"}
{"number_samples":23,"species":"human","abstract":"To define molecular mechanisms underlying rod and cone differentiation, we generated H9 human embryonic stem cell line carrying a GFP reporter that is controlled by the promoter of cone-rod homeobox (CRX) gene, the first known marker of post-mitotic photoreceptor precursors. CRXp-GFP reporter in H9 line replicates endogenous CRX expression when induced to form self-organizing 3-D retina-like tissue. We define temporal transcriptome dynamics of developing photoreceptors during the establishment of cone and rod cell fate. Our studies provide an essential framework for delineating molecules and cellular pathways that guide human photoreceptor development and should assist in chemical screening and cell-based therapies of retinal degeneration. Overall design: Undifferentiated CRXp-GFP HP hES cells and 3D-neural retina were collected at days 37, 47, 67 and 90 and dissociated into single cells. Cells were sorted at 4°C and by FACSAria (Becton Dickinson). GFP+ and GFP- cells were separately collected. Total RNA was extracted by RNA purification kit (Norgen Biotek) and analyzed by 2100 Bioanalyzer (Agilent Technologies Genomics). High quality of total RNA (RIN: 7.7-9.2) was subjected to libraries construction using 40-60 ng of total RNA as input. Libraries were constructed using a stranded modification of the Illumina TruSeq mRNA (Brooks, et al. Meth Mol Biol 2012). Each library was single-end sequenced in an independent lane of a GAIIx at a length of 76 bases.  Fastq files were generated from reads passing chastity filter.","project":"SRP056957"}
{"number_samples":20,"species":"human","abstract":"RNA sequencing of total RNA from 20 human tissues","project":"SRP056969"}
{"number_samples":1,"species":"human","abstract":"Knowledge of the expression profile and spatial landscape of the transcriptome in individual cells is essential for understanding the rich repertoire of cellular behaviors. Here we report multiplexed error-robust fluorescence in situ hybridization (MERFISH), a single-molecule imaging approach that allows the copy numbers and spatial localizations of thousands of RNA species to be determined in single cells. Using error-robust encoding schemes to combat single-molecule labeling and detection errors, we demonstrated the imaging of 100 – 1000 unique RNA species in hundreds of individual cells. Correlation analysis of the ~10^4 – 10^6 pairs of genes allowed us to constrain gene regulatory networks, predict novel functions for many unannotated genes, and identify distinct spatial distribution patterns of RNAs that correlate with properties of the encoded proteins. Overall design: A single sample is analyzed","project":"SRP056989"}
{"number_samples":15,"species":"human","abstract":"<p/>","project":"SRP057017"}
{"number_samples":24,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: Pilot Dual RNA-seq: Infection of HeLa-S3 cells with wild-type Salmonella; 2 time points (4 h, 24 h p.i.; each sorted into invaded host cells [GFP+] and non-infected bystanders [GFP-]) and the respective human (4 h mock, 24 h mock) or bacterial reference controls (0 h LB,  0 h input), respectively. Three biological replicates were taken.","project":"SRP057064"}
{"number_samples":51,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: High-resolution comparative Dual RNA-seq time-course","project":"SRP057065"}
{"number_samples":2,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: rRNA depletion establishment","project":"SRP057066"}
{"number_samples":20,"species":"human","abstract":"We discovered that two mitotic regulators, BuGZ and Bub3, involved in splicing regulation during interphase Overall design: 8 samples from primary Human foreskin fibroblast cells (HFFs) , 12 samples from TOV21G cells. Control siRNA. BuGZ siRNA or Bub3 siRNA were transfected for 48 h before sample collection. Cells treated with pladienolide B served as positive controls.  For each RNAi experiment, we had two biological replicates.","project":"SRP057074"}
{"number_samples":28,"species":"human","abstract":"Background: Psoriasis is a chronic disease characterized by the development of scaly red skin lesions and possible co-morbid conditions. The psoriasis lesional skin transcriptome has been extensively investigated, but mRNA levels do not necessarily reflect protein abundance.  Methods: Lesional (PP) and uninvolved (PN) skin samples from 14 patients were analyzed using high-throughput complementary DNA sequencing (RNA-seq) and liquid chromatography-tandem mass spectrometry (LC-MS/MS).  Results: We identified 4122 differentially expressed genes (DEGs) along with 748 differentially expressed proteins (DEPs). Global shifts in mRNA were modestly correlated with changes in protein abundance (r = 0.40). We identified similar numbers of increased and decreased DEGs, but 4-fold more increased than decreased DEPs. Ribosomal subunit and translation proteins were elevated within lesions, without a corresponding shift in mRNA expression (RPL3, RPS8, RPL11). We identified 209 differentially expressed genes/proteins (DEGPs) with corresponding trends at the transcriptome and proteomic levels. Most DEGPs were similarly altered in at least one other skin disease. Psoriasis-specific and non-specific DEGPs had distinct cytokine-response patterns, with only the former showing disproportionate induction by IL-17A in cultured keratinocytes.  Conclusions: Our findings reveal global imbalance between the number of increased and decreased proteins in psoriasis lesions, consistent with heightened translation. This effect could not have been discerned from mRNA profiling data alone. We have also identified high-confidence DEGPs and shown that only those most specific to psoriasis are enriched with IL-17A targets. Overall design: RNA-seq-based comparison between gene expression in psoriasis lesions and uninvolved skin from 14 patients","project":"SRP057087"}
{"number_samples":15,"species":"human","abstract":"Primordial germ cells (PGCs), the embryonic precursors of sperm and ova, undergo comprehensive epigenetic reprogramming, including DNA demethylation. The detailed dynamics of epigenome resetting in the human germline is largely unknown. Here, we studied transcriptome transitions and epigenetic reprogramming in developmental week 5-9 in vivo human hPGCs (hPGCs) and nascent hPGC-like cells hPGCLCs by RNA-sequencing (RNA-Seq) and whole-genome bisulfite sequencing (BS-Seq). We found that the transcriptional program of hPGCs is distinct from that in mice, with co-expression of somatic specifiers and naïve pluripotency genes. Base-resolution hPGC methylome analysis revealed progressive DNA demethylation to basal levels by week 7. Despite global hypomethylation, some loci associated with metabolic and neurological disorders are resistant to DNA demethylation, revealing potential for transgenerational epigenetic inheritance","project":"SRP057098"}
{"number_samples":6,"species":"human","abstract":"Analysis of alternative splicing in  heart (left ventricles) samples of 3 adult DM1 patients versus 3 adult controls Overall design: PolyA RNA from left ventricles (heart) of 3 controls and 3 DM1 patients were analysed by massive parrallel sequencing","project":"SRP057118"}
{"number_samples":9,"species":"human","abstract":"Purpose: Common genetic variation at chromosome 4q25 lead to the strongest locus associated with atrial fibrillation (AF), the most frequent arrhythmia.  The mechanism of association is currently unknown.  We recently have identified a novel noncoding RNA expressed in the left atria.  To determine the potential functional roles of the LNCRNA adjacent to PITX2 (PANCR) via knockdown in cardiomyocytes Methods and Results: H9 differentiated cardiomyocytes were treated with siRNA targeting PANCR and PITX2c and a scrambled control in triplicate.  RNA and small RNA extracted and sent for sequencing.  Approximately 50 million read fragments mapped to the transcriptome using STAR aligner to hg19 and fragments counted with htseq.  EdgeR was used to quantify the differences between treatment groups.   There were significant changes upon knocking down PITX2c or PANCR in the RNA sequencing with high concordance of effect sizes between the two treatments (r2 = 0.85).  Similar to the RNAseq analysis, this miRNAseq analysis shows that the effects of PANCR knockdown on miRNA expression may be largely mediated by through its effect on PITX2c expression. Conclusion: PANCR knockdown decreased PITX2c expression in H9 differentiated cardiomyocytes, and altered the transcriptome similar to PITX2c knockdown. Overall design: H9 derived cardiomyoctyes were treated with siRNA knockdown (scrambled control, PANCR, PITX2c) in triplicate and RNA and smallRNAs extracted for sequencing.","project":"SRP057147"}
{"number_samples":16,"species":"human","abstract":"Analysis of transcriptional differences between control and RA-treated cells during cardiac differentiation. The hypothesis tested in these samples is that addition of RA during differentiation towards atrial-like cardiomyocytes while control cells treated with DMSO result in ventricular-like cardiomyocytes. Overall design: NKX2.5 (eGFP/w)-hESCs were differentiated to cardiomyocytes with spin EB protocol, with the addition of RA or DMSO. Cells were sorted at day-31 based on GFP resulting in CTplus, CTminus, RAplus or RAminus goups. RNA was isolated from each of these fractions for sequencing.","project":"SRP057156"}
{"number_samples":466,"species":"human","abstract":"We used single cell RNA sequencing on 466 cells to capture the cellular complexity of the adult and fetal human brain at a whole transcriptome level. Healthy adult temporal lobe tissue was obtained from epileptic patients during temporal lobectomy for medically refractory seizures. We were able to classify individual cells into all of the major neuronal, glial, and vascular cell types in the brain. Overall design: Examination of cell types in healthy human brain samples.","project":"SRP057196"}
{"number_samples":56,"species":"human","abstract":"Reprogramming of somatic cells produces induced pluripotent stem cells (iPSCs) that are invaluable resources for biomedical research. Transcriptional and epigenetic changes have been investigated to facilitate our understanding of the reprogramming process. Here, we extended the previous transcriptome studies by performing RNA-seq on cells defined by a combination of multiple cellular surface markers. We found that transcriptome changes during early reprogramming occur independently from the opening of closed chromatin by OCT4, SOX2, KLF4 and MYC (OSKM). Furthermore, our data identify multiple spliced forms of genes uniquely expressed at each progressive stage of reprogramming. In particular, we found a pluripotency-specific spliced form of CCNE1 that significantly enhances reprogramming. In addition, single nucleotide polymorphism (SNP) expression analysis reveals that monoallelic gene expression is induced in the intermediate stages of reprogramming while biallelic expression is recovered upon completion of reprogramming. Our transcriptome data provide unique opportunities in understanding human iPSC reprogramming. Overall design: RNA samples from intermediates of hiPSC reprogramming were obtained. Gene expression of those cells were analyzed.","project":"SRP057205"}
{"number_samples":4,"species":"human","abstract":"MicroRNAs (miRNAs) are small regulatory RNAs processed from stem-loop regions of primary transcripts (pri-miRNAs), with the choice of stem-loops for initial processing largely determining what becomes a miRNA. To identify sequence and structural features influencing this choice, we determined cleavage efficiencies of >50,000 variants of three human pri-miRNAs, focusing on the regions intractable to previous high-throughput analyses. Our analyses revealed a mismatched motif in the basal stem region, a preference for maintaining or improving base-pairing throughout the remainder of the stem, and a narrow stem-length preference of 35±1 base pairs. Incorporating these features with previously identified features, including three primary-sequence motifs, yielded a unifying model defining mammalian pri-miRNAs, in which motifs help orient processing and increase efficiency, with the presence of more motifs compensating for structural defects. This model enables generation of artificial pri-miRNAs, designed de novo, without reference to any natural sequence, yet processed more efficiently than natural pri-miRNAs. Overall design: Three major experiments are included in the submitted data. 1) Pools of synthetic DNA containing barcodes and pri-miRNA variants were sequenced to infer the barcode-pri-miRNA linkages, and the pri-miRNA variants were assayed for their cleavage efficiency (\"selection\"), which can be quantified by comparing their representations in the input and after cleavage. 2) Artificial miRNAs produced in HEK293T cells were sequenced.  3) Transcriptomes of artificial miRNA-expressed cells were sequenced to quantify mRNA changes.","project":"SRP057213"}
{"number_samples":42,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: sRNA profiling in various cell types","project":"SRP057244"}
{"number_samples":12,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: Pulse-expression of pinT","project":"SRP057248"}
{"number_samples":8,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: Host response to pinT*","project":"SRP057249"}
{"number_samples":42,"species":"human","abstract":"Bacteria possess many small noncoding RNAs whose regulatory roles in pathogenesis are little understood due to a paucity of macroscopic phenotypes in standard virulence assays. Here, we use a novel Dual RNA-seq approach for a single-step simultaneous RNA profiling in both pathogen and host to reveal molecular phenotypes of sRNAs during infection with Salmonella Typhimurium. We identify a new PhoP/Q-activated small RNA which upon bacterial internalization acts to temporally control the expression of both, invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity is shown to adjust the human response to replicating Salmonella, and have a pervasive impact on host RNA expression both inside and outside protein-coding regions including infection-specific alterations of an array of long noncoding RNAs. Our study provides a paradigm for a comprehensive RNA-based analysis of intracellular bacterial pathogens without their physical purification from a host and a new discovery route for hidden functions of pathogen genes. Overall design: Dual RNA-seq of further sRNA mutants","project":"SRP057250"}
{"number_samples":12,"species":"human","abstract":"The goal of this study was to determine if fibroblasts from different origin (skin, colon, tumors) were keeping their characteristic while extracted and cultured ex vivo for several passages. HUVEC was used as a control, being cells from a different background. Surprisingly, fibroblasts from different origins are losing their independant characteristic to cluster in a similar way after 5-6 passages in culture in vitro, showing an activated status. Overall design: Fibroblasts were extracted from human skin, colon normal stroma and colon tumor stroma. HUVECs were extracted from human samples at the same time. All cells, each group from 3 different patients, were grown on plastic for 5 passages and mRNA was extracted to perform RNASeq analysis.","project":"SRP057251"}
{"number_samples":6,"species":"human","abstract":"Entry into and exit from mitosis is driven by precisely-timed changes in protein abundance, and involves transcriptional regulation and protein degradation. However, the role of translational regulation in modulating cellular protein content during mitosis remains poorly understood. Here, using ribosome profiling, we show that translational, rather than transcriptional regulation is the dominant mechanism for modulating protein synthesis at mitotic entry. The vast majority of regulated mRNAs are translationally repressed, which contrasts previous findings of selective mRNA translational activation at mitotic entry. One of the most pronounced translationally repressed genes in mitosis is Emi1, an inhibitor of the anaphase promoting complex (APC), which is degraded during mitosis. We show that Emi1 degradation is insufficient for full APC activation and that simultaneous translational repression is required. These results provide a genome-wide view of protein translation during mitosis and suggest that translational repression may be used to ensure complete protein inactivation Overall design: Ribosome profiling and mRNA-seq from 3 time points in the cell cycle","project":"SRP057253"}
{"number_samples":6,"species":"human","abstract":"Transiently transfected Huh-7.5 cells (hepatocellular carcinoma cell line)","project":"SRP057280"}
{"number_samples":3,"species":"human","abstract":"Whole transcriptome sequencing (RNA-seq) of KMS11, MM.1S, and RPMI8226 multiple myeloma cell lines","project":"SRP057322"}
{"number_samples":7,"species":"human","abstract":"Mammalian cells contain copious amounts of RNA including both coding and non-coding RNA (ncRNA). Generally the ncRNAs function to regulate gene expression at the transcriptional and post-transcriptional level. Among ncRNA, the long ncRNA and small ncRNA can affect histone modification, DNA methylation targeting and gene silencing. Here we show that endogenous DNA methyltransferase 1 (DNMT1) co-purifies with inhibitory ncRNAs. MicroRNAs (miRNAs) binds directly to DNMT1 with high affinity. The binding of miRNAs, such as miR-155, leads to inhibition of DNMT1 enzyme activity. Exogenous miR-155 in cells induces aberrant DNA methylation of the genome, resulting in hypomethylation of low to moderately methylated regions. And small shift of hypermethylation of previously hypomethylated region was also observed. Furthermore, hypomethylation led to activation of genes. Based on these observations, we propose that overexpression of specific miRNAs in human cancer may lead to aberrant DNA methylation and altered gene-expression.  Overall design: Examine of the DNA methylation and mRNA profile of HCT 116 cells transfected by random 23-mer or miR-155 RNA","project":"SRP057437"}
{"number_samples":4,"species":"human","abstract":"Chromosome arm deletions are frequently observed in human cancers. Such large-scale aberrations may trigger phenotypes distinct from those that arise by loss of single genes. Using TALEN-based genomic engineering, we generated cell models with targeted 8p loss-of-heterozygosity (LOH) that mimics an 8p deletion commonly found in breast cancer patients. 8p LOH triggered up-regulation of the mevalonate pathway. The alterations in lipid metabolism led to increased metastatic potential and drug resistance of 8p-deleted cells that are in agreement with poorer survival rates of breast cancer patients harbouting 8p LOH. Our findings also suggest novel therapeutic strategies for treatment of 8p LOH tumors. Overall design: RNASeq Analysis of two Wt 8p Cell clones vs two 8p LOH cell clones generated via TALEN Approach","project":"SRP057448"}
{"number_samples":290,"species":"human","abstract":"We report RNA-sequencing data of 283 blood platelet samples, including 228 tumor-educated platelet (TEP) samples collected from patients with six different malignant tumors (non-small cell lung cancer, colorectal cancer, pancreatic cancer, glioblastoma, breast cancer and hepatobiliary carcinomas). In addition, we report RNA-sequencing data of blood platelets isolated from 55 healthy individuals. This dataset highlights the ability of TEP RNA-based 'liquid biopsies' in patients with several types with cancer, including the ability for pan-cancer, multiclass cancer and companion diagnostics. Overall design: Blood platelets were isolated from whole blood in purple-cap BD Vacutainers containing EDTA anti-coagulant by standard centrifugation. Total RNA was extracted from the platelet pellet, subjected to cDNA synthesis and SMARTer amplification, fragmented by Covaris shearing, and prepared for sequencing using the Truseq Nano DNA Sample Preparation Kit. Subsequently, pooled sample libraries were sequenced on the Illumina Hiseq 2500 platform. All steps were quality-controlled using Bioanalyzer 2100 with RNA 6000 Picochip, DNA 7500 and DNA High Sensitivity chips measurements. For further downstream analyses, reads were quality-controlled using Trimmomatic, mapped to the human reference genome using STAR, and intron-spanning reads were summarized using HTseq. The processed data includes 285 samples (columns) and 57736 ensemble gene ids (rows). The supplementary data file (TEP_data_matrix.txt) contains the intron-spanning read counts, after data summarization by HTseq.","project":"SRP057500"}
{"number_samples":6,"species":"human","abstract":"Technical advances have enabled the collection of genome and transcriptome data sets with single-cell resolution. However, single-cell characterization of the epigenome has remained challenging. Furthermore, because cells must be physically separated prior to biochemical processing, conventional single-cell preparatory methods scale linearly. We applied combinatorial cellular indexing to measure chromatin accessibility in thousands of single cells per assay, circumventing the need for compartmentalization of individual cells. We report chromatin accessibility profiles from over 15,000 single cells and use these data to cluster cells on the basis of chromatin accessibility landscapes. We identify modules of coordinately regulated chromatin accessibility at the level of single cells both between and within cell types, with a scalable method that may accelerate progress toward a human cell atlas. Overall design: 3 replicates from GM12878 and HL-60 cell lines collected for differential gene expression analysis.","project":"SRP057508"}
{"number_samples":10,"species":"human","abstract":"We performed RNA-seq and ChIP-seq on clones of human cell lines carrying an inducible XIST transgene on 1p, 8p, or 12q to study the effects of allelic silencing in cis Overall design: Total gene expression and allelic changes were examined in HT1080 clones carrying an inducible XIST transgene on 1p, 8p, or 12q after induction by doxycycline. A replicate was done for the 8p clone treated with DOX. An additional 1p clone integrated with an empty vector, and an 1p, 8p, and 12q clone without induction were included as controls. ChIP was performed on the 8p clone to investigate the changes in H3K27 acetylation and trimethylation.","project":"SRP057512"}
{"number_samples":2,"species":"human","abstract":"Cohesin ensures correct chromosome segregation by holding sister chromatids together from the S phase until their separation in anaphase. Mutations in cohesin and regulatory cohesin genes are associated with Cornelia de Lange syndrome (CdLS), a rare human developmental disorder. Since CdLS cell lines show no clear defects in sister chromatid cohesion, the molecular basis underlying CdLS remains elusive. Interestingly, in recent years data have been accumulated showing that cohesin is also involved in gene expression regulation. To gain insight into the effect of cohesin mutations on transcriptional mechanisms, we analyzed the recruitment of RNA Polymerase II (Pol II) and phosphorylated Pol II at the regulatory regions of active genes by a genome-wide approach","project":"SRP057545"}
{"number_samples":16,"species":"human","abstract":"Objective: Amniotic fluid (AF) is a proximal fluid to the fetus containing higher amounts of cell-free fetal RNA/DNA than maternal serum, thereby making it a promising source for novel biomarker discovery of fetal development and maturation. Our aim was to compare AF transcriptomic profiles at different time points in pregnancy to demonstrate unique genetic signatures that would serve as potential biomarkers indicative of fetal maturation. Methods: We isolated AF RNA from 16 women at different time points in pregnancy: 4 from 18-24 weeks, 6 from 34-36 weeks, and 6 from at 39-40 weeks. RNA-sequencing was performed on cell-free RNA. Gene expression and splicing analyses were performed in conjunction with cell-type and pathway inference.  Results: Sample-level analysis at different time points in pregnancy yielded a strong correlation with cell types found in the intrauterine environment and fetal respiratory, digestive and external barrier tissues of the fetus, using high-confidence cellular molecular markers. While some genes and splice variants were present throughout pregnancy, an abundant number of transcripts were uniquely expressed at different time points in pregnancy and associated with distinct fetal co-morbidities (respiratory distress and gavage feeding), indicating fetal immaturity. Conclusions: The AF transcriptome exhibits unique cell- and organ-selective expression patterns at different time points in pregnancy that can potentially identify fetal organ maturity and predict neonatal morbidity. Developing novel biomarkers indicative of the maturation of multiple organ systems can improve upon our current methods of fetal maturity testing which focus solely on the lung, and better inform obstetrical decisions regarding delivery timing. Overall design: RNA-Seq from cell-free was used to idenitfy mRNA transcripts indicative of overall fetal maturity.","project":"SRP057586"}
{"number_samples":21,"species":"human","abstract":"Purpose: To determine global gene expression changes following siRNA knockdown of Myc, Kcnk1, and Snta1 compared to non-targeting siRNAs or mock-transfected cells Methods: Total RNA was processed using the Illumina TruSeq Stranded mRNA Sample Preparation Kit according to manufacturer’s protocol. Generated cDNA libraries were sequenced using an Illumina HiSeq 2000 sequencer with four biological replicates sequenced per condition using single read, 50 cycle runs. Quality of sequencing reads were assessed using FastQC (Babraham Bioinformatics) and then aligned to a reference genome (hg19, UCSC Genome Browser) using TopHat. Sequencing yielded, on average, 23.7 million unique reads per sample with a 60.7 - 65.7% mapping rate. Cufflinks was used to generate transcript abundance for each annotated protein-coding gene as Fragments Per Kilobase of transcript per Million mapped reads (FPKM), and statistical analysis and comparison of FPKM values was calculated using R (Bioconductor). Results: Fold changes comparing mock and a non-targeting siRNA were highly congruent. Myc RNAi induced numerous changes, with 955 downregulated genes and 1214 upregulated genes. The effect on Myc itself was relatively modest, possibly reflecting its ability to negatively auto-regulate its own expression. Gene ontology analysis highlighted ribosome biogenesis, metabolism, gene expression, cell cycle, and apoptosis pathways, consistent with known Myc functions. The Kcnk1 siRNA affected 424 genes, with KCNK1 itself one of the most repressed. While gene ontology analysis also highlighted metabolism and biosynthesis pathways, the p-values and fold enrichment scores were substantially lower, indicating that DiM can be suppressed without major effects on metabolism and biosynthesis pathways. The Snta1 siRNA deregulated 575 genes, with SNTA1 itself the most repressed gene. Cell cycle and mitosis-related gene ontology terms feature heavily, consistent with this siRNA accelerating mitotic exit. Interestingly, FoxM1, which drives G2/M gene expression was reduced 1.75-fold, indicating that this siRNA may disrupt mitotic controls by deregulating FoxM1. Conclusions: Global gene expression profiling identifies Egr1 as regulator of mitotic cell fate. Overall design: Total RNA was extracted from at least four replicates of siRNA-transfected cells and libraries for sequencing was prepared for each replicate.","project":"SRP057613"}
{"number_samples":6,"species":"human","abstract":"Here we report a novel role for H2A.Z.2 (H2AFV) as a mediator of cell proliferation and sensitivity to targeted therapies in malignant melanoma. While both H2A.Z.1 and H2A.Z.2 are highly expressed in metastatic melanoma and correlate with decreased patient survival, only H2A.Z.2 deficiency results in impaired cellular proliferation of melanoma cells, which occurs via a G1/S arrest. Integrated gene expression and ChIP-seq analyses revealed that H2A.Z.2 positively regulates E2F target genes, and that such genes acquire a distinct H2A.Z occupancy signature over the promoter and gene body in metastatic melanoma cells. We further identified the BET family member BRD2 as an H2A.Z-interacting protein in melanoma cells, and demonstrate that H2A.Z.2 silencing cooperates with BET inhibition to induce cell death. Overall design: Expression levels for non tumorigenic (Melanocytes) and human melanoma cell line SKmel147, before and after JQ1 treatement","project":"SRP057616"}
{"number_samples":2,"species":"human","abstract":"In this study, combining 1) RNA-Sequencing on MCF-7 cell line, 2) a new computational approach to identify splice isoforms and 3) an extensive experimental validation (in vitro - on MCF-10, MCF-7 and MDA-MB-231 – and ex vivo – on human breast cancer tissues) we investigated simultaneously gene expression and alternative splicing in genes encoding for adhesion and motility-related molecules-, including Semaphorins and their receptors and co-receptors. Overall design: Transcriptome profiles of MCF7 cell line were generated by deep sequencing, in duplicate, using Hi-Seq Illumina","project":"SRP057627"}
{"number_samples":10,"species":"human","abstract":"About 50% of human malignancies exhibit unregulated signalling through the Ras-ERK1/2 (ERK) pathway, as a consequence of activating mutations in members of Ras and Raf families. However, the quest for alternative Ras-ERK pathway-directed therapies is desirable. Upon phosphorylation ERK dimerize. We had previously demonstrated that dimerization is essential for ERK extranuclear but not nuclear signaling. Furthermore, by molecular biology approaches, we showed that specifically inhibiting ERK extranuclear component, by impeding ERK dimerization, is sufficient for curtailing tumor progression. Here, we have identified a small molecule inhibitor for ERK dimerization in vitro and in vivo that, without affecting ERK phosphorylation, prevents tumorigenesis driven by Ras-ERK pathway oncogenes, both in cellular and animal models. Importantly, this compound is unaffected by resistance-acquisition processes that hamper “classical” Ras-ERK pathway inhibitors. Thus, ERK dimerization inhibitors provide the proof of principle for two novel concepts in cancer therapy: 1) The blockade of sublocalization-specific sub-signals, rather than total signals, as a means of effectively counteracting oncogenic Ras-ERK signaling. 2) Targeting regulatory protein-protein interactions such as dimerization, rather than catalytic activities, within a signaling route, as an approach for producing effective anti-tumoral agents. Strategies aimed at preventing aberrant flux through this route remain an attractive option for therapeutic intervention in cancer. In this respect, drugs inhibiting the kinase activities of BRaf and MEK have yielded promising results. Overall design: A375p cells treated with10 µM of either DEL22379, SCH772984 or DMSO as a control for two hours. mRNA from A375p cells was extrated using RNeasy mini kit (Qiagen, Germany) according to the manufacturer's instructions. Cells were previously treated with10 µM of either DEL22379, SCH772984 or DMSO as a control for two hours.","project":"SRP057629"}
{"number_samples":8,"species":"human","abstract":"The goal of this study is to generate RNA-seq data sets for assembling the structures of primary microRNA transcripts.","project":"SRP057660"}
{"number_samples":16,"species":"human","abstract":"We report the effects of 1,25(OH)2D3 treatment on the mRNA expression in human muscle cells Overall design: Primary cultures of human muscle cells were treated with 1,25(OH)2D3 or vehicle for 48 hours.","project":"SRP057721"}
{"number_samples":13,"species":"human","abstract":"Synthetic DNA-binding proteins have found broad application in gene therapies and as tools for interrogating biology. Engineered proteins based on the CRISPR/Cas9 and TALE systems have been used to alter genomic DNA sequences, control transcription of endogenous genes, and modify epigenetic states. Although the activity of these proteins at their intended genomic target sites have been assessed, the genome-wide effects of their action have not been extensively characterized.  Additionally, the role of chromatin structure in determining the binding of CRISPR/Cas9 and TALE proteins to their target sites and the regulation of nearby genes is poorly understood.  Characterization of the activity these proteins using modern high-throughput genomic methods would provide valuable insight into the specificity and off-target effects of CRISPR- and TALE-based genome engineering tools. We have analyzed the genome-wide effects of TALE- and CRISPR-based transcriptional activators targeted to the promoters of two different endogenous human genes in HEK293T cells using a variety of high-throughput DNA sequencing methods. In particular, we assayed the DNA-binding specificity of these proteins and their effects on the epigenome. DNA-binding specificity was evaluated by ChIP-seq and RNA-seq was used to measure the specificity of these activators in perturbing the transcriptome. Additionally, DNase-seq was used to identify the chromatin state at target sites of the synthetic transcriptional activators and the genome-wide chromatin remodeling that occurs as a result of their action. Our results show that these genome engineering technologies are highly specific in both binding to their promoter target sites and inducing expression of downstream genes when multiple activators bind to a single promoter. Moreover, we show that these synthetic activators are able to induce the expression of silent genes in heterochromatic regions of the genome by opening regions of closed chromatin and decreasing DNA methylation. Interestingly, the transcriptional activation domain was not necessary for DNA-binding or chromatin remodeling in these regions, but was critical to inducing gene expression. This study shows that these CRISPR- and TALE-based transcriptional activators are exceptionally specific. Although we detected limited binding of off-target sites in the genome and changes to genome structure, these off-target event did not lead to any detectable changes in gene regulation.  Collectively, these results underscore the potential for these technologies to make precise changes to gene expression for gene and cell therapies or fundamental studies of gene function. Overall design: HEK293T cells were transfected in triplicate with plasmids expressing synthetic transcription factors. The synthetic TFs were either (a) dCas9-VP64 fusion protein and a targeting guide RNA (gRNA), or (b) a TALE-VP64 fusion protein engineered to bind to a specific target site in the genome. As a control, cells were transfected with plasmids expressing GFP. After transfection, RNA-seq was used to identify both on-target and off-target binding sites for the synthetic TFs. The data in this submission were generated using the TALE transfection experiments.","project":"SRP057745"}
{"number_samples":192,"species":"human","abstract":"Exploring effect of progesterone/progestin treatment on gene expression Overall design: Two cell lines, three conditions (Full Media with E2, E2+ Progesterone, Full Media + R5020 Progestin)","project":"SRP057758"}
{"number_samples":10,"species":"human","abstract":"Purpose: Oxygen (O2) levels in cell culture conditions is typically 2-5 fold higher than the physiological O2 levels that most tissues experience in vivo. The ambient atmospheric O2 (21%) is known to induce cell proliferation defects and cellular senescence in stem cell and primary cell cultures. Therefore, culturing these cells under lower O2 levels (2-9%) is currently a standard practice. However, the non-cancerous immortalized cells and cancer cells, which evade cellular senescence are normally cultured under 21% O2 levels and the effects of higher O2 levels on these cells are not fully understood. Methods: Gene expression (RNA seq transcriptomics) analysis of immortalized human bronchial epithelial (BEAS-2B) cells cultured at ambient 21% O2 and lower 10% O2 levels for 3 days and 3 weeks. Further the beneficial effects of cuturing cells under lower oxygen tension is evalulated Results: Our results show NF-?B/RelA mediated activation of pro-inflammatory cytokines as a major outcome of cells being cultured 21% O2. Moreover, we demonstrate increased RelA binding at the NF-?B1/RelA target gene promoters at 21% O2. Interestingly, contrary to cells cultutred at 21% O2, external stress induced by H2O2 exposure did not induce inflammatory response in cells grown at 10% O2, suggesting increased ability to handle external stress in cells cultured at lower O2 levels. Overall design: RNA Seq gene expression comparision done in replicates","project":"SRP057766"}
{"number_samples":18,"species":"human","abstract":"We investigated the acetylation of H3K27, in normal condition and after stimulation of EGF and before or after the depletion of INTS11. Additionally, we examined transcription by sequencing the chromatin-bound fraction of RNA and by fractionating total RNA into polyadenylated and non-polyadenylated compartement. Overall design: Comparison of the genome wide profiles of H3K27ac and RNA in HeLa cells","project":"SRP057804"}
{"number_samples":8,"species":"human","abstract":"We showed that Bmal1 mainly regulates clock gene initiation, with this functioning being highly conserved across species. Overall design: compare RNA-seq and chip-seq between wildtype and mutant cells","project":"SRP057814"}
{"number_samples":2,"species":"human","abstract":"RNAseq data from control and TDP-43 knockdown in HeLa cells","project":"SRP057819"}
{"number_samples":5,"species":"human","abstract":"Microvesicles (MVs) are extracellular vesicles released by most cells and play critical roles in the progression and relapse of cancers. Our previous work demonstrated that MVs derived from K562 chronic myeloid leukemia cell lines could transform mononuclear cells (MNCs) from normal hematopoietic transplants to acute leukemia-like cancer cells. During the transformation, a new group of leukemia-like cells could be observed after 14 days consecutive incubation with MVs and most of them were transformed to leukemia-like cells after 21 days. To study the gene expression change and the mechanism of the process of K562-MVs transforming MNCs, we performed RNA-seq and small RNA-seq for 5 samples of the key time points, which were samples of K562-MVs, MNCs, 1 week/2 weeks/3 weeks cells after MVs incubation. The K562 cell line was purchased from the China Center for Type Culture Collection (Wuhan, China). MNCs were extracted from the peripheral blood mobilization provided by healthy volunteers. Sequencing of 5 samples were performed by WuXi AppTec Co Shanghai, China.","project":"SRP057826"}
{"number_samples":32,"species":"human","abstract":"We performed gene expression analysis using the SCRB-Seq method from cervical and peripheral blood antigen presenting cells that were collected from women with different cervicovaginal bacterial community types. Overall design: Cervical and peripheral blood antigen presenting cells were FACS sorted using the gating strategy: FSC vs. SSC  -> Singlets -> Live CD45+ -> HLA-DR+ -> CD11c+ or CD14+. The genital bacterial microbiomes were characterized in the same women and categorized into Lactobacillus dominance, Gardnerella dominance, or Prevotella dominance.","project":"SRP057852"}
{"number_samples":14,"species":"human","abstract":"In glioblastoma multiforme (GBM), brain tumor initiating cells (BTICs) with cancer stem cell characteristics have been identified and proposed as primordial cells responsible for disease initiation, recurrence and therapeutic resistance. However, the extent to which individual, patient-derived BTIC lines reflect the heterogeneity of GBM remains poorly understood. Here, we applied a stem cell biology approach and compared self-renewal, marker expression, label retention and asymmetric cell division in 20 BTIC lines. Through cluster analysis, we identified two subgroups of BTIC lines with distinct precursor states, stem- or progenitor-like, predictive of survival after xenograft. Moreover, stem and progenitor transcriptome signatures were identified, which showed a strong association with the proneural and mesenchymal subtypes, respectively, in the TCGA cohort. This study proposes a new framework for the study and use of BTIC lines and provides novel precursor biology insights into GBM.","project":"SRP057855"}
{"number_samples":2,"species":"human","abstract":"Existing methods for detecting RNA intermediates resulting from exonuclease degradation are low-throughput and laborious. In addition, mapping the 3’ ends of RNA molecules to the genome after high-throughput sequencing is challenging, particularly if the 3’ ends contain posttranscriptional modifications. To address these problems, we developed EnD-Seq, a high-throughput sequencing protocol that preserves the 3’ end of RNA molecules, and AppEnD, a computational method for analyzing high-throughput sequencing data. Together these allow determination of the 3’ ends of RNA molecules, including nontemplated additions. Applying EnD-Seq and AppEnD to histone mRNAs revealed that a significant fraction of cytoplasmic histone mRNAs end in one or two uridines, which have replaced the 1-2 nts at the 3’ end of mature histone mRNA maintaining the length of the histone transcripts. Histone genes in fly embryos and ovaries show the same pattern, but with different tail nucleotide compositions. We increase the sensitivity of EnD-Seq by using cDNA priming to specifically enrich low-abundance tails of known sequence composition allowing identification of degradation intermediates. In addition, we show the broad applicability of our computational approach by using AppEnD to gain insight into 3’ additions from diverse types of sequencing data, including data from small capped RNA sequencing and some alternative polyadenylation protocols. Overall design: Libraries were prepared essentially as described in Slevin et al. (Mol. Cell  53: 1020-1030, 2014)","project":"SRP057878"}
{"number_samples":12,"species":"human","abstract":"CFBE cells with flip-in constructs to introduce WT, F508del, and G551D CFTR products underwent RNA-sequencing in hopes of elucidating their expression profiles.","project":"SRP057964"}
{"number_samples":4,"species":"human","abstract":"The targeting of oncogenic ‘driver’ kinases with small molecule inhibitors has proven to be a highly effective therapeutic strategy in selected non-small cell lung cancer (NSCLC) patients. However, acquired resistance to targeted therapies invariably arises and is a major limitation to patient care. ROS1 fusion proteins are a recently described class of oncogenic driver, and NSCLC patients that express these fusions generally respond well to ROS1-targeted therapy. In this study, we sought to determine mechanisms of acquired resistance to ROS1 inhibition. To accomplish this, we generated a ROS1 inhibition-resistant derivative of the initially sensitive NSCLC cell line HCC78.","project":"SRP058026"}
{"number_samples":19,"species":"human","abstract":"Gene expression is the most fundamental level at which the genotype leads to the phenotype of the organism. Enabled by ultra-high-throughput next-generation DNA sequencing, RNA-Seq involves shotgun sequencing of fragmented RNA transcripts by next-generation sequencing followed by in silico assembly, and is rapidly becoming the most popular method for gene expression analysis. Poly[A]+ RNA-Seq analyses of normal human adult tissue samples such as Illumina s Human BodyMap 2.0 Project and the RNA-Seq atlas have provided a useful global resource and framework for comparisons with diseased tissues such as cancer. However, these analyses have failed to provide information on poly[A]- RNA, which is abundant in our cells. The most recent advances in RNA-Seq analyses, use ribosomal RNA-depletion to provide information on both poly[A]  and poly[A]- RNA. In this paper, we describe the use of Illumina s HiSeq 2000 to generate high quality rRNA-depleted RNA- Seq sequence datasets from human fetal and adult tissues. The datasets reported here will be useful in demonstrating the different expression profiles in different tissues.","project":"SRP058036"}
