Mismatch recognition and subsequent processing have distinct effects on mitotic recombination intermediates and outcomes in yeast

Abstract

The post-replicative mismatch repair (MMR) system has anti-recombination activity that limits interactions between diverged sequences by recognizing mismatches in strand-exchange intermediates. In contrast to their equivalent roles during replication-error repair, mismatch recognition is more important for anti-recombination than subsequent mismatch processing. To obtain insight into this difference, ectopic substrates with 2% sequence divergence were used to examine mitotic recombination outcome (crossover or noncrossover; CO and NCO, respectively) and to infer molecular intermediates formed during double-strand break repair in Saccharomyces cerevisiae. Experiments were performed in an MMR-proficient strain, a strain with compromised mismatch-recognition activity (msh6Δ) and a strain that retained mismatch-recognition activity but was unable to process mismatches (mlh1Δ). While the loss of either mismatch binding or processing elevated the NCO frequency to a similar extent, CO events increased only when mismatch binding was compromised. The molecular features of NCOs, however, were altered in fundamentally different ways depending on whether mismatch binding or processing was eliminated. These data suggest a model in which mismatch recognition reverses strand-exchange intermediates prior to the initiation of end extension, while subsequent mismatch processing that is linked to end extension specifically destroys NCO intermediates that contain conflicting strand-discrimination signals for mismatch removal.