Saccharomyces cerevisiae Raw sequence reads. Saccharomyces cerevisiae

Acetic acid tolerance of Saccharomyces cerevisiae is an important trait in sourdough fermentation processes, where the accumulation of acid by the growth of lactic acid bacteria reduces the yeast metabolic activity. In this work, we have carried out adaptive laboratory evolution (ALE) experiments in two sourdough isolates of S. cerevisiae exposed to acetic acid, or alternatively to acetic acid and myriocin, an inhibitor of sphingolipid biosynthesis that sped-up the evolutionary adaptation. Evolution approaches resulted in acetic tolerance, and surprisingly, increased lactic susceptibility. Four evolved clones, one from each parental strain and evolutionary scheme, were selected based on their potential for CO2 production in sourdough conditions. Among them, two showed phenotypic instability characterized by strong lactic sensitivity after several rounds of growth under unstressed conditions, while two others, displayed increased constitutive acetic tolerance with no loss of growth in lactic medium. Genome sequencing and ploidy level analysis of all strains revealed aneuploidies, which could account for phenotypic heterogeneity. In addition, copy number variations (CNVs), affecting specially to genes involved in ion transport or flocculation, and single nucleotide polymorphisms (SNPs) were identified. Mutations in several genes, ARG82, KEX1, CTK1, SPT20, IRA2, ASG1 or GIS4, were confirmed as involved in acetic and/or lactic tolerance, and new determinants of these phenotypes, MSN5 and PSP2, identified.

酿酒酵母（Saccharomyces cerevisiae）的乙酸耐受性是酸面团（sourdough）发酵过程中的重要性状——乳酸菌增殖所积累的有机酸会抑制酵母的代谢活性。本研究以两株分离自酸面团的酿酒酵母为亲本菌株，分别设置乙酸胁迫、乙酸+多球壳菌素（myriocin）两种胁迫体系开展适应性实验室进化（adaptive laboratory evolution, ALE）实验，其中多球壳菌素是一种鞘脂生物合成抑制剂，可显著加速进化适应进程。进化实验成功获得了乙酸耐受菌株，但意外的是这些菌株对乳酸的敏感性均有所升高。本研究依据菌株在酸面团环境下的二氧化碳（CO₂）产生潜力，从每种亲本菌株和每种进化方案的进化群体中各筛选1株，共获得4株进化克隆。其中2株菌株表现出表型不稳定性：在无胁迫条件下经过多轮传代培养后，其对乳酸的敏感性显著增强；而另外2株则呈现组成型乙酸耐受特性，在乳酸培养基中的生长未受任何影响。对所有菌株进行基因组测序与倍性水平分析后发现，非整倍性变异是导致表型异质性的重要原因。此外，本研究还鉴定出大量拷贝数变异（copy number variations, CNVs）与单核苷酸多态性（single nucleotide polymorphisms, SNPs），其中拷贝数变异主要靶向影响离子转运或絮凝相关基因。经验证，ARG82、KEX1、CTK1、SPT20、IRA2、ASG1及GIS4等多个基因的突变与乙酸和/或乳酸耐受性密切相关；同时本研究还发现了MSN5和PSP2这两个调控该类性状的全新决定因子。