Table_5_Differential Gene Expression and Allele Frequency Changes Favour Adaptation of a Heterogeneous Yeast Population to Nitrogen-Limited Fermentations.XLSX

Alcoholic fermentation is fundamentally an adaptation process, in which the yeast Saccharomyces cerevisiae outperforms its competitors and takes over the fermentation process itself. Although wine yeast strains appear to be adapted to the stressful conditions of alcoholic fermentation, nitrogen limitations in grape must cause stuck or slow fermentations, generating significant economic losses for the wine industry. One way to discover the genetic bases that promote yeast adaptation to nitrogen-deficient environments are selection experiments, where a yeast population undergoes selection under conditions of nitrogen restriction for a number of generations, to then identify by sequencing the molecular characteristics that promote this adaptation. In this work, we carried out selection experiments in bioreactors imitating wine fermentation under nitrogen-limited fermentation conditions (SM60), using the heterogeneous SGRP-4X yeast population, to then sequence the transcriptome and the genome of the population at different time points of the selection process. The transcriptomic results showed an overexpression of genes from the NA strain (North American/YPS128), a wild, non-domesticated isolate. In addition, genome sequencing and allele frequency results allowed several QTLs to be mapped for adaptation to nitrogen-limited fermentation. Finally, we validated the ECM38 allele of NA strain as responsible for higher growth efficiency under nitrogen-limited conditions. Taken together, our results revealed a complex pattern of molecular signatures favouring adaptation of the yeast population to nitrogen-limited fermentations, including differential gene expression, allele frequency changes and loss of the mitochondrial genome. Finally, the results suggest that wild alleles from a non-domesticated isolate (NA) may have a relevant role in the adaptation to the assayed fermentation conditions, with the consequent potential of these alleles for the genetic improvement of wine yeast strains.

酒精发酵本质上是一种适应性过程，酿酒酵母（Saccharomyces cerevisiae）在此过程中优于其竞争者并主导发酵进程。尽管葡萄酒酵母菌株似乎已适应酒精发酵的胁迫环境，但葡萄汁中的氮限制会导致发酵停滞或发酵迟缓，给葡萄酒产业造成显著经济损失。解析酵母适应低氮环境的遗传基础的方法之一为选择实验：将酵母种群置于氮限制条件下历经多代选择，随后通过测序鉴定介导该适应性的分子特征。本研究采用异质性SGRP-4X酵母种群，在模拟葡萄酒发酵的氮限制发酵条件（SM60）下的生物反应器中开展选择实验，并在选择过程的不同时间点对种群的转录组与基因组进行测序。转录组分析结果显示，野生非驯化分离株NA（North American/YPS128）的基因呈现上调表达。此外，基因组测序与等位基因频率分析结果帮助定位到多个与氮限制发酵适应性相关的数量性状位点（QTL）。最后，我们验证了NA菌株的ECM38等位基因是氮限制条件下提升酵母生长效率的关键因子。综上，本研究结果揭示了助力酵母种群适应氮限制发酵的复杂分子特征谱，涵盖差异基因表达、等位基因频率改变以及线粒体基因组丢失。本研究结果还表明，非驯化分离株（NA）携带的野生等位基因在受试发酵条件的适应性过程中发挥重要作用，这些等位基因因此具备用于葡萄酒酵母菌株遗传改良的潜在价值。