{"number_samples":1,"species":"human","abstract":"Mammalian interphase chromosomes interact with the nuclear lamina (NL) through hundreds of large Lamina Associated Domains (LADs). We report a method to map NL contacts genome-wide in single human cells. Analysis of ~400 maps reveals a core architecture of gene-poor LADs that contact the NL with high cell-to-cell consistency, interspersed by LADs with more variable NL interactions. The variable contacts are more sensitive to a change in genome ploidy than the consistent contacts. Single-cell maps indicate that NL contacts involve multivalent interactions over hundreds of kilobases. Moreover, we observe extensive intra-chromosomal coordination of NL contacts, even over tens of megabases. Such coordinated loci exhibit preferential interactions as detected by Hi-C. Finally, single-cell gene expression and chromatin accessibility analysis shows that loci with consistent NL contacts are expressed at lower levels and are more consistently inaccessible than loci with lower contact frequencies. These results highlight fundamental principles of single cell chromatin organization. Overall design: In this dataset, single-cell mRNA sequencing results from 96 single KBM7 cells have been deposited","project":"SRP058046"}
{"number_samples":4,"species":"human","abstract":"RNA interference (RNAi) is an evolutionarily conserved phenomenon of post-transcriptional gene silencing mediated by small interfering RNAs (siRNAs) generated from short hairpin RNAs (shRNAs) by Dicer cleavage. Here, we report that siRNA precursors can be divided into two categories with different processing mechanisms and silencing activities that are dependent on stem loop length. We designed an alternative siRNA precursor for triggering RNAi named single-stranded Argonaute2 (Ago2)-processed interfering RNA (saiRNA). saiRNA is composed of a 16-18 bp stem and a loop complementary to the target transcript. The introduction of a self-cleavage ribozyme derived from hepatitis delta virus (HDV) to the 3’ end of the saiRNA dramatically improved its silencing activity. Unlike classical shRNA, the strand-specific cleavage of saiRNA by Ago2 during its processing not only eliminated the passenger strand but also avoided the association of mature siRNA with non-nucleolytic Ago proteins, thereby further reducing the risk of off-target effects. Additionally, saiRNA exhibited less competition with the biogenesis of endogenous miRNAs. Therefore, HDV ribozyme-enhanced saiRNA provides a reliable tool for RNAi applications. Overall design: A total of 400 ng plasmids of control, shRNA, shRNA-LC, saiRNA-RZ were transiently transfected into HEK293 cells in a 6-well plate for 48 hr. For RNA-seq, total cellular RNA was extracted to construct a cDNA library using the NEBNext Ultra Directional RNA Library Prep Kit (NEB) according to the manufacturer’s protocol. The cDNA libraries were sequenced using Illumina Hi-seq 2000 with 2 × 100 running circles. For small-RNA seq, total cellular RNA was extracted to construct a small RNA library according to the Illumina protocol,  and cDNA libraries was sequencing by Hi-seq 2000 with 50 running circles.","project":"SRP058087"}
{"number_samples":4,"species":"human","abstract":"This study was aimed at understanding the genome-wide binding and regulatory role of the DAXX transcriptional repressor, recently implicated in PCa. ChIP-Seq analysis of genome-wide distribution of DAXX in PC3 cells revealed over 59,000 DAXX binding sites, found at regulatory enhancers and promoters. ChIP-Seq analysis of DNA methyltransferase 1 (DNMT1), which is a key epigenetic partner for DAXX repression, revealed that DNMT1 binding was restricted to a small number of DAXX sites. DNMT1 and DAXX bound close to transcriptional activator motifs. DNMT1 sites were found to be dependent on DAXX for recruitment by analyzing DNMT1 ChIP-Seq following DAXX knockdown (K/D), corroborating previous findings that DAXX recruits DNMT1 to repress its target genes. Massively parallel RNA sequencing (RNA-Seq) was used to compare the transcriptomes of WT and DAXX K/D PC3 cells. Genes induced by DAXX K/D included those involved in autophagy, and DAXX ChIP-Seq peaks were found close to the transcription start sites (TSS) of autophagy genes, implying they are more likely to be regulated by DAXX. Overall design: Determine changes in gene expression levels between WT and DAXX K/D prostate cancer cells by RNA-Seq (PC3 Cells).","project":"SRP058098"}
{"number_samples":12,"species":"human","abstract":"RNA sequencing of tumor transcriptomes","project":"SRP058107"}
{"number_samples":30,"species":"human","abstract":"Alternative Cleavage and Polyadenylation (APA) plays a crucial role in the regulation of gene expression across eukaryotes. Although APA is extensively studied, its regulation within cellular compartments and its physiological impact remains largely enigmatic. Here, we employed a rigorous subcellular fractionation approach to compare APA profiles of cytoplasmic and nuclear RNA fractions from human cell lines. This comparison allowed us to extract APA isoforms that are subjected to differential regulation and provided us with a platform to interrogate the molecular regulatory pathways that shape the APA profiles in the subcellular locations. Subsequently, we depleted these cells of Dicer to compromise the miRNA biogenesis pathway to isolate the differentially regulated APA events that were regulated by miRNA. Overall design: Two biological replicates of Dicer-depleted and non-depleted HEK293 subcellular fractions were analysed using 3'region extraction and deep sequencing to quantitatively measure the usage of each cleavage and polyadenylation site transcriptome-wide. In addition, 3'READS was performed on the subcellular RNA fractions of HBL melanocyte cells to see if the pattern of differential APA regulation holds between cell lines. Furthermore, standard RNA-seq was performed using the Ion Proton system on HEK293 subcellular RNA fractions. Please note that the raw data from two replicates of Dicer-depleted and non-depleted HEK293 subcellular fractions samples were merged to generate processed data, which is linked to the first replicate sample record (i.e. *rep1.1). For example, the processed data files, C0d.p.bw and C0d.m.bw, were generated from both 3'READs cyto_rep1.1 and 3'READs cyto_rep1.2 sample data and linked to the  '3'READs cyto_rep1.1' sample records as supplementary data file.","project":"SRP058120"}
{"number_samples":73,"species":"human","abstract":"Parkinson disease (PD) is a neurodegenerative disease characterized by the accumulation of alpha-synuclein (SNCA) and other proteins in aggregates termed “Lewy Bodies” within neurons. PD has both genetic and environmental risk factors, and while processes leading to aberrant protein aggregation are unknown, past work points to abnormal levels of SNCA and other proteins. Although several genome-wide studies have been performed for PD, these have focused on DNA sequence variants by genome-wide association studies (GWAS) and on RNA levels (microarray transcriptomics), while genome-wide proteomics analysis has been lacking. After appropriate filters, proteomics identified 3,558 unique proteins and 283 of these (7.9%) were significantly different between PD and controls (q-value<0.05). RNA-sequencing identified 17,580 protein-coding genes and 1,095 of these (6.2%) were significantly different (FDR p-value<0.05), but only 166 of the FDR significant protein-coding genes (0.94%) were present among the 3,558 proteins characterized. Of these 166, eight genes (4.8%) were significant in both studies, with the same direction of effect. Functional enrichment analysis of the proteomics results strongly supports mitochondrial-related pathways, while comparable analysis of the RNA-sequencing results implicates protein folding pathways and metallothioneins. Ten of the implicated genes or proteins co-localized to GWAS loci. Evidence implicating SNCA was stronger in proteomics than in RNA-sequencing analyses. Notably, differentially expressed protein-coding genes were more likely to not be characterized in the proteomics analysis, which lessens the ability to compare across platforms. Combining multiple genome-wide platforms offers novel insights into the pathological processes responsible for this disease by identifying pathways implicated across methodologies. Overall design: The study consists of mRNA-Seq (29 PD, 44 neurologically normal controls) and three-stage Mass Spectrometry Tandem Mass Tag Proteomics (12 PD, 12 neurologically normal controls) performed in post-mortem BA9 brain tissue. The proteomics samples are a subset of the RNA-Seq samples.","project":"SRP058181"}
{"number_samples":12,"species":"human","abstract":"The RNA-binding protein RC3H1 (also known as ROQUIN) promotes TNFalpha mRNA decay via a 3'UTR constitutive decay element (CDE). Here, we applied PAR-CLIP to human RC3H1 to identify about 3800 mRNA targets with more than 16000 binding sites. A large number of sites are distinct from the consensus CDE and revealed a structure-sequence motif with U-rich sequences embedded in hairpins. RC3H1 binds preferentially short-lived and DNA damage induced mRNAs, indicating a role of this RNA-binding protein in the posttranscriptional regulation of the DNA damage response. Intriguingly, RC3H1 affects expression of NF-kB pathway regulators such as IkBalpha and A20. RC3H1 uses roquin and Zn-finger domains to contact a binding site in the A20 3'UTR, demonstrating a not yet recognized mode of RC3H1 binding. Knockdown of RC3H1 resulted in increased A20 protein expression, thereby interfering with IkB kinase and NF-kB activities, demonstrating that RC3H1 can modulate the activity of the IKK/NF-kB pathway. Overall design: We measured global mRNA decay rates in mock and RC3H1/RC3H2-depleted HEK293 cells. Transcription was blocked by Actinomycin D zero, one or two hours before harvesting. Total RNA was isolated in two biological replicates and subjected to polyA selection followed by high-throughput sequencing.","project":"SRP058191"}
{"number_samples":12,"species":"human","abstract":"46BR.1G1 cell line is impaired in DNA ligase 1 (LIG1) activity resulting in an increased level of endogenous single (SSBs) and double stranded DNA breaks (DSBs). 46BR.1G1 fibroblastoid cells represent a suitable model system to investigate how cells cope with low levels of chronic DNA damage, a condition frequently encountered in tumors. Transcriptional alterations in 46BR.1G1 cells were determined by RNAseq by comparison with 7A3, a cell line in which the defect was rescued by stable expression of ectopic wild-type Lig1. The identification of genes differentially expressed in 46BR.1G1 cells would contribute to the elucidation of DNA damage response (DDR) mechanisms.","project":"SRP058222"}
{"number_samples":17,"species":"human","abstract":"Lung cancer is the leading cause of cancer related mortality worldwide, with non-small cell lung cancer (NSCLC) as the most prevalent form.  Despite advances in treatment options including minimally invasive surgery, CT-guided radiation, novel chemotherapeutic regimens, and targeted therapeutics, prognosis remains dismal. Therefore, further molecular analysis of NSCLC is necessary to identify novel molecular targets that impact prognosis and the design of new-targeted therapies. In recent years, tumor “activated/reprogrammed” stromal cells that promote carcinogenesis have emerged as potential therapeutic targets. However, the contribution of stromal cells to NSCLC is poorly understood. Here, we show increased numbers of bone marrow (BM)-derived hematopoietic cells in the tumor parenchyma of NSCLC patients compared with matched adjacent non-neoplastic lung tissue. By sorting specific cellular fractions from lung cancer patients, we compared the transcriptomes of intratumoral myeloid compartments within the tumor bed with their counterparts within adjacent non-neoplastic tissue from NSCLC patients. The RNA sequencing of specific myeloid compartments (immature monocytic myeloid cells and polymorphonuclear neutrophils) identified differentially regulated genes and mRNA isoforms, which were inconspicuous in whole tumor analysis. Genes encoding secreted factors, including osteopontin (OPN), chemokine (C-C motif) ligand 7 (CCL7) and thrombospondin 1 (TSP1) were identified, which enhanced tumorigenic properties of lung cancer cells indicative of their potential as targets for therapy. This study demonstrates that analysis of homogeneous stromal populations isolated directly from fresh clinical specimens can detect important stromal genes of therapeutic value. Overall design: We sorted pure populations of the immature monocytic myeloid cells (IMMCs), neutrophils (Neu), and epithelial cells (Epi) from tumors and adjacent lung tissues of stage I-III lung adenocarcinoma patients. RNA samples (totally 17 samples) were sequenced: from tumor IMMC (n=3), Neu (n=2), Epi (n=2); from adjacent lung IMMC (n=3), Neu (n=4), Epi (n=3).","project":"SRP058237"}
{"number_samples":6,"species":"human","abstract":"The conserved RNA-binding protein Musashi1 (MSI1) has emerged as a key oncogenic factor in numerous solid tumors, including glioblastoma. However, its mechanism of action has not yet been established comprehensively. We set out to map its impact on the transcriptome in U251 cells using RNA-seq and iCLIP. Overall design: Examination of gene expression and splicing changes upon KD of Musashi1 in U251 cells and link to iCLIP-identified Musashi1 RNA binding sites","project":"SRP058244"}
{"number_samples":3,"species":"human","abstract":"We used RNA-seq and RRBS sequence to characterize the transcriptomic and methylome changes in chemoresistant and sensitive breast cancer cells Overall design: Analysis of a transcriptome and methylome breast cancer cells","project":"SRP058260"}
{"number_samples":4,"species":"human","abstract":"To further our understanding of the RNAi machinery within the human nucleus, we analyzed the chromatin and RNA binding of Argonaute 2 (AGO2) within human cancer cell lines. Our data indicated that AGO2 binds directly to nascent tRNA and 5S rRNA, and to the genomic loci from which these RNAs are transcribed, in a small RNA- and DICER-independent manner. AGO2 chromatin binding was not observed at non-TFIIIC-dependent RNA polymerase (Pol) III genes or at extra-TFIIIC (ETC) sites, indicating that the interaction is specific for TFIIIC-dependent Pol III genes. A genome-wide analysis indicated that loss of AGO2 caused a global increase in the mRNA expression level among genes that flank AGO2-bound tRNA genes. This effect was shown to be distinct from that of the disruption of DICER, DROSHA, or CTCF. We propose that AGO2 binding to tRNA genes has a novel and important regulatory role in human cells. Overall design: ChIP-seq for AGO2 was performed from K562 cells using 2 commercially available monoclonal antibodies (mAbs) and 5 replicates.  Replicates 1-3 were performed with Millipore 04-642, and replicates 4 and 5 were performed with Abcam 57113.  RNA-seq was carried out on 2 replicates of shMock and 2 replicates of shAGO2.  Lentiviral vectors GIPZ Lentiviral #RHS4531-EG271611.","project":"SRP058300"}
{"number_samples":12,"species":"human","abstract":"Facioscapulohumeral Muscular Dystrophy (FSHD) is caused by chromatin relaxation that results in aberrant expression of the transcription factor Double Homeobox 4 (DUX4). DUX4 protein is present in a small subset of FSHD muscle cells, making its detection and analysis of its effects historically difficult. Using a DUX4-activated reporter we demonstrate the burst expression pattern of endogenous DUX4, its method of signal amplification in the unique shared cytoplasm of the myotube, and FSHD cell death that depends on its activation. Transcriptome analysis of DUX4 expressing cells revealed that DUX4 activation disrupts RNA metabolism including RNA splicing, surveillance, and transport pathways. Cell signaling, polarity, and migration pathways were also disrupted. Thus, DUX4 expression is sufficient for myocyte death and these findings suggest mechanistic links between DUX4 expression and cell migration, supporting recent descriptions of phenotypic similarities between FSHD and an FSHD-like condition caused by FAT1 mutations.","project":"SRP058319"}
{"number_samples":18,"species":"human","abstract":"For some neurological disorders, disease is primarily RNA-mediated due to expression of non-coding microsatellite expansion RNAs (RNAexp). Toxicity is thought to result from enhanced binding of proteins to these expansions and depletion from their normal cellular targets. However, experimental evidence for this sequestration model is lacking. Here, we use HITS-CLIP and pre-mRNA processing analysis of human control versus myotonic dystrophy (DM) brains to provide compelling evidence for this RNA toxicity model. MBNL2 binds directly to DM repeat expansions in the brain resulting in depletion from its normal RNA targets with downstream effects on alternative splicing and polyadenylation. Similar RNA processing defects were detected in Mbnl compound knockout mice, highlighted by dysregulation of Mapt splicing and fetal tau isoform expression in adults. These results demonstrate that MBNL proteins are directly sequestered by RNAexp in the DM brain and introduce a powerful experimental tool to evaluate RNA-mediated toxicity in other expansion diseases. Overall design: HITS-CLIP analysis was performed to identify RNA binding sites of MBNL2 in control, DM type 1 (DM1), and DM type 2 (DM2) autopsy-derived brain (n=3).  Two regions of the brain selected for the study included the frontal cortex and hippocampus. Libraries were sequenced and wiggle files were generated for each biological replicate as well as three pooled biological replicates (3BRs) per group (control, DM1, and DM2).  In addition, differential CLIP analysis (dCLIP) was performed to normalize binding data between groups and identify statistically significant changes in binding.  The dCLIP analysis generated bedgraph files representing normalized binding profiles of MBNL2 in each group (control, DM1, and DM2) for visualization and comparative analysis. PolyA-seq was performed on control, DM1, and DM2 autopsy-derived brain samples (hippocampus and frontal cortex, n=3) as well as wild-type (WT) and Mbnl1; Mbnl2 conditional double knockout (Mbnl1-/-; Mbnl2c/c; Nestin-Cre+/- or DKO) brain (n=3).  Libraries were sequenced and the resultant files were processed and aligned to the reference genomes (hg19 and mm10).  Further computational processing was performed to remove internal oligo(dT) mis-priming events, identify valid polyA sites, and trim to the exact polyA sites.  BedGraph files were generated for each group in human (control, DM1, and DM2) and mouse (WT and Mbnl DKO) for comparative visualization.","project":"SRP058351"}
{"number_samples":8,"species":"human","abstract":"Stephen Paget first proposed, in 1889, that organ distribution of metastases is a non-random event, yet metastatic organotropism remains one of the greatest mysteries in cancer biology. Here, we demonstrate that exosomes released by lung-, liver- and brain-tropic tumor cells fuse preferentially with resident cells at their predicted destination, such as fibroblasts and epithelial cells in the lung, Kupffer cells in the liver, and endothelial cells in the brain. We found that exosome homing to organ-specific cell types prepares the pre-metastatic niche and that treatment with exosomes derived from lung tropic models can redirect metastasis to the lung. Proteomic profiling of exosomes revealed distinct integrin expression patterns associated with each organ-specific metastasis. Whereas exosomal integrins a6ß4 and a6ß1 were associated with lung metastasis, exosomal integrins avß5 and avß3 were linked with liver and brain metastases, respectively. Targeting a6ß4 and avß5 integrins decreased exosome uptake and metastasis in the lung and liver, respectively. Importantly, we demonstrate that exosome uptake activates a cell-specific subset of S100 family genes, known to support cell migration and niche formation. Finally, our clinical data indicate that integrin-expression profiles in circulating plasma exosomes from cancer patients could be used to predict organ-specific metastasis. Overall design: Education of human von Kupffer cells in vitro with human pancreatic cancer exosomes","project":"SRP058375"}
{"number_samples":7,"species":"human","abstract":"Acute myeloid leukemia (AML) is characterized by a marked genetic heterogeneity, which complicates the development of novel therapeutics. The delineation of pathways essential within the patient-individual mutational background might overcome this limitation and facilitate personalized treatment. We report the results of a large-scale lentiviral loss-offunction RNA-interference-(RNAi)-screen in primary leukemic cells. Stringent validation identified six genes (BNIPL1, ROCK1, RPS13, STK3, SNX27, WDHD1) whose knockdown impaired growth and viability of the cells. Dependence on these genes was not caused by mutation or overexpression and while some of the candidates seemed to be rather patientspecific, others were essential in cells isolated from other AML patients. In addition to the phenotype observed after ROCK1 knockdown, treatment with the approved ROCK-inhibitor fasudil resulted in increased apoptosis and decreased viability of primary AML cells. In contrast to observations in some other malignancies, ROCK1-inhibition did not foster growth of immature malignant progenitors; but was also toxic to this cell fraction in feeder-co-culture and xenotransplantion experiments, indicating a distinct effect of ROCK1 inhibition on leukemic progenitors. We conclude that large scale RNAi screens in primary patient-derived cells are feasible and can complement other methods for personalized cancer therapies, such as expression and mutation profiling. Overall design: Large-scale lentiviral loss-of-function shRNA screen in primary leukemic cells with two time points, exome sequencing of primary leukemic cells, and a gene expression profiling of primary cells from four leukemic and three healthy samples.","project":"SRP058383"}
{"number_samples":8,"species":"human","abstract":"Mapping of RNA:DNA hybrids in human cells reveals a number of characteristics of these non-canonical  nucleic  acid  structures.    A  directional  sequencing  approach  reveals  the  RNA component of the RNA:DNA hybrid to be purine-rich, indicating a thermodynamic contribution to the stability of these structures.  The RNA:DNA hybrids are enriched at loci with decreased DNA methylation  and  increased  DNase  hypersensitivity,  and  within  larger  domains  with characteristics of heterochromatin formation.  Studies of chromatin at RNA:DNA hybrids shows the  presence  of  the  ILF2  and  ILF3  transcription  factors,  supporting  a  model  of  certain transcription factors binding preferentially to the RNA:DNA conformation.  Overall, there is little to indicate a dependence for RNA:DNA hybrids forming co-transcriptionally, with results from the ribosomal DNA repeat unit instead supporting a model of RNA generating these structures in trans.  The results of the study indicate heterogeneous functions of these genomic elements and new insights into their formation and stability in vivo. Overall design: Investigation of expression data genome-wide in HEK293T and IMR-90 cells through RNA-seq, including rRNA","project":"SRP058387"}
{"number_samples":27,"species":"human","abstract":"RNA-seq was performed to analyze whole genome transcriptome in pre-B cell acute lymphoblastic leukemia","project":"SRP058414"}
{"number_samples":2,"species":"human","abstract":"Mutations such as gene fusion, translocation and focal amplification are a frequent cause of proto-oncogene activation during tumorigenesis, but such mutations do not explain all cases of proto-oncogene activation. Here we show that disruption of local chromosome conformation can also activate proto-oncogenes in human cells. We mapped chromosome structures in T-cell acute lymphoblastic leukemia (T-ALL), and found that active oncogenes and silent proto-oncogenes generally occur within insulated neighborhoods formed by the looping of two interacting CTCF sites co-occupied by cohesin. Recurrent microdeletions frequently overlap neighborhood boundary sites in T-ALL genomes, and we demonstrate that site-specific perturbation of loop boundaries is sufficient to activate the respective proto-oncogenes in non-malignant cells. We found somatic genomic rearrangements affecting loop boundaries in many cancers. These results suggest that chromosome structural organization is fundamental to identify functional somatic alterations in cancer genomes. Overall design: Paired-end 80x80 Poly-A RNA-seq in Jurkat T-ALL","project":"SRP058435"}
{"number_samples":4,"species":"human","abstract":"Aqueous humor from patients undergoing intraocular surgery is screened for miRNA in order to detect possible variations in miRNA-expressions associated with different ocular diseases.","project":"SRP058476"}
{"number_samples":6,"species":"human","abstract":"The AF4/FMR2 proteins AFF1 and AFF4 act as a scaffold to assemble the Super Elongation Complex (SEC) that strongly activates transcriptional elongation of HIV-1 and cellular genes. Although they can dimerize, it is unclear whether the dimers exist and function within a SEC in vivo. Furthermore, it is unknown whether AFF1 and AFF4 function similarly in mediating SEC-dependent activation of diverse genes. Providing answers to these questions, our current study shows that AFF1 and AFF4 reside in separate SECs that display largely distinct gene target specificities. While the AFF1-SEC is more potent in supporting HIV-1 transactivation by the viral Tat protein, the AFF4-SEC is more important for HSP70 induction upon heat shock. The functional difference between AFF1 and AFF4 in Tat-transactivation has been traced to a single amino acid variation between the two proteins, which causes them to enhance the affinity of Tat for P-TEFb, a key SEC component, with different efficiency. Finally, genome-wide analysis confirms that the genes regulated by AFF1- and AFF4-SEC are largely non-overlapping and perform distinct functions. Thus, the SEC represents a family of related complexes that exist to increase the regulatory diversity and gene control options during transactivation of diverse cellular and viral genes. Overall design: RNA-seq in HeLa cells of wild-type and after RNAi of AFF1 or AFF4.","project":"SRP058479"}
{"number_samples":6,"species":"human","abstract":"We compared Sox9-association at chondrocyte targets to a broad catalogue of regulatory indicators of chromatin organization and transcriptional activity to determine Sox9’s direct regulatory actions in normal developing chondrocytes. Sox9-associated regions resolve into two distinct regulatory categories. Class I regions closely associate with transcriptional start sites (TSSs). Their targets reflect general regulators of basal cell activities that Sox9 engages indirectly through a likely association with the basal transcriptional complex. In contrast, Class II regions outside of the local TSS domains highlight evolutionarily conserved, active enhancers directing expression of chondrocyte specific target genes, though DNA binding of Sox9-dimers at target sites with sub-optimal binding affinity. The level of associated chondrocyte gene expression correlates with the number of enhancer modules around the target gene and grouping into super-enhancer clusters. Comparison of Sox9 programs between neural crest and mesoderm-derived chondrocytes points to similar modes of chondrocyte specification in distinct chondrocyte lineages. These data provide the first insight into mammalian Sox family actions at the genome scale in the vivo setting. The resulting enhancer sets provide a key resource for further dissection of the regulatory programs of mammalian chondrogenesis. Overall design: Incorportation of ChIP-seq data of Sox9 and histone modification marks for chromatin status together with micorarray gene expression profiling in neonatal mice chondrocytes to uncover Sox9 regulatory system. Overexpression of Sox9 with a control of EGFP in human fibroblasts to identify the direct targets of Sox9 regulatory system","project":"SRP058567"}
{"number_samples":10,"species":"human","abstract":"Basal-like and luminal breast tumors have distinct clinical behavior and molecular profiles, yet the underlying mechanisms are poorly defined. To interrogate processes that determine these distinct phenotypes and their inheritance pattern, we generated somatic cell fusions and performed integrated genetic and epigenetic (DNA methylation and chromatin) profiling. We found that the basal-like trait is generally dominant and it is largely defined by epigenetic repression of luminal transcription factors. Definition of super-enhancers highlighted a core program common in luminal cells but high degree of heterogeneity in basal-like breast cancers that correlates with clinical outcome. We also found that protein extracts of basal-like cells is sufficient to induce luminal-to-basal phenotypic switch implying a trigger of basal-like autoregulatory circuits. We determined that KDM6A might be required for luminal-basal fusions, and identified EN1, TBX18, and TCF4 as candidate transcriptional regulators of luminal-to-basal switch. Our findings highlight the remarkable epigenetic plasticity of breast cancer cells. Overall design: RNA-Seq in breast cancer cell-lines","project":"SRP058571"}
{"number_samples":6,"species":"human","abstract":"This study identified genomwide KCl inducible readthrough transcription. The project also includes a Cap-Seq experiment to identify transcriptional start sites, demonstrating that KCl does not activate downstream transcriptional start sites, but indeed does induce readthrough","project":"SRP058633"}
{"number_samples":4,"species":"human","abstract":"Spinal Muscular Atrophy (SMA) is well-known to be caused by mutations in the gene Survival of Motor Neuron 1 (SMN1). Because this gene is ubiquitously expressed, it remains poorly understood why motor neurons (MNs) are one of the most affected cell types. To begin to address this question, we carried out RNA-sequencing studies using fixed, antibody-labeled and purified MNs produced from control and SMA patient-derived induced pluripotent stem cells (iPSCs). We found SMA-specific changes in MNs, including hyper-activation of the endoplasmic reticulum (ER) stress pathway and enhanced apoptosis. Functional studies demonstrated that inhibition of ER stress improves overall MN health and survival in vitro even in MNs with low SMN levels. In SMA mice, we show that systemic delivery of an ER stress inhibitor that crosses the blood-brain-barrier led to preservation of MNs in the spinal cord and prolonged survival of these mice. Therefore, our study implies that selective activation of ER stress underlies MN death in SMA. Moreover, the approach we have taken would be broadly applicable to studying disease-prone human cells in heterogeneous cultures. Overall design: total RNA collected from ES cell and patient ES cell derived spinal motor neurons","project":"SRP058640"}
{"number_samples":8,"species":"human","abstract":"Neuroblastoma cell lines can differentiate upon retinoic acid (RA) treatment, a finding that provided the basis for the clinical use of RA to treat neuroblastoma. However, resistance to RA is often observed, which limits its clinical utility. Using a gain-of-function genetic screen we identify the transcriptional coactivator Mastermind-like 3 (MAML3) as a gene whose ectopic expression confers resistance to RA. We find that MAML3 expression leads to loss of activation of a subset of RA target genes, which hampers RA-induced differentiation. The regulatory DNA elements of this subset of RA target genes show overlap in binding of MAML3 and the retinoic acid receptor, suggesting a role for MAML3 in the regulation of these genes. In addition, MAML3 has RA independent functions, including the activation of IGF1R and downstream AKT signaling via upregulation of IGF2, resulting in increased proliferation. Our results indicate an important role for MAML3 in differentiation and proliferation of neuroblastomas. Overall design: RNA-seq of SK-N-SH control and MAML3 overexpressing (SD3.23) cells, either untreated (UT) or treated with 1 µM RA (RA).","project":"SRP058647"}
{"number_samples":1,"species":"human","abstract":"In platelets, splicing and translation occur in the absence of a nucleus. However, the integrity and stability of mRNAs derived from megakaryocyte progenitor cells remain poorly quantified on a transcriptome-wide level. As circular RNAs (circRNAs) are resistant to degradation by exonucleases, their abundance relative to linear RNAs can be used as a surrogate marker for mRNA stability in the absence of transcription. Here we show that circRNAs are enriched in human platelets 17-188 fold relative to nucleated tissues, and 14-26 fold relative to samples digested with RNAseR to selectively remove linear RNA. We compare RNAseq read depths inside and outside circRNAs to provide in silico evidence of transcript circularity, show that exons within circRNAs are enriched ~13X in platelets relative to nucleated tissues, and identify 3162 genes significantly enriched for circRNAs including some where all RNAs appear to be derived from circular molecules. We also confirm that this is a feature of other anucleate cells through transcriptome sequencing of mature erythrocytes, demonstrate that circRNAs are not enriched in megakaryocytes, and that linear RNAs decay more rapidly than circRNAs in platelet preparations. Collectively, these results suggest that circulating platelets have lost on aveage over 90% of their progenitor mRNAs, and that translation in platelets occurrs against the backdrop of a highly degraded transcriptome. Finally, we find that transcripts classified as products of reverse transcriptase template switching are both enriched in platelets and resistant to decay, countering the recent suggestion that up to 50% of rearranged RNAs are artefacts. Overall design: A single rRNA depleted total RNA sample was sequenced. This together with 25 publicly available rRNA depleted total RNA samples (including 3 from platelets) were analysed using PTESFinder v 1 (http://sourceforge.net/projects/ptesfinder-v1/) to identify back-splice junctions, characteristic of circRNA transcripts. The contribution of circRNA producing exons was analysed on a gene by gene basis as follows: All circRNA transcripts identified in any sample were first pooled to define exons which can contribute to circRNA generation using custom scripts (available on request). For each sample, expression estimates (RPKMI) across all circRNA producing exons were computed for each locus using the total size of exons (in bp) and the read counts mapping to them. Similarly, total size and exonic read counts for exons for which no circRNA were detected in any sample were used to compute expression estimates (RPKME) for non-circRNA producing exons for each locus. Abundance ratios (RPKMI/RPKME and RPKMI/RPKMI+RPKME) were calculated and compared between Platelets and human tissues using Wilcoxon signed-rank test. Please note that the '25sample_info_accn_no.txt' contains the accession numbers and tissue/cell type information for 25 samples analyzed together.","project":"SRP058654"}
{"number_samples":2,"species":"human","abstract":"We aim to establish a highly spine metastasis mouse model and research its potential molecular mechanisms.","project":"SRP058664"}
{"number_samples":6,"species":"human","abstract":"To address the need to study frozen clinical specimens using next-generation RNA, DNA, chromatin immunoprecipitation (ChIP) sequencing and protein analyses, we developed a biobank work flow to prospectively collect biospecimens from patients with renal cell carcinoma (RCC). We describe our standard operating procedures and work flow to annotate pathologic results and clinical outcomes. We report quality control outcomes, nucleic acid yields of our RCC submissions (N=16) to The Cancer Genome Atlas (TCGA) project, as well as newer discovery platforms by describing mass spectrometry analysis of albumin oxidation in plasma and 6 ChIP sequencing libraries generated from nephrectomy specimens after histone H3 lysine 36 trimethylation (H3K36me3) immunoprecipitation. From June 1, 2010, through January 1, 2013, we enrolled 328 patients with RCC. Our mean (SD) TCGA RNA integrity numbers (RINs) were 8.1 (0.8) for papillary RCC, with a 12.5% overall rate of sample disqualification for RIN <7. Banked plasma had significantly less albumin oxidation (by mass spectrometry analysis) than plasma kept at 25°C (P<.001). For ChIP sequencing, the FastQC score for average read quality was at least 30 for 91-95% of paired-end reads. In parallel, we analyzed frozen tissue by RNA sequencing and after genome alignments, only 0.2-0.4% of total reads failed the default quality check steps of Bowtie2, which was comparable to the disqualification ratio (0.1%) of the 786-O RCC cell line, prepared under optimal RNA isolation conditions. The overall correlation coefficients for gene expression between the Mayo Clinic vs. TCGA tissues ranged from 0.75 to 0.82. These data support the generation of high-quality nucleic acids for genomic analyses from banked RCC. Importantly, the protocol does not interfere with routine clinical care. Collections over defined time points during disease treatment further enhance collaborative efforts to integrate genomic information with outcomes. Overall design: Examination of RNA expression in ccRCC","project":"SRP058667"}
{"number_samples":2,"species":"human","abstract":"Whole transcription RNA-Seq, H3K4me3 and H3K27ac ChIP-Seq, and ALK-locus hybrid capture bisulfite sequencing of ALK-ATI positive tumors and ALK-ATI negative controls","project":"SRP058714"}
{"number_samples":20,"species":"human","abstract":"To elucidate the transcriptional ‘landscape’ that regulates human lymphoid commitment during postnatal life, we used RNA sequencing to assemble the long non-coding transcriptome across human bone marrow and thymic progenitor cells spanning the earliest stages of B lymphoid and T lymphoid specification. Over 3,000 genes encoding previously unknown long non-coding RNA (lncRNAs) were revealed through the analysis of these rare populations. Lymphoid commitment was characterized by lncRNA expression patterns that were highly stage specific and were more lineage specific than those of protein-coding genes. Protein-coding genes co-expressed with neighboring lncRNA genes showed enrichment for ontologies related to lymphoid differentiation. The exquisite cell-type specificity of global lncRNA expression patterns independently revealed new developmental relationships among the earliest progenitor cells in the human bone marrow and thymus. Overall design: We performed RNA-Seq of 10 distinct cell types isolated by fluorescence activated cell sorting (FACS). From BM, we isolated CD34+CD38neglinneg cells, a population highly enriched for HSC, as well as three lymphoid progenitor populations; LMPP (CD34+CD45RA+CD38+CD10neg CD62Lhilinneg), CLP (CD34+CD38+CD10+CD45RA+linneg ) and fully B cell committed progenitors (BCP, CD34+CD38+CD19+). From thymus we isolated three CD34+ subsets; Thy1 (CD34+CD7neg CD1aneg CD4negCD8neg), Thy2 (CD34+CD7+CD1aneg CD4negCD8neg), and Thy 3 (CD34+CD7+CD1a+CD4negCD8neg), as well as fully T cell committed populations CD4+CD8+ (Thy 4), CD3+CD4+CD8neg (Thy5) and CD3+CD4neg CD8+ (Thy6).","project":"SRP058719"}
{"number_samples":9,"species":"human","abstract":"We performed deep strand-specific sequencing of poly-adenylated RNA (polyA+ RNAseq) from human, chimpanzee, macaque and mouse tissues, with the goal of detecting numerous non-annotated poorly expressed and antisense genes. We identified thousands of annotated and novel genes, especially in testis. We discovered that ~2% of the human and chimpanzee multiexonic genes were specific from such species. Overall design: We generated RNA-Seq data (~2.10 billion paired-end reads, 25-100 bp length) for the polyadenylated RNA fraction of brain (cerebral cortex), heart, liver and testis. In human and chimpanzee, we generated 2 samples per tissue corresponding to different individuals. In macaque, only 1 sample per tissue was generated. In mouse, considered as the evolutionary outgroup, we generated three pools of brain samples, and one pool of heart, liver and testis samples.  We generated an additional sample in Testis without including reverse transcriptase as a control of DNA contamination.","project":"SRP058740"}
{"number_samples":18,"species":"human","abstract":"gene expression profiling by RNA-seq in THP-1 cells treated with 1,25(OH)2D3 for 2.5-24 h Overall design: three independent experiments of 1,25(OH)2D3 time course in THP-1 cells","project":"SRP058771"}
{"number_samples":52,"species":"human","abstract":"Induced pluripotency is a promising avenue for disease modeling and therapy, but the molecular principles underlying this process, particularly in human cells, remain poorly understood due to donor-to-donor variability and intercellular heterogeneity. Here we constructed and characterized a clonal, inducible human reprogramming system that provides a reliable source of cells at any stage of the process. This system enabled integrative transcriptional and epigenomic analysis across the human reprogramming timeline at high resolution. We observed distinct waves of gene network activation, including the ordered re-activation of broad developmental regulators followed by early embryonic patterning genes and culminating in the emergence of a signature reminiscent of pre-implantation stages. Moreover, complementary functional analyses allowed us to identify and validate novel regulators of the reprogramming process. Altogether, this study sheds light on the molecular underpinnings of induced pluripotency in human cells and provides a robust cell platform for further studies. Overall design: single cell RNA-seq profiles from 52 unfractionated hiF-T cells after 10 days of reprogramming","project":"SRP058773"}
{"number_samples":4,"species":"human","abstract":"Activating Transcription Factor 4 (ATF4) is a transcription factor induced by the integrated stress response (ISR). This experiment is a genome-wide profiling of ATF4-dependent RNA expression in human HAP-1 cells. HAP-1 is a near-haploid human cell line that was derived from KBM-7 cells isolated from a patient with Chronic Myelogenous Leukemia. We analyzed WT and ATF4 KO cells. We induced ATF4 expression by mimicking amino acid starvation with the drug histidinol. Overall design: RNA expression profiles were generated for WT and ATF4 KO HAP1 cells. ATF4 genes were mutated using Cas9 genome editing technology. Amino acid starvation was mimicked by treating WT and ATF4 KO cells with 2 mM histidinol for 24 hours, which increases ATF4 expression.","project":"SRP058783"}
{"number_samples":8,"species":"human","abstract":"Transcription factor-mediated reprogramming is a powerful method to study cell fate changes. In this work, we demonstrate that the transcription factor Gata6 can initiate reprograming of multiple cell types to induced extraembryonic endoderm (iXEN) cells. Intriguingly, Gata6 is sufficient to drive iXEN cells from mouse pluripotent cells and differentiated neural cells. Furthermore, GATA6 induction in human ES (hES) cells also downregulates pluripotency gene expression and upregulates extraembryonic endoderm genes, revealing a conserved function in mediating this cell fate switch. Profiling transcriptional changes following Gata6 induction in mES cells reveals step-wise pluripotency factor disengagement, with initial repression of Nanog and Esrrb, then Sox2 and finally Oct4, alongside step-wise activation of extraembryonic endoderm genes. Chromatin immunoprecipitation and subsequent high-throughput sequencing analysis shows Gata6 enrichment near both pluripotency and endoderm genes, suggesting that Gata6 functions as both a direct repressor and activator. Together this demonstrates that Gata6 is a versatile and potent reprogramming factor that can act alone to drive a cell fate switch from diverse cell types. Overall design: (1) Microarray analysis of Gata6 overexpressing cells from 12 to 144 hours of doxycycline treatment in mouse embryonic stem (mES) cells compared to uninduced mES cells, embryo-derived XEN cells and Sox7 overexpressing mES cells after 144 hours of doxycycline treatment. (2) ChIP-seq analysis of Gata6 binding 36 hours following doxycycline treatment. (3) ChIP-seq analysis of Gata6 binding in embryo-derived XEN cells. (4) RNA-seq analysis of GATA6 overexpressing cells following 144 hours of induction in hES cells.","project":"SRP058804"}
{"number_samples":7,"species":"human","abstract":"5-hydroxymethylcytosine (5-mC) and gene expression profiling of T84 cell differentiation; 5-hmC profiling of colon cancer colonocyte fractions Overall design: T84 cell differentiation was monitored as cells formed confluent monolayers in tissue culture","project":"SRP058822"}
