P300-mediated Acetylation of Histone H3 Lysine 56 Functions in DNA Damage Response in Mammals

Rahul K Vempati,Ranveer S Jayani,Dimple Notani,Amrita Sengupta,Sanjeev Galande,Devyani Haldar

doi:10.1074/jbc.m110.149393

Abstract

The packaging of newly replicated and repaired DNA into chromatin is crucial for the maintenance of genomic integrity. Acetylation of histone H3 core domain lysine 56 (H3K56ac) has been shown to play a crucial role in compaction of DNA into chromatin following replication and repair in Saccharomyces cerevisiae. However, the occurrence and function of such acetylation has not been reported in mammals. Here we show that H3K56 is acetylated and that this modification is regulated in a cell cycle-dependent manner in mammalian cells. We also demonstrate that the histone acetyltransferase p300 acetylates H3K56 in vitro and in vivo, whereas hSIRT2 and hSIRT3 deacetylate H3K56ac in vivo. Further we show that following DNA damage H3K56 acetylation levels increased, and acetylated H3K56, which is localized at the sites of DNA repair. It also colocalized with other proteins involved in DNA damage signaling pathways such as phospho-ATM, CHK2, and p53. Interestingly, analysis of occurrence of H3K56 acetylation using ChIP-on-chip revealed its genome-wide spread, affecting genes involved in several pathways that are implicated in tumorigenesis such as cell cycle, DNA damage response, DNA repair, and apoptosis.

Highlights

Genomic integrity is maintained by a complex interplay of DNA replication, repair, and checkpoint signaling
H3 Lysine 56 Is Acetylated and Up-regulated during S Phase of the Cell Cycle in Mammals—The histone H3 lysine 56 has been reported to be acetylated in yeast and Drosophila; it was not detected in mammals [18]
Immunoblot of histones from untreated and histone deacetylase (HDAC) inhibitor-treated cells probed with antibodies specific to H3K56ac showed that histone H3 is acetylated at lysine 56 in human cells, and this acetylation increases upon class III HDAC inhibition (Fig. 1A, top panel)

Summary

Introduction

Genomic integrity is maintained by a complex interplay of DNA replication, repair, and checkpoint signaling. HEK 293T, Jurkat, and HeLa cells were treated with shown in Fig. 5A (top row, lane 4), damage-induced acetylated various DNA-damaging agents such as MMS, hydroxyurea, H3K56 foci completely colocalized with ␥-H2AX foci. We wanted level of H3K56 acetylation in mammalian cells seems to be to determine whether both ␥-H2AX and histone H3 acetylated required for cellular response to DNA damage.

Results

Conclusion