The rate of spontaneous mutations in yeast deficient for MutSβ function

Abstract

Mutations in simple sequence repeat loci underlie many inherited disorders in humans, and are increasingly recognized as important determinants of natural phenotypic variation. In eukaryotes, mutations in these sequences are primarily repaired by the MutSβ mismatch repair complex. To better understand the role of this complex in mismatch repair and the determinants of simple sequence repeat mutation predisposition, we performed mutation accumulation in yeast strains with abrogated MutSβ function. We demonstrate that mutations in simple sequence repeat loci in the absence of mismatch repair are primarily deletions. We also show that mutations accumulate at drastically different rates in short (<8 bp) and longer repeat loci. These data lend support to a model in which the mismatch repair complex is responsible for repair primarily in longer simple sequence repeats.

Spontaneous single-nucleotide substitutions and microsatellite mutations have distinct distributions of fitness effects

“…8 msh3Δ MA strains had no identified mutation. Many of the msh3Δ MA strains with significant s values did not have any identified SSR mutations ( S4 Fig ), and there was no significant effect of the most severe impact type on mean s value (Kruskal-Wallis test p-value = 0.47); neither finding is unexpected considering the relatively low number of SSRs that we are able to call in these strains [38].…”

“…One likely source of variation in s values across strains is differences in the effect of these mutations on protein-coding genes. To test whether strains' s values were explained by the predicted severity of the substitutions found in these strains, we sequenced the haploid MA strains and identified SNMs and short indels in non-repetitive regions relative to the ancestral strain, as described in [38]. We identified a total of 307 SNMs and 3 indels across 68 strains.…”

“…This strain was then passaged independently in 151 replicates by streaking a single colony every 2 days for ~2000 generations on YPD media ('2000-generation' experiment) [41]. To generate MA lines deficient in SSR slippage repair, the MSH3 gene was deleted in a strain (MAT0.a1) derived from a spore of the diploid ancestor of the 2000-generation experiment, and the resulting strain was passaged independently in 36 replicates as above for ~200 generations [38]. Because respiring ade2 mutant colonies are red, petites could be detected by their white color and were not passaged to the next generation.…”

“…To call mutations, we followed the freebayes mutation-calling pipeline described in [38], but using the S288C sacCer3 genome assembly as a reference. Briefly, mutations were called as differences relative to the ancestral-strain derived MAT0.a1 sequence.…”

“…Petite proportion estimated for strains from [38] using plate-based colony color assay versus by modeling observed colony growth rates in the microscope assay as a mixture of Gaussians.…”