{"number_samples":78,"species":"human","abstract":"Induced pluripotency is a promising avenue for disease modeling and therapy, but the molecular principles underlying this process, particularly in human cells, remain poorly understood due to donor-to-donor variability and intercellular heterogeneity. Here we constructed and characterized a clonal, inducible human reprogramming system that provides a reliable source of cells at any stage of the process. This system enabled integrative transcriptional and epigenomic analysis across the human reprogramming timeline at high resolution. We observed distinct waves of gene network activation, including the ordered re-activation of broad developmental regulators followed by early embryonic patterning genes and culminating in the emergence of a signature reminiscent of pre-implantation stages. Moreover, complementary functional analyses allowed us to identify and validate novel regulators of the reprogramming process. Altogether, this study sheds light on the molecular underpinnings of induced pluripotency in human cells and provides a robust cell platform for further studies. Overall design: hiF-T cells are secondary reprogrammable cells derived from hiF cells  obtained thorough the directed differentiation of a dox-inducible OKMS  hIPSC line obtained from reprogramming BJ fibroblasts. After directed  differentiation, hiF cells were infected with a lentivirus expressing  the human telomerase and after clonal expansion individual hiF-T  clones were obtained. The multiplexed data is available at: https://refinery.stemcellcommons.org/data_sets/GSE69351_DGE/","project":"SRP058840"}
{"number_samples":18,"species":"human","abstract":"Eukaryotic genes generate multiple mRNA transcript isoforms though alternative transcription, splicing, and polyadenylation. However, the relationship between human transcript diversity and protein production is complex as each isoform can be translated differently. We fractionated a polysome profile and reconstructed transcript isoforms from each fraction, which we term Transcript Isoforms in Polysomes sequencing (TrIP-seq). Analysis of these data revealed regulatory features that control ribosome occupancy and translational output of each transcript isoform. We extracted a panel of 5' and 3' untranslated regions that control protein production from an unrelated gene in cells over a 100-fold range. Select 5' untranslated regions exert robust translational control between cell lines, while 3' untranslated regions can confer cell-type-specific expression. These results expose the large dynamic range of transcript-isoform-specific translational control, identify isoform-specific sequences that control protein output in human cells, and demonstrate that transcript isoform diversity must be considered when relating RNA and protein levels. Overall design: Total cytoplasmic and eight polysomal fractions of RNA were purified from HEK 293T cells in biological duplicate. Ribosomal RNA was depleted using Ribo-Zero (Human/Mouse/Rat; Epicenter) and libraries were prepared using the TruSeq RNA v2 kit (RS-122-2001; Illumina) skipping the polyA selection step. Reads are paired-end 75bp and sequencing adapters are GATCGGAAGAGCACACGTCTGAACTCCAGTCAC (read1) and AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT (read2).","project":"SRP058841"}
{"number_samples":88,"species":"human","abstract":"Small molecule inhibitors of the bromodomain and extraterminal (BET) family of proteins are in clinical trials for a variety of cancers, but patient selection strategies are limited.  This is due in part to the heterogeneity of response following BET inhibition (BETi), which includes differentiation, senescence, and cell death in subsets of cancer cell lines.  To elucidate the dominant features defining response to BETi, we carried out phenotypic and gene expression analysis of both treatment naïve cell lines and engineered tolerant lines.  We found that both de novo and acquired tolerance to BET inhibition are driven by the robustness of the apoptotic response and that genetic or pharmacological manipulation of the apoptotic signaling network can modify the phenotypic response to BETi.  We further identify that ordered expression of the apoptotic genes BCL2, BCL2L1, and BAD significantly predicts response to BETi.  Our findings highlight the role of the apoptotic network in response to BETi, providing a molecular basis for patient stratification and combination therapies. Overall design: Gene expression profiling of A375 melanoma cells or NOMO-1 AML cells treated with DMSO or the BET inhibitor, CPI203.  Also, gene expression profiling of the respective derived BETi-tolerant cells treated with DMSO or CPI203.","project":"SRP058856"}
{"number_samples":13,"species":"human","abstract":"The genomic landscape of cutaneous T cell lymphoma","project":"SRP058948"}
{"number_samples":2,"species":"human","abstract":"Next Generation Sequencing has proven to be an exceptionally powerful tool in the field of genomics and transcriptomics. With recent development it is nowadays possible to analyze ultra-low input sample material down to single cells. Nevertheless investigating such sample material still limits the analysis to either the genome or transcriptome, hence a combined analysis of both types of nucleic acids from the same sample material is still in demand.We developed a protocol which enables the combined analysis of the genome as well as the transcriptome by Next Generation Sequencing from ultra-low input samples. The protocol was evaluated by sequencing sub-colony structures from human embryonic stem cells containing 150 to 200 cells. The method can be adapted to any available sequencing system (e.g. Illumina, SOLiD, 454, etc.).To our knowledge, this is the first report where sub-colonies of human embryonic stem cells have been analyzed both at the genomic as well as transcriptome level. The method of this proof of concept study may find useful practical applications for cases where only a limited number of cells are available, e.g. for tissues samples from biopsies, circulating tumor cells and cells from early embryonic development. The results we present demonstrate that a combined analysis of genomic DNA and messenger RNA from ultra low input samples is feasible and can readily be applied to other cellular systems with limited material available. Overall design: sequencing analysis of ultra-low amounts of RNA (in duplicate) and DNA from the same sample","project":"SRP058977"}
{"number_samples":201,"species":"human","abstract":"To address how intratumoral heterogeneity affects anti-cancer drug responses, we profiled transcriptomes of single cancer cells originating from lung adenocarcinoma patient-derived xenograft (PDX) tumors. Overall design: We performed single-cell RNA sequencing (scRNA-Seq) together with bulk sequencing by applying Smart-Seq protocol (Ramsköld et al., Nat Biotechnol 2012). Enrichment of cancer cells in PDX from primary tumor (LC-PT-45: bulk RNA-Seq, n=1) was identified by histopathological examination and genomic signatures. Tumor cell-enriched PDX cells (LC-PT-45: scRNA-Seq, n=34; bulk RNA-Seq, n=9) were analyzed, and additional batch (LC-Pt-45-Re: scRNA-Seq, n=43; bulk RNA-Seq, n=7) was obtained to check comparable results. H358 human lung cancer cells (scRNA-Seq, n=50; bulk RNA-Seq, n=1) were used as cell line controls. Another lung cancer PDX case (LC-MBT-15: scRNA-Seq, n=49; bulk RNA-Seq, n=7) was prepared to validate our analytical strategy applied in the LC-PT-45 case.","project":"SRP059035"}
{"number_samples":207,"species":"human","abstract":"Diarrhea remains a major cause of death in children. Current diagnostic methods largely rely on stool culture and suffer from low sensitivity and inadequate specificity, often leading to inappropriate treatment. The objective of the present study was to use RNA sequencing (RNAseq) analysis to determine blood transcriptional profiles specific for several common pathogenic bacteria and viruses that cause diarrhea in children. We collected whole blood samples from children in Mexico having diarrhea associated with a single pathogen and without systemic complications. Our RNAseq data suggested that the blood signatures can differentiate children with diarrhea from healthy children either with or without bacterial colonization. Moreover, we detected different expression profiles from bacterial and viral infection, demonstrating for the first time the use of RNAseq to identify the etiology of infectious diarrhea. Overall design: 255 whole blood samples from 246 children including children with diarrhea caused by rotavirus (n=60 total; 5 repeated; 55 unique), E.coli (n=55), Salmonella (n=36), Shigella (n=37), adenovirus (n=8), norovirus (n=7), and control children (n=52 total; 4 repeated; 48 unique).","project":"SRP059039"}
{"number_samples":74,"species":"human","abstract":"We used RNA sequencing to profile over 70 transcriptomes of CD4+ T cells, a cell type crucial for Coeliac Disease CD pathogenesis, in both stimulated and resting samples from individuals with CD and unaffected controls The data gave us the opportunity to (i) compare gene expression between cases and controls; (ii) specifically assess whether genes that have been genetically associated with the disease were being DE; (iii) and also look for known and novel aspects of pathogenesis in the transcriptome of this specific cellular compartment. Overall design: RNA sequencing was performed on mRNA  extracted from the CD4+ T cells of 15 Coeliac patients and 11 Controls that had been stimulated with anti-CD3/anti-CD28, PMA and left unstimulated. In total we sequenced 74 transcriptome samples using 50bp reads on an Illumina HiSeqâ„¢ 2000.","project":"SRP059057"}
{"number_samples":19,"species":"human","abstract":"The precise makeup of chromatin remodeling complexes is important for determining cell type and cell function. The SWI/SNF chromatin remodeling complex is made up of multiple subunits that can be filled by mutually exclusive proteins. Inclusion or exclusion of these proteins has profound functional consequences, yet we currently understand little about the direct functional relationship between these biochemically distinct forms of remodeling complexes. Here we combine chromatin immunoprecipitation, transcriptome analysis, and transcription factor binding information from the ENCODE project to determine the functional relationship between three biochemically distinct forms of SWI/SNF. We find widespread overlap in transcriptional regulation and the genomic binding of the three ARID (AT-Rich Interacting Domain) subunits of SWI/SNF. Despite the numerous similarities in their transcriptional regulation and the co-factors bound with each ARID we identify several novel interaction modalities. Previous work has found examples of competition or subunit switching at individual loci, and we find this functional relationship is widespread, and in these cases gene expression changes following loss of one ARID depend on the function of another ARID. We also identify a previously unknown cooperative interaction between ARID1B and ARID2 in the repression of a large number of genes. Together these data help untangle the complicated combinatorial relationships between a highly heterogenous chromatin remodeling family. Overall design: We performed depletion of ARID subunits (ARID1A , n=5; ARID1B, n=3, ARID2, n=5) of SWI/SNF using siRNA or a Non-Targeting control (N=6) and performed expression analysis using polyA+ selected RNA and a strand-specific dUTP incorporation library protocol.","project":"SRP059066"}
{"number_samples":8,"species":"human","abstract":"Innate lymphoid cell (ILC) subsets that mirror helper T cells in their effector cytokine profiles have recently emerged as central players in both homeostatic and inflammatory conditions.  Like their Th1, Th2 and Th17/Th22 helper T cell counterparts, ILC subsets are categorized based on their expression of specific transcription factors and effector cytokines:  group 1 ILC (ILC1) express T-bet and IFN-?; group 2 ILC (ILC2) express GATA-3 and type 2 effector cytokines such as IL-13 and IL-5; and group 3 ILC (ILC3) express RORgt and the cytokines IL-22 and/or IL-17.  Under this nomenclature, natural killer (NK) cells and lymphoid tissue inducers (LTi) are considered ILC1 and ILC3, respectively.  ILC1 contain both CD4+ and CD4- populations, but whether this phenotypic characteristic reflects functional differences between these two populations is unknown.  These studies examine the gene expression profiles of CD4+ vs CD4- ILC1 in a cohort of healthy control subjects. Overall design: ILC subsets were isolated from the peripheral blood of healthy control subjects.  cDNA was isolated and amplified from sorted populations, and gene expression was analyzed by RNAseq","project":"SRP059170"}
{"number_samples":169,"species":"human","abstract":"Diagnosis of brucellosis remains challenging for several reasons, including lack of culture sensitivity, nonspecific symptomatology, and high prevalence of positive serology in endemic areas. The main objectives of this study were to identify blood biomarkers specific to brucellosis compared to other endemic infections and to monitor changes in blood biomarkers during treatment. To obtain a global profile of the disease, we employed RNA sequencing (RNAseq) of whole blood RNA to measure host response against brucellosis infection in patients from Macedonia and Spain. Long-term follow up of patients was used to classify patients as having acute or chronic/reinfection brucellosis and as treatment responders and non-responders. We observed distinct gene expression differences between samples from acute brucellosis and control donors. The magnitude of gene expression changes was associated with antibody titers determined by standard serological tests for brucellosis, including the rose Bengal and standard agglutination tests. The expression signature characteristic of acute brucellosis was also different from that of subjects with leishmaniasis. In depth integration of clinical data and serological findings with our gene expression data will be performed to provide insight into cellular and molecular mechanisms of brucellosis infection. Overall design: Whole blood from 169 subjects including 105 patients with suspected brucellosis, 17 patients with leishmaniasis, and 47 healthy controls.","project":"SRP059172"}
{"number_samples":34,"species":"human","abstract":"We report a time course of RNA-seq data from wild-type embryonic stem cells and embryonic stem cells in which the cardiogenic transcription factors ZNF503, ZEB2 and NKX2-5 are depleted with shRNAs differentiating along the cardiac lineage. Overall design: Biological replicates of RNA-seq data from embryonic stem cells differentiating along the cardiac lineage.","project":"SRP059197"}
{"number_samples":20,"species":"human","abstract":"Granulopoietic differentiation of myeloid progenitor cells derived from control iPSCs  was performed in a two-step liquid culture. At the end of culture, stages of differentiation were identified by morphological analysis and submitted for RNA-sequencing analysis in order to provide insight into the genomic landscape of myeloid lineage hematopoiesis as modeled by the in vitro induced differentiation of iPSCs as compared to in vivo bone marrow-derived promyelocytes. Overall design: Peripheral blood from healthy controls was obtained and iPSC were generated from peripheral blood mononuclear cells.  Hematopoietic progenitors generated from control iPSCs when cultured in myeloid expansion medium containing 50ng/mL SCF, 10ng/mL IL-3 and 10ng/mL GM-CSF for 5 days at which point cells were stained for CD45-Pacific blue, CD34-PECy7, CD33-AP, CD11b-APC-Cy7, CD15-FITC.  7-AAD was used to eliminate the dead cells. The promyelocytic population (CD45+CD34-CD33+CD11b-CD15+/lo) was sorted and the RNA from control iPSC promyelocytes was isolated using QIAGEN RNAeasy mini kit. The RNA samples were processed for RNA-seq analyses using RNA-seq protocol from NuGEN and Illumina. The amplified products were sequenced to analyze the gene expression profile of each replicate sample.  A total of 20 samples were used in this analysis to characterize and compare iPSC in vitro differentiated myeloid cells with those isolated from human bone marrow.","project":"SRP059205"}
{"number_samples":8,"species":"human","abstract":"Alternative splicing (AS) generates extensive transcriptomic and proteomic complexity. However, the functions of species- and lineage-specific splice variants are largely unknown. Here, we show that mammalian-specific skipping of exon 9 of PTBP1 alters its splicing regulatory activities and affects the inclusion levels of numerous exons. During neurogenesis, skipping of exon 9 reduces PTBP1 repressive activity so as to facilitate activation of a brain-specific AS program. Engineered skipping of the orthologous exon in chicken cells induces a large number of mammalian-like AS changes in PTBP1 target exons. These results thus reveal that a single exon skipping event in an RNA binding regulator directs numerous AS changes between species. The results further suggest that these changes contributed to evolutionary differences in the formation of vertebrate nervous systems. Overall design: This study contains two sets of samples: (Set 1) mRNA profiling of human 293 cells subjected to four different conditions in two biological replicates: non-targetting control siRNA, PTBP1 and PTBP2 siRNA, PTBP1 and PTBP2 siRNA with overexpression of full-length human PTBP1, PTBP1 and PTBP2 siRNA with overexpression of exon-excluded human PTBP1. (Set 2) mRNA profiling of chicken DT40 cells with 3 genotypes in two bioligcal replicates: wildtype cells, cells with PTBP1 exon 8 (orthologous to human PTBP1 exon 9) deleted in one allele, and cells with PTBP1 exon 8 deleted in both alleles.","project":"SRP059242"}
{"number_samples":13,"species":"human","abstract":"Personalized anti-cancer treatment based on our proprietary geno-proteomic approach that hinges on a combination of next generation sequencing and high-throughput mass spectrometry (MS) peptide identification.","project":"SRP059244"}
{"number_samples":6,"species":"human","abstract":"Novel analytic tools are needed to elucidate the molecular basis of leukemia-relevant gene mutations in the post-genome era. We generated isogenic leukemia cell clones in which the FLT3 gene was disrupted in a single allele using TALENs. Isogenic clones with mono-allelic disrupted FLT3 were compared to an isogenic wild-type control clone and parental leukemia cells for transcriptional expression, downstream FLT3 signaling and proliferation capacity. The global gene expression profiles of mutant K562 clones and corresponding wild-type controls were compared using RNA-seq. The transcriptional levels and the ligand-dependent autophosphorylation of FLT3 were decreased in the mutant clones. TALENs-mediated FLT3 haplo-insufficiency impaired cell proliferation and colony formation in vitro. These inhibitory effects were maintained in vivo, improving the survival of NOD/SCID mice transplanted with mutant K562 clones. Cluster analysis revealed that the gene expression pattern of isogenic clones was determined by the FLT3 mutant status rather than the deviation among individual isogenic clones. Differentially expressed genes between the mutant and wild-type clones revealed an activation of nonsense-mediated decay pathway in mutant K562 clones as well as an inhibited FLT3 signaling. Our data support that this genome-editing approach is a robust and generally applicable platform to explore the molecular bases of gene mutations. Overall design: Global gene expression profiles of three isogenic K562 mutant clones (clones k20, k112, k324) and three randomly selected wild-type clones (clones kw1, kw2, kw3) were generated by RNA-seq, using Illumina Hiseq 2000.","project":"SRP059266"}
{"number_samples":8,"species":"human","abstract":"Enhanced sensitivity to Wnts is an emerging hallmark of a subset of cancers, defined in part by mutations regulating the abundance of their receptors. Inhibition of Wnt secretion by blocking an essential post-translational modification, palmitoleation, provides a useful therapeutic intervention. Inhibition of PORCN in RSPO3-translocated cancers via treatment with ETC-159 causes a marked remodeling of the transcriptome, with loss of cell cycle, stem cell, and proliferation genes and an increase in differentiation markers. Inhibition of Wnt signaling by PORCN inhibition holds promise as differentiation therapy in genetically defined human cancers. Overall design: RNA-seq of colorectal PDXs with confirmed R-spondin fusion genes. RNA-seq was performed in 2 conditions (Vehicle and treatment with ETC-159 (an inhibitor of PORCN) with 4 replicates per condition.","project":"SRP059275"}
{"number_samples":4,"species":"human","abstract":"The control of cell identity is orchestrated by transcriptional and chromatin regulators in the context of specific chromosome structures. With the recent isolation of human naive embryonic stem cells (ESCs) representative of the ground state of pluripotency, it is possible to deduce this regulatory landscape in one of the earliest stages of human development. Here we generate cohesin ChIA-PET chromatin interaction data in naive and primed human ESCs and use it to reconstruct and compare the 3D regulatory landscapes of these two stages of early human development. The results reveal shared and stage-specific regulatory landscapes of topological domains and their subdomains, which consist of CTCF-CTCF/cohesin loops and enhancer-promoter/cohesin loops. The enhancer-promoter loop data reveal that genes with key roles in pluripotency are nearly always regulated by one or more super-enhancers, and show that these genes tend to occur in insulated neighborhoods. Our results reveal the key features of the 3D regulatory landscape of early human cells that form the foundation for embryonic development. Overall design: Polyadenylated RNA-seq from naive and primed human embroynic stem cells.","project":"SRP059279"}
{"number_samples":8,"species":"human","abstract":"The purpose of this study is to identify the transcriptome-wide functional targets of the splicing factor hnRNP L using mRNA sequencing and bioinformatic analysis.","project":"SRP059357"}
{"number_samples":16,"species":"human","abstract":"Induced pluripotent stem cell (iPSC)-derived cortical neurons present a powerful new model of neurological disease. Previous work has established that differentiation protocols produce cortical neurons but little has been done to characterise these at cellular resolution. In particular, it is unclear to what extent in vitro two-dimensional, relatively disordered culture conditions recapitulate the development of in vivo cortical layer identity. Single cell multiplex RT-qPCR was used to interrogate the expression of genes previously implicated in cortical layer or phenotypic identity in individual cells. Unexpectedly, 22.7% of neurons analysed frequently co-expressed canonical fetal deep and upper cortical layer markers, and this co-expression was also present at the level of translated protein. By comparing our results to available single cell RNA-seq data from human fetal and adult brain, we observed that this co-expression of layer markers was also seen in primary tissue. These results suggest that establishing neuronal layer identity in iPSC-derived or primary cortical neurons using canonical marker genes transcripts is unlikely to be informative. Overall design: Single cell RNA-seq of 16 iPSC-derived cortical neurons. This dataset was used for normalization purposes for GSE67835.","project":"SRP059379"}
{"number_samples":75,"species":"human","abstract":"PIP3 is synthesized by PI3Ks and regulates complex cell responses, such as growth and migration. Signals that drive long-term reshaping of cell phenotypes are difficult to resolve because of complex feedback networks that operate over extended times. It is clear PIP3-dependent modulation of mRNA accumulation is important in this process but is poorly understood. We have quantified the genome-wide mRNA-landscape of non-transformed, breast epithelium-derived MCF10a cells and its response to transient (EGF or PI3Ka-selective inhibitor) or chronic (isogenic cells expressing an oncomutant PI3Ka allele or lacking the PIP3-phosphatase /tumour-suppressor, PTEN) perturbations of PIP3.These results show that whilst many mRNAs are changed by long-term  genetic perturbation of PIP3 signaling (“butterfly effect”), a much smaller number change with a directional logic that aligns with different PIP3 perturbations, allowing discrimination of more directly regulated mRNAs. Our results also indicate that mRNAs can be differentially sensitive to specific features of PIP3 signals, that PIP3-sensitive mRNAs encode PI3K pathway components and identify the transcription factor binding motifs SRF and PRDM1 as important regulators of PIP3-sensitive mRNAs involved in cell movement. Overall design: RNA-seq on WT MCF10a, treated or not with A66 (Pi3Kalpha inhibitor), PIK3CA H1047R MCF10a and PTEN KO MCF10a. EGF time course stimulation applied (0, 15, 40, 90, 180, 300 min). A66 no EGF when A66 was applied for 300min w/o EGF simulation. All samples made in triplicate. Total of 75 samples.","project":"SRP059431"}
{"number_samples":95,"species":"human","abstract":"The goal of this study was to assess the single-cell transcriptional profiles Mucosal Associated Invariant T-cells (MAIT) stimulated with IL-12, IL-15 and IL-17 compared to non-stimulated cells","project":"SRP059458"}
{"number_samples":18,"species":"human","abstract":"The molecular clock is a transcriptional oscillator present in brain and peripheral cells that coordinates behavior and physiology with the solar cycle. Here we reveal that the clock gates insulin secretion through genome-wide transcriptional control of the pancreatic exocyst across species. Clock transcription factors bind to unique enhancer sites in cycling genes in beta cells that diverge from those in liver, revealing the dynamics of inter-tissue clock control of genomic and physiologic processes important in glucose homeostasis. Overall design: Transcriptome profiling in mouse and human islets at serial 4-hour time intervals by polyA RNA-Seq","project":"SRP059509"}
{"number_samples":14,"species":"human","abstract":"During its long infection cycle, human cytomegalovirus (HCMV) extensively manipulates cellular gene expression to maintain conditions favorable for viral propagation. In order to reveal the signature of cellular genes that are manipulated by HCMV, we measured RNA abundance and rate of protein production through the course of HCMV infection. We characterized changes for most expressed cellular genes and although much of the regulation was transcriptional we uncover diverse and dynamic translational regulation for subsets of host genes, revealing unappreciated coordination in translational control that suggests common regulators Overall design: Ribosome profiling and mRNA-seq along HCMV infection","project":"SRP059531"}
{"number_samples":8,"species":"human","abstract":"Human Brain CAGE and RNA-seq data","project":"SRP059579"}
{"number_samples":47,"species":"human","abstract":"The MYC transcription factor is an unstable protein and its turnover is controlled by the ubiquitin system. Ubiquitination enhances MYC-dependent transactivation, but the underlying mechanism remains unresolved. Here we show that proteasomal turnover of MYC is dispensable for recruitment of RNA polymerase II (RNAPII), but is required to promote transcriptional elongation at MYC target genes. Degradation of MYC stimulates histone acetylation and recruitment of BRD4 and P-TEFb to target promoters, leading to phosphorylation of RNAPII CTD and the release of elongating RNAPII. In the absence of degradation, the RNA polymerase II-associated factor (PAF) complex associates with MYC via interaction of its CDC73 subunit with a conserved domain in the amino-terminus of MYC (\"MYC box I\"), suggesting that a MYC/PAF complex is an intermediate in transcriptional activation. Since histone acetylation depends on a second highly conserved domain in MYCs amino-terminus (\"MYC box II\"), we propose that both domains co-operate to transfer elongation factors onto paused RNAPII. Overall design: RNA-Seq Experiments were performed in a primary breast epithelial cell line (IMEC).The cell line expressed doxycycline-inducible versions of MYC (WT;Kless,Swap=WTN-KC). Where indicated cells were transfected with siRNAs (siCtrl;siSKP2). Where indicated cells were treaed with the proteasome inhibitor MG132 or EtOH as solvent control. DGE was performed by comparing Dox-treated populations expressing either Dox-inducible MYC or a vector control or comparing Dox-induced cells with EtOH (solvent control) treated cells.","project":"SRP059643"}
{"number_samples":12,"species":"human","abstract":"To identify HNRNPA2B1 binding sites on endogenous nuclear RNAs, we performed HITS-CLIP for endogenous HNRNPA2B1 and RNA-seq to analyze the nuclear RNA under either METTL3 or HNRNPA2B1 depletion. Overall design: Wild type MDA-MB-231 cells were subjected to the HITS-CLIP procedure on immunoprecipitated HNRNPA2B1 associated RNA obtained from the nuclear fraction (Licatalosi D, et al. 2008, Nature 456:464-U22). For RNA-seq, nuclear RNA was extracted from MDA-MB-231 or Hela cells knocked down for METTL3 or HNRNPA2B1.","project":"SRP059692"}
{"number_samples":12,"species":"human","abstract":"We report gene expression in human neutrophils isolated by two methods: Polymorphprep (~95% purity) and negative selection (~99% purity) from two healthy donors - one donor with low eosinophil contamination of neutrophils and one donor with high eosinophil contamination of neutrophils.  We report the effect of the presence of contaminating leukocytes in neutrophil preparations, and in reponse to inflammatory cytokines TNF-alpha and GM-CSF. Overall design: Healthy human neutrophils were isolated using Polymorphprep or negative selection, and incubated for 1h in the absence or presence of TNF-alpha or GM-CSF.  RNA was analysed by Illumina HiSeq 2000.  The results from n=2 donors were analysed as biological replicates for differential expression analysis.","project":"SRP059695"}
{"number_samples":8,"species":"human","abstract":"The androgen receptor (AR), a nuclear transcription factor (TF), is consistently reprogrammed during prostate tumorigenesis Overall design: Gene expresion profiles when LHSAR with overexpressed FOXA1, HOXB13 or FOXA1 and HOXB13 together compared with LacZ control","project":"SRP059701"}
{"number_samples":20,"species":"human","abstract":"High grade serous ovarian cancers (HGSC) are deadly malignancies that relapse despite carboplatin chemotherapy. Here we show that 16 independent primary HGSCs contain a CA125 negative population enriched for carboplatin resistant cancer initiating cells. Transcriptome analysis reveals up-regulation of homologous recombination DNA repair and anti-apoptotic signals in this population. While treatment with carboplatin enriches for CA125 negative cells, co-treatment with carboplatin and birinapant eliminates these cells in HGSCs expressing high levels of the inhibitor of apoptosis protein cIAP in the CA125 negative population. Birinapant sensitizes CA125 negative cells to carboplatin by mediating degradation of cIAP causing cleavage of caspase-8 and restoration of apoptosis. This co-therapy significantly improved disease free survival in vivo compared to either therapy alone in tumor-bearing mice. These findings suggest that therapeutic strategies that target CA125 negative cells may be useful in the treatment of HGSC. Overall design: mRNA profiles of CA125 positive and negative populations, generated by next generation sequencing of populations FACS isolated from 10 independent dissociated primary human high grade serous ovarian cancers, were compared.","project":"SRP059732"}
{"number_samples":6,"species":"human","abstract":"High grade serous ovarian cancers (HGSC) are deadly malignancies that relapse despite carboplatin chemotherapy.  Many commercially ovarian cancer cell lines are not good models for HGSC. Here we demonstrate that 3 low passage cell lines derived from HGSC have similar transcriptomes to their parental bulk tumors. These cell lines recapitulated tumor characteristics of the primary cancer and had responded to therapy in the same manner as primary HGSC cells, demonstrating they are accurate models for HGSCs. Overall design: mRNA profiles of low passage high grade serous tumor cell lines and their parental tumors, generated by next generation sequencing, were compared.","project":"SRP059733"}
{"number_samples":12,"species":"human","abstract":"BACKGROUND: Combination antiretroviral therapy (cART) is able to control HIV-1 viral replication, however long-lived latent infection in resting memory CD4+ T-cells persist. The mechanisms for establishment and maintenance of latent infection in resting memory CD4+ T-cells remain unclear. Previously we have shown that HIV-1 infection of resting CD4+ T-cells co-cultured with CD11c+ myeloid dendritic cells (mDC) produced a population of non-proliferating T-cells with latent infection. Here we asked whether different antigen presenting cells (APC), including subpopulations of DC and monocytes, were able to induce post-integration latent infection in resting CD4+ T-cells, and examined potential cell interactions that may be involved using RNA-seq. RESULTS: mDC (CD1c+), SLAN+ DC and CD14+ monocytes were most efficient in stimulating proliferation of CD4+ T-cells during syngeneic culture and in generating post-integration latent infection in non-proliferating CD4+ T-cells following HIV-1 infection of APC-T-cell co-cultures. In comparison, plasmacytoid DC (pDC) and B-cells did not induce latent infection in APC-T-cell co-cultures. We compared the RNA expression profiles of APC subpopulations that could and could not induce latency in non-proliferating CD4+ T-cells. Gene expression analysis, comparing the mDC, SLAN+ DC and CD14+ monocyte subpopulations to pDC identified 53 upregulated genes that encode proteins expressed on the plasma membrane that could signal to CD4+ T-cells via cell-cell interactions (32 genes), immune checkpoints (IC) (5 genes), T-cell activation (9 genes), regulation of apoptosis (5 genes), antigen presentation (1 gene) and through unknown ligands (1 gene). CONCLUSIONS: APC subpopulations from the myeloid lineage, specifically mDC subpopulations and CD14+ monocytes, were able to efficiently induce post-integration HIV-1 latency in non-proliferating CD4+ T-cells in vitro. Inhibition of key pathways involved in mDC-T-cell interactions and HIV-1 latency may provide novel targets to eliminate HIV latency. Overall design: mRNA profiles of sorted, pure antigen presenting cells including, CD1c+ myleoid dendirtic cells (mDC), SLAN+ mDC, CD14+ monocytes and plasmacytoid DC (pDC), were generated using next generation sequencing in triplicate, using Illumina Illumina Hiseq 2000.","project":"SRP059735"}
{"number_samples":192,"species":"human","abstract":"A cell supsension containing an equal mix of HEK and 3T3 cells was used in the Fluidigm C1 Overall design: Suspensions of 3T3 and HEK cells were diluted down to a concentration of 250,000 per mL and mixed 1:1, then loaded onto two medium C1 cell capture chips.","project":"SRP059775"}
{"number_samples":6,"species":"human","abstract":"To define ncRNA expression in hypoxic endothelial cells, we applied pro-angiogenic hypoxia to cultured endothelial cells. Afterwards, total RNA was isolated and underwent RNA-seq analysis. Overall design: HUVECs were subjected to normoxic or hypoxic (0.1-0.2% O2) cell culture conditions.","project":"SRP059925"}
{"number_samples":4,"species":"human","abstract":"Purpose: identify sites in endogenous mRNAs that are cut by KSHV SOX; Method: parallel analysis of RNA ends (PARE, following Zhai et al., 2014); Results: SOX cuts at discrete locations in mRNAs Overall design: human Xrn1 was knocked down in HEK293T cells by shRNAs or siRNAs to stabilize degradation fragments with free 5' ends; GFP-SOX or GFP were transfected for ~24 hrs; total RNA samples were collected and subjected to PARE protocol (Zhai et al., 2014)","project":"SRP059948"}
{"number_samples":12,"species":"human","abstract":"Developing animal models representating the cancer biology of advanced prostate cancer patients is challenging but essential for delivering individualized medical therapies. In an effort to develop patient derived xenograft (PDX) models, we took the metastatic site tissue from the rib lesion twice (ie, before and after enzalutamide treatment) over a twelve week period  and implanted subcutaneously and under the renal capsule in immuno-deficient mice.  To characterize and compare the genome and transcriptome landscapes of patient tumor tissues and the corresponding PDX models, we performed whole exome and transcriptome sequencing for metastatic tumor tissue as well as its derived PDXs. We demonstrated the feasibility of developping PDX models from patient who developed castrate-resistant prostate cancer. Our data  suggested PDX models preserve the patient’s genomic and transcriptomic alterations in high fidelity, as illustrated by somatic mutation, copy number variation, gene fusion and gene expression. Overall design: RNA sequencing of prostate cancer tumor tissue and derived xenograft using Illumina HiSeq 2000.","project":"SRP059950"}
{"number_samples":8,"species":"human","abstract":"Total RNA sequencing of human and murine myoblasts and myotubes was extracted, depleted of ribosomal RNA and subjected to Illumina stranded paired end library prep and sequencing. Samples from Duchenne Muscular Dystrophy patients-derived myoblasts were included  in this study Overall design: 12 samples were analysed: murine C2C12 myoblasts and myotubes differentiated for 4 days, each in two biological replicates; human 9208 myoblasts and myotubes differentiated for 6 days, each in two biological replicates; human Duchenne Muscular Dystrophy patients derived myoblasts and myotubes differentiated for 10 days.","project":"SRP059957"}
{"number_samples":58,"species":"human","abstract":"Background: Long non-coding RNAs (lncRNAs) are increasingly implicated as gene regulators and may ultimately be more numerous than protein-coding genes in the human genome. Despite large numbers of reported lncRNAs, reference annotations are likely incomplete due to their lower and tighter tissue-specific expression compared to mRNAs. An unexplored factor potentially confounding lncRNA identification is inter-individual expression variability. Here, we characterize lncRNA natural expression variability in human primary granulocytes. Results: We annotate granulocyte lncRNAs and mRNAs in RNA-seq data from ten healthy individuals, identifying multiple lncRNAs absent from reference annotations, and use this to investigate three known features (higher tissue-specificity, lower expression, and reduced splicing efficiency) of lncRNAs relative to mRNAs. Expression variability was examined in seven individuals sampled three times at one or more than one month intervals. We show that lncRNAs display significantly more inter-individual expression variability compared to mRNAs. We confirm this finding in 2 independent human datasets by analyzing multiple tissues from the GTEx project and lymphoblastoid cell lines from the GEUVADIS project. Using the latter dataset we also show that including more human donors into the transcriptome annotation pipeline allows identification of an increasing number of lncRNAs, but minimally affects mRNA gene number. Conclusions: A comprehensive annotation of lncRNAs is known to require an approach that is sensitive to low and tight tissue-specific expression. Here we show that increased inter-individual expression variability is an additional general lncRNA feature to consider when creating a comprehensive annotation of human lncRNAs or proposing their use as prognostic or disease markers. Overall design: We used PolyA+ RNA-seq data from human primary granulocytes of 10 healthy individuals to de novo annotate lncRNAs and mRNAs in this cell type and ribosomal depleted (total) RNA-seq data from seven of these individuals sampled three times to analyze lncRNA amd mRNA expression variability","project":"SRP059959"}
{"number_samples":10,"species":"human","abstract":"By means of 3' end sequencing we provide a genome-wide, high-resolution polyadenylation map of the human heart. By sequencing 5 control en 5 dilated cardiomyopathy (DCM) myocardial specimens we investigate the difference in alternative polyadenylation (APA) in healthy and diseased hearts.","project":"SRP059989"}
{"number_samples":4,"species":"human","abstract":"The control of promoter-proximal pausing and the release of RNA polymerase II (RNA Pol II) is a widely used mechanism for regulating gene expression in metazoans, especially for genes that respond to environmental and developmental cues. Here, we identify Pol II associated Factor 1 (PAF1) as a major regulator of promoter-proximal pausing. Knockdown of PAF1 leads to increased release of paused Pol II into gene bodies at thousands of genes. Genes with the highest levels of paused Pol II exhibit the largest redistribution of Pol II from the promoter-proximal region into the gene body in the absence of PAF1. PAF1 depletion results in increased nascent transcription and increased levels of phosphorylation of Pol II’s c-terminal domain on serine 2 (Ser2P). These changes can be explained by the recruitment of the Ser2P kinase Super Elongation Complex (SEC) effecting increased release of paused Pol II into productive elongation, thus establishing a novel function for PAF1 as a major regulator of pausing in metazoans. Overall design: ChIP-seq of Pol II of different forms, SEC subunits, PAFc subunits and H2Bub in human cell lines targeted by PAF1 or scramble shRNA. ChIP-seq of total Pol II in HCT116 cells targeted by BRE1A or scramble shRNA. ChIP-seq of total Pol II in S2 cells targeted by Paf1 or LacZ RNAi. Total RNA-seq, nascent RNA-seq and GRO-seq in HCT116 cells targeted by PAF1 or scramble shRNA.","project":"SRP060227"}
{"number_samples":6,"species":"human","abstract":"NCCIT cell with EtOH treatment","project":"SRP060284"}
{"number_samples":4,"species":"human","abstract":"RNA Sequencing performed on EBV positive gastric cancer biopsies and cells lines to study expression of EBV specific genes. Overall design: Examination of two EBV postitive gastric carcinoma biopsies and two EBV positive gastric cancer cell lines, NCC24 and YCCEL1 by RNA-Seq.","project":"SRP060348"}
{"number_samples":4,"species":"human","abstract":"Hep3B and Huh7 are two types of human hepatoma cell lines (HCC). In our laboratory, we cultured their stem-like cancer cells (HCSCs), Hep3B-C and Huh7-C. And we have demonstrated that these cells had enhanced stem cell properties, drug resistance, properties of EMT, and stronger tumor-initiating capabilities. To explore functionally crucial mRNAs in HCSCs, 2 samples of HCSCs and 2 samples of HCCs were sequenced by the Illumina Genome Analyzer II. Through differential expression analysis, we finally identified 115 up- and 402 down-regulated miRNAs which were consistently up- and down-regulated in two stem cells compared to the cancer cells. Overall design: Expression analysis using total RNAs extracted from 2 HCSC cell lines (Hep3B-C and Huh7-C), and 2 HCC cell lines (Hep3B and Huh7).","project":"SRP060359"}
{"number_samples":12,"species":"human","abstract":"To identify RNA transcripts involved in acute and chronic renal epithelial injury, we performed unbiased whole transcriptome profiling of human proximal tubular epithelial cells (PTECs) in hypoxic and inflammatory conditions. RNA sequencing (RNA-seq) revealed that the protein-coding and noncoding transcriptomic landscape differed between hypoxia-stimulated and cytokine-stimulated human PTECs. Overall design: Examination of transcriptomic response of human PTECs to hypoxic or inflammatory injury","project":"SRP060370"}
{"number_samples":648,"species":"human","abstract":"Single cell RNA-sequencing of human tonsil Innate lymphoid cells (ILCs) from three independent tonsil donors. Overall design: Sequencing libraries were prepared from FACS sorted individual ILCs with the Smart-Seq2 protocol  (Picelli et al. Nature Methods 2013)","project":"SRP060416"}
{"number_samples":6,"species":"human","abstract":"Oncogene-induced senescence (OIS) is a tumor suppression mechanism that blocks cell proliferation in response to oncogenic signalling. OIS is frequently accompanied by multinucleation; however, the origin of this is unknown. Here we show that multinucleate OIS cells originated mostly from failed mitosis. Prior to senescence, mutant RasV12 activation in primary human fibroblasts compromised mitosis, associated with abnormal expression of mitotic genes that enter M-phase.  Simultaneously, RasV12 activation enhanced survival of damaged mitoses, culminating in extended mitotic arrest and aberrant exit from mitosis via mitotic slippage. ERK-dependent transcriptional up-regulation of Mcl1 was responsible for enhanced slippage of cells with mitotic defects and subsequent cell survival. Importantly, mitotic slippage and oncogene signalling synergistically induced senescence and key senescence regulators p21 and p16. We propose that activated Ras induces transcriptional changes that predispose cells undergoing OIS to mitotic stress and multinucleation. Overall design: We used RNA-seq of IMR90 cells with inducible expression of oncogenic RasV12 that were synchronised in mitosis, to characterise the nature of mitotic defects that lead to multinucleation of oncogene-induced senescent cells","project":"SRP060598"}
