Roles for Mismatch Repair Proteins in Maintaining Genome Stability

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Project Summary DNA mismatch repair (MMR) systems act to excise misincorporation errors that occur during DNA replication. In eukaryotes MSH proteins recognize these errors in the context of base-base and insertion/deletion mismatches and recruit MLH complexes to form ternary complexes that work with replication factors and Exo1 to excise the newly replicated DNA strand through the mismatch site. This is followed by DNA re-synthesis steps. MMR factors also recognize mismatches that form during strand invasion steps in homologous recombination; they recruit a helicase complex that unwinds (rejects) recombination intermediates and allows a new homology search. Sydney Rosen's summer research is aimed at understanding the decision steps that take place during homologous recombination when a double-strand break occurring in a DNA sequence is repaired using a donor sequence containing low levels of sequence divergence (Project 3 in parent grant R35GM134872). More specifically, she is interested in understanding how the kinetics and subnuclear location(s) of homologous recombination in S. cerevisiae compare when an identical donor sequence is used as a template for repair versus a divergent donor template. She plans to track and measure the timing of repair of an induced DSB via a fluorescence-marked protein involved in homologous recombination (Rad52) as well as nuclear pore components. Her hypothesis is that recombination between divergent sequences will result in slower repair and possibly differences in repair location. Results from this time course will provide a better understanding of how the rejection decision is facilitated, and how chromosomal rearrangements can lead to disease. Sydney has expressed strong interest in entering a Ph.D. program in molecular biology after graduating from Cornell. She will become experienced transforming baker's yeast and analyzing transformants using western blot analysis and fluorescence and confocal microscopy. She will also learn the appropriate statistical analyses for each of her experiments. Her mentors will meet with her daily to discuss her research progress and plan future experiments. She is already an active participant in group meeting and journal club, presenting research summaries and formal presentations. Over the summer Sydney will join our department's summer Research Experience for Undergraduates (REU) program where she will gain experience presenting her work in written and oral formats.