The proposed research aims at identifying mitotic targets of the p34cdc2/CDC28 protein kinase. The genetic approach intended to be used will make use of the budding yeast Saccharomyces cerevisiae as the most tractable experimental system. In particular we will take advantage of the M-phase specific defect of the cdc28-1N allele to screen for mutations that enhance the thermosensitive phenotype conferred by this allele. The rationale behind this approach is that in a context where the M-phase activity of cdc28 kinase is drastically compromised by growing the parental cdc28-IN strain at its semi-permissive temperature, novel mutations that affect the abundance or activity of downstream components must yield a more severe growth defect phenotype, thereby allowing a specific screen for such mutations to be carried out. The project will consist of the isolation and genetic analysis of such mutations, followed by the molecular characterization of the corresponding gene products. Because the cell cycle control machinery has been highly conserved through evolution that, for example, the human homologue of the cdc28 gene can function and be properly regulated in yeast cells, it can reasonably be anticipated that among the various cdc28 mitotic targets this study will hopefully identify, at least some of them might have a structural and functional equivalent in mammalian cells. This study should therefore contribute to a better understanding of both the cell cycle control mechanism in human cells and their disruption during the oncogenic process.