{"number_samples":24,"species":"human","abstract":"RNA-Seq after Cas9-gRNA transfection with different length gRNAs Overall design: we performed PolyA Selection and RNA-Seq on cells transfected with dCas9-VPR and a gRNA of each length (20nt, 16nt, or 14nt) targeting ACTC1, MIAT, or HBG1/2","project":"SRP060637"}
{"number_samples":6,"species":"human","abstract":"Here we derive human and chimpanzee cranial neural crest cells (CNCCs) and profile histone modifications, transcription factors, chromatin accessibility and gene expression to systematically and quantitatively annotate evolutionary divergence of craniofacial cis-regulatory landscapes. Overall design: Histone modifications (H3K27ac, H3K4me1, H3K4me3, H3K27me3), chromatin modifiers (p300), transcription factors (NR2F1, TFAP2A), chromatin accessibility (ATACseq) and gene expression (RNAseq) were assayed in CNCCs derived from iPSCs/ESCs from 2 chimpanzee and 3 human individuals.","project":"SRP060650"}
{"number_samples":3,"species":"human","abstract":"CTCF and BORIS (CTCFL), two paralogous mammalian proteins sharing nearly identical DNA binding domains, are thought to function in mutually exclusive manners in DNA binding and transcriptional regulation. Here we show that these two proteins co-occupy a specific subset of regulatory elements consisting of clustered CTCF binding motifs (termed 2xCTSes). BORIS occupancy at 2xCTSes is largely invariant in BORIS-positive cancer cells, with the genomic pattern recapitulating the germline-specific BORIS binding to chromatin. In contrast to the single-motif CTCF target sites (1xCTSes), the 2xCTS elements are preferentially found at active promoters and enhancers, both in cancer and germ cells. 2xCTSes are also enriched in genomic regions that escape histone to protamine replacement in human and mouse sperm. Depletion of BORIS gene leads to altered transcription of a large number of genes and the differentiation of K562 cells, while the ectopic expression of this CTCF paralog leads to specific changes in transcription in MCF7 cells. In summary, we discover two functionally and structurally different classes of CTCF binding regions, 2xCTSes and 1xCTSes, revealed by their predisposition to bind BORIS. We propose that 2xCTSes play key roles in the transcriptional program of cancer and germ cells Overall design: Genome-wide mapping of CTCF and BORIS occupancies in both germ and cancer cells. ChIP-seq and expression profiling by high throughput sequencing","project":"SRP060661"}
{"number_samples":3,"species":"human","abstract":"Comparison of ccRCC cells with high HIF or low HIF activity Overall design: Cells expressing control (VEC) were compared to cells with ARNT1 loss using sh/sgRNA targeting ARNT1 The samples labeled 'con' are parental Vhl-null cells and represent the 'high HIF' state. The other 2 cell lines represent the 'low HIF' state. The cell line labeled shArnt have loss of a protein named ARNT by virtue of RNAi-mediated knockdown using a shRNA targeting ARNT. The sample labeled lcARNT has loss of ARNT protein using a lenti-crispr mediated sgRNA.","project":"SRP060708"}
{"number_samples":15,"species":"human","abstract":"Background: To determine differentially expressed and spliced RNA transcripts in chronic lymphocytic leukemia specimens a high throughput RNA-sequencing (HTS RNA-seq) analysis was performed. Methods:  Ten CLL specimens and five normal peripheral blood CD19+ B cells were analyzed by HTS RNA-seq. The library preparation was performed with Illumina TrueSeq RNA kit and analyzed by Illumina HiSeq 2000 sequencing system. Results: An average of 48.5 million reads for B cells, and 50.6 million reads for CLL specimens were obtained with 10396 and 10448 assembled transcripts for normal B cells and primary CLL specimens respectively. With the Cuffdiff analysis, 2091 differentially expressed genes (DEG) between B cells and CLL specimens based on FPKM (fragments per kilobase of transcript per million reads and false discovery rate, FDR q<0.05, fold change >2) were identified. Expression of selected DEGs (n=32) with up regulated and down regulated expression in CLL from RNA-seq data were also analyzed by qRT-PCR in a test cohort of CLL specimens. Even though there was a variation in fold expression of DEG genes between RNA-seq and qRT-PCR; more than 90% of analyzed genes were validated by qRT-PCR analysis. Analysis of RNA-seq data for splicing alterations in CLL and B cells was performed by Multivariate Analysis of Transcript Splicing (MATS analysis). Skipped exon was the most frequent splicing alteration in CLL specimens with 128 significant events (P-value <0.05, minimum inclusion level difference >0.1). Conclusion: The RNA-seq analysis of CLL specimens identifies novel DEG and alternatively spliced genes that are potential prognostic markers and therapeutic targets. High level of validation by qRT-PCR  for a number of DEG genes supports the accuracy of this analysis. Global comparison of transcriptomes of B cells, IGVH non-mutated CLL (U-CLL) and mutated CLL specimens (M-CLL) with multidimensional scaling analysis was able to segregate CLL and B cell transcriptomes but the M-CLL and U-CLL transcriptomes were indistinguishable. The analysis of HTS RNA-seq data to identify alternative splicing events and other genetic abnormalities specific to CLL is an added advantage of RNA-seq that is not feasible with other genome wide analysis. Overall design: Ten CLL specimens and five normal peripheral blood CD19+ B cells were analyzed by HTS RNA-seq. The library preparation was performed with Illumina TrueSeq RNA kit and analyzed by Illumina HiSeq 2000 sequencing system.","project":"SRP060715"}
{"number_samples":14,"species":"human","abstract":"Nascent transcription profiles are shown for scaled megadomains and 100kb flanking regions before BRD4-NUT induction (0h) and at different time points (2h, 3h, 7h) following induction in 293T cells. Increase of the transcription from 0h to 7h after induction. Average level of transcriptional activity is reduced within the megadomains and their flanking regions following JQ1 treatment of TC-797 cells. Profile of nascent RNA-seq is shown for cells without JQ1 treatment, and for cells 1hr, 2.5hr and 4hr following JQ1 treatment. Overall design: Recovery and analysis of nascent RNA","project":"SRP061033"}
{"number_samples":2,"species":"human","abstract":"To analyze transcriptome data for the mouse xenograft cells derived from a human lung adenocarcinoma patient, reference RNA-seq data were generated for the tracking of gene expression changes associated with the xeno-engraftment and culture. Overall design: RNA-seq data were generated as a supplementary to GSE69405. The data include RNA-seq of patient normal tissue (matched normal, mN) and of mouse xenograft tissue (xenoT) of the lung adenocarcinoma.","project":"SRP061207"}
{"number_samples":384,"species":"human","abstract":"Extracellular vesicles such as exosomes are selectively enriched in RNA that has potential for use as disease biomarkers. To systemically characterize circulating extracellular RNA profiles, we performed RNA sequencing analysis on plasma extracellular vesicles derived from 192 individuals including 100 colon cancer, 36 prostate cancer and 6 pancreatic cancer patients along with 50 healthy individuals. Of ~12.6 million raw reads for each of these subjects, the number of mappable reads aligned to RNA references was ~5.4 million including microRNAs(miRNAs) (~40.4%), piwi-interacting RNAs(piwiRNAs) (~40.0%), pseudo-genes (~3.7%), long noncoding RNAs (lncRNAs) (~2.4%), tRNAs (~2.1%), and mRNAs (~2.1%). To select the best candidates for potential extracellular RNA reference controls, we performed abundant level stability testing and identified a set of miRNAs showing relatively consistent expression. To estimate biological variations, we performed association analysis of expression levels with age and sex in healthy individuals. This analysis showed significant sex association with seven small noncoding RNAs (false discovery rate, or FDR<0.05), while no small noncoding RNAs were statistically associated with age. To identify disease-associated RNA transcripts, we performed analysis of covariance by including disease status, age, sex, RNA isolation and gel size selection dates. We observed a gradual increase of significantly associated RNAs (in particular, miRNAs) with disease advancement as denoted by cancer staging. We found significant association of miR-125a-5p and miR-1246-3p with all cancer types tested (FDR<0.05). Based on the disease associations, we developed cancer type-specific multivariate statistical models to predict disease status with an area under the ROC curve from 0.67 in stage I colon cancer to 0.92 in advanced prostate cancer. To date, this is the largest RNA-seq study to systematically profile extracellular RNA species, which has not only provided a baseline reference profile for circulating extracellular RNA, but also a set of RNA candidates for reference controls and disease biomarkers. Overall design: RNAs fro plasma circulating microviscles in 192 individuals were sequenced and quantified. RNA expression stability testing was performed to identify stably expressed RNAs. Distribution of RNA species and individual RNA transcripts were compared in normal and cancer patients.","project":"SRP061240"}
{"number_samples":2,"species":"human","abstract":"The goals of this study were to identify LIN28 downstream gene targets in breast cancer cells. We use a subclone of the MCF-7 breast cancer cell line, MCF-7M as our model system. Methods: mRNA-protein complexes (mRNP) lysates were prepared from MCF-7M cells and incubated with Protein-A Sepharose beads (Sigma-Aldrich) and either LIN28 (Abcam) or control normal rabbit serum IgG antibodies. LIN28 interacting mRNAs were identified by whole genome sequencing. Results: Using an optimized data analysis workflow, we mapped approximately 13 million sequence reads for LIN28-IP and CTL- IP (IgG), respectively to the to the human genome (build h19). Conclusions: mRNA were significantly bound by LIN28 if LIN28 RIP had 2.5 fold increase in normalized reads compared to IgG. We found that LIN28 was predominantly bound at coding exons and 3'UTRs, 38% & 45% respectively, in the 843 mRNAs within MCF-7M genome. Overall design: LIN28 mRNA enriched regions identified from LIN28/RNA complexes prepared from MCF-7M cells.","project":"SRP061241"}
{"number_samples":8,"species":"human","abstract":"The goals of this study were to identify LIN28 downstream gene targets in breast cancer cells. We use a subclone of the MCF-7 breast cancer cell line, MCF-7M as our model system. Methods: mRNA profiles from MCF-7M breast cancer cells treated with siRNA against non-targeting control (NT), LIN28, hnRNP A1, LIN28 and hnRNPA1 (LIN28A1) for 72 hrs were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays Results: Using an optimized data analysis workflow, we mapped over 200 million sequence reads per sample to the human genome (build h19). Each of the four groups had two biological replicates. We developed a custom method to identify alternative splicing events and identified 111 genes with significant (FDR<0.05) differential splicing for LIN28 depleted cells compared to non-targeting siRNA control, as well as 249 and 182 genes for hnRNP A1 and LIN28A1 respectively. RNA-seq data were validated with by qRT–PCR analysis of a subset of genes. Conclusions: Results reveal that LIN28 regulates alternative splicing and steady state mRNA expression of genes implicated in aspects of breast cancer biology. Notably, cells lacking LIN28 undergo significant isoform switching of the ENAH gene, resulting in a decrease in the expression of ENAH exon 11a isoform. Expression of ENAH isoform 11a has been shown to be elevated in breast cancers that express HER2. Overall design: mRNA profiles of MCF-7M cells treated with siRNA for NT control, LIN28, hnRNP A1, and LIN28 plus hnRNP A1 (A1) (LIN28A1) were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000","project":"SRP061243"}
{"number_samples":1,"species":"human","abstract":"MicroRNAs (miRNAs) have been shown to play a considerable role in the development and progression of nasopharyngeal carcinoma(NPC). Despite accumulating studies on the molecular mechanisms of NPC, the miRNA regulatory networks in cancer progression remain largely unknown. Laser capture microdissection(LCM) and deep sequencing are powerful tools that can help us to detect the integrated view of miRNA-target network.","project":"SRP061255"}
{"number_samples":6,"species":"human","abstract":"The analysis investigates the impact of methyl donor S-adenosylmethionin on transcription and methylation profiles of prostate carcinoma cells. Overall design: PC-3 cells (Prostate carcinoma cells) were treated with 160 micromolar S-adenosylmethionine or vehicle","project":"SRP061292"}
{"number_samples":12,"species":"human","abstract":"We sequenced polyA mRNA from OVCAR8-ADR-Cas9 cells in which one or two of 3 epigenetic regulators (BRD4, KDM4C, KDM6B) had been knocked out to examine how global gene expression was affected and evaluate potential synergistic effects at a molecular level. Overall design: Gene expression data (RNA-Seq) in OVCAR8-ADR-Cas9 cells infected with control vector or vectors expressing gRNAs targeting one of 4 epigenetic regulators (BRD4, KDM4C, KDM6B) with biological replicates.","project":"SRP061310"}
{"number_samples":6,"species":"human","abstract":"MYC is a major oncogenic driver of Multiple Myeloma (MM) and yet almost no therapeutic agents exist that target MYC in MM. Here we report that the let-7 biogenesis inhibitor LIN28B correlates with MYC expression in MM and is associated with adverse outcome. We also demonstrate that the LIN28B/let-7 axis modulates the expression of MYC, itself a let-7 target. Further, perturbation of the axis regulates the proliferation of MM cells in vivo in a xenograft tumor model. RNA sequencing and gene set enrichment analyses of CRISPR-engineered cells further suggest that the LIN28/let-7 axis regulates MYC and cell cycle pathways in MM. We provide proof-of-principle for therapeutic regulation of MYC through let-7 with an LNA-GapmeR containing a let-7b mimic in vivo, demonstrating that high levels of let-7 expression repress tumor growth by regulating MYC expression. These findings reveal a novel mechanism of therapeutic targeting of MYC through the LIN28B/let-7 axis in MM that may impact other MYC dependent cancers as well. Overall design: RNA sequencing of MOLP-8 cells transduced with lentiCRISPRv2 scrambled control or containing a sgRNA against LIN28B. Both control and LIN28B KO cells were sequenced in triplicate.","project":"SRP061329"}
{"number_samples":12,"species":"human","abstract":"High uniform fluid shear stress (FSS) is atheroprotective and preserves the endothelial phenotype and function through activation of downstream mediators such as MAPK7 (Erk5). Endothelial cells respond to FSS thanks to mechanotransduction. However, how the resulting signaling is integrated and resolved at the epigenetic level, remains elusive. We hypothesized that Polycomb methyltransferase EZH2 is involved in the effects of FSS in human endothelial cells. We showed that FSS decreases the expression of the Polycomb methyltransferase EZH2. Despite simultaneous activation of MAPK7, MAPK7 pathway does not directly influence the transcription of EZH2. Interestingly though, the knock down of EZH2 activates the protective MAPK7 signaling in endothelial cells, even in the absence of FSS. To understand the influence of the FSS-decreased expression of EZH2 on endothelial transcriptome, we performed RNA-seq and differential gene expression analysis. We identified candidate groups of genes dependent on both EZH2 and FSS. Among those, Gene Ontology overrepresentation analysis revealed highly significant enrichment of the cell cycle-related genes, suggesting changes in proliferation. Indeed, the depletion of EZH2 strongly inhibited endothelial proliferation, indicating cell cycle arrest. The concomitant decrease in CCNA expression suggests the transition of endothelial cells into a quiescent phenotype. Further bioinformatical analysis suggested TXNIP as a possible mediator between EZH2 and cell cycle-related gene network. Our data show that EZH2 is a FSS-responsive gene. Decreased EZH2 levels enhance the activation of the atheroprotective MAPK7 signaling. Decrease in EZH2 under FSS mediates the decrease in the expression of the network of cell cycle-related genes, which allows the cells to enter quiescence. EZH2 is therefore important for the protective effects of FSS in endothelium. Overall design: Puromycin-selected HUVEC (Human Umbilical Vein Endothelial Cells, Lonza, Switzerland) cells, expressing either scrambled control (SCR) or anti-EZH2 short-hairpin (shEZH2) constructs (at total 7 days after the first viral transduction), were used in FSS experiments (72h of control static culture or exposure to 20 dynes/cm2 of fluid shear stress, using Ibidi pump system (in µ-Slides I 0.4 Luer, Ibidi, Planegg/Martinsried, Germany)). Each replicate experiment consisted of viral transductions and puromycin selection of a separate HUVEC batch, followed by the FSS experiment. Two FSS experimental sets of the same HUVEC batch were run every time in parallel and lysed at the same end time point, one in RNAse-free conditions with RNA-Easy Mini Plus kit RLT Plus lysis buffer (QIAGEN, Venlo, The Netherlands), and one with RIPA buffer. The RIPA-lysates were analyzed with Western blotting and confirmed the complete (no protein present) knock-down of EZH2. From the RNA-lysates, RNA was isolated using the RNA-Easy Mini Plus kit (QIAGEN, Venlo, The Netherlands). High quality RNA samples (pre-assessed by Nanodrop measurements) were further processed in the Genome Analysis Facility of the University Medical Center Groningen. The RNA quality and integrity were verified using PerkinElmer Labchip GX with a cut-off value of 9 (scale 1 to 10, where 9 is very high quality RNA). RNA library was created in accordance with the TruSeqTM RNA Sample Preparation v2 Guide (Illumina, San Diego, CA, USA), using the PerkinElmer Sciclone liquid handler, resulting in 330bp cDNA fragments. The paired-end sequencing (100bp reads) was performed using the Illumina HiSeqTM 2500. (Quoted from the Materials and Methods of the related manuscript, with adjustments).","project":"SRP061380"}
{"number_samples":3,"species":"human","abstract":"Circular RNAs (circRNAs) are widespread circular forms of non-coding RNAs with largely unknown function. Because stimulation of mammary cells with the epidermal growth factor (EGF) leads to dynamic changes in the abundance of both coding and non-coding RNA molecules, and culminates in the acquisition of a robust migratory phenotype, this cellular model might disclose functions of circRNAs. Here we show that circRNAs of EGF-stimulated mammary cells are stably expressed, while mRNAs and micro-RNAs change within minutes. In general, the circRNAs we detected are relatively long-lived and weakly expressed. Interestingly, they are almost ubiquitously co-expressed with the corresponding linear transcripts, and the respective, shared promoter regions are more active compared to genes producing linear isoforms only. These findings imply that altered abundance of circRNAs, unlike changes in the levels of other RNAs, might not play critical roles in signaling cascades and downstream transcriptional networks that rapidly commit cells to specific outcomes. Overall design: Detection of circRNAs from RNA-Seq – triplicate","project":"SRP061405"}
{"number_samples":12,"species":"human","abstract":"MicroRNAs (miRNAs) have been implicated in regulating multiple processes during brain development in various species. However, the function of miRNAs in human brain development remains largely unexplored. Here, we provide a comprehensive analysis of miRNA expression of regionalized neural progenitor cells derived from human embryonic stem cells and human fetal brain. We found mir-92b-3p and mir-130b-5p to be specifically associated with neural progenitors and several miRNAs that display both age-specific and region-specific expression patterns. Among these miRNAs, we identified miR-10 to be specifically expressed in the human hindbrain and spinal cord, while absent from rostral regions. We found that miR-10 regulates a large number of genes enriched for functions including transcription, actin cytoskeleton and ephrin receptor signaling. When overexpressed, miR-10 influences caudalization of human neural progenitors cells. Together, these data confirms a role for miRNAs in establishing different human neural progenitor populations. This data set also provides a comprehensive resource for future studies investigating the functional role of different miRNAs in human brain development. Overall design: Human embryonic stem cells (hESCs) were transduced with lentiviral vectors expressing either miR10a-GFP or miR10b-GFP. The expression of the vectors is Tet-regulated and they will only be expressed in the presence of Doxycycline. In order to detect direct targets of the miR10a and miR10b, we differentiated the trasduced hESCs for 14 days, and added doxycycline to only half of the groups - resulting in groups that are overexpressing miR10a or miR10b and some groups that are not overexpressing these miRNAs.","project":"SRP061416"}
{"number_samples":8,"species":"human","abstract":"We report nuclear receptor Esrrb's responsive genes with or without Esrrb ligand DY131 in DU145 cells. Using Esrrb-null cells, we used RNA-Seq to find Esrrb responsive genes. In addition, we tested DY131-driven Esrrb-dependent genes to test the ligand dependency of Esrrb in regulating gene expression. Overall design: Control vector transfected cells with vehicle treatment, Esrrb expression vector transfected cell with vehicle treatment, control vector transfected cells with DY131 treatment, Esrrb expression vector transfected cell with DY131 treatment.","project":"SRP061425"}
{"number_samples":4,"species":"human","abstract":"Human umbilical vein vascular endothelial cells (HUVECs) are crucial for angiogenesis that benefits functional recovery after cerebral infarction. This study aims to investigate the mechanisms underlying the effects of vascular endothelial growth factor (VEGF) on HUVECs. Overall design: HUVECs were treated with 16 ng/mL VEGF165 for 4 days","project":"SRP061426"}
{"number_samples":4,"species":"human","abstract":"The goals of this study are to analyze the transcriptome profiling (RNA-seq) after PBX3 overexpression in SMMC-7721 cells. Overall design: SMMC-7721 cells were transfected by PBX3 expression lentivirus and blank lentivirus as control. Total RNA was extraced by Qiagen kit and the mRNA profiles were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000.","project":"SRP061455"}
{"number_samples":4,"species":"human","abstract":"CTCF/cohesin play a central role in insulator function and higher-order chromatin organization of mammalian genomes. Recent studies identified a correlation between the orientation of CTCF-binding sites (CBSs) and chromatin loops. To test the functional significance of this observation, we combined CRISPR/Cas9-based genomic-DNA-fragment editing with chromosome-conformation-capture experiments to show that the location and relative orientations of CBSs determine the specificity of long-range chromatin looping in mammalian genomes, using protocadherin (Pcdh) and ß-globin as model genes. Inversion of CBS elements within the Pcdh enhancer reconfigures the topology of chromatin loops between the distal enhancer and target promoters, and alters gene-expression patterns. Thus, although enhancers can function in an orientation-independent manner in reporter assays, in the native chromosome context the orientation of at least some enhancers carrying CBSs can determine both the architecture of topological chromatin domains and enhancer/promoter specificity. The findings reveal how 3D chromosome architecture can be encoded by genome sequence. Overall design: HEC-1B mRNA profiles of HS5-1 Inversion","project":"SRP061462"}
{"number_samples":16,"species":"human","abstract":"Many thousand long non-coding (lnc) RNAs are mapped in the human genome. Time consuming studies using reverse genetic approaches by post-transcriptional knock-down or genetic modification of the locus demonstrated diverse biological functions for a few of these transcripts. The Human Gene Trap Mutant Collection in haploid KBM7 cells is a ready-to-use tool for studying protein-coding gene function. As lncRNAs show remarkable differences in RNA biology compared to protein-coding genes, it is unclear if this gene trap collection is useful for functional analysis of lncRNAs. Here we use the uncharacterized LOC100288798 lncRNA as a model to answer this question. Using public RNA-seq data we show that LOC100288798 is ubiquitously expressed, but inefficiently spliced. The minor spliced LOC100288798 isoforms are exported to the cytoplasm, whereas the major unspliced isoform is nuclear localized. This shows that LOC100288798 RNA biology differs markedly from typical mRNAs. De novo assembly from RNA-seq data suggests that LOC100288798 extends 289kb beyond its annotated 3' end and overlaps the downstream SLC38A4 gene. Three cell lines with independent gene trap insertions in LOC100288798 were available from the KBM7 gene trap collection. RT-qPCR and RNA-seq confirmed successful lncRNA truncation and its extended length. Expression analysis from RNA-seq data shows significant deregulation of 41 protein-coding genes upon LOC100288798 truncation. Our data shows that gene trap collections in human haploid cell lines are useful tools to study lncRNAs, and identifies the previously uncharacterized LOC100288798 as a potential gene regulator. Overall design: We cultured and processed 8 KBM7 cell lines in one batch. These cell lines were: two wild type KBM7 cells (WT2 and WT3), two monoclonal KBM7 cell lines with gene trap cassette insertions outside of the body of LOC100288798 (C1 and C2), two independently obtained KBM7 clones with gene trap cassette insertion 3kb downstream LOC100288798 transcriptional start site (TSS) (3kb1 and 3kb2), one independently obtained KBM7 clone with gene trap cassette insertion 100kb downstream LOC100288798 TSS replicated twice at the thawing step (100kb1 and 100kb2). We isolated total RNA from all th 8 cell lines, applied DNAseI treatment and ribosomal RNA depletion, and thhen prepared strand-specific RNA-seq libraries, which were pooled in equal molarities and sequenced using Illumina HiSeq 2000 (8 pooled samples were sequence on 2 lanes). We performed 50bp single-end RNA-seq. We used these 8 samples (4 untreated: WT2, WT3, C1, C2 and 4 treated:3kb1, 3kb2, 100kbk1, 100kb2) to analyze genome-wide gene deregulation associated with LOC100288798 lncRNA truncation","project":"SRP061522"}
{"number_samples":9,"species":"human","abstract":"SRSF2 is an RNA binding protein that plays important roles in splicing of mRNA precursors. Mutations in SRSF2 are frequently found in patients with myelodysplastic syndromes and certain leukemias, but how they affect SRSF2 function has only begun to be examined. Here we used CRISPR/Cas9 to introduce the P95H mutation to SRSF2 in K562 leukemia cells, generating an isogenic model so that splicing alterations can be attributed solely to mutant SRSF2. We found that SRSF2 (P95H) misregulates 548 splicing events (<1% of total). Of these, 374 involve the inclusion of cassette exons, and the inclusion was either increased (206) or decreased (168). We detected a specific motif (UCCA/UG) enriched in the more included exons and a distinct motif (UGGA/UG) in the more excluded exons. RNA gel shift assays showed that a mutant SRSF2 derivative bound more tightly than its wild-type counterpart to RNA sites containing UCCAG, but less tightly to UGGAG sites. The pattern of exon inclusion or exclusion thus correlated in most cases with stronger or weaker RNA binding, respectively. We further show that the P95H mutation does not affect other functions of SRSF2, i.e., protein-protein interactions with key splicing factors. Our results thus demonstrate that the P95H mutation positively or negatively alters the binding affinity of SRSF2 for cognate RNA sites in target transcripts, leading to misregulation of exon inclusion. Our findings not only shed light on the mechanism of the disease-associated SRSF2 mutation in splicing regulation, but also reveal a group of mis-spliced mRNA isoforms for potential therapeutic targeting. Overall design: Examination of differentially spliced events in K562 CRISPR cell clones (with wild-type or mutant SRSF2) by RNA sequencing","project":"SRP061539"}
{"number_samples":6,"species":"human","abstract":"Constitutively active AR variants are truncated proteins lacking the c-terminal region containing the ligand binding domain (LBD) and the activation function 2 (AF-2). The expression of these AR variants in CRPC was also associated with the resistance to novel therapies such as enzalutamide and abiraterone acetate. These variants are also involved in tumor progression. Overall design: LNCaP cells were transduced with lentival particles for expression of AR-WT or AR-V7 for 72 hours in complete medium containing 10 nM DHT. Libraries were sequenced using the Illumina Hiseq2500 technology The goal of this experiment was to delineate differential gene expression upon the expression of the AR-V7 in LNCaP cells compared with cells expressing the wild-type AR.","project":"SRP061566"}
{"number_samples":28,"species":"human","abstract":"We evaluated changes in mRNA stability and transcription using 4sU metabolic pulse labeling across a four hour time course following activation of Jurkat T cells with PMA and PHA Overall design: Measurement of total mRNA (T) and 4sU labeled mRNA (IP) in three biological replicates at five time points: prior to activation (U) and the first four hours after activation (1-4)","project":"SRP061639"}
{"number_samples":5,"species":"human","abstract":"We differentiated hESC into retinal pigment epithelial cells using two methods (three-dimensional culture and spontaneous differentiation methods). We investigated which kind of RPE cells derived from hESC showed similar gene expression patterns to those of human fetal native RPE.","project":"SRP061670"}
{"number_samples":42,"species":"human","abstract":"Purpose: Genetic and clinical association studies have identified disrupted-in-schizophrenia 1 (DISC1) as a candidate risk gene for major mental illness. DISC1 is interrupted by a balanced chr(1;11) translocation in a Scottish family, in which the translocation predisposes to psychiatric disorders. We investigate the consequences of DISC1 interruption in human neural cells using TALENs or CRISPR-Cas9 to target the DISC1 locus. We sought to compare the gene expression profiles of human neural progenitor cells (NPCs) and neurons with interruption of the DISC1 gene in exon 2 (affecting all known coding transcripts) or exon 8 (near the site of the Scottish translocation, affecting longer transcripts). Methods: Wild-type and DISC1-targeted iPSCs (wild-type = \"WT\", exon 8 single allelic frameshift mutant = \"ex8_wm\", exon 8 biallelic frameshift mutant = \"ex8_mm\", exon 2 biallelic frameshift mutant = \"ex2mm\") were differentiated to NPCs and neurons using an embryoid aggregate method. NPC or neuronal cultures were used for RNA harvest and subsequent paired-end stranded sequencing of >50M reads/sample and 3-6 biological replicates per group. Results: We find that a subset of genes related to neuronal differentiation and development are dysregulated with DISC1 disruption at the NPC timepoint, whereas expression of genes related to neuronal function and signaling are altered at the neuronal timepoint. This study implicates DISC1 as a regulator of neuronal development. Overall design: mRNA profiles of wild-type and DISC1-targeted human iPSC-derived neural progenitor cells (day 17) and neurons (day 50) by paired-end sequencing, with 3-6 biological replicates, using Illumina HiSeq","project":"SRP061682"}
{"number_samples":2,"species":"human","abstract":"Notch pathway antagonists such as gamma-secretase inhibitors (GSI) are being tested in diverse cancers, but exceptional responses have yet to be reported. We describe the case of a patient with relapsed/refractory early-T-cell progenitor acute lymphoblastic leukemia (ETP-ALL) who achieved a complete hematologic response following treatment with the GSI BMS-906024. Whole exome sequencing of leukemic blasts revealed heterozygous gain-of-function driver mutations in NOTCH1, CSF3R, and PTPN11, and a homozygous/hemizygous loss-of-function mutation in DNMT3A. The three gain-of-function mutations were absent from remission marrow cells, but the DNMT3A mutation persisted in heterozygous form in remission marrow, consistent with an origin for the patient’s ETP-ALL from clonal hematopoiesis. Ex vivo culture of ETP-ALL blasts confirmed high levels of activated NOTCH1 that were repressed by GSI treatment, and RNA-Seq documented that GSI downregulated multiple known Notch target genes. Surprisingly, one potential target gene that was unaffected by GSI was MYC, a key Notch target in GSI-sensitive T-ALL of cortical T cell type. H3K27ac superenhancer landscapes near MYC showed a pattern previously reported in acute myeloid leukemia (AML) that is sensitive to BRD4 inhibitors, and in line with this ETP-ALL blasts downregulated MYC in response to the BRD4 inhibitor JQ1. To our knowledge, this is the first example of complete response of a Notch-mutated ETP-ALL to a Notch antagonist and is also the first description of chromatin landscapes associated with ETP-ALL. Our experience suggests that additional attempts to target Notch in Notch-mutated ETP-ALL are merited. Overall design: RNAseq and H3K27Ac ChIP seq of primary leukemic blasts treated in vitro with vehicle control or gamma-secretase inhibitor BMS-096024","project":"SRP061685"}
{"number_samples":6,"species":"human","abstract":"Using UNC0638 and genetic assays to inhibit EHMT1/2 and derepress fetal hemoglobin in adult hematopoietic cells. Overall design: RNA-Seq in primary adult human erythroid cells treated with UNC0638 or the vehicle control (DMSO) in biological triplicates.","project":"SRP061689"}
{"number_samples":12,"species":"human","abstract":"The constitutive androstane receptor (CAR, NR1I3) modulates the transcription of numerous genes involving drug metabolism, energy homeostasis, and cell proliferation. Most functions of CAR however were defined from animal studies. Given the known species difference of CAR and the significant cross-talk between CAR and the pregnane X receptor (PXR), it is extremely difficult to decipher the exact role of human CAR (hCAR) in gene regulation, relying predominantly on pharmacological manipulations.  Here, utilizing a newly generated hCAR-knockout (KO) HepaRG cell line, we carried out RNA-seq analysis of the global transcriptomes in wild-type (WT) and hCAR-KO HepaRG cells treated with CITCO, a selective hCAR agonist, phenobarbital (PB), a dual activator of hCAR and hPXR, or vehicle control. Real-time PCR assays in separate experiments were used to validate RNA-seq findings. Our results indicate that genes encoding drug-metabolizing enzymes are among the main clusters altered by both CITCO and PB. Specifically, CITCO significantly changed the expression of 135 genes in an hCAR-dependent manner, while PB altered the expression of 227 genes in WT cells of which 94 were simultaneously modulated in both cell lines reflecting the dual effects of PB on hCAR/PXR. Notably, we found that many genes promoting cell proliferation and tumorigenesis were up-regulated in hCAR-KO cells, suggesting that hCAR may play an important role in cell growth that differs from mouse CAR. Together, our results reveal both novel and known targets of hCAR and support the role of hCAR in maintaining the homeostasis of metabolism and cell proliferation in the liver. Overall design: Compare the mRNA profiles of HepRG WT and hCAR-KO cells after DMSO, PB and CITCO treatment, each sample was done in duplicate.","project":"SRP061701"}
{"number_samples":111,"species":"human","project":"SRP061801"}
{"number_samples":18,"species":"human","abstract":"Epigenome editing with the CRISPR/Cas9 platform is a promising technology to modulate gene expression to direct cell phenotype and to dissect the causal epigenetic mechanisms that direct gene regulation.  Fusions of the nuclease-inactive dCas9 to the KRAB repressor domain (dCas9-KRAB) can effectively silence target gene expression.  We targeted dCas9-KRAB to the HS2 enhancer, a distal regulatory element that orchestrates the expression of multiple globin genes. Genome-wide analyses demonstrated that localization of dCas9-KRAB to HS2 specifically induced H3K9 tri-methylation (H3K9me3) at that enhancer and reduced the chromatin accessibility of both the enhancer and its promoter targets.  Targeted epigenetic modification of HS2 silenced the expression of multiple globin genes, with minimal off-target changes in gene expression. These results demonstrate that repression mediated by dCas9-KRAB is sufficiently specific to disrupt the activity of individual enhancers via local modification of the epigenome. This approach enables precise modulation of epigenetic function without modifying the underlying genome sequence. Overall design: K562 cells were transduced with in triplicate lentivirus encoding dCas9-KRAB with gRNA targeted to the HS2 globin enhancer.  Cells transduced with dCas9-KRAB without gRNA or dCas9 with gRNA targeted to the HS2 globin enhancer were included as controls.  RNA-seq was used to identify differential expression at on-target and off-target sites.","project":"SRP061840"}
{"number_samples":56,"species":"human","abstract":"Through RNA sequencing of CD4+ Tmemory/effector cells derived from the synovium of JIA patients and healthy controls, we analyzed the JIA gene expression signature. Treatment of autoinflammatory site-derived patient T cells with the BET-inhibitor JQ1 inhibited immune-related super-enhancers and preferentially reduced disease-associated gene expression, including cytokine-related processes. Overall design: RNA-sequencing of CD4+ memory/effector T cells derived from Healthy Controls (HC) and JIA patients upon JQ1 treatment.","project":"SRP061881"}
{"number_samples":8,"species":"human","abstract":"The knowledge of an expression network signature in end-stage heart failure (HF) diseased hearts may offer important insights into the complex pathogenesis of advanced cardiac failure, as well as it may provide potential targets for therapeutic intervention. In this study, the NGS sequencing of RNA (RNA-Seq) method was employed to obtain the whole transcriptome of cardiac tissues from transplant recipients with advanced stage of HF. The analysis of RNA-Seq data presents novel challenges and many methods have been developed for the purpose of mapping reads to genomic features and quantifying gene expression. The main goal of this work was to identify, characterize and catalogue all the transcripts expressed within cardiac tissue and to quantify the differential expression of transcripts in both physio- and pathological conditions through whole transcriptome analyses. Expression levels, differential splicing, allele-specific expression, RNA editing and fusion transcripts constitute important information when comparing samples for disease related studies. Analysis methods for RNA-Seq data are continuing to evolve. Thus, in order to find the best solution for filter generated list of differentially expressed genes, an informatic approach of NOISeq BIO method has been applied in this RNA-Seq analysis. Most of the genes obtained by filtering differentially expressed gene list, have been experimentally validated by Real time RT-PCR. Noteworthy, these findings provide valuable resources for further studies of the molecular mechanisms involved in heart ischemic response thus leading to potential novel biomarkers and targets for therapeutic intervention in the onset and progression of cardiomyopathies. Overall design: Heart biopsies from candidates for solid organ transplantation were collected and their RNA samples were used for high-throughput sequencing purposes. Libraries were sequenced on the Illumina HiSeq2000 NGS platform.","project":"SRP061888"}
{"number_samples":12,"species":"human","abstract":"The aim of the dataset was to identify genome-wide regulators of gene expression in early differentiation of human cord blood derived CD4+ T cells cultured under Th1 (Act+IL12) and Th2 (Act+IL4) polarizing conditions. Overall design: Total RNA from naive CD4+ T cells was compared to total RNA from cells cultured in the following three conditions: activating (antiCD3+antiCD28)+antiIL4+antiIFNG; activating (antiCD3+antiCD28)+IL12+antiIL4; activating (antiCD3+antiCD28) +IL4+antiIFNG. Samples from 3 biological replicates were analysed.","project":"SRP061932"}
{"number_samples":6,"species":"human","abstract":"bone marrow-drived mesenchymal stem cells","project":"SRP061971"}
{"number_samples":9,"species":"human","abstract":"Mutations in the poly(A) ribonuclease (PARN) gene cause telomere diseases including familial idiopathic pulmonary fibrosis (IPF) and dyskeratosis congenita (DC)1,2, but how PARN deficiency impacts telomere maintenance is unclear. Here, using somatic cells and induced pluripotent stem (iPS) cells from DC patients with PARN mutations, we show that PARN is required for the 3' end maturation of the telomerase RNA component (TERC). Patient cells as well as immortalized cells in which PARN is disrupted show decreased levels of TERC. Deep sequencing of TERC RNA 3' termini reveals that PARN is required for removal of posttranscriptionally acquired oligo(A) tails that target nuclear RNAs for degradation. Diminished TERC levels and the increased oligo(A) forms of TERC are normalized by restoring PARN, which is limiting for TERC maturation in cells. Our results reveal a novel role for PARN in the biogenesis of TERC, and provide a mechanism linking PARN mutations to telomere diseases. Overall design: mRNA sequencing of fibroblasts, induced pluripotent stem cells, and 293 cell line.","project":"SRP062010"}
{"number_samples":73,"species":"human","abstract":"Gene expression analysis of purified hematopoietic stem and progenitor cells isolated from low to intermediate risk MDS patients and age-matched normal healthy controls. Overall design: Analysis of lineage associated genes and PCA clustering of populations","project":"SRP062025"}
{"number_samples":45,"species":"human","abstract":"The human neocortex is created from diverse progenitors that are intermixed with multiple cell types in the prenatal germinal zones. These progenitors have been difficult to profile with unbiased transcriptomics since progenitors-particularly radial glia (RG)-are rare cell types, defined by a combination of intracellular markers, position and morphology. To circumvent these problems, we developed a method called FRSCR for transcriptome profiling of individual fixed, stained, and sorted cells. After validation of FRSCR with human embryonic stem cells, we profiled primary human RG that constitute only 1% of the mid-gestation cortex. These data showed that RG could be classified into ventricle zone-enriched RG (vRG) that expressed ANXA1 and CRYAB, and outer subventricular zone-localized RG (oRG) that expressed HOPX. Our study identified the first markers and molecular profiles of vRG and oRG cells, and provides an essential step for understanding molecular networks that control the development and lineage of human neocortical progenitors. Furthermore, FRSCR allows targeted single-cell transcriptomic profiling of many tissues that currently lack live-cell markers. Overall design: 26 Llive and 19 Fixed cultured hESCs were prepared and sequenced using both FRISCR and TritonX-100 Lysis as proof of principal for FRSCR.","project":"SRP062177"}
{"number_samples":20,"species":"human","abstract":"RNA-Seq of Endemic Burkitt Lymphoma","project":"SRP062178"}
