Sem1p and Ubp6p orchestrate telomeric silencing by modulating histone H2B ubiquitination and H3 acetylation

Nucleic Acids Research, Apr 2009

Histone covalent modifications and 26S proteasome-mediated proteolysis modulate many regulatory events in eukaryotes. In Saccharomyces cerevisiae, heterochromatin mediates transcriptional silencing at telomeres, HM loci and rDNA array. Here, we show that proteasome-associated Sem1p and its interacting partner, Ubp6p (a deubiquitinating enzyme), are essential to maintain telomeric silencing. Simultaneous deletion of SEM1 and UBP6 induces dramatic silencing defect accompanied by significantly increased level of ubiquitinated-histone H2B and markedly reduced levels of acetylated-lysine 14 and 23 on histone H3 at the telomeres. Further, the loss of Sem1p and Ubp6p triggers relocation of silencing factors (e.g. Sir proteins) from telomere to HM loci and rDNA array. Such relocation of silencing factors enhances gene silencing at HM loci and rDNA array, but diminishes telomeric silencing. Interestingly, both Sem1p and Ubp6p participate in the proteolytic function of the proteasome. However, we find that the telomeric silencing is not influenced by proteolysis. Taken together, our data demonstrate that Sem1p and Ubp6p maintain telomeric heterochromatin structure (and hence silencing) through modulation of histone covalent modifications and association of silencing factors independently of the proteolytic function of the proteasome, thus offering a new regulatory mechanism of telomeric silencing.

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Sem1p and Ubp6p orchestrate telomeric silencing by modulating histone H2B ubiquitination and H3 acetylation

Song Qin 1 2 Qien Wang 2 Alo Ray 2 Gulzar Wani 2 Qun Zhao 2 Sukesh R. Bhaumik 0 Altaf A. Wani 1 2 3 0 Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine , Carbondale, IL 62901 1 Comprehensive Cancer Center, The Ohio State University , Columbus, OH 43210 2 Department of Radiology 3 Department of Molecular and Cellular Biochemistry, College of Medicine, The Ohio State University , Columbus, OH 43210, USA - Histone covalent modifications and 26S proteasome-mediated proteolysis modulate many regulatory events in eukaryotes. In Saccharomyces cerevisiae, heterochromatin mediates transcriptional silencing at telomeres, HM loci and rDNA array. Here, we show that proteasome-associated Sem1p and its interacting partner, Ubp6p (a deubiquitinating enzyme), are essential to maintain telomeric silencing. Simultaneous deletion of SEM1 and UBP6 induces dramatic silencing defect accompanied by significantly increased level of ubiquitinated-histone H2B and markedly reduced levels of acetylated-lysine 14 and 23 on histone H3 at the telomeres. Further, the loss of Sem1p and Ubp6p triggers relocation of silencing factors (e.g. Sir proteins) from telomere to HM loci and rDNA array. Such relocation of silencing factors enhances gene silencing at HM loci and rDNA array, but diminishes telomeric silencing. Interestingly, both Sem1p and Ubp6p participate in the proteolytic function of the proteasome. However, we find that the telomeric silencing is not influenced by proteolysis. Taken together, our data demonstrate that Sem1p and Ubp6p maintain telomeric heterochromatin structure (and hence silencing) through modulation of histone covalent modifications and association of silencing factors independently of the proteolytic function of the proteasome, thus offering a new regulatory mechanism of telomeric silencing. In Saccharomyces cerevisiae, genes in three distinct genomic regions: telomeres, mating-type loci (HMR and HML) and rDNA array, are transcriptionally silenced in a nonspecific fashion. Establishment of silencing requires formation of silent chromatin and the proper function of the silent information regulatory (SIR) complex composed of Sir2p, Sir3p and Sir4p (1,2). This locus-specific silencing is known to be sensitive to the status changes in methylation, acetylation and ubiquitination of the core histones (35). These changes alter the binding of silencing proteins to chromatin. Many histone modification enzymes, such as Set1p, Dot1p, Ubp10p and Rad6p, have been shown to be involved in silencing. Rad6p, an E2 ubiquitinconjugating enzyme, functions together with E3 ubiquitin ligase Bre1p to attach ubiquitin to lysine 123 (K123) of histone H2B (47). H2B ubiquitination is required for functions of Set1p and Dot1p (810). Set1p methylates K4 in histone H3 N-terminal tail (46,11), while Dot1p methylates K79 in the core domain of histone H3 (46,12). Simultaneous deletion of DOT1 and SET1 significantly reduces the binding of Sir proteins to telomeres, indicating that these two modifications function together to mediate silencing. Recently, a deubiquitinating enzyme Ubp10p was found to be involved in silencing (13,14). Either ubp10D or mutation in the catalytic domain of Ubp10p results in reduced silencing, especially at telomeres. Ubp10p has been implicated to participate in H2B deubiquitination which influences H3K4 and H3K79 methylation in silent chromatin regions (13,14). Thus, a delicate equilibrium between H2B ubiquitination and deubiquitination is critical for establishing methylation pattern of H3K4 and H3K79 in silent chromatin domains. Several studies implicate acetylation of lysine residues on histone N-terminal tails to transcriptional activation while deacetylation is more frequently associated with silent chromatin. The status of histone acetylation is controlled by a dynamic equilibrium between histone acetyltransferases (HATs) and histone deacetylases (HDACs). Many enzymes modulating the status of histone acetylation, such as Esa1p, Sas2p, Sir2p and Hat1p, contribute to silencing in budding yeast (1518). Among the four acetylable lysines in the N-terminal tail of histone H4, only mutation of H4K16 significantly affects telomeric silencing (19). Among the five acetylable lysine residues in the N-terminal tail of histone H3, K14 and K23 (H3K14/K23) are more important than K9 or K18 in telomeric silencing (17). Recently, Taverna et al. (20) have shown that histone H3 K14 acetylation is correlated with histone H2B ubiquitination via H3 K4 methylation. Thus, the enzymes involved in histone H2B deubiquitination can potentially regulate telomeric silencing. Ubp6p is one of the two deubiquitinating enzymes associated with the lid subcomplex of the 26S proteasome (1,2125). Association of Ubp6p with the proteasome is critical for the deubiquitinating activity of Ubp6p (26) and for the half-life of ubiquitin (27). Although the exact roles of Ubp6p remain to be discovered (...truncated)


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Song Qin, Qien Wang, Alo Ray, Gulzar Wani, Qun Zhao, Sukesh R. Bhaumik, Altaf A. Wani. Sem1p and Ubp6p orchestrate telomeric silencing by modulating histone H2B ubiquitination and H3 acetylation, Nucleic Acids Research, 2009, pp. 1843-1853, 37/6, DOI: 10.1093/nar/gkn1049