Studies in the model eukaryote Saccharomyces cerevisiae have revealed that homologous recombination provides a major mechanism for eliminating DNA double- stranded breaks (DSBs) induced by ionizing radiation or are associated with injured DNA replication forks. During the repair process, the ends of the DNA breaks are processed nucleolytically to yield 3'ssDNA tails, which are bound by recombination factors. The nucleoprotein complex thus formed then conducts a search to locate an undamaged DNA homologue, and catalyzes the formation of a DNA joint, called D-loop, with the homologue. Resolution of the D-loop can proceed via at least three mechanistically distinct pathways, two of which generate only non-crossover recombinants and are therefore more adept at genome preservation, with the remaining pathway able to produce crossovers frequently. Proteins encoded by evolutionarily conserved genes of the RAD52 epistasis group catalyze the HR reaction. Our studies have provided insights into the mechanistic underpinnings of the HR machinery that harbors proteins of this gene group. In this renewal project, a combination of biochemical, genetic, and other cell-based approaches will be employed to (i) define the mechanism of action of the DNA motor-driven path of DSB end resection, and (ii) delineate the roles of two novel protein complexes in subsequent stages of the HR reaction. Knowing that the structure and function of the RAD52 group of genes and proteins have been conserved highly, the results from our endeavors will allow us to formulate detailed models to elucidate HR mechanisms in other eukaryotes, including humans. Given the importance of HR-mediated chromosome damage repair in tumor suppression, our work also has direct relevance to cancer biology.
Cancers are characterized by genomic instability and rearrangements, which can be induced by exposure of cells to DNA damaging agents such as ionizing radiation or are associated with the stalling or collapse of DNA replication forks. The proposed studies will continue to delineate the mechanism by which cells avoid deleterious genome rearrangements via the process of homologous recombination. The results from our research endeavors have direct relevance to understanding the role of homologous recombination in cancer suppression.
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