{"number_samples":6,"species":"human","abstract":"These RNA-seq data were generated to correlate with genomic interaction data in a related Hi-C analysis.  MCF10A is a normal-like mammary epithelial cell line and MCF7 is a transformed estrogen responsive breast cancer cell line derived from a metastatic site; both are commonly used in models of breast cancer progression.  Analysis revealed a set of genes related to repression of WNT signalling that were both up-regulated in MCF7 and located in genomic regions that had transitioned from closed to open structure in MCF7. Overall design: RNA-seq of MCF10A and MCF7 cells. 3 replicates each.  Sequencing was strand-specific and conducted on ribo-depleted RNA.","project":"SRP062188"}
{"number_samples":6,"species":"human","abstract":"Induced pluripotent stem cells (iPSC) were prepared from multiple subjects with Ataxia-telangiectasia (A-T).  iPSC were prepared from activated T-cells using commercial Sendai virus to deliver reprogramming factors.  ATM protein-expressing and non-expressing cultures would found in multiple sub-lines (sub-lines are generated from distinct founder colonies of cells) from a single subject (Q3).  Genetic analysis determined that all sub-lines originated from the same subject and were likely the product of spontaneous reversion by gene correction.  To compare gene expression profiles, three iPSC samples were assayed: (1) A reverted ATM+/- subline from subject Q3, (2) An ATM-/- subline from subject Q3, and (3) An iPSC line from an unrelated ATM-/- subject, Q1.  Analysis reveals that the majority of significantly-different genes between the unrelated ATM-/- line (Q1SA) and the reverted ATM+/- line (Q3SC) was due to genetic variation between individuals.  A more focused set of contrasting genes could be identified between the isogenic Q3-derived lines (Q3SA, ATM-/-, vs. Q3SC, ATM+/-).  The 206 transcripts that are significantly different point to a differential regulation of p53-associated pathways. Overall design: Total cellular RNA was prepared from each iPSC culture.  Different passage numbers or sister cultures were used as replicates (n=2).","project":"SRP062214"}
{"number_samples":8,"species":"human","abstract":"Small-molecule inhibitors of AKT signaling are being in evaluated in patients with various cancer types, but have so far proven therapeutically disappointing for reasons that remain unclear. Here, we treat cancer cells with sub-therapeutic doses of Akti-1/2, an allosteric small molecule AKT inhibitor, in order to experimentally model pharmacologic inhibition of AKT signaling in vitro. We then apply a combined RNA, protein, and metabolite profiling approach to develop an integrated, multi-scale, molecular snapshot of this “AKTlow” cancer cell state. We find that AKT-inhibited cancer cells suppress thousands of mRNA transcripts, and proteins related to the cell cycle, ribosome, and protein translation. Surprisingly, however, these AKT-inhibited cells simultaneously up-regulate a host of other proteins and metabolites post-transcriptionally, reflecting activation of their endo-vesiculo-membrane system, secretion of inflammatory proteins, and elaboration of extracellular microvesicles. Importantly, these microvesicles enable rapidly proliferating cancer cells of various types to better withstand different stress conditions, including serum deprivation, hypoxia, or cytotoxic chemotherapy in vitro and xenografting in vivo. These findings suggest a model whereby cancer cells experiencing a partial inhibition of AKT signaling may actually promote the survival of neighbors through non-cell autonomous communication. Overall design: Profiles of MCF7 and HCT116 Akti-1/2 treated cells and MCF7 and HCT116 vehicle (i.e. DMSO) treated cells were generated, each in duplicate, using RNA-Seq.","project":"SRP062230"}
{"number_samples":1,"species":"human","abstract":"Characterization of a novel fusion gene EML4-NTRK3 in an aggressive case of congenital fibrosarcoma","project":"SRP062247"}
{"number_samples":4,"species":"human","abstract":"COAD L1","project":"SRP062284"}
{"number_samples":12,"species":"human","abstract":"The goal of this work was to identify all estrogen receptor beta target genes using RNA sequencing in MDA-MB-468 triple negative breast cancer cells engineered with inducible expression of full length estrogen receptor beta. Overall design: MDA-MB-468 breast cancer cells with inducible ERb expression (MDA-468-ERb cells) were treated in triplicate with vehicle (control, no ERb) or doxycycline (plus ERb) for 48 hr prior to treatment with 0.1% DMSO vehicle or 10 nM 17b-estradiol for 4 hr.","project":"SRP062287"}
{"number_samples":8,"species":"human","abstract":"Using RNA-seq to identify genes regulated by dCas9-KRAB mediated enhancer repression in NCI-H2009 lung adenocarcinoma cells Overall design: NCI-H2009 cells were first transfected with dCas9-KRAB fusion and subsequently transfected with no guide RNA (Empty), a guide RNA that has no target in the human genome (Dummy), and two separate guide RNAs (e3 #1 and e3 #2) recognizing the enhancer region of our study.","project":"SRP062332"}
{"number_samples":33,"species":"human","abstract":"RNA sequencing of HeLa cells treated with siRNA against the RNA exosome components hRRP40, hRRP6, hDIS3, and hRRP6/hDIS3 or the splicing inhibitors Isoginkgetin and spliceostatin A, respectively. Overall design: Stranded, ribo-depleted RNA seq profiles of HeLa cells treated with exosome targeting siRNAs or splicing inhibitors using Illumina HiSeq. All experiments were carried out in triplicate starting with independent cell cultures","project":"SRP062389"}
{"number_samples":15,"species":"human","abstract":"Identification of the counterpart protein of Nef during HIV infection","project":"SRP062444"}
{"number_samples":4,"species":"human","abstract":"The goals of this study were to compare the distribution of H3K4me3 before and after methionine restriction to understand how nutrient availability and alterations in methionine metabolism affect the epigenetic state and regulate gene transcription.  To do this we performed ChIP-seq with an H3K4me3 antibody and RNA-seq using PolyA+ mRNA. Overall design: We compared genome-wide H3K4me3 and mRNA profiles generated by Next Generation Sequencing (NGS) between +Methionine and -Methionine conditions after 24 hours in HCT116","project":"SRP062511"}
{"number_samples":6,"species":"human","abstract":"Pioneer transcription factors bind to silent chromatin, and initiate cell fate conversion. One potential pioneer factor, GATA3, is a critical component for multiple cellular programs. GATA3 is of particular interest as it regulates gene expression in breast cancers, and low expression correlates with poor prognosis. Here we demonstrate the pioneering activity of GATA3 utilizing a cellular reprogramming system (the mesenchymal-epithelial transition) in breast cancer cells. During the epithelial transition, GATA3 catalyzes chromatin reprogramming by inducing chromatin opening, active enhancer modifications, and nucleosome remodelling. We determined that the transactivation domain is required for this chromatin reprogramming. Importantly, a mutant lacking the transactivation domain possessed the chromatin binding ability but failed to create open chromatin, suggesting binding alone is not sufficient to induce open chromatin. These data illustrate a fundamental mechanism of GATA3-mediated gene regulation, and provide evidence for a pre-engagement state of closed chromatin bound by a pioneer factor. Overall design: Genome-wide mapping of chromatin state in GATA3-mediated mesenchymal-epithelial transition","project":"SRP062544"}
{"number_samples":6,"species":"human","abstract":"SRSF1-regulated splicing events in breast cancer","project":"SRP062609"}
{"number_samples":6,"species":"human","abstract":"Glioblastomas display hierarchies with self-renewing cancer stem-like cells (CSCs). RNA sequencing and enhancer mapping revealed regulatory programs unique to CSCs causing upregulation of the iron transporter transferrin, the top differentially expressed gene compared to tissue-specific progenitors. Direct interrogation of iron uptake demonstrated CSCs potently extract iron from the microenvironment more effectively than other tumor cells. Systematic interrogation of iron flux determined that CSCs preferentially require transferrin receptor and ferritin - two core iron regulators - to propagate and form tumors in vivo.  Depleting ferritin disrupted CSC mitotic progression, through the STAT3-FoxM1 regulatory axis, revealing an iron-regulated CSC pathway. Iron is a unique, primordial metal fundamental for earliest life forms, and on which CSCs have an epigenetically programmed, targetable dependence. Overall design: RNA-seq of primary patient-derived GBM cancer stem cells and normal human neural progenitor cells","project":"SRP062617"}
{"number_samples":7,"species":"human","abstract":"Chromatin immunoprecipitation followed by next-generation DNA sequencing (ChIP-seq) is a widely used technique for identifying transcription factor (TF) binding events throughout an entire genome. However, ChIP-seq is limited by the availability of suitable ChIP-seq grade  antibodies, and the vast majority of commercially available antibodies fail to generate usable  datasets. To ameliorate these technical obstacles, we present a robust methodological  approach for performing ChIP-seq through epitope tagging of endogenous TFs. We used Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9-based genome  editing technology to develop CRISPR Epitope Tagging ChIP-seq (CETCh-seq) of DNA-binding  proteins. We assessed the feasibility of CETCh-seq by tagging several TFs spanning a wide  range of endogenous expression levels in the hepatocellular carcinoma cell line HepG2. Our  data exhibit strong correlations between both replicate types as well as with standard ChIP-seq  approaches that use TF antibodies. Notably, we also observed minimal changes to the cellular  transcriptome and to the expression of the tagged TF. To examine the robustness of our  technique, we further performed CETCh-seq in the breast adenocarcinoma cell line MCF7 as  well as mouse embryonic stem cells and observed similarly high correlations. Collectively,  these data highlight the applicability of CETCh-seq to accurately define the genome-wide  binding profiles of DNA-binding proteins, allowing for a straightforward methodology to  potentially assay the complete repertoire of TFs, including the large fraction for which ChIPquality  antibodies are not available. Overall design: CRISPR/Cas9 mediated epitope tagging of transcription factors in human cell types","project":"SRP062873"}
{"number_samples":4,"species":"human","abstract":"The Swedish mutation (K595N/M596L) of amyloid precursor protein (APP-sw) has been known to increase abnormal cleavage of cellular APP by Beta-secretase (BACE), which causes tau protein hyperphosphorylation and early-onset Alzheimer's disease (AD). Here, we analyzed the effect of APP-sw in global gene expression using deep transcriptome sequencing technique. We found 283 genes were down-regulated and 348 genes were up-regulated in APP-sw expressing H4-sw cells compared to H4 wild-type cells from a total of approximately 74 million reads of 38 base pairs from each transcriptome. Two independent mechanisms such as kinase and phosphatase signaling cascades leading hyperphosphorylation of tau protein were regulated by the expression of APP-sw. Expressions of catalytic subunit as well as several regulatory subunits of protein phosphatases 2A were decreased. In contrast, expressions of tau-phosphorylating glycogen synthase kinase 3ß (GSK-3ß), cyclin dependent kinase 5 (CDK5), and cAMP-dependent protein kinase A (PKA) catalytic subunit were increased. Moreover, the expression of AD-relatedAquaporin 1 and presenilin 2 expression was regulated by APP-sw. Taken together, we propose that the expression of APP-sw modulates global gene expression directed to AD pathogenesis.","project":"SRP062936"}
{"number_samples":8,"species":"human","abstract":"Despite many years of study of inversions, very little is known about their functional consequences, especially in humans. A common hypothesis is that the selective value of inversions stems in part from their effects on nearby genes, although evidences of this in natural populations are almost nonexistent. Here we present a global analysis of a new 415-kb polymorphic inversion that is among the longest ones found in humans and is the first with clear position effects. This inversion is located in chromosome 19 and has been generated by non-homologous end joining between blocks of transposable elements with low identity. PCR genotyping in 541 individuals from eight different human populations allowed the detection of tag SNPs and inversion genotyping in multiple worldwide populations, showing that the inverted allele is mainly found in East-Asia with an average frequency of 4.7%. Interestingly, one of the breakpoints disrupts the transcription factor gene ZNF257, causing a significant reduction in the total expression level of this gene in lymphoblastoid cell lines. RNA-Seq analysis of the effects of this expression change in standard homozygotes and inversion heterozygotes revealed distinct expression patterns that were validated by quantitative RT-PCR. Moreover, we have found a new fusion transcript that is generated exclusively from inverted chromosomes around one of the breakpoints. Finally, by the analysis of the associated nucleotide variation, we have estimated that the inversion was generated approximately 43,450 years ago and, while a neutral evolution cannot be ruled out, its current frequencies are more consistent with those expected for a deleterious variant, although no significant association with phenotypic traits has been found so far.","project":"SRP062950"}
{"number_samples":10,"species":"human","abstract":"RNA-seq of Ro60-null GM12878 cell lines in order to determine the gene expression changes resulting from loss of Ro60. Overall design: 3 separate clones of Ro60(Trove2)-null cells derived from zinc finger nuclease targeting of exon 2, two wildtype biological replicates, +/- IFNa for 6 hours.","project":"SRP062956"}
{"number_samples":6,"species":"human","abstract":"Determine the gene expression profile in peripheral blood monocytes isolated from 3 healthy donors +/- 6 hours of interferon-alpha treatment. Overall design: 3 healthy donor PBMCs +/- interferon-alpha.","project":"SRP062958"}
{"number_samples":117,"species":"human","abstract":"RNA-seq of systemic lupus erythematosus (SLE) whole blood and healthy controls to determine the gene expression changes in these patients. Overall design: RNA-seq of PAXgene blood from SLE and healthy donors.","project":"SRP062966"}
{"number_samples":26,"species":"human","abstract":"The goal of this study is to analyzed transcriptome changes caused by POLA1 deficiency. Our data represents the first detailed analysis of molecular basis of XLPDR syndrome. We report than POLA1 deficiency leads to over-activation of IRF and NF-kB pathways with overexpression of typical markers of autoimmune syndromes. Overall design: Wild type and XLPDR-derived dermal fibroblasts are analyzed under non-stimulated (basal) conditions, after TNF treatment (2 and 12 h, 1000 U/mL), and poly(dA:dT) stimulation (16h, 1 mkg/mL). Obtained data were confirmed using the cellular model of XLPDR - normal dermal fibroblasts pretreated with control or anti-POLA1 siRNA and stimulated in analogous way.","project":"SRP063059"}
{"number_samples":15,"species":"human","abstract":"Epidermal Growth Factor Receptor (EGFR) gene amplification and mutations are the most common oncogenic events in Glioblastoma (GBM), but the mechanisms by which they promote aggressive tumor growth are not well understood. Here, through integrated epigenome and transcriptome analyses of cell lines, genotyped clinical samples and TCGA data, we show that EGFR mutations remodel the activated enhancer landscape of GBM, promoting tumorigenesis through a SOX9 and FOXG1-dependent transcriptional regulatory network in vitro and in vivo. The most common EGFR mutation, EGFRvIII, sensitizes GBM cells to the BET-bromodomain inhibitor JQ1 in a SOX9, FOXG1-dependent manner. These results identify the role of transcriptional/epigenetic remodeling in EGFR-dependent pathogenesis and suggest a mechanistic basis for epigenetic therapy. Overall design: ChIP-Seq for H3K27ac, H3K4me1, and H3K4me3, and RNA-seq for Glioblastoma (GBM) cells and/or tissues with or without EGFRvIII mutation.","project":"SRP063070"}
{"number_samples":26,"species":"human","abstract":"Genome-wide association studies (GWAS) have identified single nucleotide polymorphisms (SNPs) associated with increased risk for colorectal cancer (CRC). A molecular understanding of the functional consequences of this genetic variation is complicated because most GWAS SNPs are located in non-coding regions. After identifying H3K27Ac peaks in HCT116 colon cancer cells that harbor SNPs associated with an increased risk for CRC, we used CRISPR/Cas9 nuclease to delete 2 CRC risk-associated H3K27Ac peaks (E7 and E24), a peak that is not associated with CRC risk (18qE) and a region that doesn?t have H3K27Ac peak (18qNE) from HCT116 cells and analyzed effects on the transcriptome and epigenome. We also deleted E7 region from HEK 293 cells and analyzed effects on the transcriptome of HEK 293. We also confirmed the physical interaction between enhancers of our interest and their putative target genes. Overall design: Analysis of RNA-seq data and ChIP-seq between control clones and enhancer deleted clones in HCT116 cell. For Control, gRNA empty vectorplasmid was transfected with Cas9-GFP. For Deletion, gRNAs that have enhancer target sequences were transfected along with Cas9-GFP. Cells with high GFP expression were identified using fluorescence-activated cell sorting. Sorted cells were plated into individual wells of a 24 well plate and then re-plated as single cells in 10cm dishes and subsequently expanded for further analyses. Samples used in this study were clonal populations. Interaction profiling of enhancers of interest with putative target genes using 4C-seq","project":"SRP063339"}
{"number_samples":6,"species":"human","abstract":"Guillain-Barré syndrome (GBS) is an immune-mediated peripheral neuropathy that debilitates the voluntary and autonomous response of the patient. In this study the transcriptome of peripheral blood mononuclear cells from a GBS patient and her healthy twin were compared to discover possible correlates of disease progression and recovery. Overall design: Blood samples were collected simultaneously from the Guillain-Barré patient (A) and from her control healthy twin (B) at three different time points during disease progression from hospitalization in the intensive care unit (T1), passing to intermediate care (T2),  and at conclusion of locomotion rehabilitation program when the patient was close to abandon the hospital (T3).","project":"SRP063363"}
{"number_samples":4,"species":"human","abstract":"In order to globally identify targets of the nonsense-mediated mRNA decay pathway (NMD), RNA-seq was performed on HeLa cells where NMD was inhibited by shRNA knockdown of UPF1 and on control cells (scrambled shRNA).","project":"SRP063462"}
{"number_samples":66,"species":"human","abstract":"Recurrent mutations in the spliceosome are observed in several human cancers but their functional and therapeutic significance remain elusive. SF3B1, the most frequently mutated component of the spliceosome in cancer, is involved in the recognition of the branch point sequence (BPS) during selection of the 3’ splice site (ss) in RNA splicing. Here, we report that common and tumor-specific splicing aberrations are induced by SF3B1 mutations and establish aberrant 3’ ss selection as the most frequent splicing defect. Strikingly, mutant SF3B1 utilizes a BPS that differs from that used by wild-type SF3B1 and requires the canonical 3’ ss to enable aberrant splicing during the second step. Approximately 50% of the aberrantly spliced mRNAs are subjected to nonsense-mediated decay resulting in downregulation of gene and protein expression. These findings ascribe functional significance to the consequences of SF3B1 mutations in cancer. Overall design: 72 samples, including two sets of patient data and cell lines with two additional technical replicates each","project":"SRP063493"}
{"number_samples":73,"species":"human","abstract":"RNA sequencing was performed on RNA isolated from baseline biopsies from UC patients enrolled in the Phase II EUCALYPTUS study of etrolizumab. Gene expression differences were identified in a subset of anti-TNF naïve patients that achieved clinical remission at 10 weeks in response to etrolizumab. Overall design: Baseline colonic biopsies from UC patients treated with etrolizumab were sequenced by the Illumina HiSeq 2000 Sequencing System.","project":"SRP063496"}
{"number_samples":27,"species":"human","abstract":"Mycobacterium abscessus is an emerging pathogen causing pulmonary infections in those with inflammatory lung disorders, such as Cystic Fibrosis (CF), and is associated with the highest fatality rate among rapidly growing mycobacteria (RGM). Phenotypically, MAB manifests as either a Smooth (MAB-S) or a Rough (MAB-R) morphotype, which differ in their levels of cell wall glycopeptidolipids (GPLs) and in their pathogenicity in vivo. As one of the primary immune cells encountered by MAB, we sought to examine the early transcriptional events within macrophages, following infection with both MAB-S or MAB-R. We sampled the small RNA (sRNA) transcriptome of THP-1-derived macrophages infected with both MAB-R and MAB-S at 1, 4 and 24 hours post-infection (hpi) using RNA-seq. MAB-S elicited a more robust transcriptional response at the miRNA level, reflecting higher cytokine levels in culture supernatants. However, and a direct comparison identified no differentially expressed miRNAs between MAB-R- and MAB-S-infected cells. Most of the induced miRNAs have previously been associated with mycobacterial infection and overall miRNA expression patterns were similarly highly correlated between the morphotypes. Overall design: THP-1-derived macrophages were infected in parallel with the MAB-R and MAB-S morphotypes. Poly-A selected RNAs were purified and sequenced at 1, 4 and 24 hours post-infection, and compared with uninfected controls.","project":"SRP063500"}
{"number_samples":5,"species":"human","abstract":"MIcrophthalmia-associated Transcription Factor (MITF) regulates melanocyte and melanoma physiology. ShRNA-mediated silencing of the NURF subunit BPTF revealed its essential role in several melanoma cell lines and in untransformed melanocytes in vitro. Comparative RNA-seq shows that MITF and BPTF co-regulate overlapping gene expression programs in cell lines in vitro. Somatic and specific inactivation of Bptf in developing murine melanoblasts in vivo shows that Bptf regulates their proliferation, migration and morphology. Once born, Bptf-mutant mice display premature greying where the second post-natal coat is white. This second coat is normally pigmented by differentiated melanocytes derived from the adult melanocyte stem cell (MSC) population that is stimulated to proliferate and differentiate at anagen. An MSC population is established and maintained throughout the life of the Bptf- mutant mice, but these MSCs are abnormal and at anagen, give rise to reduced numbers of transient amplifying cells (TACs) that do not express melanocyte markers and fail to differentiate into mature melanin producing melanocytes. MSCs display a transcriptionally repressed chromatin state and Bptf is essential for reactivation of the melanocyte gene expression program at anagen, the subsequent normal proliferation of TACs and their differentiation into mature melanocytes. Overall design: 5 samples corresponding to mRNA profiles of 501Mel and Hermes3A after BPTF shRNA-mediated knockdown were generated by deep sequencing in triplicate (Hermes 3A) or duplicate (501Mel), using HiSeq2500.","project":"SRP063573"}
{"number_samples":3,"species":"human","abstract":"To profile gene expression affected by AKT3 p.E17K expression in human neural progenitor cells","project":"SRP063581"}
{"number_samples":9,"species":"human","abstract":"Toca 511 is a modified Retroviral Replicating Vector based on Moloney g-retrovirus with an amphotropic envelope. As an investigational cancer treatment, Toca 511 preferentially infects cancer cells without direct cell lysis and encodes an enhanced yeast cytosine deaminase that converts the antifungal drug 5-fluorocytosine to the anticancer drug, 5-fluorouracil. A panel of established human cancers cell lines, derived from glioblastoma, colon, and breast cancer tissue was used to evaluate parameters critical for effective anticancer activity. As part of these analyses, we profiled relative mRNA levels across these cell lines via RNA sequencing. Overall design: mRNA expression profiles across nine human cancer cell lines.","project":"SRP063620"}
{"number_samples":12,"species":"human","abstract":"Here, we describe a strategy in which candidate functional risk variants are evaluated using genome-editing to create isogenic cell lines representing all possible genotypes. We created a panel of isogenic 22Rv1 prostate cancer cell lines representing all three rs339331 genotypes (TT, TC, CC). The risk allele causally increased transcription of the RFX6 gene, increased HOXB13 binding, and increased deposition of the enhancer-associated H3K4me2 histone mark. The cell lines also differed in cellular morphology: TT cells were more adhesive than the CC cells consistent with a cancer-related phenotype. Pathway analysis of differentially expressed genes between the CC and TTlines was predicted to be androgen regulated, a key pathway in prostatecancer biology.","project":"SRP063631"}
{"number_samples":3,"species":"human","abstract":"The RNA-seq results reveal TMPyP4 and TPyP4 could promote cancer cell migration. Overall design: Examination of the gene expression of the following cells:A549-untreated and TMPyP4-treated or TPyP4-treated.","project":"SRP063658"}
{"number_samples":15,"species":"human","abstract":"Purpose: To compare the transcriptome profiles (RNA-seq) of cultured human epididymis cells and tissue from the caput, corpus and cauda regions of the human epididymis. Methods: Human epididymis tissue was obtained with Institutional Review Board approval from 3 patients (UC05, UC06, UC09, range: 22 - 36 years) undergoing inguinal radical orchiectomy for a clinical diagnosis of testicular cancer. None of the epididymides had extension of the testicular cancer. The three anatomical regions: caput, corpus and cauda, were separated and segments of each snap frozen. Adult human epididymis epithelial (HEE) cultures were also established from  tissue. RNA was extracted from both tissue and cultured HEE cells and RNA-seq libraries prepared (TruSeq RNA Sample Preparation Kit v2, Low-Throughput protocol, Illumina). Libraries were sequenced on Illumina HiSeq2500 machines. Data were analyzed using TopHat and Cufflinks. Results: Libraries generated ~19-39 million reads per library from the cells (95-99% mapping to the human genome) and ~14-39 million reads from the tissue samples (84-99% mapped). Raw reads were aligned to the genome with Tophat and gene expression values were processed using Cufflinks as Fragments Per Kilobase per Million mapped fragments (FPKM). FPKM values were subject to principle component analysis, which revealed that though caput, corpus and cauda cell samples respectively from UC05, UC06 and UC09 clustered together. RNA-seq data from the 3 biological replicas (UC05, UC06 and UC09) of caput, corpus and cauda were pooled for further analysis. Cufflinks was used to determine differentially expressed genes (DEGs) between caput, corpus and cauda cells, combined from the 3 donors. The gene expression profiles of corpus and cauda are remarkably similar and both differ from the caput to a similar degree. We identified ~40 genes differentially expressed between corpus and cauda and more than 1600 DEGs between caput and cauda. The DEGs for each comparison (caput and corpus/cauda) were analysed using a gene ontology process enrichment analysis (DAVID, Huang et al., NAR 2009;37:1-13, Huang et al., 2009 Nat Prot 4:44-57). Conclusions: Here we describe an in depth analysis of the gene expression repertoire of primary cultures of epithelial cells and intact tissues from each region of the adult human epididymis. These data will be valuable to decipher pathways of normal epididymis function and aspects of epididymis disease that cause male infertility. Overall design: RNA-seq was performed on libraries generated from caput, corpus and cauda-derived cultured cells (passage 2 or 3) from 3 donors and on caput, corpus and cauda tissue from 2 of the same donors. Donor age range: 22 - 36 years.","project":"SRP063661"}
{"number_samples":7,"species":"human","abstract":"Neuronal migration defects (NMDs) are among the most common and severe brain abnormalities in humans. Lack of disease models in mice or in human cells has hampered the identification of underlying mechanisms. From patients with severe NMDs we generated iPSCs then differentiated neural progenitor cells (NPCs). On artificial extracellular matrix, patient-derived neuronal cells showed defective migration and impaired neurite outgrowth. From a cohort of 107 families with NMDs, sequencing identified two homozygous C-terminal truncating mutations in CTNNA2, encoding aN-catenin, one of three paralogues of the a-catenin family, involved in epithelial integrity and cell polarity. Patient-derived or CRISPR-targeted CTNNA2- mutant neuronal cells showed defective migration and neurite stability. Recombinant aN-catenin was sufficient to bundle purified actin and to suppress the actin-branching activity of ARP2/3. Small molecule inhibitors of ARP2/3 rescued the CTNNA2 neurite defect. Thus, disease modeling in human cells could be used to understand NMD pathogenesis and develop treatments for associated disorders. Overall design: 2 biological replicates per individual (2 iPSC clone differentiations), excluding 1263A, which has one sample","project":"SRP063669"}
{"number_samples":3,"species":"human","abstract":"Discordant growth is a common complication of monochorionic/diamniotic pregnancies; in approximately 50% of cases, the cause is unknown. The case presented here suggests that discordant growth of monozygotic twins could start during preimplantation development. Two inner cell masses (ICMs) within the same blastocyst may originate in uneven splitting of a single “parental” ICM or the two ICMs may be formed independently de novo. We studied the transcriptomes of two morphologically distinct ICMs within a single blastocyst, using high-resolution RNA sequencing. The data indicated that the two ICM were at different stages of development; one was in the earliest stages of lineage commitment, whilst the other had already differentiated into epiblast and primitive endoderm. IGF1-mediated signalling is likely to have a key role in ICM growth, and to be the major driver behind these differences.The expanded Day 6 blastocyst, donated for research, displayed two distinct inner cell masses (ICM) four hours post-thawing. Single cells and cell borders were clearly visible in ICM1, whereas the cells in ICM2 were more tightly packed and no single cell could be distinguished indicating that ICM2 is either better quality or more advanced than ICM1. To explore further this morphological discrepancy, we separated ICM1 and ICM2 from TE and by NGS evaluated the transcriptome of each of the three fractions.","project":"SRP063754"}
{"number_samples":16,"species":"human","abstract":"The etiology of fibrolamellar hepatocellular carcinoma (FL-HCC), a liver cancer occurring increasingly in children to young adults, is poorly understood. We performed high-throughput sequencing on mRNA isolated from a newly developed model of FL-HCC and 4 different maturational lineages of the hepatobiliary system. Our results indicate that the transcriptome of FL-HCC is most similar to that of biliary tree stem cells. Overall design: mRNA profiles of a newly developed model of FL-HCC and 4 maturational stages of the human hepatobiliary system (biliary tree stem cells, hepatic stem cells, hepatoblasts, and adult hepatocytes) were generated by high throughput sequencing using the Illumina HiSeq 2500.","project":"SRP063834"}
{"number_samples":12,"species":"human","abstract":"We report abonormally expressed genes of idiopathic interstitial pneumonia by RNA-seq analysis. BMP3 was found down-regulated in idiopathic intestital pneumonia patients, and it closely correlated with pathogenesis of disease. The role of BMP3 in advancement of idiopathic interstitial pneumonia was verified by a series of experiments. Overall design: Examination of differentially expressed genes in idiopathic interstitial pneumonia. Verification the function of selected gene in pathogenesis of idiopathic interstitial pneumonia in vivo and invitro.","project":"SRP063838"}
{"number_samples":121,"species":"human","abstract":"Clear cell renal cell carcinoma (ccRCC) initiated from the renal epithelium is the most prevalent histological type of adult kidney cancers. Dissecting intratumoral heterogeneity (ITH) of ccRCC has leveraged to extend our knowledge on how primary tumors harboring driver mutations evolve and spread to other sites. The cellular fractions within and across the primary (pRCC) and metastatic RCC (mRCC) are heterogeneous in both their genetic and biological features determining the variability in clinical aggressiveness and sensitivity to the therapy. To achieve sustainable therapeutic benefit with targeted agents in mRCC, the effective target should focus on signaling pathways that are related to driver mutations occurred early in the clonal evolution of the disease and thus should be common to primary tumor and metastatic sites. Considering that extensive genetic heterogeneity may result in drug response variability among patients and treatment resistance, the tailored strategies for metastatic RCC is urgently needed. Here, we analyze single-cell RNA-seq (scRNA-seq) data from a matched primary RCC (pRCC) and lung metastasis (mRCC) to dissect ITH at the highest resolution to date with the objective of discovering the better therapeutic regimen. Overall design: In order to identify successful clonal propagation from patient to PDX samples and understand pathogenesis from primary to metastatic RCC, we performed whole-exome sequencing (WES, n=4) and matched aCGH (n=4) on bulk tumor samples. And we utilized single-cell RNA sequencing (scRNA-seq) to model and dissect functional heterogeneity acroass primary and metastatic RCC tumors. We checked whether of capturing live one cell, not more cells, in microfluidics by fluorescent microscopic observation. To construct RNA sequencing libraries, we performed further quality controls including adequate quantities and qualities of amplified transcriptomes respectively from single cells. Tumor cells from the parental mRCC (n=34), PDX-mRCC (n=36) and PDX-pRCC (n=46) were finally analyzed in this study after filtering out poor quality cells.","project":"SRP063840"}
{"number_samples":32,"species":"human","abstract":"The specialized corneal epithelium requires differentiated properties, specific for its role at the anterior surface of the eye, thus tight maintenance of the differentiated qualities of the corneal epithelial is essential.  Our studies have focused on pinin (PNN), an exon junction component (EJC) that has dramatic implications on corneal epithelial cell differentiation and may act as a stabilizer of the corneal epithelial cell phenotype. Our studies revealed that PNN is involved in both transcriptional repression complexes and the spliceosomal complexes, placing PNN at the fulcrum between chromatin and mRNA splicing.  Transcriptome analysis of PNN-knockdown cells revealed clear and reproducible alterations in transcript profiles and splicing patterns of a subset of genes that would significantly impact the epithelial cell phenotype.  Here, we further investigate PNN’s role in the regulation of gene expression and alternative splicing (AS) in a corneal epithelial context.  We used human corneal epithelial cells (HCET cells) that carry doxycycline-inducible PNN-knockdown shRNA vector and performed RNA-seq to determine differential gene expression and differential AS events. Multiple genes and AS events were identified as differentially expressed between PNN-knockdown and controls cells. Genes up-regulated by PNN-knockdown included a large proportion of genes that are associated with processes associated with enhanced cell migration and ECM remodeling including: MMPs, ADAMs, HAS2, LAMA3, CXCRs and UNC5C.  Genes down-regulated in response to PNN depletion included: IGFBP5. FGD3, FGFR2, PAX6, RARG and SOX10. AS events in PNN compared to controls was also more likely to be detected, and uregulated in PNN-knockdowns.  In particular, 60% of exon skipping events detected in only one condition were detected in PNN-knockdowns and of the shared exon skipping events, 92% of those differentially expressed were more frequent in the PNN-knockdown. This suggests that in the absence of PNN the epithelial cells are dramatically transformed in the amount and composition of isoforms and that PNN plays a crucial role in the selection of which isoforms differentiating cells produce.   Many of the genes affected by PNN-knockdown are known to affect epithelial phenotype. This window into the complexity of RNA splicing in the corneal epithelium implies that PNN exerts broad influence over the regulation and maintenance of epithelial cell phenotype. Overall design: We used HCET cells that carry doxycycline-inducible PNN knockdown shRNA vector and performed RNA-seq to determine differential gene expression and differential alternative splicing events.","project":"SRP063860"}
{"number_samples":73,"species":"human","abstract":"The equivalency of human induced pluripotent stem cells (hiPSCs) with human embryonic stem cells (hESCs) remains controversial. Here, we devised a strategy to assess the contribution of clonal growth, reprogramming method and genetic background to transcriptional patterns in hESCs and hiPSCs. Surprisingly, transcriptional variation originating from two different genetic backgrounds was dominant over variation due to the reprogramming method or cell type of origin of pluripotent cell lines. Moreover, the few differences we detected between isogenic hESCs and hiPSCs neither predicted functional outcome, nor distinguished an independently derived, larger set of unmatched hESC/hiPSC lines. We conclude that hESCs and hiPSCs are transcriptionally and functionally highly similar and cannot be distinguished by a consistent gene expression signature. Our data further imply that genetic background variation is a major confounding factor for transcriptional comparisons of pluripotent cell lines, explaining some of the previously observed expression differences between unmatched hESCs and hiPSCs. Overall design: Expression profiling of human embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs) and fibroblasts, mostly in triplicates.","project":"SRP063867"}
{"number_samples":17,"species":"human","abstract":"Inhibition of H3K27 methyltransferase EZH2 enhances osteogenic commitment of human mesenchymal progenitors and Ezh2 inactivation in mouse calvarial cells induces a post-proliferative state concomitant with increased production of a bone-related mineralizing extra-cellular matrix. Overall design: Expression of genes of interest, including Ezh2, was assessed during osteogenic differentiation of human adipose-derived mesenchymal (AMSCs) on plastic (2D) and on porous-sintered titanium discs (3D). We also assessed gene expression when AMSCs were differentiated into the adipogenic lineage. In addition, the effect of GSK126 on osteogenic differentiation of AMSCs was also evaluated. Finally, we compared gene expression in calvarial bones obtained from wild type mice and mice lacking functional Ezh2 in the mesenchymal lineage (Prrx1-Cre, 3 day old female mice).","project":"SRP063889"}
{"number_samples":6,"species":"human","abstract":"We sequenced mRNA from 6 human cell lines stably over-expressed specific gene or empty vector, and searched for differently expressed genes after gene over-expression as compared to empty vector. Overall design: mRNA levels were compared as following pairs: U2OS+hTERT vs U2OS+vector; U2OS+hTERTmut vs U2OS+vector; VA-13+hTERT vs VA-13+vector; VA-13+hTERTmut vs VA-13+vector.","project":"SRP063948"}
{"number_samples":18,"species":"human","abstract":"RNAseq is performed (50bp single end reads) on HT-29 and HCT-116 cell lines utilizing two independent shRNAs against BRD4 and a non-targeting control shRNA (NTC) Overall design: Examination of transcriptomic changes after knockdown of BRD4","project":"SRP063978"}
{"number_samples":36,"species":"human","abstract":"RNAseq is performed (50bp single end reads) on SW480, HT-29, HCT-15, HCT-116, COLO 205, and COLO 320 cell lines after DMSO or JQ1 treatment Overall design: Examination of transcriptomic changes after JQ1 treatment","project":"SRP063980"}
{"number_samples":12,"species":"human","abstract":"RNA-seq of liver samples from biliary atresia patients and controls","project":"SRP064138"}
{"number_samples":2,"species":"human","abstract":"Universal human reference RNA (UHRR) was used for library construction by RNaseIII and an alternative library preparation protocol based on mRNAs chemical fragmentation for Proton sequencing.","project":"SRP064142"}
{"number_samples":45,"species":"human","abstract":"The Microprocessor complex (DGCR8/Drosha) is required for microRNA (miRNA) biogenesis but also binds and regulates the stability of several types of cellular RNAs. Of particular interest, DGCR8 controls the stability of mature small nucleolar RNA (snoRNA) transcripts independently of Drosha, suggesting the existence of alternative DGCR8 complex/es with other nucleases to process a variety of cellular RNAs. Here, we found that DGCR8 co-purifies with subunits of the nuclear exosome, preferentially associating with its hRRP6-containing nucleolar form. Importantly, we demonstrate that DGCR8 is essential for the recruitment of the exosome to snoRNAs and to human telomerase RNA. In addition, we show that the DGCR8/exosome complex controls the stability of the human telomerase RNA component (hTR/TERC). Altogether, these data suggests that DGCR8 acts as a novel adaptor to recruit the exosome complex to structured RNAs and induce their degradation. Overall design: [i] Examination of the RNA binding profile of hRRP6 (also known as EXOSC10) via iCLIP. [ii] HeLa cells were transiently depleted of hRRP6 or DGCR8 using siRNAs. For a control an non-targetting (siNon) siRNA was used. Three biological replicates of each samples were sent for RNA sequencing.","project":"SRP064143"}
{"number_samples":82,"species":"human","abstract":"Large-scale genomic studies have identified multiple somatic aberrations in breast cancer, including copy number alterations, translocations, and point mutations. Still, identifying causal variants and emergent vulnerabilities that arise as a consequence of genetic alterations remain major challenges. We performed whole genome shRNA “dropout screens” on 77 breast cancer cell lines. Using a new hierarchical linear regression algorithm to score our screen results and integrate them with accompanying detailed genetic and proteomic information, we identify novel vulnerabilities in breast cancer, including new candidate “drivers,” and reveal general functional genomic properties of cancer cells. Comparisons of gene essentiality with drug sensitivity data suggest potential resistance mechanisms, novel effects of existing anti-cancer drugs, and new opportunities for combination therapy.  Finally, we demonstrate the utility of this large dataset by identifying BRD4 as a potential target in luminal breast cancer, and PIK3CA mutations as a resistance determinant for BET-inhibitors. Additional formatted data can be found at http://neellab.github.io/bfg/. Code and tutorials for the siMEM algorithm can be found at http://neellab.github.io/simem/. Overall design: RNA-Seq expression profiling of 82 breast cancer cell lines without replicates or control samples","project":"SRP064259"}
{"number_samples":32,"species":"human","abstract":"The present study used RNA sequencing to identify the transcriptional landscape of two human neural progenitor cell lines, SK-N-SH and ReNcell CX, as they differentiate into human cortical projection neurons. These cells were collected at two time-points of differentiation (24 h and 72h post transfection with an empty pIRES2-AcGFP vector).","project":"SRP064264"}
{"number_samples":1,"species":"human","abstract":"To determine the extent to which psychoactive pharmaceuticals altered expression of neuronal genes associated with neurological disorders (including autism) in humans, we analyzed the gene expression profiles with respect to neuronal systems in human neuroblastoma cell cultures. We used valproate as a positive control in this study as it has been found strongly associated with Autism Spectrum Disorders (ASD). Identifying altered gene expression profiles in treated cells and finding a pattern similar to valproate-induced gene expressions would reveal the extent to which psychoactive pharmaceuticals at very low concentrations could induce gene expression potentially associated with neurological disorders such as ASD","project":"SRP064270"}
