SUMOylation and DeSUMOylation at a Glance - Journal of Cell ...
SUMOylation and DeSUMOylation at a Glance - Journal of Cell ...
SUMOylation and DeSUMOylation at a Glance - Journal of Cell ...
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<strong>Journal</strong> <strong>of</strong> <strong>Cell</strong> Science<br />
4250<br />
<strong>Journal</strong> <strong>of</strong> <strong>Cell</strong> Science 122 (23)<br />
dynamic than the other SUMO paralogues, <strong>and</strong><br />
its p<strong>at</strong>tern <strong>of</strong> conjug<strong>at</strong>ion responds differently to<br />
he<strong>at</strong> shock <strong>and</strong> stress (Ayaydin <strong>and</strong> Dasso, 2004;<br />
Saitoh <strong>and</strong> Hinchey, 2000).<br />
Smt3p, SUMO-2 <strong>and</strong> SUMO-3 can form<br />
conjug<strong>at</strong>ed chains through a single conserved<br />
acceptor lysine (Bylebyl et al., 2003; T<strong>at</strong>ham<br />
et al., 2001). SUMO-1 does not have an<br />
equivalent lysine residue, <strong>and</strong> thus probably<br />
does not act as a link in elong<strong>at</strong>ing chains<br />
in vivo. However, SUMO-1 might termin<strong>at</strong>e<br />
chains th<strong>at</strong> are elong<strong>at</strong>ed through serial<br />
conjug<strong>at</strong>ion <strong>of</strong> SUMO-2/3 (M<strong>at</strong>ic et al., 2008).<br />
Notably, although <strong>SUMOyl<strong>at</strong>ion</strong> does not seem<br />
to rely upon the geometry <strong>of</strong> chain linkages to<br />
confer inform<strong>at</strong>ion, as ubiquityl<strong>at</strong>ion does<br />
(Pickart <strong>and</strong> Fushman, 2004), SUMO<br />
conjug<strong>at</strong>es built from different paralogues or<br />
with different chain lengths can specify distinct<br />
target f<strong>at</strong>es, as discussed below.<br />
SUMO-interacting motifs<br />
SUMO-interacting motifs (SIMs) play a central<br />
role in both the enzymology <strong>of</strong> the SUMO<br />
p<strong>at</strong>hway <strong>and</strong> in the f<strong>at</strong>e <strong>of</strong> conjug<strong>at</strong>ed species.<br />
The best-characterized class <strong>of</strong> SIM consists <strong>of</strong> a<br />
hydrophobic core ([V/I]-x-[V/I]-[V/I]) flanked<br />
by a cluster <strong>of</strong> neg<strong>at</strong>ively charged amino acids<br />
(Kerscher, 2007). The SIM hydrophobic core<br />
can bind to an interaction surface on SUMO<br />
proteins in a parallel or antiparallel orient<strong>at</strong>ion.<br />
The acidic residues adjacent to the core might<br />
contribute to the affinity, orient<strong>at</strong>ion or<br />
paralogue specificity <strong>of</strong> binding (Hecker et al.,<br />
2006; Meulmeester et al., 2008). A variant SIM<br />
was recently defined within the transcriptional<br />
repressor CoREST1, consisting <strong>of</strong> [I/V/L]-<br />
[D/E]-[I/V/L]-[D/E]-[I/V/L] with N-terminal<br />
acidic residues (Ouyang et al., 2009). This SIM<br />
is highly selective for SUMO-2/3 binding, <strong>and</strong><br />
differs from previously identified SIMs because<br />
its core lacks a hydrophobic residue <strong>at</strong> position<br />
4. Notably, the diversity <strong>of</strong> SIMs identified to<br />
d<strong>at</strong>e is much less than the 16 known ubiquitinbinding<br />
domains (Grabbe <strong>and</strong> Dikic, 2009); it is<br />
reasonable to specul<strong>at</strong>e th<strong>at</strong> additional SIMs<br />
remain to be discovered.<br />
SUMO-processing, -activ<strong>at</strong>ing <strong>and</strong><br />
-conjug<strong>at</strong>ing enzymes<br />
Newly transl<strong>at</strong>ed SUMO proteins are cleaved to<br />
reveal C-terminal diglycine motifs. This<br />
processing is medi<strong>at</strong>ed by a family <strong>of</strong><br />
proteases known as ubiquitin-like-proteinspecific<br />
proteases (Ulps) in yeast <strong>and</strong><br />
sentrin-specific proteases (SENPs) in mammals<br />
