|Ph.D Student||Yudkovsky Yana|
|Subject||The Mechanisms of the Regulation of Cyclosome Activity|
in Somatic Human Cells
|Department||Department of Medicine||Supervisor||? 18? Avram Hershko|
The cyclosome/Anaphase Promoting Complex (APC) is a multi-subunit ubiquitin ligase, which targets for degradation specific cell cycle regulatory proteins, such as mitotic cyclins and securin, an inhibitor of anaphase initiation. It is activated at the end of mitosis by phosphorylation and association with the WD-40 protein Cdc20/Fizzy and is kept active in the G1 phase by association with Cdh1/Fizzy-related. The mitotic checkpoint system that keeps cells with defective spindles from leaving mitosis interacts with Cdc20 and prevents its stimulatory action on the cyclosome/APC. The aim of this study was to examine the mode of the regulation of the cyclosome/APC in somatic mammalian cells.
It was found that, in contrast to Cdh1, Cdc20 specifically activated cyclosome from M-phase cells and that the phosphorylation of mammalian cyclosome was required for this activation. It was furthermore observed that activity of Cdc20 is negatively regulated by Cdk-mediated phosphorylation, and this may provide a safeguard, in addition to the mitotic checkpoint system to prevent the premature or inappropriate activation of the cyclosome/APC in early mitosis. This notion was supported by the observation that a non-phosphorylatable derivative of Cdc20 stimulated cyclin-ubiquitin ligation in extracts from nocodazole-arrested cells.
Next, an attempt was made to gain some insight into the molecular mechanisms of the cyclosome/APC regulation by the checkpoint proteins Mad2 and BubR1. It was found that combination of low concentrations of Mad2 and BubR1 specifically inhibited the activity of the mitotic (phosphorylated) form of the cyclosome-Cdc20 in an additive mode.