Different phosphorylation states of the anaphase promoting complex in response to antimitotic drugs: A quantitative proteomic analysis

doi:10.1073/pnas.0709807104

Different phosphorylation states of the anaphase promoting complex in response to antimitotic drugs: A quantitative proteomic analysis

Departments of *Neurobiology and
^†Pathology, Harvard Medical School and Children's Hospital Boston, Boston, MA 02115;
^‡Department of Systems Biology, Harvard Medical School, Boston, MA 02115;
^§Department of Neurology, Harvard Medical School and Brigham and Women's Hospital, Boston, MA 02115; and
^¶Interdisciplinary Center for Scientific Computing, University of Heidelberg, D-69120 Heidelberg, Germany

Contributed by Marc W. Kirschner, October 16, 2007 (received for review June 27, 2007)

Abstract

The anaphase promoting complex (APC) controls the degradation of proteins during exit from mitosis and entry into S-phase. The activity of the APC is regulated by phosphorylation during mitosis. Because the phosphorylation pattern provides insights into the complexity of regulation of the APC, we studied in detail the phosphorylation patterns at a single mitotic state of arrest generated by various antimitotic drugs. We examined the phosphorylation patterns of the APC in HeLa S3 cells after they were arrested in prometaphase with taxol, nocodazole, vincristine, or monastrol. There were 71 phosphorylation sites on nine of the APC subunits. Despite the common state of arrest, the various antimitotic drug treatments resulted in differences in the phosphorylation patterns and phosphorylation stoichiometries. The relative phosphorylation stoichiometries were determined by using a method adapted from the isotope-free quantitation of the extent of modification (iQEM). We could show that during drug arrest the phosphorylation state of the APC changes, indicating that the mitotic arrest is not a static condition. We discuss these findings in terms of the variable efficacy of antimitotic drugs in cancer chemotherapy.

Footnotes

^‖To whom correspondence should be addressed. E-mail: marc{at}hms.harvard.edu
Author contributions: J.A.J.S., H.S., A.G., and M.W.K. designed research; J.A.J.S., H.S., and A.G. performed research; J.A.J.S., K.P., M.S., M.K., and F.H. contributed new reagents/analytic tools; J.A.J.S., H.S., K.P., and M.K. analyzed data; and J.A.J.S., H.S., and M.W.K. wrote the paper.
The authors declare no conflict of interest.
This article contains supporting information online at www.pnas.org/cgi/content/full/0709807104/DC1.
Freely available online through the PNAS open access option.