(Mukhopadhyay <strong>and</strong> Dasso, 2007). Ulps <strong>and</strong><br />
SENPs also medi<strong>at</strong>e de<strong>SUMOyl<strong>at</strong>ion</strong> (see<br />
below).<br />
All SUMO paralogues share the same<br />
activ<strong>at</strong>ing (E1) <strong>and</strong> conjug<strong>at</strong>ing (E2) enzymes.<br />
These enzymes are structurally similar to E1 <strong>and</strong><br />
E2 enzymes <strong>of</strong> ubiquitin, <strong>and</strong> they share many <strong>of</strong><br />
the properties th<strong>at</strong> have been demonstr<strong>at</strong>ed for<br />
those enzymes (Hochstrasser, 2009). The<br />
yeast SUMO E1 enzyme is a heterodimer<br />
consisting <strong>of</strong> Aos1p (also known as Sae1 in<br />
vertebr<strong>at</strong>es) <strong>and</strong> Uba2p (also known as Sae2<br />
in vertebr<strong>at</strong>es), which show sequence similarity<br />
to the N-terminus <strong>and</strong> C-terminus <strong>of</strong> the<br />
monomeric ubiquitin E1 enzyme, respectively<br />
(Johnson et al., 1997). Aos1p-Uba2p c<strong>at</strong>alyzes<br />
the form<strong>at</strong>ion <strong>of</strong> a high-energy thioester bond<br />
between Uba2p <strong>and</strong> the SUMO C-terminus,<br />
with ATP hydrolysis to AMP <strong>and</strong> pyrophosph<strong>at</strong>e<br />
(Johnson et al., 1997; Lois <strong>and</strong> Lima, 2005). The<br />
activ<strong>at</strong>ed SUMO is subsequently passed to a<br />
cysteine in the active site <strong>of</strong> the E2 enzyme,<br />
Ubc9, through an intermolecular thiol-transfer<br />
reaction.<br />
Residues <strong>of</strong> Ubc9 th<strong>at</strong> are directly involved in<br />
the transfer <strong>of</strong> SUMO act to orient the lysine<br />
<strong>of</strong> the target protein <strong>and</strong> to decrease its pKa,<br />
resulting in a higher occurrence <strong>of</strong> its activ<strong>at</strong>ed,<br />
de-proton<strong>at</strong>ed st<strong>at</strong>e (Yunus <strong>and</strong> Lima, 2006).<br />
SUMO transfer from Ubc9 to some target<br />
proteins can occur through <strong>at</strong> least two ligaseindependent<br />
mechanisms. First, many<br />
SUMOyl<strong>at</strong>ed lysines lie within a consensus<br />
motif, �-K-x-[D/E] (where � is an aliph<strong>at</strong>ic<br />
branched amino acid <strong>and</strong> x is any amino acid).<br />
Ubc9 can directly recognize this motif <strong>and</strong><br />
conjug<strong>at</strong>e the lysine residue within it (Bernier-<br />
Villamor et al., 2002). Second, some SUMO<br />
substr<strong>at</strong>es contain SIMs th<strong>at</strong> promote their own<br />
conjug<strong>at</strong>ion (Meulmeester et al., 2008; Zhu<br />
et al., 2008). These SIMs bind to the SUMO<br />
moiety to which Ubc9 is <strong>at</strong>tached, thereby<br />
increasing its local concentr<strong>at</strong>ion <strong>and</strong><br />
facilit<strong>at</strong>ing <strong>SUMOyl<strong>at</strong>ion</strong>. Because SIMs show<br />
paralogue preference, this mechanism allows<br />
targets to be modified in a paralogue-selective<br />
manner. Notably, mammalian Ubc9 can itself be<br />
SUMOyl<strong>at</strong>ed on a nonconsensus lysine in its<br />
N-terminal helix (Knipscheer et al., 2008). This<br />
modific<strong>at</strong>ion does not inhibit its activity per se,<br />
but alters its target preference, increasing the<br />
conjug<strong>at</strong>ion <strong>of</strong> substr<strong>at</strong>es th<strong>at</strong> contain SIMs,<br />
which bind to the SUMO th<strong>at</strong> is conjug<strong>at</strong>ed to<br />
Ubc9.<br />
SUMO ligases<br />
SUMO ligases (E3 enzymes) facilit<strong>at</strong>e the<br />
majority <strong>of</strong> <strong>SUMOyl<strong>at</strong>ion</strong> under physiological<br />
conditions (Meulmeester et al., 2008). A number<br />
<strong>of</strong> SUMO ligases have been described, most <strong>of</strong><br />
which seem to be specific to metazoans.<br />
Siz/PIAS-family proteins<br />
