Cell Cycle Regulation of Glc7p Protein Phosphatase

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The orderly steps of eukaryotic cell division are regulated in part by timely protein degradation and reversible protein phosphorylation. Protein phosphatase-1 (PP1) enzymes are important for cell cycle dephosphorylation reactions. The budding yeast, Saccharomyces cerevisiae, offers the simplest known PP1 system because it has only a single gene for PP1, GLC7 (gene product is GlcTp). Glc7p dephosphorylates proteins critical for two transitions in the cell cycle of yeast. The first, G1 to S-phase is regulated by the nutritional status of the cells. The second transition, prior to anaphase, appears to use GlcTp to monitor the mitotic spindle assembly, which is necessary for faithful chromosome segregation. The proteins dephosphorylated by GlcTp, which control the cell cycle, are unknown.Recent results of this investigator show that Glc7p protein levels oscillate during the cell cycle. Peak levels of Glc7p were found in late G1 and S-phases. Conversely, low levels were detected during cell cycle intervals in which G1 or mitotic cyclins are destroyed by ubiquitin-dependent proteolysis. Therefore, they hypothesize that Glc7p is a target of ubiquitin-dependent proteolysis by the ubiquitin ligase complexes that destroy cyclins. The first aim of this project is to analyze potential ubiquitin-dependent Glc7p proteolysis. The second aim of this proposal defines the role of Glc8p, a noncatalytic subunit of Glc7p, in its regulation, to see if it shields Glc7p from ubiquitination and destruction during G1 to allow Glc7p to dephosphorylate substrate proteins . The high cyclin-dependent protein kinase (CDK) activity found in Glc7p-depleted cells suggests that S-phase or mitotic cyclins are not destroyed appropriately when Glc7p activity is diminished. This finding further implies the ubiquitin ligase known as the anaphase-promoting complex (APC) may be activated by Glc7p dephosphorylation. The last aim seeks to identify the responsible regulatory phosphoproteins that are Glc7p substrates and APC regulators.Although several phosphoproteins play important roles in cell division, the protein kinases and phosphatases that control their phosphorylation state are largely unknown. The control of cell division is a complicated process, which involves many participants. Preliminary analysis indicates that Glc7p protein phosphatase plays a part in this process. The goal of this project is to integrate Glc7p function with other known aspects of cell division.