The regulation of meiotic recombination by DNA damage response proteins

Homologous recombination (HR) is a vital aspect of meiotic cell division, required for the correct segregation of chromosomes, and generation of genetic diversity. DNA double strand breaks (DSBs) are formed by Spo11 and repaired by HR to produce crossovers (COs) between homologous chromosomes. DSBs and COs are subject to interference to ensure even spacing of recombination events. The DNA damage response checkpoint, mediated by Mec1 and Tel1, is partly responsible for the regulation of HR and its outcomes. To investigate HR outcomes, hybrid budding yeast containing ~65,000 variants were induced to undergo meiosis and the resulting spores were sequenced. Recombination events were categorized via comparison to the parental genomes, using a custom-designed bioinformatic pipeline. Changes in recombination event type and distribution were examined in Mec1 and Tel1 mutants, revealing that these proteins are required to limit Spo11-DSB formation, reduce DSB clustering and maintain DSB interference, and to promote CO formation and mediate CO interference, likely by altering the ratio of interfering to non-interfering CO formation. An activator of Mec1, Rad24, is shown to have a meiotic recombination phenotype similar to that of the Mec1 mutant, but displays an increase in event length, and a reduction in CO interference and CO formation. The roles of Mec1, Tel1 and Rad24 are further examined by testing genetic interaction with factors that influence the outcome of meiotic recombination. Separable roles of Mec1 and Rad24 are also identified by examining the different effects of prophase extension on these strains. During these analyses, it is determined that the mismatch repair protein Msh2 has opposing roles in both improving and reducing spore survival in hybrids, and in the promotion of interfering CO formation. Collectively, this study reveals the consequences of the loss of important DNA damage sensor proteins on the meiotic genome, at a high resolution conferred by the use of deep-sequencing and bioinformatic tools.