Unbiased proteomic screen for binding proteins to modified lysines on histone H3

Abstract

We report a sensitive peptide pull-down approach in combination with protein identification by LC-MS/MS and qualitative abundance measurements by spectrum counting to identify proteins binding to histone H3 tail containing dimethyl lysine 4 (H3K4me2), dimethyl lysine 9 (H3K9me2), or acetyl lysine 9 (H3K9ac). Our study identified 86 nuclear proteins that associate with the histone H3 tail peptides examined, including seven known direct binders and 16 putative direct binders with conserved PHD finger, bromodomain, and WD40 domains. The reliability of our proteomic screen is supported by the fact that more than one-third of the proteins identified were previously described to associate with histone H3 tail directly or indirectly. To our knowledge, the results presented here are the most comprehensive analysis of H3K4me2, H3K9me2, and H3K9ac associated proteins and will provide a useful resource for researchers studying the mechanisms of histone code effector proteins.

PHD Finger Protein 2 (PHF2) Represses Ribosomal RNA Gene Transcription by Antagonizing PHF Finger Protein 8 (PHF8) and Recruiting Methyltransferase SUV39H1

“…Through a proteomics approach, we first identified PHF2 as a H3K4me2/3-specific binding protein (40). Wen et al (17) later elegantly demonstrated via both biochemical and crystallographic studies that the PHD finger of PHF2 is responsible for the recognition of H3K4me2/3 mark.…”

Identification and Characterization of Nardilysin as a Novel Dimethyl H3K4-binding Protein Involved in Transcriptional Regulation

“…Identification of NRDc as a H3K4me2-binding Protein-We reported recently an effort to identify H3K4me2 or H3K9me2 effector proteins from HeLa nuclear extracts using an unbiased pull-down assay (35). NRDc was identified among the proteins that bound to H3K4me2 peptide by LC-MS/MS mass spectrometry analysis (data not shown).…”

Structure and function of WD40 domain proteins

“…However, a recent report suggested that WDR61 is a histone methyllysine binder (Chan et al, 2009). Nevertheless, the crystal structure of WDR61 (PDB ID:3OW8) shows that this WD40 does not harbor an aromatic cage.…”