Translesion Polymerases Drive Microhomology-Mediated Break-Induced Replication Leading to Complex Chromosomal Rearrangements

Complex genomic rearrangements (CGRs) are a hallmark of many human diseases. Recently, CGRs were suggested to result from microhomology-mediated break-induced replication (MMBIR), a replicative mechanism involving template switching at positions of microhomology. Currently, the cause of MMBIR and the proteins mediating this process remain unknown. Here, we demonstrate in yeast that a collapse of homology-driven break-induced replication (BIR) caused by defective repair DNA synthesis in the absence of Pif1 helicase leads to template switches involving 0–6 nt of homology, followed by resolution of recombination intermediates into chromosomal rearrangements. Importantly, we show that these microhomology-mediated template switches, indicative of MMBIR, are driven by translesion synthesis (TLS) polymerases Polζ and Rev1. Thus, an interruption of BIR involving fully homologous chromosomes in yeast triggers a switch to MMBIR catalyzed by TLS polymerases. Overall, our study provides important mechanistic insights into the initiation of MMBIR associated with genomic rearrangements, similar to those promoting diseases in humans. [Display omitted] •Collapse of DNA repair synthesis triggers a switch from BIR to MMBIR•Transition from BIR to MMBIR leads to chromosomal rearrangements•MMBIR is driven by translesion synthesis polymerases Polζ and Rev1•Template switches associated with MMBIR involve 0 to 6 nt of homology Microhomology-mediated break-induced replication (MMBIR) is a poorly characterized molecular mechanism leading to the formation of complex chromosomal rearrangements associated with human diseases. Here, Sakofsky and colleagues demonstrate in yeast that MMBIR is initiated by the collapse of homology-mediated BIR and is driven by translesion synthesis polymerases Polζ and Rev1.