Genome-wide analysis of H3.3 dissociation in mouse embryonic stem cells
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE63641 | GEO | 2015/05/14
SECONDARY ACCESSION(S): PRJNA268514
REPOSITORIES: GEO
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Project description:Using Tet-inhibited expression of epitope-tagged H3.3 combined with ChIP-Seq we undertook genome-wide measurements of H3.3 dissociation rates across the ESC genome and examined the relationship between H3.3-nucleosome turnover and ESC-specific transcription factors, chromatin modifiers and epigenetic marks. To measure dissociation rates of H3.3, we utilized a TET-repressible ESC line, ES[MC1R(20)], with the expression cassette integrated at the ROSA26 locus. We transfected MC1R ESCs with HA/FLAG-tagged H3.3 controlled by tetracycline response elements. For ‘TET-OFF’ experiments, ESCs were cultured over several passages (weeks) on feeder cells in the absence of DOX and were subsequently passaged onto feeder-free plates prior to the inhibition of HA-H3.3 expression. To repress HA/FLAG-H3.3 expression we treated cells with 2 ?g/ml doxycycline hyclate before crosslinking with formaldehyde at various time points. Measurement of H3.3-HA enrichment over time-course was performed using two replicates of ChIP-seq experiments. As a control, input DNA sequencing was performed for every time point
2015-05-14 | E-GEOD-63641 | biostudies-arrayexpress
Project description:We developed a system to study the DNA replication-independent turnover nucleosomes containing the histone variant H3.3 in mammalian cells. By measuring the genome-wide incorporation of H3.3 at different time points following epitope-tagged H3.3 expression, we find three categories of H3.3-nucleosome turnover in vivo: rapid turnover, intermediate turnover and, specifically at telomeres, slow turnover. Our data indicate that H3.3-containing nucleosomes at enhancers and promoters undergo a rapid turnover that is associated with active histone modification marks including H3K4me1, H3K4me3, H3K9ac, H3K27ac and the histone variant H2A.Z. The rate of turnover is negatively correlated with H3K27me3 at regulatory regions and with H3K36me3 at gene bodies. Examination of incorporation dynamics of histone variant H3.3
2013-10-23 | E-GEOD-51505 | biostudies-arrayexpress
Project description:We developed a system to study the DNA replication-independent turnover nucleosomes containing the histone variant H3.3 in mammalian cells. By measuring the genome-wide incorporation of H3.3 at different time points following epitope-tagged H3.3 expression, we find three categories of H3.3-nucleosome turnover in vivo: rapid turnover, intermediate turnover and, specifically at telomeres, slow turnover. Our data indicate that H3.3-containing nucleosomes at enhancers and promoters undergo a rapid turnover that is associated with active histone modification marks including H3K4me1, H3K4me3, H3K9ac, H3K27ac and the histone variant H2A.Z. The rate of turnover is negatively correlated with H3K27me3 at regulatory regions and with H3K36me3 at gene bodies.
2013-10-23 | GSE51505 | GEO
Project description:Histone variants and their chaperones are key regulators of eukaryotic transcription, and are critical for normal development. The histone variant H3.3 has been shown to play important roles in pluripotency and differentiation, and although its genome-wide patterns have been investigated, little is known about the role of its dynamic turnover in transcriptional regulation. To elucidate the role of H3.3 dynamics in embryonic stem cell (ESC) biology, we generated mouse ESC lines carrying a single copy of a doxycycline (Dox)-inducible HA-tagged version of H3.3, and monitored the rate of incorporation of the HA-tagged variant by ChIP-seq at varying time points following Dox induction. Active H3K4me3-enriched genes display high turnover dynamics in transcription start sites (TSS); H3K27me3-enriched genes have a lower turnover dynamics in the TSS, but higher turnover rates in gene bodies; developmental bivalent genes marked by both H3K4me3 and H3K27me3 show a mixed behavior with high TSS dynamics and high gene body dynamics, especially in genes bound by both PRC1 and PRC2. In contrast, genes proximal to super-enhancers show the most reduced gene-body turnover dynamics, while genes proximal to other enhancers show no unique pattern. In RA-treated cells, while the overall profile in expressed genes remains largely unaltered over the TSS and gene body, a significant and dramatic decrease in turnover occurs at the -1 nucleosome position immediately before the TSS, revealing a hyperdynamic nucleosome in promoters of genes expressed in ESCs. We suggest that histone turnover dynamics provides an additional mechanism involved in expression regulation, that gene-body turnover dynamics is distinct from that of promoter regions, and that a hyperdynamic -1 nucleosome marks promoters in ESCs. Our data provide evidence for regional regulation of H3.3 turnover in ESC promoters, and calls for testing, in high resolution, the dynamic behavior of additional histone variants and other structural chromatin proteins.
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Project description:Nucleosome turnover concomitant with incorporation of the replication-independent histone variant H3.3 is a hallmark of regulatory regions in the animal genome. In our current understanding, nucleosome turnover is universally linked to DNA accessibility and histone acetylation. In mouse embryonic stem cells, H3.3 is also highly enriched at interstitial heterochromatin, most prominently intracisternal-A particle endogenous retroviral elements. Interstitial heterochromatin is established over confined domains by the TRIM28/SETDB1 corepressor complex and has stereotypical features of repressive chromatin, such as H3K9me3 and recruitment of all HP1 isoforms. Here, we demonstrate that fast histone turnover and H3.3 incorporation is compatible with these hallmarks of heterochromatin. We identify Smarcad1 to be a chromatin remodeler that generates nucleosome-free regions which are subsequently filled with histone H3.3, generating a surprisingly dynamic heterochromatin state. Loss of histone H3.3 elicits a highly specific opening of interstitial heterochromatin demonstrating that in the wake of Smarcad1 remodeling, H3.3 is required to maintain minimal DNA accessibility by reassembling nucleosomes.
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Project description:A mouse knock-in model engineered for Cre recombinase-activated expression of the endogenous mouse H3f3a allele generating an epitope-tagged H3.3 equipped with or without a K27M mutation to investigate H3.3 K27M effects on brain cell and tumor growth, gene expression and epigenetics.
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Project description:A mouse knock-in model engineered for Cre recombinase-activated expression of the endogenous mouse H3f3a allele generating an epitope-tagged H3.3 equipped with or without a K27M mutation to investigate H3.3 K27M effects on brain cell and tumor growth, gene expression and epigenetics. These samples are matched with H3K27me3 ChIPseq and RNAseq in GSE108364.
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Project description:We report high-throughput profiling of transgenic histone H3.3 in zebrafish cardiomyocytes. The replacement histone H3.3 is deposited at sites of nucleosome turnover including regions of active chromatin. We profile H3.3 in uninjured cardiomyocytes, those undergoing regeneration 14 days after genetic ablation and those proliferating 7 days after Nrg1 stimulated hyperplasia. This study provides a framework for understanding chromatin transitions during adult models of regeneration.
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