Abstract 3770: The spindle assembly checkpoint requires phosphatase activity

Abstract

Abstract The spindle assembly checkpoint (SAC) prevents activation of the anaphase-promoting complex (APC/C) until all chromosomes are correctly attached to the mitotic spindle. A defective SAC contributes to errors in chromosome segregation, which promotes aneuploidy and may eventually lead to cancer. Early in mitosis, the mitotic checkpoint complex (MCC) inactivates the APC/C by binding the APC/C activating protein cdc20 until the chromosomes are properly aligned, at which point the MCC dissociates and releases cdc20 to activate the APC/C. Once the APC/C is activated, it targets many substrates for degradation, including cyclin B and securin, and the cell progresses into anaphase. The precise molecular mechanisms regulating SAC inactivation are still poorly understood. We sought to determine the role of mitotic phosphatases in the SAC. To address this question, we treated mitotic cells with various phosphatase inhibitors to examine their effect on APC/C activation. Using this approach we identified two inhibitors, calyculin A and okadaic acid (1uM), that cause MCC dissociation and APC/C activation leading to cyclin A and cyclin B degradation in nocodazole-arrested cells. Although the cells are able to degrade cyclin B, they do not exit mitosis as evidenced by high levels of cdk1 substrate phosphorylation and chromosome condensation. Our results demonstrate that a calyculin A and okadaic acid (1uM) sensitive phosphatase is required for SAC activation. Citation Format: Kristen Foss, Liguo Zhang, Alexander C. Robeson, Sally Kornbluth. The spindle assembly checkpoint requires phosphatase activity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3770. doi:10.1158/1538-7445.AM2015-3770