The Histone Chaperone Anti-silencing Function 1 Stimulates the Acetylation of Newly Synthesized Histone H3 in S-phase

Abstract

Anti-silencing function 1 (Asf1) is a highly conserved chaperone of histones H3/H4 that assembles or disassembles chromatin during transcription, replication, and repair. We have found that budding yeast lacking Asf1 has greatly reduced levels of histone H3 acetylated at lysine 9. Lysine 9 is acetylated on newly synthesized budding yeast histone H3 prior to its assembly onto newly replicated DNA. Accordingly, we found that the vast majority of H3 Lys-9 acetylation peaked in S-phase, and this S-phase peak of H3 lysine 9 acetylation was absent in yeast lacking Asf1. By contrast, deletion of ASF1 has no effect on the S-phase specific peak of H4 lysine 12 acetylation; another modification carried by newly synthesized histones prior to chromatin assembly. We show that Gcn5 is the histone acetyltransferase responsible for the S-phase-specific peak of H3 lysine 9 acetylation. Strikingly, overexpression of Asf1 leads to greatly increased levels of H3 on acetylation on lysine 56 and Gcn5-dependent acetylation on lysine 9. Analysis of a panel of Asf1 mutations that modulate the ability of Asf1 to bind to histones H3/H4 demonstrates that the histone binding activity of Asf1 is required for the acetylation of Lys-9 and Lys-56 on newly synthesized H3. These results demonstrate that Asf1 does not affect the stability of the newly synthesized histones per se, but instead histone binding by Asf1 promotes the efficient acetylation of specific residues of newly synthesized histone H3.The eukaryotic genome is packaged into a nucleoprotein structure known as chromatin. The basic repeating unit of chromatin, the nucleosome, is made up of 147 base pairs of DNA wrapped around a histone octamer (two molecules each of histone proteins H2A, H2B, H3, and H4) (1). Chromatin provides a formidable obstacle to the cellular machinery gaining access to the DNA. Indeed, it is becoming apparent that chromatin is a highly dynamic structure that tightly regulates all nuclear processes that use DNA as a substrate; including transcription, DNA replication, repair, and recombination (2-4). Thus, the mechanisms by which chromatin structures are made and modified are fundamental questions of broad interest.The entire eukaryotic genome is assembled into chromatin following DNA replication, and this occurs in a stepwise manner; a tetramer of histones H3/H4 is deposited first followed by two dimers of H2A/H2B on the outside of the tetramer. Histone-binding proteins, termed histone chaperones, mediate chromatin assembly. It is known that when this process is coupled to DNA replication in vitro, it is mediated by the histone H3/H4 chaperone chromatin assembly factor 1 (CAF-1) 2 (5) together with another H3/H4 chaperone termed anti-silencing function 1 (Asf1) (6). CAF-1 and Asf1 have also been shown to localize to the sites of DNA replication (7, 8), supporting a role in assembling chromatin following DNA replication in vivo. The fact that CAF-1 and Asf1 co-purify with the replicationspecific histone variant H3.1 from HeLa cells is further evidence that Asf1 ...

Acetylated histone H3K56 interacts with Oct4 to promote mouse embryonic stem cell pluripotency

“…The histone chaperone Asf1 is required for the acetylation of H3K56, whereas the effect of Asf1 deletion TS4_compacted morula TS4_inner cell mass Theiler_stage_4 TS4_embryo TS4_second polar body TS4_zona pellucida Theiler_stage_5 TS5_inner cell mass TS4_extraembryonic TS5_embryo TS3_zona pellucida Theiler_stage_3 TS3_second polar body TS3_4-8 on other histone acetylation sites is relatively minor as determined by Western blots and MS (18,36,37). Using Western blots, we found that knockdown of Asf1a with shAsf1a in mouse E14Tg2a cells decreases H3K56ac significantly but has very little effect on other acetylation sites in histone H3 (Fig.…”

Regulation of Histone H3 Lysine 56 Acetylation in Schizosaccharomyces pombe

“…H3 K56R), rtt109⌬ mutants display sensitivity to replicate stress (18,19,21). Interestingly, loss of the H3/H4 histone chaperone ASF1 also results in the absence of H3 Lys-56-Ac (13,16,18,19,21,22). Taken together, these data suggest an intimate, but poorly understood, functional relationship between Rtt109, Asf1, and H3 Lys-56-Ac.…”

Autoacetylation of the Histone Acetyltransferase Rtt109

“…Rtt109, with the histone chaperone Asf1 (anti-silencing function 1), acetylates lysine 56 (Lys-56) on the histone H3 core domain (12,14,15,19). Asf1 is thought to form a transient complex with Rtt109 and stimulate activity by properly presenting H3-H4 dimers for Lys-56 acetylation (23)(24)(25)(26)(27). This mark occurs on newly synthesized H3 in Saccharomyces cerevisiae and Schizosaccharomyces pombe during S-phase and is required for the assembly of H3-H4 dimers onto DNA during cell replication (12,15,28).…”