Proteolytic Control of Early Events in Mitosis

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DESCRIPTION (provided by applicant): The timing of events in mitosis is critical to ensure accurate chromosome segregation and genomic stability. Critical to these timing events is the Anaphase Promoting Complex, an E3 ubiquitin ligase that directs the ordered destruction of cyclin A, the chromosome segregation regulator Securin, and cyclin B. The known controllers of APC activity are the components of the spindle assembly checkpoint and the zinc binding protein Emi1. Emi1 functions to restrain the APC in S and G, thereby allowing the accumulation of cyclins. Emi1 is transcriptionally activated in G1 by the cyclin D/Rb/E2F pathway and destroyed in early mitosis following phosphorylation specific binding of the SCFI3TrCP ubiquitin ligase. One aim of this grant is to identify the critical events triggering Emi1 destruction including the kinases that trigger Emi 1 destruction. We have also begun to characterize a large number of Emi1 interacting proteins (kIPs) to better understand the network of regulation controlling the APC. Here, we find the protein encoded by the Evi5 oncogene interacts with Emi1 and functions upstream of Emi1 to direct the accumulation of both Emi1 and cyclin A. Evi5 is a frequent site of proviral insertion in mouse T-cell lymphomas and is mutated in human neuroblastoma. Additionally, it is highly expressed in a variety of tumors. The Evi5 protein is predicted to be a GTPase activating protein, but the GTPase remains unknown. We find that Evi5 is required for accumulation of Emi1 and cyclin A in early G1 and new evidence suggests that it may participate in steps regulating the post-ubiquitination delivery of ubiquitinated proteins to the proteasome. Thus, Evi5 may promote oncogenesis by a previously unknown mechanism. Our aims here are to (1) define factors controlling Evi5 function at G1-S and in mitosis; (2) to identify the mechanism for Evi5 control of proteolysis; (3) to identify the critical small GTPase regulated by Evi5; and (4) to define the factors controlling Emi1 destruction in mitosis.