title: Efficient readout of posttranslational codes on the 50-residue tail of histone H3 by high-resolution MS/MS.
abstract: Histone modifications are highly linked to DNA methylation and together they exert epigenetic control over many activities in the cell including gene transcription. Using a streamlined mass spectrometric approach to determine changes in modification states in the first 50 residues of histone H3, we found a decrease in the global methylation states of H3.1 at Lys 9, Lys 14, and Lys 27 after inhibition of DNA methyltransferases by 5-aza-2'-deoxycytidine. Collisional ion dissociation methods proved adequate to determine site-specific H3 posttranslational modifications (PTMs) because ample backbone bonds are cleaved between each modification site and PTMs were stable to MS/MS using threshold fragmentation in a linear ion trap (LTQ). Our assay allows for a quick profiling and site-specific interrogation of modification states on the first 50 residues of H3 and is directly applicable to H3.1, H3.2, or H3.3 using most OrbiTrap, FT ICR, or TOF mass spectrometers.
authors.affiliation: Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
authors.forename: Nertila
authors.initials: N
authors.lastname: Siuti
authors.forename: Neil L
authors.initials: NL
authors.lastname: Kelleher
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000077209:Decitabine; D006367:HeLa Cells; D006657:Histones / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008745:Methylation; D011499:Protein Processing, Post-Translational; D012697:Serine Endopeptidases / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000964:Antimetabolites, Antineoplastic; D006657:Histones; D000077209:Decitabine; D004248:DNA (Cytosine-5-)-Methyltransferases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D001374:Azacitidine