Acetylation of Rsc4p by Gcn5p Is Essential in the Absence of Histone H3 Acetylation

Abstract

Rsc4p, a subunit of the RSC chromatin-remodeling complex, is acetylated at lysine 25 by Gcn5p, a wellcharacterized histone acetyltransferase (HAT). Mutation of lysine 25 does not result in a significant growth defect, and therefore whether this modification is important for the function of the essential RSC complex was unknown. In a search to uncover the molecular basis for the lethality resulting from loss of multiple histone H3-specific HATs, we determined that loss of Rsc4p acetylation is lethal in strains lacking histone H3 acetylation. Phenotype comparison of mutants with arginine and glutamine substitutions of acetylatable lysines within the histone H3 tail suggests that it is a failure to neutralize the charge of the H3 tail that is lethal in strains lacking Rsc4p acetylation. We also demonstrate that Rsc4p acetylation does not require any of the known Gcn5p-dependent HAT complexes and thus represents a truly novel function for Gcn5p. These results demonstrate for the first time the vital and yet redundant functions of histone H3 and Rsc4p acetylation in maintaining cell viability.Posttranslational modifications can augment protein function extending the diversity of proteins produced by the cell. For example, many thousands of proteins in a typical eukaryotic cell are modified by the covalent addition of a phosphate group (22), which can serve to either directly alter protein structure or mediate protein-protein interactions. Another well-studied modification is protein acetylation. Amino-terminal acetylation is one of the most common protein modifications in eukaryotes, occurring on ca. 85% of proteins. In addition, the acetylation of the epsilon-amino group of internal lysines occurs on ␣-tubulin, high-mobility group proteins, transcription factors, nuclear import factors, and histones (28).Histones H2A, H2B, H3, and H4 are the best-characterized substrates for posttranslational acetylation of internal lysines, with the majority of histone acetylation occurring on the unstructured amino-terminal "tails" of these proteins. These modifications are proposed to have two functions: to directly alter chromatin structure by weakening histone-DNA, as well as internucleosome interactions (1,2,11,33,34), and to act as a "molecular dock" for recruitment of factors that modify chromatin structure (42). Histone acetylation is catalyzed by histone acetyltransferases (HATs), which are comprised of a catalytic subunit complexed with accessory proteins that serve to either target or potentiate HAT activity. The best-studied catalytic subunit is Gcn5p, a component of multiple histone H3-specific HAT complexes in Saccharomyces cerevisiae. These complexes are responsible for acetylation of lysines 9, 14, 18, 23, 27, and 36 of histone H3 (14, 26, 37). All Gcn5p-dependent HAT complexes share the accessory proteins Ada2p and Ada3p, and several studies have demonstrated that ADA2 and ADA3 are essential for both the nucleosomal HAT activity of Gcn5p and its incorporation into HAT complexes (5, 7, 13). Indeed, the majority of p...

Histone H3K4 demethylation is negatively regulated by histone H3 acetylation in Saccharomyces cerevisiae

“…To test whether H3K4me3 was dependent on H3 acetylation, we used immunoblot analysis of S. cerevisiae whole cell extracts to examine levels of H3K4me3 in a mutant strain lacking the HATs responsible for histone H3 acetylation. In previously published work, we demonstrated that concomitant deletion of ADA2 and SAS3 abolishes the majority of histone H3 acetylation, without disrupting acetylation of the nonhistone substrate Rsc4 (20). ADA2 encodes a noncatalytic component of all Gcn5-dependent HATs, whereas SAS3 codes for the catalytic subunit of the NuA3 HAT complex.…”

Histone H3K4 and H3K36 Methylation Independently Recruit the NuA3 Histone Acetyltransferase in Saccharomyces cerevisiae

“…Indeed, proteomic studies show that Nto1 and Taf14 both contain acetylated lysines (Henriksen et al 2012), and so it is possible that the YEATS domain is binding to a modified lysine in the NuA3 complex. Such a role is seen for the Rsc4 bromodomain, which binds to an acetylated lysine in the complex to regulate its function (VanDemark et al 2007;Choi et al 2008).…”

Acetylome Profiling Reveals Overlap in the Regulation of Diverse Processes by Sirtuins, Gcn5, and Esa1

“…7B). Intriguingly, however, Ada2 is not required for Gcn5-dependent acetylation of Rsc4 (48), suggesting that Gcn5 may target some substrates independently of its known interaction partners. In support of this idea, strains lacking ADA2 grew better than those lacking GCN5, relative to control strains, especially on synthetic media (supplemental Fig.…”