Homologous recombination (HR) and non-homologous end joining (NHEJ) are conserved pathways for the repair of DNA double stranded breaks (DSBs) induced by ionizing radiation and genotoxic chemicals. HR utilizes a homologous template strand to repair damaged DNA in an error-free manner. NHEJ, which entails processing of the broken chromosome ends and their ligation, is often error-prone. HR is initiated by nucleolytic processing of the DNA break to yield 3' ssDNA tails for the recruitment of the repair machinery. Studies in the model eukaryote Saccharomyces cerevisiae have revealed multiple pathways of DNA break resection, two of which are responsible for long-range resection. DNA break processing is regulated during the cell cycle. Specifically, NHEJ proteins suppress it in G1, but this suppression is relieved during S and G2. In the K99 phase of this project, I will determine the mechanisms of polarity control and DNA end engagement during the end resection process. In the R00 phase, I will elucidate the functional crosstalk between the long-range resection paths that are dependent on Sgs1/Dna2 and Exo1 and will also examine how the restriction imposed by NHEJ proteins is overcome by DNA motor proteins in conjunction with the Mre11 complex. Given the remarkable degree of conservation of the DNA end resection mechanisms, the results from this project should provide a valuable experimental framework to guide similar endeavors in other eukaryotes, including humans.
DNA damaging agents induce genetic mutations and genome rearrangements. The proposed studies will delineate the mechanism by which DNA breaks resulting from exposure to radiation and genotoxic chemicals are nucleolytically processed in order to activate the DNA damage checkpoint and to prepare for damage repair by homologous recombination. The results from our research endeavors have direct relevance to understanding the relevance of DNA repair in health and disease.
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