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¹ University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104
² Amgen Inc., Thousand Oaks, CA 91320

Address correspondence to Marko J. Kallio, University of Oklahoma Health Sciences Center, 975 N.E. 10th St., Biomedical Research Center, Rm. 266, Oklahoma City, OK 73104. Tel.: (405) 271-3404. Fax: (405) 271-7158. E-mail: marko-kallio{at}ouhsc.edu

Cdc20 is a substrate adaptor and activator of the anaphase-promoting complex/cyclosome (APC/C), the E3 ubiquitin ligase whose activity is required for anaphase onset and exit from mitosis. A green fluorescent protein derivative, Cdc20–GFP, bound to centrosomes throughout the cell cycle and to kinetochores from late prophase to late telophase. We mapped distinct domains of Cdc20 that are required for association with kinetochores and centrosomes. FRAP measurements revealed extremely rapid dynamics at the kinetochores (t_1/2 = 5.1 s) and spindle poles (t_1/2 = 4.7 s). This rapid turnover is independent of microtubules. Rapid transit of Cdc20 through kinetochores may ensure that spindle checkpoint signaling at unattached/relaxed kinetochores can continuously inhibit APC/C^Cdc20 targeting of anaphase inhibitors (securins) throughout the cell until all the chromosomes are properly attached to the mitotic spindle.

Key Words: ubiquitin; cell cycle; mitosis; spindle checkpoint; FRAP

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