Double-strand break repair in yeast requires both leading and lagging strand DNA polymerases

Mitotic double-strand break (DSB)-induced gene conversion at MAT in Sacchar omyces cerevisiae was analyzed molecularly in mutant strains thermosensitiv e for essential replication factors. The processivity cofactors PCNA and RF C are essential even to synthesize as little as 30 nucleotides following st rand invasion. Both PCNA-associated DNA polymerases delta and epsilon are i mportant for gene conversion, though a temperature-sensitive Pol epsilon mu tant is more severe than one in Pol delta. Surprisingly, mutants of lagging strand replication, DNA polymerase alpha (pol1-17), DNA primase (pri2-1), and Rad27p (rad27 Delta) also greatly inhibit completion of DSB repair, eve n in G1-arrested cells. We propose a novel model for DSB-induced gene conve rsion in which a strand invasion creates a modified replication fork, invol ving leading and lagging strand synthesis from the donor template. Replicat ion is terminated by capture of the second end of the DSB.