Characterization of a cyclin-dependent kinase inhibitor during meiosis in yeast

Meiosis is a conserved, specialized developmental process which results in the production of four haploid gametes. Meiosis differs from mitosis in three main processes: DNA replication, recombination and reductional divisions. Defects in meiosis can result in birth defects, failed fetal development, and the origin of germ cell malignancies. Due to complications in studying meiosis in mammalian systems, specific molecular mechanisms in meiosis are not well understood. In these studies we aimed to examine the specialized mechanisms of meiosis in Saccharomyces cerevisiae, specifically those of the meiotic protein kinase Ime2 and the cyclin dependent kinase (Cdk1) inhibitor Sic1. In order to elucidate the mechanism of Ime2 dependent activity during meiosis, we examined Ime2 kinase activity. Ime2 is a meiosis specific protein kinase that is required for entry into and progression through meiosis. Ime2 is proposed to functionally replace the G1 Cdk1 complexes in meiosis to promote Sic1 destruction. Based on our previous work, we hypothesized that Ime2 is not a 78 functional meiotic replacement for the G1 Cdk1 complexes. Through a series of both in vitro and in vivo studies, we found that Ime2 does not simply replace the G1 Cdk1 complexes in promoting meiotic Sic1 degradation. Separate, yet interrelated, studies were begun to further investigate the meiotic mechanism of Sic1 destruction by determining the effect of altering Sic1's phospho-acceptor sites. We found that overexpression of a mutated Sic1 protein leads to a DNA re-replication phenotype with severely disturbed meiotic progression. We found that DNA re-replication was blocked in cells with recombination defects, but occurred in cells blocked from prophase exit through the recombination checkpoint. Our studies have revealed a new branch of the meiotic recombination checkpoint that targets the machinery controlling initiation of DNA replication. These results have exposed differences in the regulatory mechanisms for preventing DNA re-replication between those in meiosis and those in mitosis. Gaining a better understanding of the mechanisms driving meiotic progression in yeast will lead to a better understanding of cell cycle and meiotic progression in humans, allowing for useful insight into diseases such as cancer that act upon these cycles in detrimental ways.