{"number_samples":6,"species":"human","abstract":"Covalently closed circular RNA molecules (circRNAs) have recently emerged as a class of RNA isoforms with widespread and tissue specific expression across animals, oftentimes independent of the corresponding linear mRNAs. circRNAs are remarkably stable and sometimes highly expressed molecules. Here, we sequenced RNA in human peripheral whole blood to determine the potential of circRNAs as biomarkers in an easily accessible body fluid. We report the reproducible detection of thousands of circRNAs. Importantly, we observed that hundreds of circRNAs are much higher expressed than corresponding linear mRNAs. Thus, circRNA expression in human blood reveals and quantifies the activity of hundreds of coding genes not accessible by classical mRNA specific assays. Our findings suggest that circRNAs could be used as biomarker molecules in standard clinical blood samples. Overall design: Sequencing of blood RNA from five healthy individuals (biological replicates) plus technical replicate of one sample and detection of circRNAs.","project":"SRP064316"}
{"number_samples":6,"species":"human","abstract":"Glioblastoma (GBM) is the most common and aggressive primary brain tumor in adults, with glioma initiating cells (GICs) implicated to be critical for tumor progression and resistance to therapy. KDM1B is involved in regulating GICs' responses to hypoxia, since over-expression of KDM1B delays the cell growth under hypoxia while knocking-down of KDM1B in GICs promotes their survival and tumorigenic abilities. Overall design: We used RNA-Sequencing to detail the global change of gene expression in GICs with knockdown of KDM1B, and identified de-regulated genes and pathways downstream of KDM1B. CD133+ D456MG GICs were infected with non-targeting control and shRNA of KDM1B. Then RNA was extracted and gene expression was profiled by RNA-Seq.","project":"SRP064317"}
{"number_samples":12,"species":"human","abstract":"Background & aims: Polycystic liver disease (PLD) is an autosomal dominantly inherited disorder caused by mutations in genes such as PRKCSH and SEC63. It has been thought that cysts develop from biliary progenitor cells due to loss-of-heterozygosity (LOH), leading to aberrant proliferation or defects in differentiation. Cyst expansion can be suppressed by somatostatin analogues such as lanreotide. There is no human in vitro model available that truly recapitulates polycystic liver disease. We hypothesize that PLD progenitors can form bipotent liver organoids that carry key features of cyst development. To find gene expression differences between Human Polycystic Liver Disease and Normal Biliary Stem Cells. Methods: Cells from normal biliary duct (n=6), cyst biliary epithelium (n=60) and cyst fluid (n=31) were isolated and placed under conditions suitable for expansion of human adult liver stem cells. We analyzed genetic LOH, gene expression, differentiation capacity, response to lanreotide and cilium formation of these organoids. Results:  Cholangiocytes from cyst biliary epithelium (47/60) and cyst fluid (9/31) proved capable of expanding as bipotent liver organoids. Multiple cyst organoids displayed LOH surrounding PRKCSH or SEC63 regions. Organoids formed cilia when proliferation was inhibited. Neither hepatocyte nor biliary differentiation of PLD organoids was impaired. RNAseq revealed no significantly dysregulated pathway in PLD organoids. Lanreotide significantly decreased expansion of liver organoids in comparison to negative control (197% ± 46% versus 547% ± 28%; p: 0.038). Conclusion & discussion: Biliary progenitor cells from patient cyst epithelium and fluid can expand into liver organoids. They recapitulate key characteristics of PLD, and are a promising human in vitro model for research, diagnostics and treatment of polycystic liver diseases and cholangiociliopathies. Overall design: 8 polycystic liver disease samples versus 4 control samples RADBOUDUMC","project":"SRP064321"}
{"number_samples":2,"species":"human","abstract":"The goal of this study was to analyze differential gene expression 8 hr after treatment of CB-5083, a small molecule inhibitor of p97 (VCP), in HCT116 cells grown in culture. Overall design: DMSO treated cells were compared to cells treated with 1uM CB-5083 after 8 hr.","project":"SRP064323"}
{"number_samples":23,"species":"human","abstract":"RNA-Seq datasets for human plasma RNAs obtained by using thermostable group II intron reverse transcriptase","project":"SRP064378"}
{"number_samples":10,"species":"human","abstract":"The human nuclear poly(A)-binding protein PABPN1 has been implicated in the decay of nuclear noncoding RNAs (ncRNAs). In addition, PABPN1 stimulates hyperadenylation by poly(A) polymerase, and this activity is thought to be required for decay. Here, we inactivated hyperadenylation by two distinct mechanisms and examined changes in gene expression in HEK293 cells by RNAseq. We observed the upregulation of various ncRNAs, including snoRNA host genes, primary miRNA transcripts, and upstream antisense RNAs, confirming that hyperadenylation is broadly required for the degradation of PABPN1-targets. In addition, we found that mRNAs with retained introns are susceptible to PABPN1 and PAPa/?-mediated decay (PPD). Transcripts are targeted for degradation due to inefficient export, which is a consequence of reduced intron number or incomplete splicing. We conclude that PPD is an important mammalian nuclear RNA decay pathway for the removal of poorly spliced and nuclear-retained transcripts. Overall design: Poly(A)+ RNA from HEK293 cells was analyzed by next generation sequencing following depletion of PAPa and PAP? or expression of a dominant negative allele of PABPN1 (LALA) designed to inhibit polyadenylation. For each condition, we collected both total RNA and a nuclear-enriched sample. Each sample was collected in duplicate.","project":"SRP064410"}
{"number_samples":41,"species":"human","abstract":"Astrocytes were purified from fetal and adult human brain tissue using an immunopanning method with the HepaCAM antibody. Samples were taken from otherwise 'healthy' pieces of tissue, unless otherwise specified. Overall design: 6 fetal astrocyte samples, 12 adult astrocyte samples, 8 GBM or sclerotic hippocampal samples, 4 whole human cortex samples, 4 adult mouse astrocyte samples, and 11 human samples of other purified CNS cell types","project":"SRP064454"}
{"number_samples":5,"species":"human","abstract":"To assess the impact of AdV-VP55 mediated degredation of host miRNAs on the cellular transcriptome. Overall design: mRNA profiles of HEK 293T cells treated with type 5 Adeno vectors expressing either GFP or GFP-VP55 for 24 hours","project":"SRP064457"}
{"number_samples":18,"species":"human","abstract":"Purpose: To determine the impact endogenous miRNAs have on the overall transcriptome profile of fibroblasts and on the intrinsic response to virus Overall design: mRNA profiles of BJ fibroblasts treated with type 5  Adeno vectors expressing either GFP or GFP-VP55 for indicated time points alone or prior to treatment with IFNB or dsRNA","project":"SRP064458"}
{"number_samples":72,"species":"human","abstract":"We report the single-cell RNA-seq based identification of 6 known human islet cell types (alpha cells, beta cells, delta cells, pp cells, acinar cells and duct cells) based on the expression of known marker genes. We further assess cell type specific gene expression and suggest novel marker genes for several cell types. Overall design: Transcriptional dissection of human pancreatic islets of one donor using single-cell RNA-seq","project":"SRP064464"}
{"number_samples":8,"species":"human","abstract":"Two human pluripotent stem cell lines (hPSC) were generated from the human embryonic stem cell line WA09. These independent hPSC lines were designated Line A and Line B for the purpose of this study. Line A and Line B hPSC cells were each cultured in 2D (monolayer) or 3D (spheroid culture) conditions (experiment 1) and this procedure was repeated in a second independent experiment (experiment 2). RNA was isolated from these culture conditions (total of 8 samples) and RNA transcripts were sequenced as described in the publication accompanying this data.","project":"SRP064475"}
{"number_samples":2,"species":"human","abstract":"Purpose: We wanted to know how histone variants H3.3 and H2AZ are deposited into genes and enhancers during gene activation Methods: U2OS cells were harvested at 70% confluency with formaldehyde crosslinking for ChIP-seq without crosslinking for RNA-seq. ChIP DNA were purified through standard chromatin immunoprecipitation using an Pol II, MED26, EP400, H3.3, H2AZ, H3k4me1 and K3K18ac antibodies. Libraries were prepared with a KAPA LTP kit and sequenced using the Illumina HiSeq 2000 platform. Total RNA was extracted with Trizol, digested with DNaseI and further purified by acid phenol. Libraries were prepared with Illumina TruSeq RNA Sample Prep Kits v2 and were sequenced on Illumina HiSeq 2000. Conclusion: Our biochemical and genomics (ChIP-Seq and  mRNA-Seq) data show that EP400 contributes to H3.3 deposition in significantly with less of an effect on H2AZ in both genes and enhancers Overall design: Enrichment of EP400, Pol II, MED26, Histone varinat H3.3 and H2AZ, and enhancers marks H3K4me1 and H3K18ac in either Mock and/or EP400 knockdown conditions on chromatin were generated  by ChIP-Seq. mRNA profiles under Mock siRNA or EP400siRNA  were generated by deep sequencing, using Illumina HiSeq 2000.","project":"SRP064481"}
{"number_samples":2,"species":"human","abstract":"The goal of the study is to identify genes whose expression are enriched in the progenitor or differentiated layers of human skin, with an ultimate aim to find new molecular regulators of skin development and differentiation.","project":"SRP064538"}
{"number_samples":2,"species":"human","abstract":"Transcriptome of CALML5-depleted human organotypic epidermis","project":"SRP064547"}
{"number_samples":41,"species":"human","abstract":"The purpose of this study, including this time course series and another dosage series, was to establish gene expression signatures representing the interaction of pathways deregulated by tumor promoting agents and pathways induced by DNA damage.  Human lymphoblastoid TK6 cells were pretreated with the protein kinase C activating tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and exposed to UVC-irradiation.  The time and dose-responsive effects of the co-treatment were captured with RNA-sequencing (RNA-seq) in two separate experiments.  TK6 cells exposed to both TPA and UVC had significantly more genes differentially regulated than the theoretical sum of genes induced by either stress alone, thus indicating a synergistic effect on global gene expression patterns.  Further analysis revealed that TPA+UVC co-exposure caused synergistic perturbation of specific genes associated with p53, AP-1 and inflammatory pathways important in carcinogenesis.  The 17 gene signature derived from this model was confirmed with other PKC-activating tumor promoters including phorbol-12,13-dibutyrate, sapintoxin D, mezerein, (-)-Indolactam V and resiniferonol 9,13,14-ortho-phenylacetate (ROPA) with quantitative real-time PCR (QPCR).  Here we show a novel gene signature that may represent a synergistic interaction in the tumor microenvironment that is relevant to the mechanisms of chemical induced tumor promotion. Overall design: mRNA profiles of TK6 cells in 0,4,8,24 hours, with individual or combined treatment between UV irradiation and tumor promoting agent;","project":"SRP064561"}
{"number_samples":21,"species":"human","abstract":"The purpose of this study, including this dosage series and another  time course series, was to establish gene expression signatures representing the interaction of pathways deregulated by tumor promoting agents and pathways induced by DNA damage.  Human lymphoblastoid TK6 cells were pretreated with the protein kinase C activating tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and exposed to UVC-irradiation.  The time and dose-responsive effects of the co-treatment were captured with RNA-sequencing (RNA-seq) in two separate experiments.  TK6 cells exposed to both TPA and UVC had significantly more genes differentially regulated than the theoretical sum of genes induced by either stress alone, thus indicating a synergistic effect on global gene expression patterns.  Further analysis revealed that TPA+UVC co-exposure caused synergistic perturbation of specific genes associated with p53, AP-1 and inflammatory pathways important in carcinogenesis.  The 17 gene signature derived from this model was confirmed with other PKC-activating tumor promoters including phorbol-12,13-dibutyrate, sapintoxin D, mezerein, (-)-Indolactam V and resiniferonol 9,13,14-ortho-phenylacetate (ROPA) with quantitative real-time PCR (QPCR).  Here we show a novel gene signature that may represent a synergistic interaction in the tumor microenvironment that is relevant to the mechanisms of chemical induced tumor promotion. Overall design: mRNA profiles of TK6 cells treated with tumor promoting agent at different dose, co-exposed to UVC","project":"SRP064562"}
{"number_samples":1,"species":"human","abstract":"Nephron endowment is determined by the self-renewal and induction of a nephron progenitor pool established at the onset of kidney development. In the mouse, the related transcriptional regulators Six1 and Six2 play non-overlapping roles in nephron progenitors. Transient Six1 activity prefigures, and is essential for, active nephrogenesis. In contrast, Six2 maintains later progenitor self-renewal from the onset of nephrogenesis. We compared Six2’s regulatory actions in mouse and human nephron progenitors by chromatin immunoprecipitation followed by DNA sequencing (ChIP-seq). Surprisingly, SIX1 was identified as a SIX2 target unique to the human nephron progenitors. Further, RNA-seq and immunostaining revealed overlapping SIX1 and SIX2 progenitor activity in the 16 week human fetal kidney. Human SIX1 ChIP-seq revealed a similar set of targets to SIX2, and predicted both factors bind DNA through an identical recognition site. In contrast to the mouse where Six2 binds its own enhancers but doesn’t interact with DNA around Six1, both human SIX1 and SIX2 bind homologous SIX2 enhancers and putative enhancers positioned around SIX1. Transgenic analysis of a putative human SIX1 enhancer in the mouse revealed a transient, mouse-like, pre-nephrogenic, Six1 regulatory pattern. Together, these data demonstrate a divergence in SIX-factor regulation between mouse and human nephron progenitors. In the human, an auto/cross-regulatory loop drives continued SIX1 and SIX2 expression during active nephrogenesis. In contrast, the mouse establishes only an auto-regulatory Six2 loop. It is tempting to speculate that differential SIX-factor regulation may contribute to species differences in the duration of progenitor programs and nephron output. Overall design: Profiling of human and mouse nephron progenitor cells with mRNA-Seq; cells were purified from human and mouse embryonic kidneys with ITGA+ Ab or transgenic Cited1TagRPF","project":"SRP064624"}
{"number_samples":5,"species":"human","abstract":"RNA-Seq profiling of MCF-7 and MDA-MB-231. We profiled RNA expression in the estrogen-receptor-positive (ER+) MCF-7 and the triple-negative MDA-MB-231 breast cancer cells. The objective was to find genes differentially expressed between these cell lines as potential drivers of invasiveness of the triple-negative MDA-MB-231. We further utilized the identified differential genes to validate expression-responsive module of non-canonical Wnt signaling pathway. Overall design: 2 biological replicates of MCF-7 and 3 biological replicates of MDA-MB-231","project":"SRP064625"}
{"number_samples":4,"species":"human","abstract":"EBNA1 is the EBV-encoded nuclear antigen required for viral episome maintenance during latency.  EBNA1 is a sequence specific DNA binding protein with high affinity binding sites for the viral genome, especially OriP.  EBNA1 can also bind sequence specifically to a large number of sites in the host cellular genome, but the function of these binding sites has remained elusive.  EBNA1 is also known to provide a host cell survival function, but the molecular mechanisms accounting for this function are not completely understood.  Here, we show by integrating ChIP-Seq and RNA-Seq with experimental validation that MEF2B, IL6R, and EBF1 are high confidence target genes of EBNA1 that are essential for viability of B-lymphocytes latently infected with EBV. We show that EBNA1 binds to ~1000 sites with many, but not all, universally bound in different cell types, including Burkitt lymphoma (BL) and nasopharyngeal carcinoma (NPC).  We find that a large subset of EBNA1 binding sites are located proximal to transcription start sites and correlate genome-wide with transcription activity.  EBNA1 bound to genes of high significance for B-cell growth and function, including MEF2B, IL6R,  EBF1, RNF145, POU2F1, KDM4C, FGR, EGFR, LAIR, CDC7, CD44, and IL17A. EBNA1 depletion from latently infected LCLs results in the loss of cell proliferation, and the loss of gene expression for some EBNA1-bound genes, including MEF2B, EBF1, and IL6R. Depletion of MEF2B, EBF1, or IL6R partially phenocopies EBNA1-depletion by decreasing EBV-positive cell growth and viability.  These findings indicate that EBNA1 binds to a large cohort of cellular genes important for cell viability, and implicates EBNA1 as a master coordinator of host cell gene expression important for enhanced survival of latently infected cells. Overall design: Examination of EBNA1 binding in Raji, MutuI, LCL and C666-1 cells and EBNA1 knockdown effect on mRNA gene expression in LCL","project":"SRP064652"}
{"number_samples":3,"species":"human","abstract":"Whole exome and RNA-seq of melanoma tumors","project":"SRP064661"}
{"number_samples":3,"species":"human","abstract":"RBM15, an RNA binding protein, determines cell-fate specification of many tissues including blood. We demonstrate that RBM15 is methylated by protein arginine methyltransferase 1 (PRMT1) at residue R578 leading to its degradation via ubiquitylation by an E3 ligase (CNOT4). Overexpression of PRMT1 in acute megakaryocytic leukemia cell lines blocks megakaryocyte terminal differentiation by downregulation of RBM15 protein level. Restoring RBM15 protein level rescues megakaryocyte terminal differentiation blocked by PRMT1 overexpression. At the molecular level, RBM15 binds to pre-mRNA intronic regions of genes important for megakaryopoiesis such as GATA1, RUNX1, TAL1 and c-MPL. Furthermore, preferential binding of RBM15 to specific intronic regions recruits the splicing factor SF3B1 to the same sites for alternative splicing. Therefore, PRMT1 regulates alternative RNA splicing via reducing RBM15 protein concentration. Targeting PRMT1 may be a curative therapy to restore megakaryocyte differentiation for acute megakaryocytic leukemia. Overall design: In order to understand how RBM15 regulates RNA metabolism, we used a leukemia cell line (MEG-01 cell) to establish stable cell lines with RBM15 knockdown. For RNA immunoprecipitation (RIP) assay, normal immunoglobulin was used as controls for sample 1,2; and anti-RBM15 antibody was used for samples 3,4.","project":"SRP064671"}
{"number_samples":6,"species":"human","abstract":"Cellular lipid requirements are achieved through a combination of biosynthesis and import programs. Using isotope tracer analysis, we show that type I interferon (IFN) signaling rapidly shifts the balance of these programs by decreasing synthesis and increasing import of cholesterol and long chain fatty acids. Genetically enforcing this metabolic shift in macrophages is sufficient to render mice resistant to viral challenge, demonstrating the importance of reprogramming the balance of these two metabolic pathways in vivo. Unexpectedly, mechanistic studies reveal that limiting flux through the cholesterol biosynthetic pathway spontaneously engages a type I IFN response in a STING-dependent manner. The upregulation of type I IFNs was traced to a decrease in the pool size of synthesized cholesterol, and could be inhibited by replenishing cells with free cholesterol. Taken together, these studies delineate a metabolic-inflammatory circuit that links perturbations in cholesterol biosynthesis with activation of innate immunity. Overall design: shRNA to SREBF1 (shSREBP1) or SREBF2 (shSREBP2) were stably introduced via 3rd generation lentivirus into human THP1 monocytic cells under puromycin selection. Non-targeting shRNA scramble was used for a control (shControl). shControl, shSREBP1 and shSREBP2 modified cell types were analyzed by RNA-seq in duplicate.","project":"SRP064735"}
{"number_samples":1,"species":"human","abstract":"ChIP-Seq for endogenous TRF2 in HT1080 cell line along with RNA-Seq of stable expressed and vector control TRF2.","project":"SRP064781"}
{"number_samples":9,"species":"human","abstract":"Toxoplasma gondii is an obligate intracellular Apicomplexan parasite capable of invading and surviving within nucleated cells in most warm-blooded animals. This remarkable task is achieved through the delivery of effector proteins from the parasite into the parasitophorous vacuole and host cell cytosol that rewire host cellular pathways, facilitating parasite evasion of the immune system. Here, we have identified a novel export pathway in Toxoplasma that involves cleavage of effector proteins by the Golgi-resident aspartyl protease 5 (ASP5) prior to translocation into the host cell. We demonstrate that ASP5 cleaves a highly constrained amino acid motif that has some similarity to the PEXEL motif of Plasmodium parasites. We show that ASP5 can mature effectors at both the N- and C-terminal ends of proteins and is also required for the trafficking of proteins without this motif. Furthermore, we show that ASP5 controls establishment of the nanotubular network and is required for the efficient recruitment of host mitochondria to the parasitophorous vacuole membrane. Global assessment of host gene expression following infection reveals that ASP5-dependent pathways influence thousands of the transcriptional changes that Toxoplasma imparts on its host cell. This work characterizes the first identified machinery required for export of Toxoplasma effectors into the infected host cell. Overall design: Three groups of human foreskin fibroblasts are compared. Each group has 3 replicates giving a total of 9 samples. The first group of samples are infected with wild type (GRA16HA) Toxoplasma gondii, the second group with Asp5 knock-out Toxoplasma gondii, and the final group remain uninfected. All fibroblasts are generated from one donor sample.","project":"SRP064783"}
{"number_samples":9,"species":"human","project":"SRP064803"}
{"number_samples":2,"species":"human","abstract":"The RNA-seq results reveal Tetra-Pt(bpy) has no effect on gene expression. Overall design: Examination of the gene expression of the following cells: MRC5-untreated and Tetra-Pt(bpy)-treated HeLa-untreated and Tetra-Pt(bpy)-treated U2OS-untreated and Tetra-Pt(bpy)-treated","project":"SRP064820"}
{"number_samples":30,"species":"human","abstract":"High-throughput sequencing of RNA (RNA-Seq) in human cancer shows remarkable potential to simultaneously identify expression levels of protein-coding genes and long non-coding RNAs (lncRNAs). We performed RNA-Seq to investigate expression level of lncRNAs and protein-coding genes in 30 esophageal samples, including 15 esophageal squamous cell carcinoma (ESCC) tissue samples and 15 paired non-tumor tissues. We further developed an integrative bioinformatics method, denoted URW-LPE (for unsupervised random walk with each dysregulated lncRNA/PCG), to identify key functional lncRNAs that regulate expression of downstream protein-coding genes in ESCC. By this method, multiple known cancer and novel potentially functional lncRNAs were effectively identified. Quantitative reverse-transcription PCR was performed to confirm the lncRNA expression level of eight novel functional lncRNAs in an additional 120 paired ESCC patient samples. Finally, we characterized lncRNA625 as a novel ESCC regulator of cell proliferation, invasion and migration. Moreover, we identified E1A-binding protein p300 (EP300) as playing a key role in executing lncRNA625-induced transcriptional responses. These findings establish the utility of integrative bioinformatics analyses of RNA-Seq to identify cancer-associated functional lncRNAs.","project":"SRP064894"}
{"number_samples":4,"species":"human","abstract":"Gene expression studies of two mammalian cell lines that display primary cilia. Primary cilia are microtubule-based organelles that are hubs for receiving and intergrating signalling cascades during embryonic development and in maintaining tissue homeostasis. Defects in the structure of function of cilia can cause a group of comparitively common human inherited conditions known as ciliopathies. The cell-lines that have been profiled are standard model systems in the ciliary biology field, and gene expression profiles are important to understand the processes of forming, maintaining and resorbing cilia.","project":"SRP064956"}
{"number_samples":2,"species":"human","abstract":"Most human transcripts are alternatively spliced, and many disease-causing mutations affect RNA splicing. Towards better modeling the sequence determinants of alternative splicing, we measured the splicing patterns of nearly 2 million (M) synthetic mini-genes, which include degenerate subsequences totaling to nearly 100M bases of variation. The massive size of these training data allowed us to improve upon current models of splicing as well as to gain new mechanistic insights. Our results show that a vast majority of hexamer sequence motifs measurably influence splice site selection when positioned within alternative exons, with multiple motifs acting additively rather than cooperatively. Intriguingly, motifs that enhance (suppress) exon inclusion in alternative 5’ splicing also enhance (suppress) exon inclusion in alternative 3’ or cassette exon splicing, suggesting a universal mechanism for alternative exon recognition.  Finally, our empirically trained models are highly predictive of the effects of naturally occurring variants on alternative splicing in vivo. Overall design: HEK293 cells were transfected with two alternatively spliced plasmid libraries. Spliced reads were sequenced to determine isoform counts for each library sequence.","project":"SRP064967"}
{"number_samples":2,"species":"human","abstract":"To understand the impact of alternative translation initiation on a proteome, we performed the first study on protein turnover using positional proteomics and ribosome profiling to distinguish between N-terminal proteoforms of individual genes. Overall, we monitored the stability of 1,941 human N-terminal proteoforms, including 147 N-terminal proteoform pairs that originate from alternative translation initiation, alternative splicing or incomplete processing of the initiator methionine. Study design: ribosome profiling of lactimidomycin and cycloheximide treated human Jurkat T-lymphocytes","project":"SRP065022"}
{"number_samples":4,"species":"human","abstract":"Normoxic (21% O2) and hypoxic (1% O2, 24 hr) U87MG human glioblastoma cells were subjected to polysome fractionation. Total RNA were isolated from individual fractions by standard phenol/chloroform extraction and ethanol precipitation following proteinase K treatment. Equal volumes of individual fractions from four independent experiments were pooled to yield the monosome/oligosomes (MO) and polysome (P) samples.","project":"SRP065114"}
{"number_samples":4,"species":"human","abstract":"UM_RNAseq_glioblastoma_supplementary","project":"SRP065127"}
{"number_samples":6,"species":"human","abstract":"RNA-seq of peripheral blood leucocyte samples from Immune Thrombocytopenia patients and controls","project":"SRP065146"}
{"number_samples":4,"species":"human","abstract":"Several studies have shown that long non-coding RNAs (lncRNAs) may play anessential role in Epithelial-mesenchymal transition (EMT), which is an important step in tumor metastasis, however, little is known about the global change of lncRNA transcriptome during EMT. To investigate how lncRNA transcriptome alteration contributes to EMT progression regulation, we performed a whole-transcriptome strand-specific RNA deep sequencing of MCF10A induced EMT by TGF-ß. Deep sequencing results showed that the long RNA (>=200-nt) transcriptome of MCF10A was undergone a global changed in EMT, and this alteration was determined as early as 8h after being induced using TGF-ß. 8703 linear novel genes with ambiguous protein-coding potential were identified, 512 of which were further determined to be novel lncRNAs. After analyzing the expression of 5473 known and novel lncRNAs, as well as 2208 known and novel circRNAs during EMT, we found a large numbers of lncRNAs might be involved in the regulation of EMT. Intriguingly, we identified 216 gene clusters constituted by lncRNAs and/ornovel genes in “gene desert” region. The expressions of all genes in these clusters were changed concurrently during EMT, indicating that these clusters might play important role in EMT. Our study reveals a global reprogramming of lncRNAs transcriptome in EMT and provides clues to the study of the molecular mechanism of EMT.","project":"SRP065153"}
{"number_samples":6,"species":"human","abstract":"We performed high throughput RNA-sequencing on blood and lymphatic HMVECs to identify differences in gene expression between these two cell types that may provide insight into their respective differentiation and roles in angiogenesis and lymphangiogenesis. Overall design: RNA was isolated from blood-specific HMVEC and lymphatic-specific HMVEC cell lines 3 separate times each","project":"SRP065196"}
{"number_samples":12,"species":"human","abstract":"Sequencing of total RNA and polysomal RNA of two cell lines, MCF7 (tumoral) and MCF10A (non-tumoral) Overall design: Polysomal and total RNA was prepared from biological triplicates from the two cell lines.  For each biological triplicate sub-confluent cell monolyers were lysed to produce cytoplasmic extracts. Half of each extract was fractionated on a sucrose gradient and the polysomal fraction recovered. Polysomal-associated RNA was recovered by Trizol extraction. From the second half of each sample total cyoplasmic RNA was recovered (Trizol extraction).","project":"SRP065202"}
{"number_samples":6,"species":"human","abstract":"Using RNA sequencing (Illumina Hi-Seq 2000 sequencer) we report the transcriptome profile of primary human chondrocytes isolated from patients with hip osteoarthritis (OA), and the transcriptome response of these cells to 4h stimulation with IL-1ß (1ng/ml).  In total, 983 long non-coding RNAs (lncRNAs) were identified, which included 642 intergenic lncRNAs (lincRNAs), 124 antisense and pseudogenes.  Less than 4% of the identified lncRNAs overlapped with putative eRNAs regions, and visual inspection showed that they were uni-directional and multi-exonic.  Upon  IL-1ß stimulation 499 protein-coding genes were differentially expressed, and 158 lncRNAs were differentially expressed, including 92 lincRNAs, 13 antisense and 18 psudogenes.  This study demonstrates that IL-1ß induces a rapid and widespread change in the transcriptome of the primary human OA chondrocyte. Overall design: RNA sequencing analysis of primary human chondrocytes isolated from n=3 patients with hip osteoarthritis, with and without 4h IL-1b (1ng/ml) stimulation","project":"SRP065219"}
{"number_samples":10,"species":"human","abstract":"Xenograft models represent an excellent method for expanding primary leukemias, but their ability to preserve the gene expression profile of the parent leukemia may also depend on providing microenvironmental factors that are not cross-reactive between human and mouse. Here we focused on leukemias with a CRLF2 mutation, and the ability of human TSLP to stimulate these cells. Overall design: Three different classes of samples were analyzed. 1) The primary leukemia sample without any culture or treatment 2) Xenograft samples (2) of the primary leukemia that were exposed to TSLP in vivo for 2 weeks, 9 weeks post-engraftment and 3) Xenograft samples (2) of the primary leukemia that were NOT exposed to TSLP in vivo for 2 weeks, 9 weeks post-engraftment.","project":"SRP065258"}
{"number_samples":6,"species":"human","project":"SRP065282"}
{"number_samples":6,"species":"human","abstract":"Transcriptomic Sequence from Caco-2 colon carcinoma cell lines grown in monolayers or in the rotating wall vessel 3D bioreactor","project":"SRP065330"}
{"number_samples":13,"species":"human","abstract":"Histiocytic neoplasms are clonal, hematopoietic disorders characterized by an accumulation of abnormal, monocyte-derived dendritic cells or macrophages in Langerhans Cell (LCH) and non-Langerhans (non-LCH) histiocytoses, respectively. The discovery of BRAFV600E mutations in ~50% of these patients provided the first molecular therapeutic target in histiocytosis. However, recurrent driving mutations in the majority of BRAFV600E-wildtype, non-LCH patients are unknown, and recurrent cooperating mutations in non-MAP kinase pathways are undefined for the histiocytic neoplasms. Through combined whole exome and transcriptome sequencing, we identified recurrent kinase fusions involving BRAF, ALK, and NTRK1, as well as recurrent, activating MAP2K1 and ARAF mutations in BRAFV600E-wildtype, non-LCH patients. In addition to MAP kinase pathway lesions, recurrently altered genes involving diverse cellular pathways were identified. Treatment of MAP2K1- and ARAF-mutated, non-LCH patients using MEK and RAF inhibitors, respectively, resulted in clinical efficacy demonstrating the importance of detecting and targeting diverse kinase alterations in these disorders. Overall design: 13 patient samples were analyzed by RNA-seq and had 2 replicates.","project":"SRP065445"}
{"number_samples":4,"species":"human","abstract":"Loss-of-function studies are fundamental for dissecting gene function. Yet, methods to rapidly and effectively perturb genes in mammalian cells are scarce. We present a novel system, deliverable with only two lentiviral vectors, which enables simultaneous control over two different proteins in the same cell. By harnessing the plant auxin and jasmonate hormone-induced degradation pathways, combined with RNA interference, this system allows constitutive depletion of two endogenous proteins and their replacement with two exogenous proteins whose degradation is rapidly and reversibly induced by external ligands, representing a dual analog molecular tuner. Focusing on NANOG, CHK1 and NOTCH1 in embryonic stem cells and p53 in cancer cells we have validated the efficiency, rapidity, reversibility, titratability and multiplicity of the engineered tuners, and demonstrated their potential to facilitate previously-unfeasible experimental approaches and to generate novel biological insights. Overall design: For mRNA-Seq preparation, coronatine/DMSO treated cells were collected.","project":"SRP065451"}
{"number_samples":1,"species":"human","abstract":"HeLa transcriptome used for benchmarking HiLive","project":"SRP065468"}
{"number_samples":10,"species":"human","abstract":"To investigate the impact of genetic and environmental factors that influence cancer cell metabolism in vivo, we conducted intra-operative 13C-glucose infusions in NSCLC patients. We determined that regional glucose metabolism was correlated with tumor perfusion as assessed by pre-surgical dynamic gadolinium enhancement.  Tumors or tumor regions showing high or low perfusion were collected and subjected to RNA-seq analysis. Overall design: RNA-seq analysis was performed to evaluate gene transcriptional levels in human non-small cell lung cancer (NSCLC) tissues showing high or low perfusion as measured by dynamic contrast-enhanced (DCE) MRI.","project":"SRP065491"}
{"number_samples":16,"species":"human","abstract":"p53 is a pivotal tumor suppressor and a major barrier against cancer. We now report that silencing of the Hippo pathway tumor suppressors LATS1 and LATS2 in non-transformed mammary epithelial cells reduces p53 phosphorylation and increases its association with the p52 NF-?B subunit. Moreover, it partly shifts p53’s conformation and transcriptional output towards a state resembling cancer-associated p53 mutants, and endow p53 with the ability to promote cell migration. Notably, LATS1 and LATS2 are frequently downregulated in breast cancer; we propose that such downregulation might benefit cancer by converting p53 from a tumor suppressor into a tumor facilitator. Overall design: MCF10A cells transfected with siRNA against LATS1/2 alone, p53 alone or LATS1/2 and p53 together. Two independent MCF10A batches provided biological replicates","project":"SRP065500"}
{"number_samples":24,"species":"human","abstract":"Half of all human cancers lose p53 function by missense mutations, with an unknown fraction of these containing p53 in a self-aggregated, amyloid-like state. Here we show that a cell-penetrating peptide, ReACp53, designed to inhibit p53 amyloid formation, rescues p53 function in cancer cell lines and in organoids derived from high-grade serous ovarian carcinomas (HGSOC), an aggressive cancer characterized by ubiquitous p53 mutations. Rescued p53 behaves similarly to its wild-type counterpart in regulating target genes, reducing cell proliferation and increasing cell death. Intraperitoneal administration decreases tumor proliferation and shrinks xenografts in vivo. Our data show the effectiveness of targeting a specific aggregation defect of p53 and its potential applicability to HGSOCs. Overall design: Vehicle vs. ReACp53 treatment in 4 different samples: 2 cell lines (MCF7 w/ WT p53 as negative control and OVCAR3 w/ R248Q p53) and 2 clinical specimens (primary cells from patient #8 w/ WT p53 as negative control and primary cells from patient #1 w/ R248Q p53)","project":"SRP065559"}
{"number_samples":6,"species":"human","abstract":"Using doxycycline-inducible IFN-kappa expression in CIN612-9E cells, which maintain extrachromosomally replicating HPV31 genomes, we demonstrate that IFN-kappa inhibits the growth of these cells and reduces viral transcription and replication. Interestingly, the initiation of viral early transcription was already inhibited 4-6h after IFN-kappa expression. This was also observed with recombinant IFN-beta suggesting a common mechanism of IFNs. RNA-seq analysis identified 1367 IFN-kappa regulated genes of which 221 were modulated >2-fold. The majority of those (71%) matched known ISGs confirming that IFN-kappa acts as a bona fide type I IFN in hr-HPV-positive keratinocytes. RNAi and co-transfection experiments indicate that the inhibition of viral transcription is mainly due to the induction of Sp100 proteins by IFN-kappa. Overall design: CIN612-9E/pInd-IFN-kappa were induced for 4h with 1µg/ml doxycyclin or not. Three biological replicates were analyzed.","project":"SRP065763"}
{"number_samples":7,"species":"human","abstract":"The resultant heat map demonstrates the maturation of CD13+/ROR2+ cells as they proceed through cardiac differentiation. Overall design: RNA-seq analysis was preformed on RNA samples from undifferentiated hESCs, 13R2+ and 13R2- populations from day 3, 13R2+/NKX2-5+ and 13R2+/NKX2-5- from day 7, and 13R2+/NKX2-5+/a-MHC+ and 13R2+/NKX2-5+/MHC- from day 14","project":"SRP065774"}
{"number_samples":52,"species":"human","abstract":"In this study, we used RNA-sequencing to profile the long non-coding RNA (lncRNA) transcriptome in lesional skin from psoriasis patients before (PP) and after treatment (PT) with adalimumab and in normal skin from healthy individuals (NN). For this we sequenced total RNA from 18 psoriasis patients (before and after treatment) and 16 healthy controls. We created our own reference set of long non-coding RNAs by merging three long non-coding RNA reference data sets. The combined reference had 67,157 lncRNA transcripts with no overlaps. We identified differential expression of 971 lncRNAs between PP and NN, 157 between PP and PT, and 377 between PT and NN. Based on differentially expressed (DE) lncRNAs between PP and NN, we identified a molecular lncRNA signature that distinguishes psoriatic skin from healthy skin . Overall design: We recruited 18 psoriatic patients in our study. We took 5 mm punch biopsies from the edge of a psoriatic plaque before treatment and after one month of treatment with adalimumab. The mean improvement in the PASI over this time period was 53.1%. We obtained sixteen normal skin samples (N=16) from healthy control surgical discard specimens.","project":"SRP065812"}
{"number_samples":6,"species":"human","abstract":"We describe an X-linked genetic syndrome associated with mutations in TAF1 presenting with global developmental delay, intellectual disability, characteristic facial dysmorphology, generalized hypotonia, and variable neurologic features, all in male cases. Simultaneous studies using diverse strategies led to nine families with overlapping clinical presentations and with de novo or maternally inherited missense mutations. Two additional families harboring large duplications involving TAF1 were also found to share phenotypic overlap with the probands harboring missense mutations, but they also demonstrated a severe neurodegeneration phenotype. Functional analysis with RNA-seq for one of the families suggest that the phenotype is associated with down regulation of a set of genes notably enriched for genes regulated by E-box proteins, and knockdown and mutant studies for this gene in zebrafish have a quantifiable, albeit small, effect on a neuronal phenotype. Our results implicate mutations in TAF1 as playing a critical role in the development of this X-linked intellectual disability syndrome.","project":"SRP065848"}
{"number_samples":9,"species":"human","abstract":"Purpose: Seek for differential gene expression in vemurafenib-resistant A375 tumors vs. untreated controls to provide a rationale for resistance mechanism Methods: mRNA profiles of vemurafenib-resistant A375 tumors and untreated control tumors were generated by transcriptome sequencing of A375 melanoma bearing mice. Since our xenograft samples contain a mixture of human and mouse RNAs we mapped RNASeq reads against a hybrid human/mouse genome. We than removed reads of potential mouse origin by taking only reads that map uniquely to human chromosomes. On average 23% of reads were removed as potential mouse reads. We than took the remaining reads (on average 77% per sample) to determine the gene expression levels for each sample. Normalized expression levels of 5 resistant samples were compared to 4 untreated control samples to detect differnetially regulated genes which may contribute to vemurfenib resistance Results: Expression levels of several genes were consistently altered in all resistant samples. Expression of e.g. genes encoding SPRY2, SPRY4, DUSP6, CCND1, PIK3R3, FGFR1, EPHA4, MCL1, and IGF1R was down-regulated, whereas expression of PDGFC, VEGFC, ABCB9 and KITLG was increased. Conclusions: Our study reports several differentially expressed genes which may contribute to vemurafenib resistance in A375 tumor bearing mice Overall design: RNA sequencing of genes expressed in A375 tumors bearing mice treated with vemurafenib until in vivo resistance appeared vs. untreated A375 tumors","project":"SRP065849"}
{"number_samples":2,"species":"human","abstract":"Intragenic 5-methylcytosine and CTCF mediate opposing affects on pre-mRNA splicing: CTCF promotes inclusion of weak upstream exons through RNA polymerase II pausing, whereas 5-methylcytosine evicts CTCF, leading to exon exclusion. However, the mechanisms governing dynamic DNA methylation at CTCF binding sites were unclear. In this study, we identify the methylcytosine dioxygenases TET1 and TET2 as active regulators of CTCF-mediated alternative splicing through conversion of 5-methylcytosine to its oxidation derivatives. Overall design: Transcriptional profiling of human T-lymphocytes in the naïve and activated states was performed by RNA-Seq.  Methylation status (5mC and 5hmC) was assayed by medIP-Seq.  CTCF binding sites were identified by ChIP-Seq.","project":"SRP065988"}
