Endless Pursuit of DNA Double-Strand Break Ends

In the pursuit of radiation-induced cellular damage that manifests in cell killing and genome instability, but is reversible by the phenomenon of liquid holding, DNA double-strand breaks (DSBs) have been shown to be the major culprit. It is now widely accepted that DSBs provide both beneficial and deleterious effects and that elaborate systems were evolved to address their detection, processing and repair. Understanding the mechanisms of DSB induction, processing and eventual biological consequences as well as just the detection of DSBs remain a challenge - especially for randomly induced breaks. Given the large number of extensive reviews available and the limited space here, I have chosen to focus on summarizing recent novel findings from our group using the budding yeast Saccharomyces cerevisiae that are relevant to many molecular and genetic aspects of DSB repair and impact on genome stability. Included are the systems we have developed that address (1) induction of random primary and secondary DSBs; (2) processing of DSB ends; (3) genetic control of DSB induction and repair; (4) genome instabilities associated with DSBs including rearrangements and hypermutability associated with resected ends; and (5) physical factors that determine the transition from DSB to chromosome break or recombination. Using a circular chromosome, we find that resection of random, dirty-end DSBs induced by ionizing radiation or derived from MMS single-strand damage is rapid and is primarily due to the MRX complex. Interestingly, the transition from DSB to chromosome break at a unique DSB in yeast is largely prevented by the nuclease function of exonuclease 1 as determined from separation of fluorescent markers that flank a DSB. Based on a tetraploid gene-dosage model, the role of the chromosome structural complex cohesin is not only to enhance DSB repair between sister chromatids, but it also directs recombinational repair events to sisters thereby preventing loss of heterozygosity. Notably, DSBs greatly sensitizes cells to localized mutability as well as gross rearrangements. Overall, these findings demonstrate the genomic vulnerability to DSBs and the genetic investment in their orderly processing.