Chemical Genetic Studies of Mitosis

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DESCRIPTION (provided by applicant): The goal of our research program is to understand how exit from mitosis is regulated in vertebrate cells and how this process is disrupted in cancer cells. Progression through the eukaryotic cell cycle is controlled by carefully timed protein phosphorylation and protein degradation. Anaphase and mitotic exit depend on the ubiquitin-dependent degradation of regulators such as securins and mitotic cyclins. These proteins are targeted for ubiquitination by a multisubunit ubiquitin ligase called the Anaphase-Promoting Complex or Cyclosome (APC). Our goal is to study how APC is controlled in vertebrate cells by finding chemicals that affect the regulation or activity of this complex. We have identified several compounds that stabilize cyclin B in Xenopus egg extracts and block exit from mitosis through a novel mechanism. Our goal is to identify the protein targets of these inhibitors using a combination of biochemical characterization and affinity-based techniques. We have also identified a compound that directly inhibits the activity of the APC in vitro, and we will use this compound to study APC regulation in cells. One of the mitotic inhibitors discovered in our Xenopus cell cycle screen paradoxically increases the rate at which cells exit mitosis in the presence of taxol. We will use this inhibitor to study the mechanism by which cells undergo this adaptation process. Understanding how exit from mitosis and adaptation is regulated will provide important insights for understanding how cancer cells respond to taxol treatment.