A KAP1 phosphorylation switch controls MyoD function during skeletal muscle differentiation
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Evarist Planet
PROVIDER: E-GEOD-62664 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:The transcriptional activator MyoD serves as a master controller of myogenesis. Often in partnership with Mef2, MyoD binds to the promoters of hundreds of muscle genes in proliferating myoblasts, yet activates these targets only upon receiving cues that launch differentiation. What regulates this off/on switch of MyoD function has been incompletely understood, although known to reflect the action of chromatin modifiers. Here, we identify KAP1/TRIM28 as a key regulator of MyoD function. In myoblasts, KAP1 is present with MyoD and Mef2 at many muscle genes, where it acts as a scaffold to recruit not only co-activators such as p300 and LSD1, but also co-repressors such as G9a and HDAC1, with promoter silencing as net outcome. Upon differentiation, MSK1-mediated phosphorylation of KAP1 releases the co-repressors from the scaffold, unleashing transcriptional activation by MyoD/Mef2 and their positive cofactors. Thus, our results reveal KAP1 as a previously unappreciated interpreter of cell signaling, which modulates the ability of MyoD to drive myogenesis. Kap1 and H3K9me3 ChIPseq in proliferating C2C12 cells
2015-02-02 | E-GEOD-62660 | biostudies-arrayexpress
Project description: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. 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.
2014-06-09 | E-GEOD-58323 | biostudies-arrayexpress
Project description: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. Analyses of epigentic effectors and marks in KAP1 WT and KD human embryonic stem cells
2014-05-29 | E-GEOD-57989 | biostudies-arrayexpress
Project description:The transcriptional activator MyoD serves as a master controller of myogenesis. Often in partnership with Mef2, MyoD binds to the promoters of hundreds of muscle genes in proliferating myoblasts, yet activates these targets only upon receiving cues that launch differentiation. What regulates this off/on switch of MyoD function has been incompletely understood, although known to reflect the action of chromatin modifiers. Here, we identify KAP1/TRIM28 as a key regulator of MyoD function. In myoblasts, KAP1 is present with MyoD and Mef2 at many muscle genes, where it acts as a scaffold to recruit not only co-activators such as p300 and LSD1, but also co-repressors such as G9a and HDAC1, with promoter silencing as net outcome. Upon differentiation, MSK1-mediated phosphorylation of KAP1 releases the co-repressors from the scaffold, unleashing transcriptional activation by MyoD/Mef2 and their positive cofactors. Thus, our results reveal KAP1 as a previously unappreciated interpreter of cell signaling, which modulates the ability of MyoD to drive myogenesis. Transcriptome profiling of control and Kap1KD in C2C12 during proliferation or at 48hours differentiation stage
2015-02-02 | E-GEOD-62659 | biostudies-arrayexpress
Project description:TRIM28 (KAP1 - KRAB-associated protein 1) is critical for the silencing of endogenous retroviruses (ERVs) in embryonic stem (ES) cells. Here, we reveal that an essential impact of this process is the protection of cellular gene expression in early embryos from perturbation by cis-acting activators contained within these genetic invaders. In TRIM28-depleted ES cells, repressive chromatin marks at ERVs are replaced by histone modifications typical of active enhancers, stimulating transcription of nearby cellular genes, notably those harboring bivalent promoters. Correspondingly, ERV-derived sequences can repress or enhance expression from an adjacent promoter in transgenic embryos depending on their TRIM28-sensitivity in ES cells. TRIM28-mediated control of ERVs is therefore crucial not just to prevent retrotransposition, but more broadly to safeguard the transcriptional dynamics of early embryos. Analyses of transcriptional profiles and chromatin state in TRIM28 WT and KO cells
2012-12-04 | E-GEOD-41903 | biostudies-arrayexpress
Project description:Polycomb group (PcG) proteins initiate the formation of repressed chromatin domains and regulate developmental gene expression. A mammalian PcG protein, Enhancer of Zeste homolog 2 (Ezh2), triggers transcriptional repression by catalyzing the addition of methyl groups onto lysine-27 of histone H3 (H3K27me2/3)1. This action facilitates the binding of other PcG proteins to histone H3 and compaction of chromatin. Interestingly, there exists a paralog of Ezh2, termed Ezh1, whose primary function remains unclear. Here, we provide evidence for genome-wide association of Ezh1 with active epigenetic marks, RNA polymerase II (PolII) and mRNA production. Ezh1 depletion reduced global PolII occupancy within gene bodies and resulted in delayed transcriptional activation during differentiation of skeletal muscle cells. Conversely, ectopic expression of wild-type Ezh1 led to premature gene activation and rescued PolII-elongation defects in Ezh1-depleted cells. Collectively, these findings reveal an unanticipated role of a PcG protein in promoting mRNA transcription. Examination of 3 different histone modifications, 3 modified forms of RNA polymerase II, Ezh1, Ezh2 and mRNA levels in a skeletal muscle cells at various developmental stages.
