Double-Strand Break Repair in Yeast Requires Both Leading and Lagging Strand DNA Polymerases

Abstract

Mitotic double-strand break (DSB)-induced gene conversion at MAT in Saccharomyces cerevisiae was analyzed molecularly in mutant strains thermosensitive for essential replication factors. The processivity cofactors PCNA and RFC are essential even to synthesize as little as 30 nucleotides following strand invasion. Both PCNA-associated DNA polymerases δ and ε are important for gene conversion, though a temperature-sensitive Pol ε mutant is more severe than one in Pol δ. Surprisingly, mutants of lagging strand replication, DNA polymerase α (pol1-17), DNA primase (pri2-1), and Rad27p (rad27 Δ) also greatly inhibit completion of DSB repair, even in G1-arrested cells. We propose a novel model for DSB-induced gene conversion in which a strand invasion creates a modified replication fork, involving leading and lagging strand synthesis from the donor template. Replication is terminated by capture of the second end of the DSB.