DNA damage in the presence of chemical genotoxic agents induce acetylation of H3K56 and H4K16 but not H3K9 in mammalian cells

Abstract

Histone covalent modifications play a significant role in the regulation of chromatin structure and function during DNA damage. Hyperacetylation of histones is a DNA damage dependent post translational modification in yeast and mammals. Although acetylation of histones during DNA damage is well established, specific lysine residues that are acetylated is being understood very recently in mammals. Here, in the present study, acetylation of three different lysine residues Histone3Lysine 9 (H3K9), Histone3Lysine 56 (H3K56) and Histone4Lysine 16 (H4K16) were probed with specific antibodies in mammalian cell lines treated with genotoxic agents that induce replication stress or S-phase dependent double strand breaks. Immunoblotting results have shown that DNA damage associated with replication arrest induce acetylation of H3K56 and H4K16 but not H3K9 in mammals. Immunofluorescence experiments further confirmed that acetylated H3K56 and H4K16 form nuclear foci at the site of DNA double strand breaks. Colocalization of H3K56ac with γ H2AX and replication factor PCNA proved the existence of this modification at the site of DNA damage and its probable role in DNA damage repair. Put together, the present data suggests that acetylation of H3K56 and H4K16 are potent DNA damage dependent histone modifications but not H3K9 in mammals.

Histone Deacetylase Inhibitors Globally Enhance H3/H4 Tail Acetylation Without Affecting H3 Lysine 56 Acetylation

“…This prompted us to determine the stoichiometry of acetylation of some of these sites using more sensitive and quantitative approaches. We were particularly interested to look for H3K56ac because several recent publications have implicated this modification in replication-coupled nucleosome assembly, regulation of gene expression and the DNA damage response38394041424344454647484950. In addition, based on immunohistochemistry, it has been argued that H3K56ac is a diagnostic and prognostic marker of several different types of tumours39.…”

“…In addition, based on immunohistochemistry, it has been argued that H3K56ac is a diagnostic and prognostic marker of several different types of tumours39. However, studies of H3K56ac in human cells have been fraught with controversy, particularly with regards to the role of this histone modification in the DNA damage response383944474850, but also the identity of the enzymes that acetylate or deacetylate H3K5639404142434445464850.…”

“…This was very intriguing because two of the HDACi that we used in our MS experiments, namely sodium butyrate and nicotinamide, were previously shown to increase H3K56ac based on immunoblotting3948. This prompted us to investigate the specificity of three commercially available antibodies that were previously used to detect H3K56ac in human cells383942434445464748. For these experiments, the same histone samples were analyzed either by MS or immunoblotting.…”

Chaperone-mediated acetylation of histones by Rtt109 identified by quantitative proteomics

“…The acetylation of specific lysine residues can also provide a chromatin mark for the regulation of gene expression [10] or to identify sites of DNA damage. For instance, recent studies have shown that acetylation of H3-K56 and H4-K16 is significantly elevated in mammalian cells in response to DNA damage [11–13]. …”

HDAC2 deregulation in tumorigenesis is causally connected to repression of immune modulation and defense escape

“…This particular histone modification shows a biphasic response to DNA damage as expression levels are initially reduced, but increase in the long term due to DNA repair. Indeed, replication stress produces an increase in the expression of histone H4 acetylated on lysine 16 [15]. Nevertheless, transformed cells lacking HDAC2 as a result of somatic mutations were recently described [16].…”