The Role of RAD51 in de novo Telomere Addition
DNA double-stranded breaks (DSBs) are detrimental to genomic stability and can be repaired by multiple mechanisms, including Homology Dependent Repair (HDR) and Non-Homologous End Joining (NHEJ). In rare cases, telomerase can act on a DSB and add a telomere in a process known as de novo telomere addition. Since telomeres primarily protect DNA ends from nucleolytic processing, de novo telomere addition can restore genomic stability, but will also lead to loss of genetic information distal to the DSB. The Saccharomyces cerevisiae genome contains ‘hotspots’ of de novo telomere addition (termed Sights of Repair-Associated Telomere Addition or SiRTAs) at which telomerase acts at an unusually high frequency following a DSB. We seek to understand why telomere addition is favored at SiRTAs and whether such events are actively promoted as a mechanism to survive a persistent DSB. Deletion of RAD51 causes a decrease in the rate of de novo telomere addition. Since Rad51 function requires Rad52 during HDR, it was hypothesized that Rad51 is also dependent on Rad52 during de novo telomere addition. RAD52 deletion mutants surprisingly do not show a decrease in rate, suggesting that RAD51 and RAD52 are independent of each other during the process. To define which aspects of Rad51 function contribute to de novo telomere addition, several separation-of-function alleles will be introduced in RAD51 deletion strains. Telomere addition at a SiRTA on Chromosome V will be monitored following induction of a single DSB and we expect to see differences in telomere addition rates between the alleles.