Genome-wide analysis of RXRa binding in mouse liver chromatin with ChIP-SEQ.
Ontology highlight
ABSTRACT: For the genome-wide analysis using ChIP-Seq on mouse liver cells, a full accounting of all RXRα binding sites and of RXRa related gene regulations is expected to address the main knowledge gap around RXRα. Liver tissue of wild type and hs-RXRa-del-exon4-/- mice, male and female for each genotype have been ChIP-Seq'ed for RXRa and Pol2.
Project description:For the genome-wide analysis using ChIP-Seq on mouse liver cells, a full accounting of all RXRα binding sites and of RXRa related gene regulations is expected to address the main knowledge gap around RXRα.
Project description:A relatively small number of proneural transcription factors specify a multitude of neural progenitor populations in the developing mammalian brain. Despite their importance, little is known about their targets, cellular processes they regulate, or what genomic sites they occupy in vivo. We used an integrated experimental and computational approach, combining RNA-seq, Histone-seq, and Atoh1 ChIP-seq in cerebellar tissue, to identify over 600 targets of Atoh1, an important developmental transcription factor. We validated 10% of these targets and found that Atoh1 directly regulates genes involved not only in early proliferation but also later differentiation, migration, cell adhesion, metabolism, and cytoskeletal organization by recognizing a novel 10 nucleotide Atoh1 E-Box associated motif (AtEAM). This Atoh1 “targetome” is not only a resource for studies aimed at understanding the molecular mechanisms involved in cerebellar development, but our integrated approach provides a framework for future in vivo studies of transcription factors and their targetomes. Examination of RNA-seq WT and null expression data (2 reps each), 2 different histone modifications (2 rep each) and IgG control, and Atoh1 DNA-binding (2 reps each) and control in the developing cerebellum
Project description:Retinoic acid (RA) triggers physiological processes by activating heterodimeric transcription factors comprising retinoic acid (RARa,b,g) and retinoid X (RXRa,b,g) receptors. How a single signal induces highly complex temporally controlled networks that ultimately orchestrate physiological processes is unclear. Using an RA-inducible differentiation model we defined the temporal changes in the genome-wide binding patterns of RARg and RXRa and correlated them with transcription regulation. Unexpectedly, both receptors displayed a highly dynamic binding, with different RXRa heterodimers targeting identical loci. Comparison of RARg and RXRa co-binding at RA-regulated genes identified putative RXRa-RARg target genes that were validated with subtype-selective agonists. Gene regulatory decisions during differentiation were inferred from transcription factor target gene information and temporal gene expression. This analysis revealed 6 distinct co-expression paths of which RXRa-RARg is associated with transcription activation, while Sox2 and Egr1 were predicted to regulate repression. Finally, RXRa-RARg regulatory networks were reconstructed through integration of functional co-citations. Our analysis provides a dynamic view of RA signalling during cell differentiation, reveals RA heterodimer dynamics and promiscuity, and predicts decisions that diversify the RA signal into distinct gene-regulatory programs. RXRa; RARg and RNA Polymerase II chromatin binding has been assessed at five time points; in addition an input control for F9 cells as well as a RXRa ChIP-seq assay from a rxra-/- ko strain has been performed.
Project description:We used RNA-seq to interrogate prostate cancer specific gene fusions, alternative splicings, somatic mutations and novel transcripts. 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 description:While a first draft of the equine genome is available and predictions are made regarding resulting genes and proteins, little is known about the actual transcriptome. So far, published expressed sequence tags (ESTs) from different horse tissues were generally rather short (?600bp) and hardly annotated, reflecting the problem that good cDNA libraries are very difficult to analyse. In this approach, we aimed to establish and analyse a normalised immune cell cDNA library (using freshly isolated and activated lymphocytes, NK cells, monocytes and DC). In particular, we wanted to test next generation sequencing combined with a series of bioinformatic approaches. The resulting cDNA library contained 2x107 clones of which 1056 were used for an initial Sanger sequencing and 4x106 for the deep sequencing analysis. Through the latter we obtained >29k sequences for which more than 5000 matches where found on the equine reference sequences. Additionally we could identify more than 3500 sequences which had matches on both - non-equine RNA sequences as well as the equine genome. In these we find both extensions of existing RefSeq models and novel mRNAs alike. Less than 2% of sequences did not have any match in the mentioned databases. 1 pooled set of samples from one animal analysed
Project description:The cJun NH2-terminal kinase (JNK) signaling pathway in the liver promotes systemic changes in metabolism by regulating PPARa-dependent expression of the hepatokine FGF21. Hepatocyte-specific gene ablation studies demonstrated that the Mapk9 gene (encodes JNK2) plays a key mechanistic role. Mutually exclusive inclusion of exons 7a and 7b yields expression of the isoforms JNK2a and JNK2b. Here we demonstrate that Fgf21 gene expression and metabolic regulation is primarily regulated by the JNK2a isoform. To identify relevant substrates of JNK2a, we performed a quantitative phosphoproteomic study of livers isolated from control mice, mice with JNK-deficiency in hepatocytes, and mice that express only JNK2a or JNK2b in hepatocytes. We identified the JNK substrate RXRa as a protein that exhibited JNK2a-promoted phosphorylation in vivo. RXRa functions as a heterodimeric partner of PPARa and may therefore mediate the effects of JNK2a signaling on Fgf21 expression. To test this hypothesis, we established mice with hepatocyte-specific expression of wild-type or mutated RXRa proteins. We found that the RXRa phosphorylation site Ser260 was required for suppression of Fgf21 gene expression. Collectively, these data establish a JNK-mediated signaling pathway that regulates hepatic Fgf21 expression.