The long term goal of this project is to elucidate the molecular basis of cell cycle regulation in eukaryotes. To achieve this goal, we have chosen to study the regulation of cell division in budding yeast, S. cerevisiae, a simple eukaryotic system eukaryotic system that is tractable to genetics, as well as cellular and molecular biology. This proposal is directed specifically toward understanding the role of cyclin-like proteins in yeast. Cyclins are conserved among eukaryotes and are thought to be rate- limiting positive elements of cell cycle control. In yeast, the CLN genes encode a family of cyclin-like proteins. We have three major aims. We will characterize the CLN gene family, first, by identifying and isolating additional members through the use of PCR or by screening of yeast genomic libraries. These genes, as well as the two CLN genes that have already been isolated, will then be subjected to mutational analysis. This will establish whether the CLN genes are essential for growth as well as providing reagents for analysis of the interaction between the CLN gene products and the Cdc28 protein kinase.
Our second aim i s to characterize the regulation of the CLN proteins during the cell cycle. Immunological and biochemical methods will be used to determine whether the abundance or phosphorylation state of these proteins is periodic during the cell cycle. Periodic production and degradation is the hallmark of animal cell cyclins. Knowledge of these parameters will dictate the design of experiments to achieve the third and primary aim of this proposal, to elucidate the involvement of these cyclin-like proteins in the regulation of cell cycle initiation in yeast. To achieve this aim we will test the hypothesis that the CLN proteins to interact with and regulate the activity of the Cdc28 protein kinase to maturation promoting factor (MPF), a proposed target of cyclins. Furthermore, like animal cell cyclins, the activity of the Cdc28 protein kinase is periodic during the cell cycle. These studies will provide a molecular description of cyclin function in yeast. This will substantially advance the understanding of cell division control in yeast as well as offering a basis for similar investigations with multicellular eukaryotes.
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