Our long-term goal is to understand the mechanisms of meiotic and mitotic recombination. A knowledge of the mechanisms by which cells create double-stranded DNA breaks (the DNA lesions that initiate recombination) and repair these breaks is essential for understanding the genomic instability associated with cancer. Although most of our proposed experiments will be done with the yeast Saccharomyces cerevisiae, we will examine DNA mismatch repair (MMR) in both yeast and C. elegans. The first Specific Aim is to examine the properties of DNA sequence and chromatin structure that affect the frequency of meiotic recombination. For most of these experiments, we will use DNA microarrays to monitor meiotic recombination activity at every locus in the genome. The second Specific Aim is to study the mechanism of reciprocal mitotic recombination. Using a system that allows selection of reciprocal mitotic crossovers, we will determine whether there are hotspots for mitotic exchanges. We will also investigate the genetic regulation of mitotic recombination using mutations that affect various pathways of recombination and DNA repair. Our third Specific Aim is to examine various aspects of DNA mismatch repair in yeast and in C. elegans. We will analyze the roles of the yeast MMR proteins Msh3p, Msh6p, MIh2p, and MIh3p, as well as the possible involvement of the DNA replication proteins DNA polymerase delta and PCNA, in the repair of mismatches generated during meiotic recombination. The proposed C. elegans studies will concentrate on determining phenotypes associated with msh-2 and msh-6 mutant worms, and on the isolation of new MMR worm mutants.
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