Regulation of Protein Kinase Dbf2 in Mitotic Exit

Citation

Mah, Angie Siu Yee (2005) Regulation of Protein Kinase Dbf2 in Mitotic Exit. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/NS1Z-3K87. https://resolver.caltech.edu/CaltechETD:etd-05242005-130533

Abstract

Cyclin-dependent kinases (Cdk) direct cell cycle transitions by associating with various cyclins throughout the cell cycle. For cells to exit mitosis, mitotic Cdk activity must be turned off. In Saccharomyces cerevisiae, the mitotic exit network, or MEN, comprises of a group of proteins that form a signaling pathway required for mitotic exit. The MEN regulates the activity of Cdc14, the protein phosphatase critical for inactivating mitotic Cdk. Components of the MEN include the protein kinases Cdc15 and Dbf2, as well as the Dbf2-associated protein Mob1. We determined how these proteins are organized within the MEN by determining the molecular mechanism of Dbf2 activation. Dbf2 requires Mob1 association in order to be active and Cdc15 phosphorylates and thereby activates the Dbf2-Mob1 protein kinase complex. We also determined that the conserved phosphorylation sites of the NDR protein kinase family are required for Dbf2 kinase activity in vitro as well as for DBF2 function in vivo. It is unknown how Dbf2-Mob1 leads to Cdc14 release or how the protein kinase complex functions in cytokinesis. As a result, we sought to identify physiological substrates of Dbf2-Mob1 which would provide insight to Dbf2-Mob1 function in both of these significant cell cycle processes. There is no known physiological substrate for Dbf2-Mob1 we first identified RXXS as the motif that Dbf2-Mob1 preferentially phosphorylates. We then identified a number of in vitro substrates for Dbf2-Mob1, of which the majority contains the RXXS motif. The mechanism of Dbf2 activity has been shown to be conserved in a number of other NDR kinase family members, which have roles in morphogenesis and cell division, and have been implicated in tumorigenesis. Studies on Dbf2 will provide insight into cell cycle processes in budding yeast as well as in higher eukaryotes.

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