New Insights into the Role of BubR1 in Mitosis and Beyond - International Review of Cell and Molecular Biology - page 245

inhibits the APC/C just enough to keep it from forcing premature exiting
mitosis before kinetochores become competent to generate the SAC signal.
Buffin et al. (2007) and Rahmani et al. (2009) showed that the timer also
exists in Drosophila and depends on Mad2 and BubR1. Even though normal
mitosis is very rapid in these cells (on the order of 9 min from NEB to ana-
phase onset), mutations in these proteins rendered it faster still (7 min). They
further showed that the onset of Cyclin B degradation was correspondingly
shifted 2 min earlier as well. Rahmani et al. did, however, detect one differ-
ence between the timer and the SAC function of BubR1. Flies with a muta-
tion in the BubR1 KEN1 box ( bubR1-KEN ) had, as expected, a
nonfunctional SAC. Basal mitotic timing was not accelerated, however,
thereby suggesting that the SAC function had a more stringent requirement
for KEN1 than does the timer function. However, fly cells simultaneously
expressing bubR1-KEN 1 and lacking Mad2 had a greatly accelerated mitotic
timer, faster than mad2 mutants alone. The biochemistry of mammalian
MCC described earlier argues that BubR1 does not stably bind CDC20
without an intact KEN1 box. One interpretation of the Drosophila study
would be that small amounts of a functional MCC can assemble even if
BubR1 lacks the KEN1 box (but only in the presence of Mad2), or if the
cell lacks Mad2, but has intact BubR1. This small amount of MCC would
then be adequate for normal timing (slightly inhibiting the APC/C), but not
for a proper SAC response (complete inhibition). But in cells lacking both
the KEN1 and Mad2, no MCC forms at all, and APC/C begins targeting
Cyclin B and Securin as soon as the cell enters M-phase.
4.5. BubR1 acetylation and sumoylation
Why is BubR1, with its two KEN boxes, not ubiquitinated by APC/C and
degraded? Choi et al. (2009, 2012) reported that BubR1 binds a complex of
tumor suppressor BRCA2 and the acetyltransferase PCAF at unattached
kinetochores, which results in acetylation of Lysine 250 (just after the
TPR fold). They presented evidence that this modification was crucial
for preventing BubR1 itself from ubiquitination. With an antibody specific
to the acetylated form, the authors reported that this acetylation is first
detectable in prometaphase on unattached kinetochores, and disappears
on kinetochores when properly attached to the spindle. A nonacetylatable
K250R mutant was SAC defective, and the protein was prematurely
ubiquitinated by the APC/C and consequently degraded. In a K250Q
mutant (which mimics constitutive acetylation), BubR1 degradation and