2011-11-23 | E-GEOD-25549 | biostudies-arrayexpress
Project description:Histone chaperones affect chromatin structure and gene expression through interaction with histones and RNA polymerase II (PolII). Here, we report that the histone chaperone Spt6 counteracts H3K27me3, an epigenetic mark deposited by the Polycomb Repressive Complex 2 (PRC2) and associated with transcriptional repression. We found that Spt6 is required for proper engagement and function of the H3K27 demethylase KDM6A (UTX) on muscle genes and regulates muscle gene expression and cell differentiation. ChIP-Seq experiments revealed an extensive genome-wide overlap of Spt6, PolII and KDM6A at transcribed regions that are devoid of H3K27me3. Mammalian cells and zebrafish embryos with reduced Spt6 display increased H3K27me3 and diminished expression of the master regulator MyoD, resulting in myogenic differentiation defects. As a confirmation for an antagonistic relationship between Spt6 and H3K27me3, inhibition of PRC2 permits MyoD re-expression in myogenic cells with reduced Spt6. Our data indicate that, through cooperation with PolII and KDM6A, Spt6 orchestrates removal of H3K27me3, thus effectively controlling developmental gene expression and cell differentiation. Examination of Spt6 and KDM6A levels in a skeletal muscle cells at various developmental stages
2013-04-01 | E-GEOD-44119 | biostudies-arrayexpress
Project description:Transcription factors and DNA regulatory binding motifs are fundamental components of the gene regulatory network (GRN). Here, by using genome-wide occupancy profiling of master regulators of MyoGenesis (MyoD and MyoGenin), we show their extensive occupancy in the extragenic enhancer regions coinciding with RNA synthesis (i.e. eRNA). In particular, multiple regions coding for eRNAs were observed within regulatory region of MYOD1, including previously characterized Distal Regulatory Regions (DRR) and Core Enhancer (CE). While CERNA enhanced RNA polymerase II (PolII) occupancy and transcription at MYOD1, DRRRNA acted in trans to activate the downstream MyoGenic GRN. The deployment of transcriptional machinery to appropriate loci is contingent on chromatin accessibility, a rate-limiting step preceding PolII assembly. By nuclease sensitivity assay, we show that eRNAs increase genomic access to the transcriptional complex at defined regulatory regions. In conclusion, our data suggest eRNAs establish a cell-type-specific transcriptional circuitry by directing chromatin-remodeling events. Examination of MyoD and MyoG binding events and production of RNA at the enhancer sites during myogenic differentiation. We performed RNAi against enhancer-derived RNA (DRRi) which resulted in reduction of chromatin accessiblity and RNA polymerase II occupancy at defined regulatory elements. In complementary experiments, overexpression of DRR-RNA (pHAN_DRR1.2) resulted in early activation of MyoG. GFPi and GFP overexpression (pHAN_GFP) were used as control in these experiments, respectively.
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Project description:The tetrapod-restricted KRAB-containing zinc finger proteins (KRAB-ZFPs) are essential early embryonic controllers of transposable elements (TEs), which they repress via their cofactor KAP1 and associated effectors through histone and DNA methylation, a process thought to result in irreversible silencing. Using a target-centered functional screen, we matched several murine TEs with their cognate KRAB-ZFP. This revealed an unexpected level of granularity in their interactions, with KRAB-ZFPs recognizing TEs from more than one subfamily, TEs recruiting more than one KRAB-ZFP, and spatially and temporally differential KRAB-ZFP-mediated regulation of TEs and nearby genes. Most importantly, we discovered that the KRAB/KAP1 system controls TEs in adult tissues, in cell culture and in vivo, where they partner up to regulate the expression of cellular genes. Therefore, TEs and KRAB-ZFPs establish widely active transcription networks that regulate not only development but probably also many physiological events. Given the high degree of species-specificity of both TEs and KRAB-ZFPs, these results have important implications for studying and understanding the biology of higher vertebrates, including humans. Analysis of transcriptional profiles of KAP1 or ZFP932/Gm15446 KO cells or tissues, and ZFP932 and Gm15446 ChIPseq in murine ES and C2C12 cells.
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Project description:The submitted files contain ChIP-seq data for the MyoD and myogenin muscle regulatory factors in diffrentiated C2C12 cells as well as two different sonicated input samples (one from a regular 1% formaldehyde fixation and one from a dual 1%FA + 1.5 mM EGS fix). Characterization of genome-wide MyoD and myogenin binding in C2C12 cells
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