{"number_samples":8,"species":"human","abstract":"Long non-coding RNAs (lncRNAs) play diverse functional roles in cancer. The HOX transcript antisense RNA (HOTAIR) lncRNA in particular is associated with metastasis, proliferation, and epithelial to mesenchymal transition in various tumor types. In this study, we find that upregulation of HOTAIR induced ovarian cancer (OC) cisplatin (CDDP) resistance in vitro and in vivo. HOTAIR expression levels were significantly increased in recurrent vs. primary OC as well as in platinum-resistant vs sensitive tumors. To investigate the role of HOTAIR during DNA damage response (DDR) induced by CDDP, we monitored double-strand breaks (DSBs) using ?-H2AX and found that the expression of HOTAIR allows for a sustained activation of DDR after CDDP treatment. We demonstrate that ectopic expression of HOTAIR induces activation of the transcription factor nuclear factor kappa B (NF-?B) during DDR as well as expression of MMP-9 and IL-6, key NF-?B target genes. We show that HOTAIR regulates NF-?B activation by decreasing NF-?B inhibitor alpha (I?-Ba) protein levels and establish that by inducing prolonged NF-?B activation and expression of NF-?B target genes during DNA damage, HOTAIR plays a critical role in OC cellular senescence and CDDP sensitivity. Our findings suggest that HOTAIR-NF-?B signaling drives a positive-feedback loop cascade during DDR and contributes to cellular senescence and chemotherapy resistance in ovarian and other cancers.","project":"SRP066008"}
{"number_samples":12,"species":"human","abstract":"Next-generation RNA sequencing (RNA-seq) has revolutionized our ability to analyze transcriptomes. Current RNA-seq methods are highly reproducible, but each has biases resulting from different modes of RNA preparation, reverse transcription, and adaptor addition, leading to variability between methods. Moreover, the transcriptome cannot be profiled comprehensively as highly structured RNAs, such as tRNAs and snoRNAs, are refractory to conventional RNA-seq methods. Recently, we developed a new method for strand-specific RNA-seq using thermostable group II intron reverse transcriptases (TGIRTs). TGIRTs have high processivity and fidelity plus a novel template-switching activity that enables addition of an RNA-seq adaptor during cDNA synthesis without using RNA-ligase. Here, we used TGIRT-seq to obtain RNA-seq datasets from well-characterized human RNA reference samples and compared them to previous datasets obtained for these RNAs by the Illumina TruSeq v2 and v3 methods. We find that TGIRT-seq recapitulates the relative abundance of human transcripts and RNA spike-ins in ribo-depleted, fragmented RNA samples similarly to the non-strand-specific TruSeq v2 and better than the strand-specific TruSeq v3 method. Moreover, TGIRT-seq is simpler and more strand-specific than TruSeq v3, gives more uniform 5’ to 3’ gene coverage than either TruSeq method, and eliminates sequence biases from random hexamer priming inherent to TruSeq. TGIRT-seq also detects more splice junctions, particularly near the 5’ ends of genes than does TruSeq v3, even in fragmented RNAs. Finally, TGIRT-seq enables simultaneous profiling of mRNAs and lncRNAs in the same RNA-seq as structured small ncRNAs, including tRNAs, which are essentially absent from TruSeq libraries.","project":"SRP066009"}
{"number_samples":9,"species":"human","abstract":"Post-translational modifications, such as poly(ADP-ribosyl)ation (PARylation), regulate chromatin-modifying enzymes, ultimately affecting gene expression. This study explores the role of poly(ADP-ribose) polymerase (PARP) on global gene expression in a lymphoblastoid B cell line. We found that inhibition of PARP catalytic activity with olaparib resulted in global gene deregulation, affecting approximately 11% of genes expressed. Gene ontology analysis revealed that PARP could exert these effects through transcription factors and chromatin-remodeling enzymes, including the Polycomb Repressive Complex 2 (PRC2) member EZH2. EZH2 mediates the trimethylation of histone H3 at lysine 27 (H3K27me3), a modification associated with chromatin compaction and gene silencing. Both pharmacological inhibition of PARP and knockdown of PARP1 induced the expression of EZH2 that resulted in increased global H3K27me3. Chromatin immunoprecipitation confirmed that PARP1 inhibition led to H3K27me3 deposition at EZH2-target genes, which resulted in gene silencing. Moreover, increased EZH2 expression is attributed to occupancy loss of the transcription repressor E2F4 at the EZH2 promoter following PARP inhibition. Together, these data show that PARP plays an important role in global gene regulation and identifies for the first time a direct role of PARP1 in regulating the expression and function of EZH2. Overall design: Examination of the effect of PARP inhibition on global gene expression in LCLs cell lines. mRNA profiles of LCLs cells lines treated at different time points with olaparib were generated by deep sequencing, in triplicate, using Illumina GAIIx.","project":"SRP066150"}
{"number_samples":16,"species":"human","abstract":"Hemoglobinopathies, including sickle cell disease and _-thalassemia, are global public health concerns. Induction of fetal-type hemoglobin (HbF) is a promising means to treat these disorders; however, precisely how HbF expression is silenced in adult erythroid cells is not fully understood. Here, we show that the LRF/ZBTB7A transcription factor is a potent repressor of HbF production. LRF inactivation derepresses embryonic/fetal _-globin expression in mouse and human adult erythroid cells. We employed genome-wide analysis of the transcriptome, chromatin accessibility and LRF occupancy sites, and demonstrate that LRF occupies the _-globin loci and maintains nucleosome density necessary for _-globin silencing. LRF confers its repressive activity through a unique NuRD repressor complex independent of BCL11A. Strikingly, human erythroid lines lacking both LRF and BCL11A exhibited almost a complete switch in expression from adult- to fetal-type globin, suggesting that these two factors cumulatively represent the near entirety of _-globin repressive activity in adult erythroid cells. Overall design: RNA-seq, LRF ChIP-seq and ATAC-seq assays were used to investigtae LRF binding, effect of LRF depletion on transcription and chromatin landscape in mouse and human cells.","project":"SRP066151"}
{"number_samples":4,"species":"human","abstract":"Expression levels of the RNA-binding protein Quaking (QKI) are low in monocytes of early, human atherosclerotic lesions, but abundant in macrophages of advanced plaques. Specific depletion of QKI protein impaired monocyte adhesion, migration, differentiation into macrophages, and foam cell formation in vitro and in vivo. RNA-seq and microarray analysis of human monocyte and macrophage transcriptomes, including those of a unique QKI haploinsufficient patient, revealed striking changes in QKI-dependent mRNA levels and splicing of RNA transcripts. Overall design: RNA-seq analysis of primary monocytes and macrophages from a  QKI haploinsufficient patient and their (control) sibling.","project":"SRP066152"}
{"number_samples":15,"species":"human","abstract":"We performed whole transcriptome sequencing of human monocytes that were co-cultured with estrogen receptor positive (ER+) or triple-negative (TNBC) breast cancer cell lines and studied the biological responses related to the differential gene activation in both cell types to understand how different cancer cells educate host cells to support tumor growth Overall design: To characterize the differences in macrophage activation under the influence of either ER+ or TNBC breast cancer cells, we cultured freshly isolated human peripheral monocytes with two breast cancer cell lines (T47D, ER+ and MDA-MB-231, TNBC) in an in vitro transwell co-culture assay. The transwell setting allowed us to investigate the effect of soluble mediators on macrophage activation since direct cell contact of these cells was inhibited by a (PET) membrane (pore size 0.4 µm).","project":"SRP066356"}
{"number_samples":6,"species":"human","abstract":"Through development of an in vivo orthotopic lung cancer model, we reveal an unanticipated pathway driving spontaneous metastasis that is orchestrated by the developmentally-regulated transcriptional repressor, Capicua (CIC). Overall design: RNAseq and DNA copy number analysis of H1975 (EGFR-mutant lung adenocarcinoma) cells in the context of drug resistance to rociletinib","project":"SRP066363"}
{"number_samples":7,"species":"human","abstract":"We identified PHF5A as a functional synthetic-lethal hit in glioblastoma stem cells compared to normal neural stem cells. We wanted to perform analysis of RNA isoforms present in glioblastoma or normal neural stem cells with or without PHF5A depletion. We performed shRNA knockdown of PHF5A or used non-silencing shRNA as a control, selected infected cells with puromycin, and isolated RNA for sequencing. Overall design: We analyzed RNA from either normal neural stem cells or two different glioblastoma specimens aster either control knockdown, or two different shRNA sequences against the PHF5A gene transcript.","project":"SRP066371"}
{"number_samples":12,"species":"human","abstract":"The ability to perturb genes in human cells is crucial for elucidating gene function and holds great potential for finding therapeutic targets for diseases such as cancer. To extend the catalog of human core and context- dependent fitness genes, we have developed a high-complexity second-generation genome-scale CRISPR-Cas9 gRNA library and applied it to fitness screens in five human cell lines. Using an improved Bayesian analytical approach, we consistently discover 5-fold more fitness genes than were previously observed. We present a list of 1,580 human core fitness genes and describe their general properties. Moreover, we demonstrate that context-dependent fitness genes accurately recapitulate pathway-specific genetic vulnerabilities induced by known oncogenes and reveal cell-type-specific dependencies for specific receptor tyrosine kinases, even in oncogenic KRAS backgrounds. Thus, rigorous identification of human cell line fitness genes using a high-complexity CRISPR-Cas9 library affords a high-resolution view of the genetic vulnerabilities of a cell.  Additional data can be found at tko.ccbr.utoronto.ca Overall design: RNAseq of five human cell lines with Cas9 knock-ins.","project":"SRP066424"}
{"number_samples":2,"species":"human","abstract":"BJAB cells over expressing KSHV PAN RNA","project":"SRP066449"}
{"number_samples":4,"species":"human","abstract":"BCBL1 cell line","project":"SRP066482"}
{"number_samples":2,"species":"human","abstract":"PBMC + KSHV","project":"SRP066484"}
{"number_samples":2,"species":"human","abstract":"293L + BAC36 WT or deltaORF50","project":"SRP066488"}
{"number_samples":20,"species":"human","abstract":"Genomic characterization and comparison of multi-regional and pooled tumor biopsy specimens","project":"SRP066596"}
{"number_samples":20,"species":"human","abstract":"Transcriptome profiling was performed on muscle biopsies from patients immediately before Total Knee Arthroplasty and two hours after TKA and tourniquet application. Overall design: RNA was isolated from 10 patients who were give vastus lateralis muscle biopsies immediately before surgery and 2 hours post surgery with tourniquet","project":"SRP066729"}
{"number_samples":734,"species":"human","abstract":"Cerebral organoids – three-dimensional cultures of human cerebral tissue derived from pluripotent stem cells – have emerged as models of human cortical development. However, the extent to which in vitro organoid systems recapitulate neural progenitor cell proliferation and neuronal differentiation programs observed in vivo remains unclear. Here we use single-cell RNA sequencing (scRNA-seq) to dissect and compare cell composition and progenitor-to-neuron lineage relationships in human cerebral organoids and fetal neocortex. Covariation network analysis using the fetal neocortex data reveals known and novel interactions among genes central to neural progenitor proliferation and neuronal differentiation. In the organoid, we detect diverse progenitors and differentiated cell types of neuronal and mesenchymal lineages, and identify cells that derived from regions resembling the fetal neocortex. We find that these organoid cortical cells use gene expression programs remarkably similar to those of the fetal tissue in order to organize into cerebral cortex-like regions. Our comparison of in vivo and in vitro cortical single cell transcriptomes illuminates the genetic features underlying human cortical development that can be studied in organoid cultures. Overall design: 734 single-cell transcriptomes from human fetal neocortex or human cerebral organoids from multiple time points were analyzed in this study. All single cell samples were processed on the microfluidic Fluidigm C1 platform and contain 92 external RNA spike-ins. Fetal neocortex data were generated at 12 weeks post conception (chip 1: 81 cells; chip 2: 83 cells) and 13 weeks post conception (62 cells). Cerebral organoid data were generated from dissociated whole organoids derived from induced pluripotent stem cell line 409B2 (iPSC 409B2) at 33 days (40 cells), 35 days (68 cells), 37 days (71 cells), 41 days (74 cells), and 65 days (80 cells) after the start of embryoid body culture. Cerebral organoid data were also generated from microdissected cortical-like regions from H9 embryonic stem cell derived organoids at 53 days (region 1, 48 cells; region 2, 48 cells) or from iPSC 409B2 organoids at 58 days (region 3, 43 cells; region 4, 36 cells).","project":"SRP066834"}
{"number_samples":12,"species":"human","project":"SRP066889"}
{"number_samples":17,"species":"human","abstract":"Purpose: PUMILIO proteins are known to repress target genes by specifically binding to PUMILIO response elements (PREs) in target mRNAs.  NORAD is a noncoding RNA that negatively regulates PUMILIO activity. The goal of this study was to determine the gene expression changes that result from knockout of NORAD or overexpression of PUMILIO and to test whether NORAD knockout causes PUMILIO hyperactivity. Methods: RNA-seq libraries were prepared using the TruSeq Stranded Total RNA with Ribo-Zero Human/Mouse/Rat Sample Preparation kit (Illumina) and sequenced using the 100 bp paired-end protocol on an Illumina HiSeq 2000.  For comparing NORAD+/+ and NORAD-/- HCT116 cells, 3 biological replicates per genotype were sequenced.  For PUM overexpression experiments, 3 replicates of GFP-expressing HCT116 cells (negative control) and 2 independent PUM1- or PUM2-overexpressing clones (2 replicates each) were sequenced. Results: Gene expression profiles show that PUMILIO target genes are downregulated in both NORAD knockout cells and PUMILIO overexpressing cells. Conclusions: These data indicate that NORAD sequesters PUMILIO, preventing excessive repression of PUMILIO target genes that are important for maintaining genomic stability. Overall design: Gene expression profiles of genetically modified HCT116 cells were determined by RNA-seq.","project":"SRP066895"}
{"number_samples":3,"species":"human","abstract":"We compared RNA-seq expression patterns in liver, an organ with high oxidative metabolism and abundant spontaneous DNA damage, from humans, naked mole rats, and mice, differing in maximum lifespan over a range of ~100, 30, and 3 years, respectively, for 130 genes involved in DNA repair. The results show that the longer-lived species, human and naked mole rat, share higher expression of these DNA repair genes, including core genes in several DNA repair pathways. A more systematic approach of signaling pathway analysis (SPA) indicates statistically significant upregulation of several DNA repair signaling pathways in human and naked mole rat compared with mouse. Overall design: Compare steady-state RNA levels from 3 samples each of adult liver tissue of human, naked mole rat, and mouse. Mouse samples available in the Short Reads Archive: SRX871336, SRX871370, SRX871395 (SRP053350) and BioProject PRJNA274780.","project":"SRP066912"}
{"number_samples":11,"species":"human","abstract":"We exained using RNA-seq the effect of targeting selected p53-bound enhancers on the p53-induced response to oncogenic stress Overall design: RNA-seq was applied to examine transcriptional response to oncogenic stress (induced by RASG12V activation) in control and genetically manipulated hTERT-immortalized BJ cells","project":"SRP066917"}
{"number_samples":9,"species":"human","abstract":"Human cancer cells are subjected to hypoxic conditions in many tumours. Hypoxia causes alterations in glycolytic pathway activation through stabilization of hypoxia-inducible factor-1. Currently, two approaches are commonly used to model hypoxia: alternatively to generating low-oxygen condition in an incubator, cells can be treated with CoCl2. We performed RNA-seq experiments to study transcriptomes of human Caki-1 cells under real hypoxia and after CoCl2-treatment. Despite causing transcriptional changes of much higher order of magnitude for the genes in hypoxia regulation pathway, CoCl2-treatment fails to induce alterations in glycolysis/gluconeogenesis pathway. Moreover, CoCl2 caused aberrant activation of other oxidoreductases in Glycine, Serine and Threonine metabolism pathways.","project":"SRP066934"}
{"number_samples":22,"species":"human","abstract":"Although mechanisms of acquired resistance of EGFR mutant non-small cell lung cancers to EGFR inhibitors have been identified, little is known about how resistant clones evolve during drug therapy. Here, we observe that acquired resistance caused by the T790M gatekeeper mutation can occur either by selection of pre-existing T790M clones or via genetic evolution of initially T790M-negative drug tolerant cells. The path to resistance impacts the biology of the resistant clone, as those that evolved from drug tolerant cells had a diminished apoptotic response to third generation EGFR inhibitors that target T790M EGFR; treatment with navitoclax, an inhibitor of BCL-XL and BCL-2 restored sensitivity. We corroborated these findings using cultures derived directly from EGFR inhibitor-resistant patient tumors. These findings provide evidence that clinically relevant drug resistant cancer cells can both pre-exist and evolve from drug tolerant cells, and point to therapeutic opportunities to prevent or overcome resistance in the clinic. Overall design: Examination of mRNA levels of PC9 parental, drug-tolerant, PC9-GR2 and PC9-GR3 cells after treatment with vehicle, gefitinib or WZ4002 for 24 hours.","project":"SRP066956"}
{"number_samples":4,"species":"human","abstract":"The special AT-rich sequence-binding protein 2 (SATB2) is a protein that binds to the nuclear matrix attachment region of the cell and regulates gene expression by altering chromatin structure.  We show that ectopic expression of SATB2 in normal human bronchial epithelial cell-line BEAS2B  increased anchorage-independent growth and cell migration,RNA sequencing analyses of SATB2 regulated genes revealed the enrichment of those involved in cytoskeleton, cell adhesion and cell-movement pathways. Overall design: Total RNA samples from 2 vector transfected clonesand 2 SATB2 transfected were converted into cDNA libraries using a Tru-seq RNA Sample Preparation v2 Kit (Illumina, San Diego, CA). Reads were aligned to Ensemble gene model (Homo_sapiens.GRCh37.71.gtf) using HTseq (0.6.1.p.1)","project":"SRP066959"}
{"number_samples":12,"species":"human","abstract":"Stem cell differentiation strategies and optimization for generating lineage-specific cells and tissues most frequently rely on a three-dimensional embryoid body (EB) intermediate. We previously applied nanotechnology tools of photolithography to generate custom microarrays that allow high throughput uniform formation of EBs of custom size for precise downstream analysis. Formation of EBs of 200 or 500 micron size revealed distinct morphological differences that are single or multicystic cores, respectively, independent of method of formation from single cells or two-dimensional (2D) clusters. Here we utilize photolithographic array generated EBs to obtain 3D cultures under a standardized platform for transcriptome analysis to compare EB size and the method of EB formation from single cells or mechanically passaged 2D clusters. Our analysis evaluates RNA expression in EBs formed from the human embryonic stem cell (hESC) line WA09 and from ethnically diverse human induced pluripotent stem cell lines (ED-iPSC) of African American and Hispanic Latino ethnicity recently derived in our laboratory. This is the first comprehensive study on EB transcriptomes including multiple size parameters, EB formation methodologies, and ethnicities. Our analysis indicates upregulation of genes involved in wound healing for mechanically passaged cells and of genes for embryonic tube formation in 500 micron multicystic EBs. We propose that EB maturation may be a longer process then previously realized. In addition, the type or extent of maturation possible may be influenced by EB size, with larger EBs capable of more extensive remodeling as revealed by multicystic morphology and initiation of early tube formation pathways while retaining pluripotency status. We anticipate that this information will be broadly useful to the stem cell and bioengineering communities in optimization of tissue engineering with pluripotent stem cells and understanding sources of variation. Overall design: mRNA profiles by RNA-seq from embryoid bodies generated by different methods","project":"SRP066994"}
{"number_samples":10,"species":"human","abstract":"LMO2 is a component of multisubunit DNA-binding transcription factor complexes that regulate gene expression in hematopoietic stem and progenitor cell development. Enforced expression of LMO2 causes leukemia by inducing hematopoietic stem cell-like features in T-cell progenitor cells, but the biochemical mechanisms of LMO2 function have not been fully elucidated.  In this study we systematically dissected the LMO2/LDB1 binding interface to investigate the role of this interaction in T-cell leukemia.  Alanine scanning mutagenesis of the LIM interaction domain of LDB1 revealed a discrete motif R320LITR required for LMO2 binding. Most strikingly, co-expression of full length, wild type LDB1 increased LMO2 steady state abundance, whereas co-expression of mutant proteins deficient in LMO2 binding compromised LMO2 stability. These mutant LDB1 proteins also exerted dominant negative effects on growth and transcription in diverse leukemic cell lines. Raw gene expression data on HSB-2 cells is presented here. Overall design: RNAseq were performed on HSB cell lines to examine their expression patterns","project":"SRP067173"}
{"number_samples":30,"species":"human","abstract":"Purpose: The goal of this study was to map the pathway of mRNA decay by human RNase L Methods: Total RNA was extracted (RNeasy kit, Qiagen). RNA integrity was verified by an RNA 6000 Nano Chip, using BioAnalyzer and 2100 Expert software (Agilent Technologies). The mRNA was enriched by oligo-dT pulldown from total RNA, followed by fragmentation, adapter ligation, PCR amplification, and finally sequencing on Illumina HiSeq 2000 platform. For sequencing introns, the oligo-dT pulldown step was replaced with Ribo-Zero rRNA removal (Illumina). Sequencing reads were mapped to the human genome hg19 using TopHat 2  set to map stranded reads with default parameters. Mapped read counts were obtained using HTseq-count run in union mode. Results: We developed an approach for transcriptome-wide profiling of RNase L activity in human cells and identified hundreds of direct RNA targets and non-targets. We show that this RNase L-dependent decay (RLDD) selectively affects transcripts regulated by miR-17/miR-29/miR-200 and other microRNAs that function as suppressors of mammalian cell adhesion and proliferation. RNase L mimics the effects of these microRNAs and acts as a suppressor of proliferation and adhesion in mammalian cells. Conclusions: Our data suggest that RLDD serves to establish an anti-proliferative state via destabilization of the microRNA-regulated transcriptome. Overall design: Human mRNA profiles from HeLa, T47D and HAP1 cells were generated by deep sequencing using Illumina Illumina HiSeq 2000.","project":"SRP067214"}
{"number_samples":8,"species":"human","abstract":"Chromodomain helicase DNA-binding protein 8 (CHD8) was previously identified as a leading autism spectrum disorder (ASD) candidate gene by whole-exome sequencing and subsequent targeted-sequencing studies. The present study used small interfering RNA-mediated knockdown of CHD8 in the human neural progenitor cell line, SK-N-SH, in order to determine the consequences of altered expression of this gene. RNA sequencing was performed following CHD8 knockdown to identify differentially expressed genes and affected molecular pathways resulting from this manipulation. This study sheds light on the functional consequences of altered CHD8 expression and may provide insight into the molecular underpinnings of autism spectrum disorder.","project":"SRP067221"}
{"number_samples":3,"species":"human","abstract":"RNAseq of human, bat and pig following ebolavirus infection","project":"SRP067312"}
{"number_samples":6,"species":"human","abstract":"A549 RNA-seqs","project":"SRP067318"}
{"number_samples":9,"species":"human","abstract":"Mesenchymal stromal cells (MSC) are increasingly used as an investigative therapeutic product for immune disorders and degenerative disease. Typically, MSC are isolated from human tissue, expanded in culture and cryopreserved until usage. The safety and efficacy of MSC therapy will depend on the phenotypical and functional characteristics of MSC. The freeze-thawing procedure may change these characteristics. Furthermore, the microenvironment that the cells encounter after administration may impact the properties of MSC. It has been demonstrated that the large majority of MSC localize to the lungs after intravenous infusion, making this the site to study the effects of the in vivo milieu on administered MSC. In the present study we investigated the effect of freeze-thawing and of the mouse lung microenvironment on human adipose tissue-derived MSC. The effect of freeze-thawing on the whole genome expression profile of MSC was small and did not exceed inter-donor differences. There were no major changes in the expression of hemostatic regulators on transcriptional level, but significantly increased expression of procoagulant tissue factor on the surface of thawed adipose MSC, correlating with increased procoagulant activity of thawed cells after blood exposure in vitro and after infusion to mice. Exposure for 2h to the lung microenvironment had a major effect on MSC gene expression and affected several immunological and metabolic pathways. This indicates that MSC undergo functional changes shortly after infusion and this may influence the efficacy of MSC to modulate inflammatory responses. The results of this study demonstrate that MSC rapidly alter in response to the local milieu and that disease specific conditions may shape MSC after administration. Overall design: 3 groups, n=3 per group + 1 negative control","project":"SRP067469"}
{"number_samples":6,"species":"human","abstract":"Specific drug treatment to inhibite the SMCC7721 liver cancer cell proliferation","project":"SRP067481"}
{"number_samples":609,"species":"human","abstract":"MicroRNA down-regulation and noise regulation","project":"SRP067502"}
{"number_samples":9,"species":"human","abstract":"We used parkin –overexpressing MRC5 fibroblasts to investigate the role of mitochondria deficiency on senescence-associated gene expression. Overall design: RNA-seq analysis on proliferating and senescent Parkin-expressing MRC5 fibroblasts treated with CCCP (treated) or DMSO (Untreated).","project":"SRP067529"}
{"number_samples":21,"species":"human","abstract":"We used total RNA-sequencing (RNA-seq) to analyze ALS MN-specific gene-expression signatures in laser capture microdissected motor neurons from post-mortem lumbar spinal cords. Overall design: To obtain transcriptome-wide gene expression measurements in adult spinal motor neurons (MNs), we performed laser capture microdissection (LCM) of lumbar spinal cord sections from 13 sporadic ALS patients and 9 controls, extracted total RNA, made RNA-seq libraries and sequenced the libraries on Illumina GA II platform","project":"SRP067645"}
{"number_samples":24,"species":"human","abstract":"In a classical view of hematopoiesis, the various blood cell lineages arise via a hierarchical scheme starting with multipotent stem cells that become increasingly restricted in their differentiation potential through oligopotent and then unipotent progenitors. We developed a cell-sorting scheme to resolve myeloid (My), erythroid (Er), and megakaryocytic (Mk) fates from single CD34+ cells and then mapped the progenitor hierarchy across human development. Fetal liver contained large numbers of distinct oligopotent progenitors with intermingled My, Er and Mk fates. However, few oligopotent progenitor intermediates were present in the adult bone marrow. Instead only two progenitor classes predominate, multipotent and unipotent, with Er-Mk lineages emerging from multipotent cells. The developmental shift to an adult 'two-tier' hierarchy challenges current dogma and provides a revised framework to understand normal and disease states of human hematopoiesis. Overall design: Using SMARTseq, the expression profile of the new subsets identified in the linked study (12 samples in duplicates) were analyzed using RNA sequencing. The indicated subsets are all derived from neonatal cord blood.","project":"SRP067661"}
{"number_samples":29,"species":"human","abstract":"Objective: Although glucagon-secreting a-cells and insulin-secreting ß-cells have opposing functions in regulating plasma glucose levels, the two cell types share a common developmental origin and have overlaps in their transcriptome and epigenome profiles. Notably, destruction of one of these cell populations can stimulate repopulation via transdifferentiation of the other cell type, at least in mice, suggesting plasticity between these cell fates. Furthermore, dysfunction of both a- and ß-cells contributes to the pathophysiology of type 1 and type 2 diabetes, and ß-cell de-differentiation has been proposed to contribute to type 2 diabetes. Our objective was to delineate the molecular properties that maintain islet cell type specification yet allow for cellular plasticity.  We hypothesized that correlating cell type-specific transcriptomes with an atlas of open chromatin will identify novel genes and transcriptional regulatory elements such as enhancers involved in a- and ß-cell specification and plasticity. Methods: We sorted human a- and b-cells and performed the “Assay for Transposase-Accessible Chromatin with high throughput sequencing” (ATAC-seq) and mRNA-seq, followed by integrative analysis to identify cell type-selective gene regulatory regions. Results: We identified numerous transcripts with either a-cell- or ß-cell-selective expression and discovered the cell type-selective open chromatin regions that correlate with these gene activation patterns. We confirmed cell type-selective expression on the protein level for two of the top hits from our screen.  The “group specific protein” (GC; or vitamin D binding protein) was restricted to a-cells, while CHODL (chondrolectin) immunoreactivity was only present in b-cells.  Furthermore, a-cell- and ß-cell-selective ATAC-seq peaks were identified to overlap with known binding sites for islet transcription factors, as well as with common single nucleotide polymorphisms (SNPs) previously identified as risk loci for type 2 diabetes. Conclusions: We have determined the genetic landscape of human a- and ß-cells based on chromatin accessibility and transcript levels, which allowed for detection of novel a- and ß-cell signature genes not previously known to be expressed in islets. Using fine-mapping of open chromatin, we have identified thousands of potential cis-regulatory elements that operate in an endocrine cell type-specific fashion. Overall design: ATAC-seq on 3 human alpha cell samples, 3 human beta cell samples, and 2 human acinar cell samples.  RNA-seq on 7 human alpha cell samples and 8 human beta cell samples.","project":"SRP067701"}
{"number_samples":8,"species":"human","abstract":"we find METTL3 associates with polyribosomes and promotes translation. METTL3 depletion inhibits translation, and both wild-type and catalytically inactive METTL3 promote translation when tethered to the 3' untranslated region (UTR) of a reporter mRNA. Mechanistically, METTL3 enhances mRNA translation through an interaction with the translation initiation machinery. Overall design: m6A seq in A549 and H1299 cells, RNA seq in METTL3 knockdown cells","project":"SRP067836"}
{"number_samples":16,"species":"human","abstract":"Assess the on- and off-target effects of dox-inducible CRISPR/Cas9 and CRISPRi constructs in a human iPS cell line.","project":"SRP067928"}
{"number_samples":30,"species":"human","abstract":"In this report, Tompkins et al describe the derivation, differentiation stage-specific purification, and genome-wide analysis of cardiomyocytes derived from hESCs. Key features of the molecular programs that define human cardiac muscle cell differentiation were described and researchers observed that cells may harbor epigenetic DNA methylation “memories” that reflect the gene activation history of important developmental genes. Overall design: For RNA-seq. Cardiomyocyte differentiation from human embryonic stem cells (H7). 11 time point pilot time series. D3 and D4 samples FACS sorted for primitive and cardiac mesoderm isolation, respectively. Data from negatives sorts (minus) included as well.","project":"SRP068078"}
{"number_samples":20,"species":"human","abstract":"Coordination of a complex series of transcriptional, structural and signaling events culminates in cellular senescence, a crucial tumor suppressor mechanism. We have discovered a repressor complex composed of TBX3 and CAPERa which functions upstream of the RB and p53 effector pathways and is required to prevent senescence of primary cells and in mouse embryos. TBX3/ CAPERa directly binds and represses transcription and chromatin structure of genes in multiple senescence pathways and the LncRNA UCA1, which we have identified as a novel tumor suppressor. The TBX3/ CAPERa  complex is physically disrupted in oncogene induced senescence, providing a new molecular mechanism for derepression of prosenescence pathways in this system. Our results provide new insight into the oncogenic properties of TBX3, and are the first demonstration of CAPERa and UCA1 function in vivo. Overall design: mRNA Seq based gene differential expression analysis of two sample types (TBX3, Caper) relative to control and two sample types (pCDNA3.1, UCA1) relative to each other.","project":"SRP068139"}
{"number_samples":7,"species":"human","abstract":"CNBP is a eukaryote-conserved nucleic-acid binding protein required in mammals for embryonic development. It contains seven CCHC-type zinc-finger domains and was suggested to act as a nucleic acid chaperone, as well as a transcription factor. Here, we identify all CNBP isoforms as cytoplasmic messenger RNA (mRNA)-binding proteins. Using Photoactivatable Ribonucleoside Enhanced Cross-linking and Immunoprecipitation, we mapped its binding sites on RNA at nucleotide-level resolution on a genome-wide scale and find that CNBP interacted with 3961 mRNAs in human cell lines, preferentially at a G-rich motif close to the AUG start codon on mature mRNAs. Loss- and gain-of-function analyses coupled with system-wide RNA and protein quantification revealed that CNBP did not affect RNA abundance, but rather promoted translation of its targets. This is consistent with an RNA chaperone function of CNBP helping to resolve secondary structures, thus promoting translation. Overall design: CNBP protein knockdown and RNA-seq","project":"SRP068194"}
{"number_samples":20,"species":"human","abstract":"Transcriptomic profiling of 19 healthy, term human placentas collected from the Rhode Island Childs Health Study (RICHS).","project":"SRP068290"}
{"number_samples":4,"species":"human","abstract":"Purpose: To characterize transcriptional changes associated with inhibition of Dot1l in 2 inv(16) patient AML samples Methods: We sequenced mRNA from patient samples that were exposed to 5 uM EPZ004777 or DMSO control for 7  days. Results: Inhibition of Dot1l leads to gene expression changes in genes related to cell growth and cell cycle. Overall design: Examination of mRNA levels between cells treated with  5 uM EPZ004777 or DMSO control","project":"SRP068318"}
{"number_samples":6,"species":"human","abstract":"The goal of this study was to determine IGF2BP3 RNA targets in human B-cell Acute Lymphocitic Leukemia cell models. Overall design: Included are iCLIP-seq libraries for IGF2BP3 from RS4;11 and REH B-ALL cell samples and RNA-seq data sets from control and IGF2BP3 knockdown RS4;11 B-ALL cell lines","project":"SRP068523"}
{"number_samples":1,"species":"human","abstract":"HUVEC were treated with either CA, ABA, or ABTAA and analyzed for their characteristics","project":"SRP068824"}
{"number_samples":18,"species":"human","abstract":"Wharton’s jelly derived Mesenchymal Stem Cells (MSCs) isolated from newborns with intrauterine fetal growth restriction were previously shown to exert anabolic features including insulin hypersensitivity. Here, we extend these observations and demonstrate that MSCs from small for gestational age (SGA) individuals have decreased mitochondrial oxygen consumption rates. Comparing normally grown and SGA MSCs using next generation sequencing studies, we measured global transcriptomic and epigenetic profiles and identified in SGA, E2F1 as an over-expressed transcription factor regulating oxidative metabolism. We further show that E2F1 regulates the differential transcriptome found in SGA and is associated with the activating histone marks H3K27ac and H3K4me3. One of the key genes regulated by E2F1 was found to be the fatty acid elongase ELOVL2, a gene involved in the endogenous synthesis of docosahexaenoic acid (DHA). Overall design: ChIP-seq of E2F1, H3K27ac, H3K27me3, H3K36me3, H3K4me1 and H3K4me3 in MSCs from 3 AGA and 3 SGA samples. RNA-seq of basal, siCTRL and siE2F1 conditions in the same 6 MSC lines.","project":"SRP068942"}
{"number_samples":2,"species":"human","abstract":"Peripheral Blood Lymphocyte","project":"SRP069016"}
{"number_samples":2,"species":"human","abstract":"N/A","project":"SRP015761"}
{"number_samples":12,"species":"human","abstract":"T-cell Acute Lymphoblastic Leukemia (T-ALL) is a heterogeneous group of hematological tumors composed of distinct subtypes that vary in their genetic abnormalities, gene expression signatures and prognoses. However, it remains unclear whether T-ALL subtypes differ at the functional level, and as such T-ALL treatments are uniformly applied across subtypes leading to variable responses between patients. Here we reveal the existence of a subtype-specific epigenetic vulnerability in T-ALL whereby a particular subgroup of T-ALL characterized by expression of the oncogenic transcription factor TAL1 is uniquely sensitive to variations in dosage and activity of the histone 3 lysine 27 (H3K27) demethylase UTX. Specifically, we identify UTX as a co-activator of TAL1. Furthermore, we demonstrate that UTX, previously described as a tumor suppressor in T-ALL, is in fact a potent oncogene essential for maintaining the leukemic phenotype of TAL1-positive (but not TAL1-negative) T-ALL. Exploiting this subtype-specific epigenetic vulnerability, we designed a novel therapeutic approach based on UTX inhibition through in vivo administration of an H3K27 demethylase inhibitor that is highly effective against TAL1-positive primary human leukemia. These findings provide the first opportunity to develop personalized epigenetic therapy for T-ALL patients. Overall design: RNA-seq expression analysis (three treatments and matched controls; two replicates of each).","project":"SRP062773"}
{"number_samples":22,"species":"human","abstract":"Background:  A subset of patients with atopic dermatitis (AD) is prone to disseminated herpes simplex virus (HSV) infection, i.e. eczema herpeticum (ADEH+). Biomarkers that identify ADEH+ are lacking. Objective: To search for novel ADEH+ gene signatures in peripheral blood mononuclear cells (PBMCs). Methods: A RNA-sequencing (RNA-seq) approach was applied to evaluate global transcriptional changes using PBMCs from ADEH+ and AD without a history of EH (ADEH-). Candidate genes were confirmed by qPCR or ELISA. Results:  ADEH+ PBMCs had distinct changes to the transcriptome when compared to ADEH- PBMCs following HSV-1 stimulation: 792 genes were differentially expressed at a false discovery rate (FDR) < 0.05 (ANOVA), and 15 type I and type III interferon (IFN) genes were among the top 20 most down-regulated genes in ADEH+.  We further validated that IFN-a and IL-29 mRNA and protein levels were significantly decreased in HSV-1 stimulated PBMCs from ADEH+ compared to ADEH- and normal. Ingenuity pathway analysis (IPA) demonstrated that the up-stream regulators of type I and type III IFNs, IRF3 and IRF7 was significantly inhibited in ADEH+ based on the down-regulation of their target genes. Furthermore, we found that gene expression of IRF3 and IRF7 were significantly decreased in HSV-1 stimulated PBMC from ADEH+ subjects. Conclusions: PBMCs from ADEH+ have a distinct immune response following HSV-1 exposure compared to ADEH- .  Inhibition of the IRF3 and IRF7 innate immune pathways in ADEH+ may be the important mechanisms for increased susceptibility to disseminated viral infection. Overall design: Expression profiles of PBMCs were taken from 5 AD patients with a history of disseminated herpes simplex virus (HSV) infection, i.e. eczema herpeticum (ADEH+) and 6 AD patients who don’t have a history of disseminated HSV infection (ADEH-).  PBMC samples from each subject were divided in two and were treated with and without HSV-1 virus for 21 hours before processing for total RNA.","project":"SRP045569"}
{"number_samples":26,"species":"human","abstract":"Transcriptome sequencing of Chronic Phase and Blast Crisis CML, normal cord blood cells, and normal cord blood cells transduced with lentiviral vectors","project":"SRP028528"}
{"number_samples":8,"species":"human","abstract":"CHD8, encoding Chromodomain helicase DNA binding protein 8, is a top autism spectrum disorders (ASDs) risk gene. To better understanding the molecular links between CHD8 functions and ASD, we have applied the CRISPR/Cas9 technology to knockout one copy of CHD8 in induced pluripotent stem cells (iPSCs) to mimic the loss of function status that would exist in the developing human embryo prior to neuronal differentiation. Transcriptome profiling (RNA-seq) in neural progenitors and early differentiating neurons revealed that CHD8 hemizygosity (CHD8+/-) affected the expression of several thousands of genes, enriched for functions of neural development, ß-catenin/Wnt signaling, extracellular matrix, and skeletal system development. Moreover, CHD8 regulates multiple genes implicated in ASD, schizophrenia and genes associated with brain volume. Overall design: iPSCs derived from a healthy subject were transduced with CRISPR/Cas9 vectors with  single guide RNA sequences to target the N-terminal of CHD8 protein to generate truncated mutation seach of the two target sequences. Two clones, one with a 2-bp (KO1) and the other with a 10-bp (KO2) heterozygous deletion were found.The CHD8+/- iPSC lines were used to generate NPCs and early differentiating neurons for RNA-seq analysis, together with samples prepared from the parental clones, for a total of 8 samples (two biological replicates of wild-type (WT) and CHD8+/- at two neurodevelopmental stages).","project":"SRP061880"}
{"number_samples":6,"species":"human","abstract":"RNA-seq of uterine adenosarcomas","project":"SRP063460"}
{"number_samples":2,"species":"human","abstract":"RNA sequencing of polymorphous low-grade adenocarcinoma of the salivary gland","project":"SRP045785"}
{"number_samples":17,"species":"human","abstract":"In excess of 12% of human cancer incidents have a causal viral cofactor. Epidemiological studies of idiopathic human cancers indicate that additional tumor viruses remain to be discovered. Recent advances in sequencing technology have enabled systematic screenings of human tumor transcriptomes for viral transcripts. However, technical problems such as low abundances of viral transcripts in large volumes of sequencing data, viral sequence divergence, and homology between viral and human factors significantly confound identification of tumor viruses.  We have developed a novel computational approach for detecting viral transcripts in human tumors and conducted the first systematic search for viruses in neuroblastoma, the most common cancer in infancy. The striking diversity in clinical progression of this malignancy as well as previous epidemiological and virological findings are highly suggestive of a viral cofactor. We computationally mapped deeply sequenced transcriptomes of neuroblastoma and several cancers with known viral cofactors to the human and all known viral genomes. We analyzed the homologous context of putative viral transcripts in a new and sensitive manner in order to detect both known and unknown viruses under consideration of the aforementioned confounds.","project":"ERP002063"}
