78 P.A. Sabelli · B.A. Larkins
CDK activity represses rereplication within one cell cycle by three overlap-
ping mechanisms that involve phosphorylation of ORC, reduced expression
of CDC6, and nuclear exclusion of the MCM2-7 proteins (Nguyen et al. 2001).
Only when all three mechanisms are inactivated does rereplication occur in
G2/M, indicating that inactivation of each mechanism is not sufficient to in-
duce rereplication. Consistent with this observation, yeast rereplication is
stimulated by mutations affecting CDC2/CycB kinase (Ravid et al. 2002). Fail-
ure to induce rereplication upon CDC6 disruption in human andXenopus
cells supports this view. However, it is conceivable that, because ORC, CDC6,
and MCM proteins work in a complex, deregulation of one type of protein
could affect the activity of the others and thereby suffice to trigger rerepli-
cation, as shown for theSchizosaccharomyces pombeCDC6 homolog Cdc18
(Nguyen et al. 2001).
CDT1 plays a key role in regulating the licensing of DNA replication, and
its overexpression results in rereplication in metazoa (DePamphilis et al.
2006) andArabidopsis(Castellano et al. 2004). In mammals, alteration in the
balance between CDT1 expression and its inhibitor, geminin, also results in
rereplication. Recent results point toa key role played by Emi1, an inhibitor
of APC/C, in the upstream regulation of both geminin/CDT1 and CycA/CDKs,
which result in rereplication through chromatin-bound, active CDT1 and hy-
pophosphorylated, and active, CDT1, CDC6, and MCMs (Machida and Dutta
2007). This work also highlights the importance of proper oscillation in
APC/C activity for pre-RC formation and coupling of S-phase to M-phase (see
below). However, plants do not possess Emi1 orthologs, and it remains to be
seen whether they utilize this pathway.
InXenopus, factors involved in nuclear import (i.e., Ran-GTP and im-
portins) prevent rereplication by sequestering MCM proteins in a CDK-
dependent manner, resulting in inhibition of pre-RCs at the G1/S-phase
transition (Yamaguchi and Newport 2003). In animals, depletion of CenA
induces endoreduplication in a fashion that might involve the coordinated
suppression of CDC6, CDT1, and ORC1 during S-phase (Mihaylov et al.
2002).
The process leading to activation of DNA synthesis is regarded as highly
conserved in eukaryotes, and plants possess orthologs ofORC(Gavin et al.
1995; Witmer et al. 2003),CDC6(de Jager et al. 2001; Ramos et al. 2001),CDT1
(Castellano et al. 2004; Masuda et al. 2004), andMCM(Sabelli et al. 1996,
1999; Springer et al. 2000) genes. InArabidopsis, a positive role forCDC6
andCDT1in endoreduplication has been reported, at least in leaf epidermis
cells (Castellano et al. 2001, 2004). Ectopic expression ofAtCDC6stimulates
endoreduplication in leaves, and the stability of CDC6 protein is enhanced in
endoreduplicating cells. This observation is intriguing because it highlights
the importance of context in determining the weight that certain licensing
factors carry in regulating checkpoints at both G/S-phase and S-phase/G tran-
sitions during endoreduplication.