Examining the potential role of DNA polymerases η and ζ in triplet repeat instability in yeast

Abstract

Triplet repeats undergo frequent mutations in human families afflicted with certain neurodegenerative diseases and also in model organisms. Although the molecular mechanisms of triplet repeat instability are still being identified, it is likely that aberrant DNA synthesis plays an important role. Many DNA polymerases stall at triplet repeat sequences, probably due to the adoption of unusual DNA secondary structures. One possible mechanism to explain triplet repeat contractions is that a triplet repeat hairpin on the template strand inhibits replicative polymerases and that one or more bypass polymerases are recruited for synthesis past the hairpin. If the translesion synthesis is mutagenic, contractions can be generated. To address this possibility, Saccharomyces cerevisiae strains lacking either pol ζ ( rev7), pol η ( rad30), or both were tested for trinucleotide repeat (TNR) contractions using three separate, sensitive genetic assays. If these bypass polymerases are important for mutagenesis, then the mutants should show a reduction in the contraction rate. Two genetic tests for triplet repeat contractions showed no significant change for the mutants compared to wild type. A third assay showed a five-fold reduction in contraction rates due to pol η ablation. Despite this modest decrease, the overall contraction rate was still high, indicating that many deletions still occur in the absence of both polymerases. Expansion rates were also unaffected in the mutant strains. These results indicate that, in yeast, pol η and pol ζ most likely have little role in triplet repeat mutagenesis.