{"number_samples":3,"species":"human","abstract":"Despite 20 years since its discovery, the gene responsible for Huntington’s Disease, HTT, has still not had its function or transcriptional profile completely characterized. In response to a recent report by Ruzo et al. of several novel splice forms of HTT in human embryonic stem cell lines, we have analyzed a set of mRNA sequencing datasets from post mortem human brain from Huntington’s disease, Parkinson’s disease, and neurologically normal control subjects to evaluate support for previously observed and to identify novel splice patterns. A custom analysis pipeline produced supporting evidence for some of the results reported by two previous studies of alternative isoforms as well as identifying previously unreported splice patterns. All of the alternative splice patterns were of relatively low abundance compared to the canonical splice form. Overall design: 29 Huntington's Disease, 29 Parkinson's Disease, and 50 Neurologically normal control samples from human post-mortem prefrontal cortex","project":"SRP061397"}
{"number_samples":7,"species":"human","abstract":"We performed differential expression analyses from RNA-seq data derived from isogenic p53 wild-type and p53-knockdown triple negative breast tumors Overall design: patient-derived isogenic human tumor lines diffring only in p53 status were implanted into mouse mammary glands. RNA-Seq was performed on 4 p53-deficient and 3 p53 wild-type tumors.","project":"SRP067934"}
{"number_samples":24,"species":"human","abstract":"A recent study by Castelo-Branco, P., et al. Methylation of the TERT promoter and risk stratification of childhood brain tumours: an integrative genomic and molecular study. Lancet Oncol 2013;14:534-542 found upstream of the transcription start site (UTSS) hypermethylation of TERT is associated with tumor progression and poor prognosis in paediatric brain tumours. They interpreted that the UTSS region of telomerase reverse transcriptase (TERT) gene is a potentially accessible biomarker for various cancers. This study, we aimed to explore the role of TERT in hepatocellular carcinoma (HCC) and to investigate whether the UTSS region of TERT promoter shows the same methylation pattern in HCC. Overall design: We analyzed a methylation assay of TERT including the UTSS region in 125 paired HCC samples using Mass Array EpiTyper (Sequenom, SanDiego, CA, USA). To obtain the relationship between TERT promoter methylation status and TERT expression level, we analysed a validation group of 12 paired HCC samples and acquired the FPKM value of TERT gene.","project":"SRP050551"}
{"number_samples":10,"species":"human","abstract":"WI-38 histone variant H2AJ function","project":"SRP048685"}
{"number_samples":11,"species":"human","abstract":"Hepatocellular carcinoma (HCC) is the most common kind of liver cancer in the world. Portal vein tumor thrombus (PVTT) is one of the most serious complications of HCC and strongly correlated to a poor prognosis for HCC patients. However, the detailed mechanisms of PVTT development still remains to be explored. In this study, we presented the large scale transcriptome analysis of 11 HCC with PVTT patients by RNA-sequencing. The dysregulated genes between HCC and PVTT suggested that the ECM-Receptor Interaction was correlated to the venous metastases of HCC. Among all the recurrent alternative splicing events, we identified exon 6 skipping of RPS24, which is likely to be a cancer driver. We also identified five common fusion genes between HCC and its corresponding PVTT samples, including ARID1A-GPATCH3, MDM1-NUP107, PTGES3-RARG, PRLR-TERT and C9orf3-TMC1. All of the findings broaden our knowledge in PVTT development and may also contribute to the diagnosis and treatment for HCC patients with PVTT. Overall design: In this study, we presented the large scale transcriptome analysis of 11 HCC with PVTT patients by RNA-sequencing.","project":"SRP058626"}
{"number_samples":19,"species":"human","abstract":"Most variants associated to diseases are located in non-coding regions of the genome, and are thought to be regulatory in nature. Cis-regulatory SNPs (cis-rSNPs) impacting transcription can be identified through the mapping of differences in allelic expression (AE). We mapped common cis-rSNPs of protein coding and non-coding genes in 3 distinct cell-types. We show 70% sharing across tissues and similar genetically controlled transcription for protein-coding genes and lincRNAs. Candidate cis-rSNPs altering the expression of 42 non-coding RNA overlap SNPs underlying GWAS associations for 39 diseases. We uncover a new class of cis-rSNPs leading to disruption of footprint-derived de novo motifs, predominantly bind by repressive factors and implicated in disease susceptibility through overlaps with GWAS SNPs. Finally, we provide proof-of-principle for a new approach for genome-wide functional validation of transcription factor – SNP interactions. We perturbed NF?B action in lymphoblasts and identified 489 cis-regulated transcripts with altered AE after NFkB perturbation. Altogether, we performed a comprehensive analysis of cis-variation in four cell-populations, and provide new tools for the identification of functional variants associated to complex diseases. Overall design: Mapping cis-rSNPs across 3 distinct cell types in humans","project":"SRP044271"}
{"number_samples":42,"species":"human","abstract":"We did mRNA profiling and  showed that an interferon-inducible gene's deletion and under-expression is significantly associated with shorter time to tumor recurrence, shorter progression-free survival, and shorter overall survival in  HCC patients. Overall design: We got the samples from HCC patients' matched primary and recurrent tumors (n=10) and primary tumors from non-recurrent patients (n=15), we conducted the RNAseq of those samples, and did the comparison.","project":"SRP040998"}
{"number_samples":1,"species":"human","abstract":"This study evaluates the fidelity of colon cancer xenografts grown in immunodeficient mice in comparison to their parental human tumor","project":"SRP027538"}
{"number_samples":12,"species":"human","abstract":"MCF7 cells exposed in 0.1% hypoxia for 72h and then stained and sorted into CA9+ve and CA9-ve populations and a differential gene expression analysis was performed between the 2 cell populations.","project":"ERP002021"}
{"number_samples":12,"species":"human","abstract":"RNA-seq of Human lung cell line A549 alone as control, or infected with human adenovirus serotype 6, or infected with human adenovirus serotoype 26","project":"SRP064863"}
{"number_samples":4,"species":"human","abstract":"Activation of CD4 T cells is a reaction to challenges such as microbial pathogens, cancer and toxins that defines adaptive immune responses. The roles of T cell receptor crosslinking, intracellular signaling, and transcription factor activation are well described, but the importance of post-transcriptional regulation by RNA-binding proteins (RBPs) has not been considered in depth. We describe a new model expanding and activating primary human CD4 T cells and applied this to characterizing activation-induced assembly of splicing factors centered on U2AF2. We immunoprecipitated U2AF2 to identify what mRNA transcripts were bound as a function of activation by TCR crosslinking and costimulation. In parallel, mass spectrometry revealed the proteins incorporated into the U2AF2-centered RNA/protein interactome. Molecules that retained interaction with the U2AF2 complex after RNAse treatment were designated as “central” interactome members (CIMs). Mass spectrometry also identified a second class of activation-induced proteins, “peripheral” interactome members (PIMs), that bound to the same transcripts but were not in physical association with U2AF2 or its partners. siRNA knockdown of two CIMs and two PIMs caused changes in activation marker expression, cytokine secretion, and gene expression that were unique to each protein and mapped to pathways associated with key aspects of T cell activation. While knocking down the PIM, SYNCRIP, impacts a limited but immunologically important set of U2AF2-bound transcripts, knockdown of U2AF1 significantly impairs assembly of the majority of protein and mRNA components in the activation-induced interactome. These results demonstrated that CIMs and PIMs, either directly or indirectly through RNA, assembled into activation-induced U2AF2 complexes and play roles in post-transcriptional regulation of genes related to cytokine secretion. These data suggest an additional layer of regulation mediated by the activation-induced assembly of RNA splicing interactomes that is important for understanding T cell activation. Overall design: RNAseq of total RNA from a primary CD4 T cell culture at rest and 48 hours after anti-CD3/CD28 bead activation","project":"SRP049462"}
{"number_samples":10,"species":"human","abstract":"Many addictive drugs such as nicotine mediate reward and reinforcing mechanisms within the mesolimbic pathway involving midbrain dopaminergic (mDA) neurons via nicotinic acetylcholine receptors (nAChRs). Previously, genome-wide association analyses (GWAs) identified several nucleotide polymorphism (SNP) associated with increased risk of addictive phenotypes including the nonsynonymous rs16969968 SNP encoding D398->N398 variation in the CHRNA5 gene (Bierut et al., 2008; Saccone et al., 2007) . However, the etiology and the pathophysiology associated with the N398 allele is unknown. Previous animal studies using a knock-in of human CHRNA5 N398 revealed increased nicotine consumption. However, given the evolutionary distance between mice and humans, identifying intrinsic factors that affect addiction may correlate poorly, limiting the conclusiveness of these studies. In this study, induced pluripotent stem cell (iPSC) lines were derived from cryopreserved lymphocytes drawn from patients with homozygous minor alleles (N398) of rs16969968 and demonstrated nicotine addiction as well as age- and gender-matched unaffected controls carrying the major allele (D398). Cultures of primarily mDA neurons were prepared from these iPSC lines. Gene expression analysis using RT-PCR and RNAseq revealed similar expression of mRNAs encoding subunits of nAChR in mDA neurons derived from both D398 and N398 iPSC lines, however, gene ontology (GO) analysis revealed an increased enrichment of genes associated with neuroactive ligand-receptor interactions and Ca2+ signaling pathways in N398 neurons. Moreover, the N398 neuronal population responded more actively to application of nicotine in both electrophysiological analysis and Ca2+-imaging analysis. Together, these results suggest that the N398 variation affects Ca2+-signaling in neurons, which may explain the predisposition of subjects carrying this mutation for addictive behavior in subjects carrying the nonsynonymous rs16969968 SNP (Sherva et al., 2008).","project":"SRP040275"}
{"number_samples":94,"species":"human","abstract":"Variation in the expression level and activity of genes involved in drug disposition and action (“pharmacogenes”) can affect drug response and toxicity, especially when in tissues of pharmacological importance.  Previous studies have relied primarily on microarrays to understand gene expression differences, or have focused on a single tissue or small number of samples. The goal of this study was to use RNA-seq to determine the expression levels and alternative splicing of 389 PGRN pharmacogenes across four tissues (liver, kidney, heart and adipose) and lymphoblastoid cell lines (LCLs), which are used widely in pharmacogenomics studies. Analysis of RNA-seq data from different individuals across the 5 tissues (N = 139 samples) revealed substantial variation in both expression levels and splicing across samples and tissue types. This in-depth exploration also revealed 183 splicing events in pharmacogenes that were previously not annotated.  Overall, this study serves as a rich resource for the research community to inform biomarker and drug discovery and use. Overall design: mRNA profile of 389 genes in 24 liver, 20 kidney (cortex), 25 heart (left ventricle), 25 adipose (subcutaneous) samples, and 45 lymphoblastoid cell lines (LCLs). The raw data for the LCL Samples will be available through dbGap under phs000481.","project":"SRP060355"}
{"number_samples":18,"species":"human","abstract":"RNA-seq study of a gastric carinoma cell line (AGS) with or without Epstein-Barr Virus infection as well as AGS cells expressing only viral noncoding RNAs (BARTs).","project":"SRP060253"}
{"number_samples":4,"species":"human","abstract":"We studied the effect of knocking down SUZ12 on the responsiveness of IFNg target genes Overall design: 12 cell lines were transfected with siRNA against SUZ12 (siSUZ12) or siCtrl and left untreated or treated with IFNg for 6 hours","project":"SRP056498"}
{"number_samples":12,"species":"human","abstract":"We performed RNA-Seq for the corpus callosum sampled from 12 individuals. The samples were dissected from the frozen postmortem brain. The 12 individuals were matched by their age, sex and ethnicity for the postmortem brain samples, but differed in their disease status with half of the subjects were diagnosed with autism spectrum disorders. Overall design: RNA-Seq for the corpus callosum sampled from 12 individuals.","project":"SRP048683"}
{"number_samples":6,"species":"human","abstract":"We performed RNA-Seq analyses on a primary xenograft tumor line derived from a chemoresistant bladder urothelial carcinoma patient. These xenografts were treated with vehicle as control, gemcitabine-cisplatin (GC) chemotherapy and celecoxib/GC combination therapy as indicated.","project":"SRP048630"}
{"number_samples":6,"species":"human","abstract":"Nicotine stressed MCF-10A cells Transcriptome Sequencing.","project":"SRP019980"}
{"number_samples":3,"species":"human","abstract":"We performed RNA-Seq for the BA9/40/AMY for a typically developing individual #5407. The samples were dissected from the frozen postmortem brain. Overall design: We performed RNA-Seq for the BA9/40/AMY for a typically developing individual #5407.","project":"SRP050089"}
{"number_samples":25,"species":"human","abstract":"Next generation sequencing of 28 thymic epithelial tumors (TETs) revealed a high frequency of GTF2I missense mutation (chr7:74146970T/A) in A thymomas, a  relatively indolent subtype. The GTF2I mutation was confirmed in 82% of A and 74% of AB thymomas in a series of 274 TETs but was rare in aggressive subtypes, where recurrent mutations of known cancer genes were identified. Therefore, GTF2I mutation correlated with a better survival. GTF2I Beta and Delta isoforms were expressed in TETs and both mutant isoforms were able to stimulate cell proliferation in vitro. Thymic carcinomas presented a higher number of mutations than thymomas (average 43.5 and 18.4, respectively). Recurrent mutations of known cancer genes, including TP53, CYLD, CDKN2A, BAP1 and PBRM1 were identified in thymic carcinomas. These findings will complement the diagnostic work up of these rare tumors, and also help the development of a molecular classification, and assessment of prognosis and treatment strategies. Overall design: Tumor samples of 286 patients were collected from 4 different institutions: National Cancer Institute (Bethesda MD), Pisa University Hospital (Pisa, Italy), Padua University Hospital (Padua, Italy) and IRCCS Istituto Clinico Humanitas (Rozzano, Italy).","project":"SRP042184"}
{"number_samples":1,"species":"human","abstract":"To study changes in gene expression and alternative splicing in obesity","project":"SRP036595"}
{"number_samples":58,"species":"human","abstract":"Here we present an RNAseq analysis of human bone samples, obtained from iliac crest needle biopsies, to yield the first in vivo interrogation of all genes and pathways that may be altered in bone with aging and E therapy in humans. Overall design: 58 healthy women were studied, including 19 young women (mean age ± SD, 30.3 ± 5.4 years), 19 old women (73.1 ± 6.6 years), and 20 old women treated with 3 weeks of E therapy (70.5 ± 5.2 years).  Using generally accepted criteria (false discovery rate [q] < 0.10), aging altered a total of 678 genes and 12 pathways, including a subset known to regulate bone metabolism (e.g., Notch).","project":"SRP063477"}
{"number_samples":1,"species":"human","abstract":"The objective of this study was to identify and analyze somatic variations in healthy human tissues. For this we obtained paired tissue from nine individuals. Four tissue pairs represented neuronal and non-neuronal parts of the brain - frontal cortex (FC) and corpus callosum (CC), respectively. These were from post-mortem, road accident victims. The other five tissue pairs were from blood and saliva of unrelated healthy adults. Exome enrichment was performed on DNA isolated from these 18 samples using Illumina Tru-Seq chemistry and then sequenced on Hi-Seq2000 (100 base pair, paired-end). The sequence reads were filtered for quality and then aligned against the reference human genome (hg19) using BWA. We observed brain to harbor more somatic variations than other tissue types – and majority of them were transversions likely to have occurred due to oxidative stress.","project":"SRP045655"}
{"number_samples":18,"species":"human","abstract":"<p/>","project":"SRP057446"}
{"number_samples":106,"species":"human","abstract":"In an effort to identify biomarkers of recurrence, we have performed global RNA-sequencing on 106 formalin-fixed, paraffin-embedded (FFPE) prostatectomy samples from 100 patients at three independent sites, and identified a new set of biomarkers of biochemical recurrence composed of a 24-gene panel. We validated this 24-gene panel on an independent publicly available dataset of 140 patients and this new panel outperformed previously published markers based on cell proliferation gene sets in terms of prediction of biochemical recurrence (BCR).  In addition we identified differences in gene expression between Gleason Pattern 4+3 and Gleason Pattern 3+4 tumors. Overall design: 106 samples from 100 patients were sequenced.  Six samples were sequenced from two independent RNA preps and libraries.  All samples were taken from FFPE Radical Prostatectomies.  Samples were obtained from Atlanta VA Medical Center, U. Toronto Sunnybrook Research Centre, and Moffitt Cancer Center.","project":"SRP036848"}
{"number_samples":35,"species":"human","abstract":"Caraterisation of tumors presenting SMARCA4 inactivation, underlying a group of undifferentiated thoracic malignancies transcriptomically related to BAF-deficient rhabdoid tumors.","project":"SRP052896"}
{"number_samples":11,"species":"human","abstract":"Recurrent gene fusions have been described in many tumor types but have been difficult to identify in high-grade serous ovarian cancer (HG-SC), a cancer known for its significant heterogeneity and genome instability. Here, we combined high-throughput transcriptome sequencing with experimental validation to identify a cancer-specific and inter-chromosomal fusion gene CDKN2D-WDFY2 that is recurrent at 25% among HG-SC patients. CDKN2D is a cell-cycle modulator that is also involved in DNA repair while WDFY2 is known to modulate AKT interactions with its substrates. Transfection of the fusion construct led to loss of CDKN2D and WDFY2 protein expression, and gain of an aberrant short WDFY2 protein isoform under the control of CDKN2D promoter. This short WDFY2 in turn led to marked alterations of the PI3K/AKT pathway, a signaling pathway with major implications in oncogenesis. Thus, CDKN2D-WDFY2 fusion could be an important molecular signature for understanding and classifying sub-lineages among heterogenous HG-SC.","project":"SRP056792"}
{"number_samples":6,"species":"human","abstract":"Superior frontal gyrus grey and white matter","project":"SRP033291"}
{"number_samples":5,"species":"human","abstract":"RNA-Sequencing of neonatal keratinocytes depleted of MPZL3 or FDXR with control samples as well","project":"SRP065120"}
{"number_samples":18,"species":"human","abstract":"The goal is to identify estrogen-dependent gene expression patterns using RNAseq for 3 different ERa forms in human osteoblasts. Overall design: We analyzed 3 different forms of estrogen receptor alpha (ERa), 1) Wild-type, 2) NERKI (DNA-binding mutant) and 3) NOER (Nuclear Only ERa, not membrane).  Each was infected into hFOB (human osteoblast) cells and treated with either vehicle or E2 (10nM) (n=3 for each group/treatment).  RNAseq was then performed.","project":"SRP039909"}
{"number_samples":6,"species":"human","abstract":"RNA-seq data from human postmortem brain from rett syndrome cases and controls.","project":"SRP066394"}
{"number_samples":11,"species":"human","abstract":"Endocrine therapies target the activation of the estrogen receptor alpha (ERa) via distinct mechanisms but it is not clear whether breast cancer cells can adapt to treatment using drug-specific mechanisms. Here we demonstrate that resistance emerges via drug-specific epigenetic reprogramming. Resistant cells display a spectrum of phenotypical changes with invasive phenotypes evolving in lines resistant to the aromatase inhibitor (AI). Orthogonal genomics analysis of reprogrammed regulatory regions identifies individual drug-induced epigenetic states involving large topological domains (TAD) and the activation of super-enhancers. AI resistant cells activate endogenous cholesterol biosynthesis (CB) through stable epigenetic activation in vitro and in vivo. Mechanistically, CB sparks the constitutive activation of estrogen receptors alpha (ERa) in AI resistant cells, partly via the biosynthesis of 27-hydroxycholesterol. By targeting CB using statins, ERa binding is reduced and cell invasion is prevented. Epigenomic-led stratification can predict resistance to AI in a subset of ERa positive patients.","project":"ERP012838"}
{"number_samples":35,"species":"human","abstract":"DCIS is a non-invasive precursor lesion to invasive breast carcinoma. We still have no understanding on why only some DCIS lesions evolve to invasive cancer while others appear not to do so during the life span of the patient. Here, we performed full exome (tumor vs. matching normal), transcriptome and methylome analysis of 30 pure high-grade DCIS (HG-DCIS) and 10 normal breast epithelial samples. Sixty two percent of HG-DCIS cases displayed mutations affecting cancer driver genes or potential drivers. Mutations were observed affecting PIK3CA (21% of cases), TP53 (17%), GATA3 (7%), MLL3 (7%) and single cases of mutations affecting CDH1, MAP2K4, TBX3, NF1, ATM and ARID1A. Significantly, 83% of lesions displayed numerous large chromosomal copy number alterations, suggesting they might precede selection of cancer driver mutations. Integrated pathway-based modeling analysis of RNA-seq data allowed us to identify two DCIS subgroups (DCIS-C1 and DCIS-C2) based on their tumor intrinsic subtypes, proliferative, immune scores and in the activity of specific signaling pathways. The more aggressive DCIS-C1 (highly proliferative, basal-like or ERBB2+) displayed signatures characteristic of activated Treg cells (CD4+/CD25+/FOXP3+) and CTLA4+/CD86+ complexes indicative of a tumor-associated immune suppressive phenotype. Strikingly, all lesions showed evidence of TP53 pathway inactivation. Similarly ncRNA and methylation profiles reproduce changes observed post-invasion. Among the most significant findings we observed upregulation of lncRNA HOTAIR in DCIS-C1 lesions and hypermethylation of HOXA5 and specific SOX genes. We conclude that most HG-DCIS lesions, in spite of representing a pre-invasive stage of tumor progression, displayed molecular profiles indistinguishable from invasive breast cancer. Overall design: RNAs from 25 out of 30 (83%) pure HG-DCIS and 10 normal breast organoids (total 35 samples) were subjected to RNA-Seq analysis by using Illumina HiSeq2000 platform Please note that description of samples employed for the NGS analyses including age, race, ER/PR immunohistochemistry results, ITIL/STIL scores and PAM50 classification is provided the 'Supplementary Data1_Samples data.xlsx' (available on Superseries record)","project":"SRP058722"}
{"number_samples":57,"species":"human","abstract":"Whole transcriptome sequencing of 19 matched patient sample trios: normal oral epithelium, oral dysplasia and oral squamous cell carcinoma from the same patent. All material is from FFPE blocks that have been stained and sections and had sample histology confirmed by 2 independent pathologists. A ScriptSeq library kit was used to create strand specific 100bp paired-end sequencing libraries following RiboZero treatment to deplete rRNA. Both coding and non-coding RNA <200bp is sequenced on an Illumina HiSeq2500 with an average of 4 samples per lane.","project":"ERP005938"}
{"number_samples":2,"species":"human","abstract":"We performed both exome and transcriptome sequencing of the LNCaP and C4-2B prostate cancer cell lines. The aggressive, castration-resistant C4-2B cell line was derived from LNCaP. Exome and transcriptome sequencing revealed numerous point mutations in both cell lines, of which about half are in common for both cell lines. Furthermore, transcriptome sequencing allowed the identification of genes that are differentially expressed in both cell lines.","project":"ERP004209"}
{"number_samples":7,"species":"human","abstract":"RNAseq of LIM2405, Human colon cancer cell line, to identify transcript.","project":"SRP016110"}
{"number_samples":6,"species":"human","abstract":"These are RNAseq reads from human cell culture (neurons and fibroblasts) infected with VZV. Thus the reads are a mixture of human and viral. The study was designed to assess whether VZV was in true latency, or whether is was simply an aborted infection.","project":"SRP040110"}
{"number_samples":5,"species":"human","abstract":"RNA-Seq and microarray platforms have emerged as important tools for detecting changes in gene expression and RNA processing in biological samples. We present ExpressionPlot, a software package consisting of a default back end, which prepares raw sequencing or Affymetrix microarray data, and a web-based front end, which offers a biologically centered interface to browse, visualize, and compare different data sets. Download and Installation instructions, user’s manual, discussion group, and a prototype are available at http://expressionplot.com/.","project":"ERP000619"}
{"number_samples":19,"species":"human","abstract":"Human pluripotent stem cell derived models that accurately recapitulate neural development in vitro and allow for the generation of specific neuronal subtypes are of major interest to the stem cell and biomedical community.  Notch signaling, particularly through the Notch effector HES5, is a major pathway critical for the onset and maintenance of neural progenitor cells (NPCs) in the embryonic and adult nervous system1-3. Use of a HES5 reporter enables the isolation distinct populations of human embryonic stem (ES) cell derived NPCs that represent building blocks of cortical development in vitro4. Here, we report the transcriptional and epigenomic analysis of six consecutive stages of human ES cell differentiation along the neural lineage aimed at modeling key cell fate decisions including specification, expansion and patterning during the ontogeny of neural stem and progenitor cells. In order to dissect the regulatory mechanisms that orchestrate the stage-specific differentiation process we developed a computational framework to infer key regulators of each cell state transition based on the progressive remodeling of the epigenetic landscape and then validated these through a pooled shRNA screen. We were also able to refine our previous observations on epigenetic priming at transcription factor binding sites and show here that they are mediated by combinations of core and stage-specific factors. Taken together, we demonstrate the utility of our reference maps and outline a general framework, not limited to the context of the neural lineage, to dissect regulatory circuits of differentiation. Overall design: hESC were differentiated in vitro into 5 different NPC populations (NE, ERG, MRG, LRG and LNP) over a time period of 220 days. The hESCs and the first 3 NPC populations (NE, ERG, MRG) were subjected to ChIP-Seq for H3K4m3, H3K4me1, H3K27ac and H3K27me3 as well as RNA-Seq with two biological replicates each as well as whole genome bisulfite sequencing in singlicate. The last two populations (LRG, LNP) were both profiled by RRBS and the LRG populations also by RNA-Seq. In addition, each of the first three NPC populations were differentiated into more mature neuronal populations (NEdN, ERGdN, MRGdN). The LRG population was differentiated along the astrocyte fate (LRGdA). Each of these more mature populations was subjected to RNA-Seq in replicate. Finally, the NE and MRG populations were profiled for the transcription facctor OTX2 by ChIP-Seq.","project":"SRP048761"}
{"number_samples":18,"species":"human","abstract":"Our aim was to investigate the interaction between epidermal differentiation and VZV infection.  By means of a calcium-induced keratinocyte differentiation model and RNA-seq we show VZV infection has a profound effect on differentiating keratinocytes and hijacks the normal process of epidermal gene expression to generate a signature resembling patterns of gene expression seen in both heritable and acquired skin-blistering disorders.  Analysis of the viral transcriptome provides evidence that VZV replication in skin is tightly linked to differentiation and critically, that late viral gene expression is associated with cellular differentiation.  The experiment was performed on human primary keratinocytes under four conditions: undifferentiated/uninfected,  uninfected/differentiated, VZV-infected/undifferentiated and VZV-infected/differentiated.","project":"ERP003467"}
{"number_samples":6,"species":"human","abstract":"The goal of the study is to identify transcriptional differences between human induced motor neurons with 2, 1, or 0 full length copies of C9ORF72.","project":"SRP064149"}
{"number_samples":1,"species":"human","abstract":"RNA-seq expression analysis of transcripts encoding proteasome subunits in human CD34+ cord blood cell-derived megakaryocytes and mouse bone marrow-derived megakaryocytes. Overall design: Analysis of transcript expression in human CD34+ cord blood cell-derived megakaryocytes and mouse bone marrow-derived megakaryocytes.","project":"SRP062023"}
{"number_samples":189,"species":"human","abstract":"We analyzed gene expression profiling of lung tissue to define molecular pathway of COPD using recent RNA sequencing technology.Lung tissue was obtained from 98 COPD subjects and 91 subjects with normal spirometry. RNA isolated from these samples was processed with RNA-seq using HiSeq 2000. Gene expression measurements were calculated using Cufflinks software. Differentially expressed genes and isoforms were chosen using t-test. Some of differentially expressed genes were validated by quantitative real-time PCR. Overall design: Examination of lung tissue in COPD patients versus normal control","project":"SRP041538"}
{"number_samples":12,"species":"human","abstract":"FOXM1 is a key transcription factor regulating cell cycle progression, DNA damage response, and a host of other hallmark cancer features, but the role of the FOXM1 cistrome in driving estrogen receptor-positive (ER+) vs. ER- breast cancer clinical outcomes remains undefined. Overall design: Chromatin immunoprecipitation sequencing (ChIP-Seq) coupled with RNA sequencing (RNA-Seq) analyses was used to identify FOXM1 target genes in breast cancer cells (MCF-7) where FOXM1 expression was either induced by cell proliferation or repressed by p53 upregulation.","project":"SRP064131"}
{"number_samples":19,"species":"human","abstract":"Purpose: Calcific aortic valve stenosis (AS) is a fatal disease with currently no medical therapy. Some genes were associated with AS, but the genetic architecture of the disease has yet to be discovered. The objective of this study was to combine genome-wide association studies (GWAS) and gene expression in human valve tissues to identify new susceptibility genes of AS. Methods: A meta-analysis was performed combining the results of two independent GWAS in 474 patients that underwent aortic valve replacement from Quebec city and 486 echocardiography cases from France. The controls consisted of 3,151 publically available individuals of European ancestry. Sixty-nine SNPs selected from the meta-analysis (p < 1 x 10-4) were followed-up for replication in a third cohort of 395 cases and 404 controls. Single marker and gene set association analyses were performed. The mRNA expression levels of susceptibility genes were evaluated in 19 human aortic valves with (n=9) and without (n=10) AS by RNA sequencing. Results: Single marker analysis identified 15 SNPs with p values lower than 1 x 10-6 in the meta-analysis near the FAR2 gene on chromosome 12. At the replication stage, none of these SNPs were confirmed and three other SNPs on chromosomes 2 and 5 reached p < 0.05. Gene set analysis revealed more meaningful association with the calcium signaling pathway enriched for genes mapped to disease-associated SNPs. Genes in this pathway including F2R, GNA14, HTR4, P2RX6, and TNNC1 were found differentially expressed in valves with and without AS. Conclusions: This integrative genomic study identified new AS susceptibility genes expressed in human valve tissue. Moderate but coordinated genetic association and expression patterns were observed for genes implicated in the calcium signaling pathway and may provide new therapeutic targets to treat this frequent and rising life-threatening disease. Overall design: Samples of aortic valves were collected from 19 male patients undergoing aortic valve replacement surgery. RNA sequencing was performed using the Illumina Hiseq 2000. Three pairwise comparison among the two kind of valves were made.","project":"SRP039338"}
{"number_samples":15,"species":"human","abstract":"To identify transcripts that are aberrantly spliced in the CD34+ CMML cells with SRSF2(P95) mutation.","project":"SRP048858"}
{"number_samples":2,"species":"human","abstract":"LNCaP RNA-Seq","project":"SRP033305"}
{"number_samples":45,"species":"human","abstract":"To understand the full range of expression variation, alternative splicing, and the genetics of regulatory variation in humans, we have sequenced the genomes, exomes, and transcriptomes of 45 individuals in the Human Genome Diversity Panel (HGDP) from seven worldwide populations that span the geographic breadth of human migration history. We find few genes that are systematically differentially expressed among populations. Further, allelic expression analysis indicates that previously mapped common regulatory variants identified in eight HapMap3 populations have similar effects across HGDP populations, suggesting that the cellular effects of common variants are shared even across very diverse populations. In contrast, comparisons of alternative splicing across pairwise populations reveal highly significant differences, with ~20% of alternatively spliced exons varying significantly by population. Greater genomic differences are associated with significantly more splicing variability overall. Together, these results provide a resource and an estimate for the degree of sharing of gene expression, alternative splicing, and regulatory genetics across populations. Overall design: Lymphoblastoid cell line mRNA profiles of 45 human samples from the Human Genome Diversity Panel sequenced on an Illumina HiSeq 2000, each sample library was sequenced in duplicate to produce a merged sample bam.","project":"SRP035599"}
{"number_samples":2,"species":"human","abstract":"Noise in gene expression is a main determinant of phenotypic variability. Increasing experimental evidence suggests that genome-wide cellular constraints largely contribute to the heterogeneity observed in gene products. It is still unclear, however, which global factors mostly affect gene expression noise, and to which extent. Since eukaryotic gene expression is an energy demanding process, differences in the energy budget of each cell could determine gene expression differences. Here, we quantify the contribution of mitochondrial variability (a natural source of ATP variation) to global variability in gene expression. We find that changes in mitochondrial content can account for ~50% of the variability observed in protein levels. This is the combined result of the effect of mitochondria dosage on transcription and translation apparatus content and activities. Moreover, we find that mitochondrial levels have a large impact on alternative splicing, thus modulating both the abundance and type of mRNAs. A simple mathematical model where mitochondrial content simultaneously affects transcription rate and splicing site choice can explain the alternative splicing data. The results of this study show that mitochondrial content (and/or probably function) influence mRNA abundance, translation and alternative splicing which ultimately impacts on cellular phenotype.","project":"ERP003617"}
{"number_samples":17,"species":"human","abstract":"We have developed a novel computational approach for detecting viral transcripts in human cancers applicable to a wide variety of viruses and tumors that takes the aforementioned confounding factors into account. We apply the approach to conducting the first systematic search for viruses in neuroblastoma, the most common cancer in infancy. The diverse clinical progression of this disease as well as related epidemiological and virological findings are highly suggestive of a pathogenic cofactor. However, a viral etiology of neuroblastoma is currently contested. We mapped 14 transcriptomes of neuroblastoma as well as several positive control transcriptomes to the human and all known viral genomes in order to detect both known and unknown viruses.  Analysis of positive controls, comparisons with related methods, and statistical estimates demonstrate the high sensitivity of our approach. However, detailed analysis of putative vi- ral transcripts within neuroblastoma samples did not provide evidence for the existence of any known human viruses. Likewise, de-novo assembly and analysis of chimeric transcripts did not result in expression signatures associated with novel human pathogens. While confounding factors such as sample dilution or viral clearance in progressed tumors may mask viral cofactors in the data, in principle, this is rendered less likely by the high sensitivity of our approach and the number of biological replicates analyzed. Therefore, our results suggest that frequent viral cofactors of metastatic neuroblastoma are unlikely.","project":"ERP003731"}
{"number_samples":13,"species":"human","abstract":"3 Commercially-available human tissues are sequenced alone and mixed in multiple proportions on two platforms as part of experiments demonstrating that linear models describe the behavior of RNA-seq mixtures, the parameters of the fit which can be used to evaluate the linear response and precision of the sequence measurement.","project":"ERP009290"}
{"number_samples":13,"species":"human","abstract":"The 8p11-p12 amplicon occurs in approximately 15% of breast cancers and occurs in aggressive luminal B-type tumors.  We have identified the WHSC1L1 oncogene as a driving oncogene from this region.  Here we demonstrate that over-expression of WHSC1L1 is linked to over-expression of ERa in SUM-44 breast cancer cells, and in primary human breast cancers.  Knockdown of WHSC1L1, particularly WHSC1L1-short, had a dramatic effect on ESR1 mRNA and ERa protein levels.  SUM-44 cells do not require exogenous estrogen for growth in vitro; however they are dependent on ERa expression as ESR1 knockdown or exposure to the selective estrogen receptor degrader (SERD) fulvestrant resulted in growth inhibition.  ChIP-seq experiments utilizing ERa antibodies demonstrated extensive ERa binding to chromatin in SUM-44 cells under estrogen-free conditions.  ERa bound ERE and FOXA1 binding motifs under estrogen-free conditions and regulated expression of estrogen responsive genes.  Short-term treatment with estradiol enhanced binding of ERa to chromatin and influenced expression of many of the same genes to which ERa was bound under estrogen-free conditions.  Finally, knockdown of WHSC1L1 in SUM-44 cells resulted in loss of ERa binding to chromatin under estrogen-free conditions, which was restored upon exposure to estradiol.  These results indicate that SUM-44 cells are a good model for a subset of luminal B breast cancers that have 8p11-p12 amplification, over-express WHSC1L1, and over-express ERa that is independent of estrogen for binding chromatin and regulating gene expression.  Breast cancers that are dependent on ERa activity but independent of estradiol are a major cause of breast cancer mortality.","project":"ERP011490"}
{"number_samples":12,"species":"human","abstract":"Androgens are key factors involved in the development and progression of prostate cancer (PCa), and PCa growth can be suppressed by androgen deprivation therapy. In a considerable proportion of men receiving androgen deprivation therapy, however, PCa progresses to castration-resistant PCa (CRPC), making the development of efficient therapies challenging. We used an orthotopic VCaP human PCa xenograft model to study cellular and molecular changes in tumors after androgen deprivation therapy (castration). Tumor growth was monitored through weekly serum prostate-specific antigen measurements, and mice with recurrent tumors after castration were randomized to treatment groups. Serum prostate-specific antigen concentrations showed significant correlation with tumor volume. Castration-resistant tumors retained concentrations of intratumoral androgen (androstenedione, testosterone, and 5a-dihydrotestosterone) at levels similar to tumors growing in intact hosts. Accordingly, castration induced up-regulation of enzymes involved in androgen synthesis (CYP17A1, AKR1C3, and HSD17B6), as well as expression of full-length androgen receptor (AR) and AR splice variants (AR-V1 and AR-V7). Furthermore, AR target gene expression was maintained in castration-resistant xenografts. The AR antagonists enzalutamide (MDV3100) and ARN-509 suppressed PSA production of castration-resistant tumors, confirming the androgen dependency of these tumors. Taken together, the findings demonstrate that our VCaP xenograft model exhibits the key characteristics of clinical CRPC and thus provides a valuable tool for identifying druggable targets and for testing therapeutic strategies targeting AR signaling in CRPC.","project":"ERP012951"}
{"number_samples":4,"species":"human","project":"SRP053791"}
{"number_samples":9662,"species":"human","abstract":"Lay Description.  The aim of the Genotype-Tissue Expression (GTEx) Project is to increase our understanding of how changes in our genes affect human health and disease with the ultimate goal of improving health care for future generations. GTEx will create a database that researchers can use to study how inherited changes in genes lead to common diseases.  GTEx researchers are studying genes in different tissues obtained from many different people. The GTEx project also includes a study of the GTEx donor consent process - this study will help ensure that the consent process and other aspects of the project effectively address the concerns and expectations of participants in the study. GTEx is a pioneering project that uses state-of-the-art protocols for obtaining and storing a large range of organs and tissues, and for testing them in the lab. Until now, no project has analyzed genetic variation and expression in as many tissues from the same person in... (for more see dbGaP study page.)","project":"SRP012682"}
{"number_samples":11284,"species":"human","abstract":"The Cancer Genome Atlas (TCGA) is a collaboration between the National Cancer Institute (NCI) and the National Human Genome Research Institute (NHGRI) that has generated comprehensive, multi-dimensional maps of the key genomic changes in 33 types of cancer. The TCGA dataset, comprising more than two petabytes of genomic data, has been made publically available, and this genomic information helps the cancer research community to improve the prevention, diagnosis, and treatment of cancer.","project":"TCGA"}