ChIP-seq of RNA polymerase II, transcription factors and histone modification in retinal pigment epithelial cells
ABSTRACT: Retinal pigment epithelial cells are critical for eye function and loss of cell function is linked to age-related blindness. Relatively little is known about the transcriptional regulatory networks in these cells. The datasets presented here are ChIP-seq experiments for RNA polymerase II , transcription factors and histone modifications in human retinal pigment epithelial cells. ChIP-Seq for transcription factors, RNA polymerase, histone modifications and CTCF in retinal pigment epithelial cells
Project description:Whole genome profiling of histone methylation (H3K4me3, H3K27me3 and RNA Pol II) in Ciliary derived stem/progenitor cells and differentiated cells. In the present study we focus on the cellular differentiation that involves loss of pluripotency and gain of lineage and cell type-specific characteristics and we show developments from adult ciliary derived stem/progenitor cells to differentiated retinal neurons/glial cells of human cadaveric eyes in vitro. Ciliary pigment epithelial cells isolated from human cadaveric eye balls were cultured in the presence of mitogens like Epidermal Growth Factor (EGF) and Fibroblast Growth Factors (FGF) to generate neurospheres (Stem/progenitor population). The generated neurospheres were plated on laminin and poly-D-lysine-coated cover slips in the presence of the Brain derived neurotrophic factor (BDNF), Retinoic acid (RA) and 10% FBS for the differentiation (Retinal neuron/glial cells). Global Methylation patterns of histone H3K4me3, RNA Pol II and H3K27me3 were analysed in the present study. This work would provide an outline of epigenetic modifications in ciliary derived stem/progenitor cells and the progeny that underwent differentiation into retinal neurons/glial cells and present that specific histone methylations are involved in gene expression reprogramming during the process of differentiation.
Project description:We generated a retinal pigment epithelial cell line with complete knockout of giantin using CRISPR. This experiment sought to define changes in the transcriptome of that cell line compared to the parental wild-type cells.
Project description:Three transcriptional states can be defined by histone modifications and RNA polymerase II enriched at promoters and across the body of genes. To gain insight into the active, poised and silent genes in human T-ALL cells, two antibodies against RNAP2, and antibodies against H3K4me3, H3K79me2, and H3K27me3 were used for chromatin immunoprecipitation coupled with massive parallel sequencing (ChIP-seq). Genomic DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. ChIP was performed using an antibody against RNAP2, H3K4me3, H3K79me2, and H3K27me3 using whole cell extract (WCE) as a background control. ChIP was performed using a two antibody against hypophosphorylated forms of RNAP2 in two biological replicates. All other ChIPs were done in biological replicates with a single lane of sequencing.
Project description:RNA Polymerase II is the enzyme responsible for active transcription. To gain insight into the genes occupied by RNA Polymerase II and actively transcribed in MEFS, chromatin immunoprecipitation coupled with massive parallel sequencing (ChIP-seq) was performed to determine the genome-wide binding targets of RNA Pol2. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. ChIP was performed using an antibody against RNA Polymerase II.
Project description:Proinflammatory stimuli rapidly and globally remodel chromatin landscape, thereby enabling transcriptional responses. Yet, the mechanisms coupling chromatin regulators to the master regulatory inflammatory transcription factor NF-kB remain poorly understood. We report in human endothelial cells (ECs) that activated NF-kB binds to enhancers, provoking a rapid, global redistribution of BRD4 preferentially at super-enhancers, large enhancer domains highly bound by chromatin regulators. Newly established NF-kB super-enhancers drive nearby canonical inflammatory response genes. In both ECs and macrophages BET bromodomain inhibition prevents super-enhancer formation downstream of NF-kB activation, abrogating proinflammatory transcription. In TNFa-activated endothelium this culminates in functional suppression of leukocyte rolling, adhesion and transmigration. Sustained BET bromodomain inhibitor treatment of LDLr -/- animals suppresses atherogenesis, a disease process rooted in pathological vascular inflammation involving endothelium and macrophages. These data establish BET-bromodomains as key effectors of inflammatory response through their role in the dynamic, global reorganization of super-enhancers during NF-kB activation. ChIP-Seq for various transcription factors, RNA Polymerase II, and histone modifications in human endothelial cells
Project description:The datasets presented here are transcription factors, transcriptional co-factors or histone modifications associated with active enhancers in a variety of mouse and human cell types. ChIP-seq of enhancer-associated transcription factors and histone modifications
Project description:Excessive expression of c-Myc occurs frequently in human cancers, where high levels are associated with tumor aggression and poor clinical outcome, but the effect of high levels of c-Myc on global gene regulation is poorly understood. We report here that in tumor cells expressing high levels of c-Myc, the transcription factor binds to E-box sequences in the core promoters of most actively transcribed genes and, unexpectedly, the enhancers of these active genes. The predominant effect of increasing c-Myc levels at both proximal and distal promoter elements is to produce higher levels of transcription at existing active genes by promoting RNA polymerase II elongation, as opposed to stimulating transcription of novel target genes. Our results argue that c-Myc overexpression drives increased transcription of growth-promoting genes, and does so by amplifying the levels of transcripts associated with the entire gene expression program of the cancer cell. Thus, excess c-Myc functions to elicit the transcriptional amplification of existing active genes through the invasion of enhancers across the cancer cell genome, thereby reducing the rate-limiting constraints required for continuous tumor growth and proliferation. ChIP-Seq of multiple factors and histone modifications in a variety of human tumor cell lines
Project description:ChIP-Seq of RNA Polymerase II, and transcriptional regulators in multiple myeloma (MM.1S), glioblastoma (U87-MG), and small cell lung carcinoma (H2171) treated with the BET bromodomain inhibitor JQ1. Cell lines (MM.1S, U87-MG, and H2171) representing multiple myeloma, glioblastoma, and small cell lung carcinoma, were treated with varying concentrations (5nM to 5µM) of the BET bromodomain inhibitor JQ1 followed by ChIP-Seq for RNA Polymerase II and transcriptional regulators. Other datasets from this series of experiments have been release as a part of GSE42355.
Project description:Retinal pigment epithelial (RPE) cells and choroidal stromal fibroblast (CSF) were isolated from healthy human donor eyes. Cells were cultured and RNA extracted. Overall design: Three human choroidal stromal fibroblast cell populations and one human retinal pigment epithelial cell population (primary cells) were cultured and analyzed by gene profiling.
Project description:Here we describe a novel class of short RNAs and high-resolution ChIP-Seq data that together indicate widespread divergent transcription at mammalian promoters and suggest the existence of novel regulatory mechanisms concerning both initiation and elongation by RNA polymerase II. ChIP-seq in murine embryonic stem cells for RNA polymerase II