All eukaryotes express proteins with Siz/PIAS<br />
RING-finger-like domains (SP-RING<br />
domains), which are known as SAP <strong>and</strong> Miz-<br />
finger domain (Siz) proteins in yeast <strong>and</strong><br />
protein inhibitor <strong>of</strong> activ<strong>at</strong>ed STAT (PIAS)<br />
proteins in vertebr<strong>at</strong>es (Hochstrasser, 2001). In<br />
budding yeast, Siz1p <strong>and</strong> Siz2p are required for<br />
most Smt3p conjug<strong>at</strong>ion (Johnson <strong>and</strong> Gupta,<br />
2001; Takahashi et al., 2001). Other SP-RING<br />
proteins, Zip3p <strong>and</strong> Mms21p, promote<br />
assembly <strong>of</strong> the synaptonemal complex<br />
between homologous chromosomes during<br />
meiosis (Cheng et al., 2006) <strong>and</strong> DNA repair<br />
(Potts, 2009), respectively. The five vertebr<strong>at</strong>e<br />
PIAS proteins (PIAS1, PIAS3, PIASx�,<br />
PIASx� <strong>and</strong> PIAS�) have been implic<strong>at</strong>ed in<br />
many processes, including gene expression,<br />
signal transduction <strong>and</strong> genome maintenance<br />
(Palvimo, 2007).<br />
Beyond their SP-RING domains, Siz/PIASfamily<br />
members share additional conserved<br />
motifs, including an N-terminal scaffold<br />
<strong>at</strong>tachment factor (SAF)-A/B, acinus, PIAS<br />
(SAP) motif, a PINIT motif, a SIM, <strong>and</strong> a<br />
C-terminal domain th<strong>at</strong> is rich in serine <strong>and</strong><br />
acidic amino acids (S/DE domain) (Palvimo,<br />
2007). The SAP domain directs the localiz<strong>at</strong>ion<br />
<strong>of</strong> Siz/PIAS proteins to chrom<strong>at</strong>in within the<br />
nucleus (Azuma et al., 2005; Palvimo, 2007).<br />
Structural analysis <strong>of</strong> a Siz1 fragment th<strong>at</strong> is<br />
sufficient for E3 activity in vitro shows th<strong>at</strong> it<br />
has an elong<strong>at</strong>ed tripartite architecture, formed<br />
by its N-terminal PINIT domain, SP-RING<br />
domain <strong>and</strong> C-terminal domain, termed the<br />
SP-CTD (Yunus <strong>and</strong> Lima, 2009). The SP-<br />
RING <strong>and</strong> SP-CTD domains are required for<br />
activ<strong>at</strong>ion <strong>of</strong> the Ubc9-SUMO thioester,<br />
whereas the PINIT domain directs<br />
<strong>SUMOyl<strong>at</strong>ion</strong> to the correct target lysine.<br />
RanBP2<br />
RanBP2 is a nuclear-pore protein th<strong>at</strong> localizes<br />
to the cytoplasmic face <strong>of</strong> the pore. RanBP2<br />
possesses a domain called the internal repe<strong>at</strong><br />
(IR) domain, which consists <strong>of</strong> two t<strong>and</strong>emly<br />
repe<strong>at</strong>ed sequences <strong>of</strong> around 50 residues (IR1<br />
<strong>and</strong> IR2), separ<strong>at</strong>ed by a 24-residue spacer (M).<br />
RanBP2 fragments containing the IR domain<br />
have SUMO ligase activity in vitro (Pichler<br />
et al., 2002). Structural analysis <strong>of</strong> a RanBP2<br />
fragment containing the IR1 <strong>and</strong> M domains<br />
indic<strong>at</strong>es th<strong>at</strong> RanBP2 enhances Ubc9 activity<br />
without direct contacts to the target protein<br />
(Reverter <strong>and</strong> Lima, 2005). It has thus been<br />
proposed th<strong>at</strong> RanBP2 promotes <strong>SUMOyl<strong>at</strong>ion</strong><br />
by aligning the Ubc9-SUMO thioester complex<br />
in an optimal configur<strong>at</strong>ion for substr<strong>at</strong>e<br />
interaction with the active site <strong>of</strong> Ubc9 <strong>and</strong> for<br />
c<strong>at</strong>alysis. Notably, the IR domain <strong>of</strong> RanBP2<br />
binds extremely stably to Ubc9 <strong>and</strong> the<br />
SUMO-1-conjug<strong>at</strong>ed form <strong>of</strong> RanGAP1,<br />
the activ<strong>at</strong>ing protein for the GTPase Ran<br />
(M<strong>at</strong>unis et al., 1998; Saitoh et al., 1998).<br />
Structural analysis yielded the puzzling result