Jennifer Cobb, Professordepartment of Biochemistry and Microbiology

A double -strand break (DSB) is one of the most deleterious forms of DNA damage. In eukaryotic cells, two main repair pathways have evolved to repair DSBs, homologous recombination (HR) and non-homologous end-joining (NHEJ). HR is the predominant pathway of repair in the unicellular eukaryotic organism, S. cerevisiae . However, during replicative aging the relative use of HR and NHEJ shifts in favor of end-joining repair. By monitoring repair events in the HO-DSB system, we find that early in replicative aging, the recovery of long-range resection factors, Dna2-Sgs1 and Exo1 decreased which coincided with a decrease in DNA resection. Subsequently, as aging progressed, the recovery of Ku70 at DSBs decreased and the break site associated with the nuclear pore complex, which is the location where DSB repair occurs through alternative pathways that are more mutagenic. End-bridging remained intact as HR and NHEJ declined, but eventually became disrupted in cells at advanced replicative age. In all, our work provides insight about the molecular changes in DSB repair pathway usage during replicative aging. HR first declined, resulting in a transient increase in the frequency of NHEJ. However, with increased cellular divisions, NHEJ subsequently declined, which coincided with the decreased recovery of Ku at DSBs. There was a high frequency of repair products with genomic deletions and microhomologies at the break junction in wild type cells of advanced replicative age, events not observed in young cells repairing primarily by HR.