Proteogenomic characterization and mapping of nucleosomes decoded by Brd and HP1 proteins.
BACKGROUND
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.


RESULTS
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.


CONCLUSIONS
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.
Gary
G
LeRoy
Iouri
I
Chepelev
Peter A
PA
DiMaggio
Mario A
MA
Blanco
Barry M
BM
Zee
Keji
K
Zhao
Benjamin A
BA
Garcia
D018797:Cell Cycle Proteins; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism; D002874:Chromosome Mapping; D002888:Chromosomes, Human, Pair 19 / Q000235:genetics / Q000378:metabolism; D044127:Epigenesis, Genetic; D023281:Genomics; D006570:Heterochromatin / Q000235:genetics / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing; D006657:Histones / Q000378:metabolism*; D006801:Humans; D005810:Multigene Family; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D009707:Nucleosomes / Q000235:genetics* / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D040901:Proteomics; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D000092468:Bromodomain Containing Proteins
D018797:Cell Cycle Proteins; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone; D006570:Heterochromatin; D006657:Histones; D009687:Nuclear Proteins; D009707:Nucleosomes; D017346:Protein Serine-Threonine Kinases; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D014157:Transcription Factors; C098415:BRD2 protein, human; D000092468:Bromodomain Containing Proteins; C109106:BRD3 protein, human; C441289:BRD4 protein, human; C000715593:CBX1 protein, human; C000715594:CBX5 protein, human