Objectives and content of the project Cell cycle progression in eukaryotes is controlled by the p34cdc2/CDC28 protein kinase and its short-lived, phase-specific regulatory subunits called cyclins. In budding yeast, Start is the major checkpoint that coordinates progress of the cell cycle engine with cell growth, nutrient availability and signals from other cells. Three redundant, unstable Gl cyclins (Clnl, Cln2 and Cln3), through their assosiation with Cdc28, allow budding yeast cells to pass Start. The aim of this project is to understand the molecular mechanisms involved in Gl cyclin degradation and their relationship to the cell cycle regulatory network using both genetic and biochemical approaches. We plan to: 1. screen for suppressors of the grrl mutant, in which Clnl is stabilized 2. test Gl cyclin stabilization in different ubc mutants 3. reconstitute active Cdc281Cln complexes for use in an in vitro degradation system Training content (objective, benefit and expected impact) The reconstitution of active Cdc28-Cln complexes will be extremely important in biochemically dissecting the Gl cyclin degradation pathway. In fact, this approach will allow us to test the direct involvement of genes, isolated by standard genetic techniques, in Gl cyclin proteolysis. The results obtained with this work can yield substancial information on the mechanism and biological role of Gl cyclin degradation in relation to the cell division cycle. Links with industry/ industrial relevance This is an industrially relevant proposal since the understanding of protein degradation (and in particular of G I cyclin degradation) might be crucial for the isolation of anti-tumor drugs. In fact it has been shown that cyclins (such as cyclin D I, a human G l cyclin) are directly involved in some human cancer. The host laboratory has ongoing collaboration, supported by a CEE Biomed2 grant, with the Ciba-Geigy (Basel-Swizerland) company.