Histone H4 Lysine 12 Acetylation Regulates Telomeric Heterochromatin Plasticity in Saccharomyces cerevisiae Bo O. Zhou1, Shan-Shan Wang 1 , Yang Zhang 1 , Xiao-Hong Fu 1 , Wei Dang1, Brian A. Lenzmeier2, Jin-Qiu 1 Zhou * 1 State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China, 2 School of Science, Buena Vista University, Storm Lake, Iowa, United States of America Abstract Recent studies have established that the highly condensed and transcriptionally silent heterochromatic domains in budding yeast are virtually dynamic structures. The underlying mechanisms for heterochromatin dynamics, however, remain obscure. In this study, we show that histones are dynamically acetylated on H4K12 at telomeric heterochromatin, and this acetylation regulates several of the dynamic telomere properties. Using a de novo heterochromatin formation assay, we surprisingly found that acetylated H4K12 survived the formation of telomeric heterochromatin. Consistently, the histone acetyltransferase complex NuA4 bound to silenced telomeric regions and acetylated H4K12. H4K12 acetylation prevented the over-accumulation of Sir proteins at telomeric heterochromatin and elimination of this acetylation caused defects in multiple telomere-related processes, including transcription, telomere replication, and recombination. Together, these data shed light on a potential histone acetylation mark within telomeric heterochromatin that contributes to telomere plasticity. Citation: Zhou BO, Wang S-S, Zhang Y, Fu X-H, Dang W, et al. (2011) Histone H4 Lysine 12 Acetylation Regulates Telomeric Heterochromatin Plasticity in Saccharomyces cerevisiae. PLoS Genet 7(1): e1001272. doi:10.1371/journal.pgen.1001272 Editor: Hiten D. Madhani, University of California San Francisco, United States of America Received June 10, 2010; Accepted December 8, 2010; Published January 13, 2011 Copyright: 2011 Zhou et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work is supported by grants from Ministry of Science and Technology of China (2005CB522402) and National Natural Science Foundation of China (90919027). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: jqzhou@sibs.ac.cn Introduction Saccharomyces cerevisiae telomeres and mating type loci (HMR and HML) are well-characterized transcriptionally silenced domains. Heterochromatin tends to initiate at a specific DNA element and propagates along a chromatin fiber to repress the expression of nearby genes [1]. The major structural components of hetero- chromatin in yeast are known as Sir (silent information regulators) proteins, including Sir2, Sir3 and Sir4 [1]. For telomeric heterochromatin, the double-stranded telomeric DNA-binding protein Rap1 interacts with Sir4 and thereby recruits the whole Sir complex to the telomeres [2]. Once recruited to telomeres, Sir2 deacetylates a critical K16 acetyl mark on histone H4 [3], a process required for Sir proteins to bind throughout the subtelomeric regions of ,3 kb proximal to the terminal ,350- bp TG-tracts [4–8]. The spreading of heterochromatin must be restricted by boundaries between silent and active chromatin [9]. Since histone H4 amino acid residues 16 to 29 are primarily required for Sir3 binding [4], histone H4K16 acetylation dominantly prevents Sir- mediated heterochromatin spreading [6,7]. Genetic and biochemical data also show that unacetylated H4 K5, 8, 12 together can substitute for the mutation of H4K16 in Sir3 binding [4,5]. Therefore, it has been generally believed that the acetylation of histone H4 at K5, 8 and 12, catalyzed by NuA4 complex [6,10], cumulatively antago- nizes heterochromatin spreading. However, it remains unclear whether H4 K5, 8 and 12 contribute equally to this process. Yeast heterochromatin possesses the ability to exchange chromatin-bound Sir3 for soluble unbound protein throughout the cell cycle [11]. In addition, heterochromatin is surprisingly permissive to activators, co-activators and transcriptional pre- initiation-complex (PIC) as well [12–14]. Moreover, heterochro- matin doesn’t prohibit active base-pair substitutions [15]. These lines of evidence have revealed the dynamic aspect of silent- and hetero-chromatin. One outstanding question of concern is what contributes to the dynamics of heterochromatin. Accordingly, it would be beneficial to know if altering the dynamics of heterochromatin structure has biological consequences. In this study, we show that the heterochromatin in yeast harbors a dynamic H4K12 acetylation mark which suppresses Sir- mediated aberrant condensation of telomeric heterochromatin and promotes telomere-related processes, including telomere transcription, replication and recombination. Thus, histone acetylation may provide a scheme for yeast cells to maintain partially flexible telomeric heterochromatin that allows for normal changes in DNA metabolism. Results A distinct system for studying de novo telomeric heterochromatin formation in yeast To better understand the orchestrated events that are involved in establishing telomeric heterochromatin, we developed a new telomeric heterochromatin formation assay, building upon the de novo telomere addition system originally established by Gottschl- ing’s lab [16]. This new assay allowed us to study telomeric heterochromatin formation under conditions where global gene expression, especially the expression of the silencing-related genes, PLoS Genetics | www.plosgenetics.org 1 January 2011 | Volume 7 | Issue 1 | e1001272