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In-Depth Information
the metaphase checkpoint
[61]
. (2) Active APC:Cdc20
promotes dissociation of the MCC. This self-activation loop
accelerates the release from the checkpoint, so that anaphase
follows soon after full chromosome alignment.
The anaphase switch (
Box 14.2
) has two stable steady
states: (1) Clb2 level high, Mad2 active, Cdc20 inactive;
and (2) Mad2 inactive, Cdc20 active, Clb2 level low. At the
metaphase/anaphase transition, the switch is flipped from
the Cdc20-inactive state to the Cdc20-active state. Conse-
quently, APC:Cdc20 degrades securin, releasing separase
to cleave cohesin rings and trigger anaphase (separation of
sister chromatids). In addition, separase has a non-catalytic
role
[62]
, inhibiting the phosphatase that has been keeping
Net1 active throughout the early stages of mitosis
[63]
.
Inhibition of the phosphatase allows Clb2-kinase and other
mitotic kinases (notably Polo kinase) to phosphorylate
Net1 and release Cdc14
[63
e
65]
. Meanwhile, APC:Cdc20
degrades Clb2, and the combination of low Clb2-kinase
activity and high Cdc14-phosphatase activity silences the
mitotic checkpoint
[66]
. (Cohesin cleavage at anaphase
creates tensionless chromosomes, but they do not reactivate
Mad2 because now Clb2:Cdk1 activity is low and Cdc14
activity is high.) Cdc14 promotes activation of the G1-
stabilizers (CKI and Cdh1). CKI inhibits any remaining
Clb-dependent kinase activity, and Cdh1 destroys Polo
kinase and Cdc20.
In
Figure 14.8
(right), we show how chromosome
alignment removes the mitotic-arrest state by a saddle-node
bifurcation, allowing the anaphase switch to flip on. Cdc20
and Cdc14 activities rise, but before they can reach the
upper steady state, they induce degradation of Clb proteins
and activation of the G1-stabilizers. The transition to G1
phase removes Cdc20 and Cdc14, restoring the newborn
cells to the beginning of the cycle.
Each of these G1 stabilizers can be phosphorylated and
neutralized by SPF and MPF, creating a fundamental
bistable switch between steady states for G1 and S-G2-M.
Progression through the mammalian cycle may also be
envisioned as flipping this switch on by a starter kinase
(Cdk2:CycE) and off by an exit pathway (Cdc20 and CAPs).
In particular, the Start transition in budding yeast is
analogous to the 'restriction point' (RP) in the mammalian
cell cycle
[70]
. The molecular mechanisms of the two
checkpoints are almost identical under the identification of
SBF with E2F, Whi5 with RB, Cln2 with CycE, Cln3 with
CycD, and Clb5 with CycA. Both checkpoints are
responsive to extracellular signals. In budding yeast cells
a
factor activates a MAP kinase pathway that upregulates
a stoichiometric inhibitor of Cln3 and blocks cells in pre-
Start. In mammalian cells, growth factor (GF) activates
a MAP kinase pathway that upregulates a transcription
factor for CycD and promotes passage through the RP. The
logic differs because, for yeast cells, the default state of the
cell cycle is vegetative growth and division, and yeast cells
need a definite signal (
a
factor) to block progression
through the cell cycle and start the mating process. For
mammalian somatic cells, on the other hand, the default
state is G1 arrest: only special cells under special
circumstances are permitted to grow and divide. The
permission is granted by specific GFs that promote passage
through the RP.
Bistability at the mammalian RP has been demonstrated
experimentally in elegant experiments by Yao et al.
[71]
(see
Figure 3
in that paper). In a later paper
[72]
, the
same authors showed experimentally that RP bistability is
due to the double-negative feedback loop (RB -
j
E2F
/
CycE -
RB).
Bistability in the mitotic exit mechanism of mammalian
cells is still a matter of some disagreement
[73,74]
,although
in our opinion the evidence is definitely in favor of a bistable
switch
[45,75]
. Our description of a bistable anaphase switch
in budding yeast, based on experiments by Uhlmann and
colleagues
[66]
, is confirmed by similar experiments with
fruit flies
[76]
and mammalian cells
[77]
.
Budding yeast cells differ from most other types of
organism (including fission yeast cells, plant cells, fruit fly
embryos, frog embryos and mammalian cells) in lacking
a checkpoint at the G2/M transition. Budding yeast cells are
unique in having many small chromosomes that need not
undergo much condensation during mitosis; hence they can
go almost directly from S phase into mitosis
[78]
. Other
organisms, on the contrary, need a gap phase (G2) between
the end of S and the onset of M, during which chromosomes
are replicated and available for transcription. When these
cells enter mitosis, their chromosomes become so highly
condensed that all transcription ceases. The duration of
G2 is determined by the G2/M checkpoint. During G2
phase, mitotic cyclins accumulate in complexes that are
j
IRREVERSIBLE TRANSITIONS IN THE
MAMMALIAN CELL CYCLE
The molecular machinery regulating progression
through the mammalian cell cycle is very similar in
principle to the yeast cell cycle. In the following para-
graphs we will highlight the most important similarities
and differences.
In mammalian cells, as in yeast, the G1 phase of the cell
cycle is stabilized by three types of interaction that keep low
the activities of S-phase promoting factor
(SPF
¼
Cdk2:CycA) and M-phase promoting factor
¼
Cdk1:CycB): (1) high activity of APC:Cdh1, which
promotes degradation of both CycA and CycB
[67]
; (2) high
abundance of CKIs that inhibit SPF and MPF heterodimers
[68]
; (3) high abundance of an inhibitor (retinoblastoma
protein, RB) of the transcription factors (E2F family) that
promote synthesis of early cyclins (CycE and CycA)
[69]
.
(MPF
