UC San Diego

Maintenance of genome stability is critical for cell viability and survival. Consequently, DNA double strand breaks (DSBs) must be faithfully repaired in order to maintain the integrity of the genome. In S. cerevisiae, the homologous recombination (HR) pathways frequently repair DSBs using a homologous chromosome as a template. However, aberrant recombination with an improper homeologous template may lead to gross chromosomal rearrangements (GCRs). It is not known how divergent homologous sequences are selected as targets for non-allelic HR, or which pathways suppress the formation of such GCRs. To study this, we devised two new GCR assays in which sequences on chromosomes 14 and 15 target imperfect homologies in the rest of the genome. The homologies targeted by these assays differ in length, number of potential rearrangement targets, and percent homology to the original sequence. The formation of GCRs detected by these assays exhibit diverse genetic requirements; surprisingly, we found that many GCRs were not suppressed by mutations in the HR pathways. We characterized the spectrum of GCR structures observed in the assays and sequenced the translocation breakpoint junctions for the most prominent GCRs. We found that mutations in some genes in the heteroduplex rejection pathway, a process that suppresses recombination between imperfect templates, shifted the location of the translocation breakpoints towards regions of lower homology. Another study involving a genome-wide systematic screen of 4 different types of GCR assays involving different types of homology identified 182 new genes involved in genome instability as well as 483 cooperatively acting genes that suppress the genome instability caused by these genes. Analysis of TCGA data revealed that 93% of ovarian and 66% of colorectal cancers had defects affecting one or more of these genes. Finally, an exploration into the role of the essential TORC2 complex in maintenance of genome stability using the temperature sensitive tor2-21 allele revealed an increase in GCRs associated with a defect in Tor2 in two different GCR assays. This mutation caused a shift in the spectrum of GCR structures observed in an assay mediated by a single copy sequence but did not distort the spectrum of GCR structures observed in an assay whose GCRs were mediated by divergent homology. Together, these results suggest that the role of divergent homology in formation of GCRs is complex and the properties of the homology as well as its recombination targets influence the mechanisms and genetics of GCR formation.