Genetic analysis of phenotypic differences between species is typically limited to interfertile species. Here, we conducted a genome-wide noncomplementation screen to identify genes that contribute to a major difference in thermal growth profile between two reproductively isolated yeast species,Saccharomyces cerevisiaeandSaccharomyces uvarum. The screen identified only a single nuclear-encoded gene with a moderate effect on heat tolerance, but, in contrast, revealed a large effect of mitochondrial DNA (mitotype) on both heat and cold tolerance. Recombinant mitotypes indicate that multiple genes contribute to thermal divergence, and we show that protein divergence inCOX1affects both heat and cold tolerance. Our results point to the yeast mitochondrial genome as an evolutionary hotspot for thermal divergence.

Mitochondrial genome origin affects the temperature tolerance of synthetic and industrial lager-brewing yeast hybrids.

Gene regulation is a ubiquitous mechanism by which organisms respond to their environment. While organisms are often found to be adapted to the environments they experience, the role of gene regulation in environmental adaptation is not often known. In this study, we examine divergence in cis-regulatory effects between two Saccharomycesspecies, S. cerevisiaeand S. uvarum, that have substantially diverged in their thermal growth profile. We measured allele specific expression (ASE) in the species’ hybrid at three temperatures, the highest of which is lethal to S. uvarumbut not the hybrid or S. cerevisiae. We find that S. uvarumalleles can be expressed at the same level as S. cerevisiaealleles at high temperature and most cis-acting differences in gene expression are not dependent on temperature. While a small set of 136 genes show temperature-dependent ASE, we find no indication that signatures of directional cis-regulatory evolution are associated with temperature. Within promoter regions we find binding sites enriched upstream of temperature responsive genes, but only weak correlations between binding site and expression divergence. Our results indicate that temperature divergence between S. cerevisiaeand S. uvarumhas not caused widespread divergence in cis-regulatory activity, but point to a small subset of genes where the species’ alleles show differences in magnitude or opposite responses to temperature. The difficulty of explaining divergence in cis-regulatory sequences with models of transcription factor binding sites and nucleosome positioning highlights the importance of identifying mutations that underlie cis-regulatory divergence between species.

A growing body of research suggests that the mitochondrial genome (mtDNA) is important for temperature adaptation. In the yeast genus Saccharomyces, species have diverged in temperature tolerance, driving their use in high or low temperature fermentations. Here we experimentally test the role of mtDNA in temperature tolerance in synthetic and industrial hybrids (Saccharomyces cerevisiae x Saccharomyces eubayanus, or Saccharomyces pastorianus), which cold-brew lager beer. We find that the relative temperature tolerances of hybrids correspond to the parent donating mtDNA, allowing us to modulate lager strain temperature preferences. The strong influence of mitotype on the temperature tolerance of otherwise identical hybrid strains provides support for the mitochondrial climactic adaptation 2 hypothesis in yeasts and demonstrates how mitotype has influenced the world's most commonly fermented beverage.One Sentence Summary: Mitochondrial genome origin affects the temperature tolerance of synthetic and industrial lager-brewing yeast hybrids.