Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Phf19 links methylated lysine 36 of histone H3 to regulation of Polycomb activity [ChIP-Seq]

ABSTRACT: Polycomb group proteins are transcriptional repressors that play essential roles in regulating genes required for differentiation and embryonic development. The Polycomb repressive complex 2 (PRC2) contains the methyltransferase activity for lysine 27 on histone 3 (H3K27me3), which is a docking site for the PRC1 complex and leads to gene repression. However, the role of other histone modifications in regulating PRC2 activity is just beginning to be understood. Here we show that direct recognition of histone H3 methylated at lysine 36 (H3K36me), an mark associated with activation, by the PRC2 subunit Phf19 is required for the full enzymatic activity of the PRC2 complex. We provide structural evidence for this interaction by nuclear magnetic resonance spectroscopy (NMR). Using genome-wide chromatin binding analyses and expression analyses, we show that Phf19 binds to a subset of PRC2 targets in embryonic stem (ES) cells, and that this is required for their repression and for H3K27me3 deposition. These findings reveal that the H3K36me2/3-Phf19 interaction is essential for PRC2 complex activity and for proper regulation of gene repression in ES cells. We determined the genome binding/occupancy profile of Phf19, H3K36me3, H3K36me2, H3K27me3 and Suz12 by high throughput sequencing in mouse embryonic stem cells. For Phf19 two independent biological replicas were performed and Phf19 binding sites were defined as those sites (ChIP-seq peaks) present in both replicas. H3K27me3 was evaluated in control ES cells and cells depleted of Phf19 (shRd and shPhf19 respectively).

ORGANISM(S): Mus musculus

SUBMITTER: Luciano Di Croce

PROVIDER: E-GEOD-41589 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Phf19 links methylated Lys36 of histone H3 to regulation of Polycomb activity.

Ballaré Cecilia C Lange Martin M Lapinaite Audrone A Martin Gloria Mas GM Morey Lluis L Pascual Gloria G Liefke Robert R Simon Bernd B Shi Yang Y Gozani Or O Carlomagno Teresa T Benitah Salvador Aznar SA Di Croce Luciano L

Nature structural & molecular biology 20121028 12

Polycomb-group proteins are transcriptional repressors with essential roles in embryonic development. Polycomb repressive complex 2 (PRC2) contains the methyltransferase activity for Lys27. However, the role of other histone modifications in regulating PRC2 activity is just beginning to be understood. Here we show that direct recognition of methylated histone H3 Lys36 (H3K36me), a mark associated with activation, by the PRC2 subunit Phf19 is required for the full enzymatic activity of the PRC2 c ...[more]

PMID: 23104054

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Project description:Osteogenesis is a highly regulated developmental process and continues during the turnover and repair of mature bone. Runx2, the master regulator of osteoblastogenesis, directs a transcription program essential for bone formation through both genetic and epigenetic mechanisms. While individual Runx2 gene targets have been identified, further insights into the broad spectrum of Runx2 functions required for osteogenesis are needed. By performing genome-wide characterization of Runx2 binding at the three major stages of osteoblast differentiation: proliferation, matrix deposition and mineralization, we identified Runx2-dependent regulatory networks driving bone formation. Using chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq) over the course of these stages, we discovered close to 80,000 significantly enriched regions of Runx2 binding throughout the mouse genome. These binding events exhibited distinct patterns during osteogenesis, and were associated with proximal promoters as well as a large percentage of Runx2 occupancy in non-promoter regions: upstream, introns, exons, transcription termination site (TTS) regions, and intergenic regions. These peaks were partitioned into clusters that are associated with genes in complex biological processes that support bone formation. Using Affymetrix expression profiling of differentiating osteoblasts depleted of Runx2, we identified novel Runx2 targets including Ezh2, a critical epigenetic regulator; Crabp2, a retinoic acid signaling component; Adamts4 and Tnfrsf19, two remodelers of extracellular matrix. We demonstrated by luciferase assays that these novel biological targets are regulated by Runx2 occupancy at non-promoter regions. Our data establish that Runx2 interactions withM-BM- chromatin across the genome reveal novel genes, pathways and transcriptional mechanisms that contribute to the regulation of osteoblastogenesis. To identiy the genome-wide occupancy of Runx2, DNA bound by Runx2 at the prolieration, matrix deposition, and mineralization stages were recovered by Runx2 ChIP. Libraries of purified DNA were generated using Illumina SR adapters (Illumina) following manufacturerM-bM-^@M-^Ys manual, and were selected for the inserted fragments of 200 M-BM-1 50 bp, and sequenced 36 bases on an Illumina Genome Analyzer II. Base calls and sequence reads were generated by Illumina CASAVA software (version 1.6, Illumina). Two independent biological repeats of Runx2 ChIP-Seq libraries were prepared for each time point, and two input libraries were prepared with sonicated DNA from day 9 MC3T3-E1 cells. We pooled the reads from two biological replicates for peaking calling using MACS (version 1.4.1) with a stringent p value threshold (p < 1e-10) in contrast to input control, and used these peaks for further bioinformatic analyses. Each sample deposited here contains three files: the sequence file with short reads combined from two biological replicates, a Bed file with peaks called from the pooled short reads, and a Wig file with peak signals.

Dataset Information

Phf19 links methylated lysine 36 of histone H3 to regulation of Polycomb activity [ChIP-Seq]

Publications

Phf19 links methylated Lys36 of histone H3 to regulation of Polycomb activity.

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