The Dephosphorylated Form of the Anaphase-Promoting Complex Protein Cdc27/Apc3 Concentrates on Kinetochores and Chromosome Arms in Mitosis
Volume 1, Issue 4
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Pages 287 - 296http://dx.doi.org/10.4161/cc.1.4.139
Authors: Leana M. Topper, Michael S. Campbell, Stuart Tugendreich, John R. Daum, Daniel J. Burke, Philip Hieter and Gary J. Gorbsky View affiliations
Cell cycle regulated protein ubiquitination and degradation within subcellular
domains may be essential for the normal progression of mitosis. Cdc27 is a conserved
component of an essential M-phase ubiquitin-protein ligase called the anaphase-promoting
complex/cyclosome. We examined the subcellular distribution of Cdc27 in greater
detail in mammalian cells and found Cdc27 concentrated at spindle poles and
on spindle microtubules as previously described, but also found Cdc27 at kinetochores
and along chromosome arms. This localization was not dependent on intact microtubules.
While the great majority of Cdc27 protein in M phase cells is highly phosphorylated,
only the dephosphorylated form of Cdc27 was found associated with isolated chromosomes.
Kinases that also associate with isolated chromosomes catalyzed the in vitro
phosphorylation of the chromosome-associated Cdc27. Microinjection of anti-Cdc27
antibody into cells causes arrest at metaphase. Microinjection of cells with
anti-Mad2 antibody normally induces premature anaphase onset resulting in catastrophic
nondisjunction of the chromosomes. However, coinjection of anti-Cdc27 antibody
with anti-Mad2 antibody resulted in metaphase arrest. The association of dephosphorylated
APC/C components with mitotic chromosomes suggests mechanisms by which the spindle
checkpoint may regulate APC/C activity at mitosis.
Centromere, Ubiquitin, Checkpoint, Cell cycle, Proteasome