Biology Reference
In-Depth Information
form of ROCK II, and thus NPM/B23 cannot bind to Rho-unbound ROCK II), and
ROCK II is super activated (5-10-fold higher than unbound ROCK II) by the
NPM/B23-binding (Ma et al.
2006
). Although unphosphorylated NPM/B23 can
bind to ROCK II, NPM/B23 acquires a significantly higher binding affinity to
ROCK II upon Thr199 phosphorylation. Under a physiological condition where
the protein concentrations are limited, especially the Rho-bound ROCK II at
centrosomes, phosphorylation-dependent upregulation of the ROCK II-NPM/B23
interaction becomes essential. Indeed, most (if not all) of the ROCK II-bound
NPM/B23 in cells are Thr199-phosphorylated. The superactivation of ROCK II by
NPM/B23 binding is critical for the timely initiation of centrosome duplication,
and is the primary downstream event of CDK2-cyclin E for the initiation of
centrosome duplication. For instance, downregulation of the CDK2 activity either
by expression of the dominant negative CDK2 or by depletion of cyclin E and
cyclin A results in complete inhibition of centrosome duplication (Hanashiro et al.
2008
), but introduction of the Rho-independent constitutively active ROCK II
mutant can override the inhibition of centrosome duplication by inactivation of
CDK2 in a NPM/B23 binding-dependent manner (Hanashiro et al.
2011
). To sum
up, in late G1, NPM/B23 acquires a high binding affinity to ROCK II by CDK2-
cyclin E mediated phosphorylation, and binds to and superactivates the Rho-bound
ROCK II, which in turn rapidly acts on the centrosomal target(s) to initiate cen-
trosome duplication. At the same time, CDK2-cyclin E targets proteins like Rb to
initiate DNA replication, and thus initiation of centrosome duplication and DNA
replication occurs in a coordinated manner.
Because CDK2-cyclin E plays a key role in the initiation of centrosome
duplication, the proteins that control the CDK2/cyclin E activity are also expected
to be critically involved in the regulation of centrosome duplication. The p53
tumor suppressor protein and its transactivation target p21
Waf1/Cip1
(p21) CDK
inhibitor are the well-known regulatory proteins of the CDK2 activity (Sherr and
Roberts,
1999
). The involvement of p53 in the regulation of centrosome dupli-
cation was initially recognized by the observations that cells and tissues from
p53-deficient mice show a high frequency of centrosome amplification resulting
from overduplication of centrosomes (Fukasawa et al.
1996
,
1997
). The sub-
sequent studies have revealed how p53 participates in the regulation of centrosome
duplication. p53 and p21 are known to present at a basal level in cycling cells,
monitoring untimely activation of CDK2-cyclin E in early to mid G1 phase
(Minella et al.
2002
; Nevis et al.
2009
). When cyclin E expression is induced at
late G1, the concentration of active CDK2-cyclin E complexes rapidly increases to
the level beyond the capacity of the p53-p21 monitoring system, leading to ini-
tiation of centrosome duplication as well as DNA replication. Indeed, overex-
pression of exogenously introduced cyclin E in cells harboring wild-type p53 (and
thus, continual activation of CDK2-cyclin E beyond the capacity of the p53-p21
monitoring system) results in initiation of centrosome duplication in early G1
phase (Mussman et al.
2000
). In the absence of p53, p21 cannot be transactivated,
hence allowing fortuitous activation of CDK2-cyclin E in early and mid-G1.
Because Rho-bound ROCK II are already available in early G1, CDK2-cyclin E
