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)