Proteo-Genomic characterization and mapping of nucleosomes decoded by Brd and HP1 proteins

Genome biology (Impact Factor: 10.81). 08/2012; 13(8):R68. DOI: 10.1186/gb-2012-13-8-r68
Source: PubMed


Histone post-translational modifications (PTMs) constitute a branch of epigenetic mechanisms that can control the expression of eukaryotic genes in a heritable manner. Recent studies have identified several PTM-binding proteins containing diverse specialized domains whose recognition of specific PTM sites leads to gene activation or repression. Here, we present a high-throughput proteogenomic platform designed to characterize the nucleosomal make-up of chromatin enriched with a set of histone PTM binding proteins known as histone PTM readers. We support our findings with gene expression data correlating to PTM distribution.

We isolated human mononucleosomes bound by the bromodomain-containing proteins Brd2, Brd3 and Brd4, and by the chromodomain-containing heterochromatin proteins HP1β and HP1α. Histone PTMs were quantified by mass spectrometry (ChIP-qMS), and their associated DNAs were mapped using deep sequencing. Our results reveal that Brd- and HP1-bound nucleosomes are enriched in histone PTMs consistent with actively transcribed euchromatin and silent heterochromatin, respectively. Data collected using RNA-Seq show that Brd-bound sites correlate with highly expressed genes. In particular, Brd3 and Brd4 are most enriched on nucleosomes located within HOX gene clusters, whose expression is reduced upon Brd4 depletion by short hairpin RNA.

Proteogenomic mapping of histone PTM readers, alongside the characterization of their local chromatin environments and transcriptional information, should prove useful for determining how histone PTMs are bound by these readers and how they contribute to distinct transcriptional states.

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Available from: Iouri Chepelev, Apr 28, 2015
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    • "BET protein ChIP-seq data obtained in HEK293T cells and CD4+ T cells were retrieved from the Gene Expression Omnibus (accession codes GSM971946-8 for BRD-2, -3, and -4 in HEK293T cells, respectively and GSE33281 for BRD4 in CD4+ T cells).28,49 Extended sequence read densities were determined in 10 kb windows around MLVIN_WT, MLVIN_dC, MLVIN_W390A integration, or MRC sites respectively. "
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    Molecular Therapy - Nucleic Acids 07/2014; 3(7):e179. DOI:10.1038/mtna.2014.33 · 4.51 Impact Factor
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    • "In 293 cells, binding sites of BET proteins (Brd2, Brd3 and Brd4) have been identified by chromatin immunoprecipitation and mapped onto the human genome (19). The correlation of these BET protein binding sites and integration sites of MLV WT IN and IN ΔC viruses was examined (Figure 4C). "
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    Nucleic Acids Research 03/2014; 42(9). DOI:10.1093/nar/gku175 · 9.11 Impact Factor
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    • "Analysis of DNA from chromatin immunoprecipitation (ChIP) by high-throughput sequencing has become a standard tool for assessing PTM localization within the genome [54]. More recently, native ChIP methodologies have been developed to allow for isolation and quantitative PTM analysis of histone proteins, a technique referred to as chromatin immunoprecipitation with quantitative MS (ChIP-qMS) [51,55,56]. Native ChIPs can be performed with either a reader protein or with a PTM-specific antibody to obtain the associated histone codes and histone variants. "
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