Adaptive evolution of yeast under heat stress and genetic reconstruction to generate thermotolerant yeast

Abstract

Saccharomyces cerevisiae is a microorganism that is widely used for the bioproduction of useful substances, including ethanol. However, during bioproduction, yeast cells are subjected to various stresses. In particular, bioproduction via fermentation generates heat, leading to decreased rates of cell growth and fermentation. Improving the thermotolerance of yeast can help to maintain metabolic activity and bioproduction efficiency at high temperatures. In this chapter, we describe the improvement of thermotolerance in yeast through stepwise adaptive evolution under heat stress. The adaptation strategy, in which the cells were selected under two selective pressures (heat stress and growth rate), improved thermotolerance while maintaining growth rate. Through this adaptation strategy, a thermotolerant yeast strain, YK60-1, was successfully isolated after adaptation to 38 °C. Transcriptome and nontargeted metabolome analyses revealed that YK60-1 induced stress-responsive genes and accumulated more trehalose than the wild-type parent strain. Furthermore, comparative genomic analysis of the intermediate populations after adaptation to each elevated temperature revealed key mutations for improving thermotolerance in the CDC25 gene. A thermotolerant yeast strain was also reconstructed by introducing CDC25 mutations in the wild-type strain. CDC25 mutation is thought to alter global transcriptional regulation through downregulation of the cAMP/PKA pathway, leading to improved stress tolerance.