Transcriptomic and metabolomic analysis: effects of different temperature conditions on higher alcohol production by Saccharomyces cerevisiae in grape must and exploration of related genes

Higher alcohols, produced during the process of alcoholic fermentation by Saccharomyces cerevisiae, significantly influence the flavor and safety of products. This study investigates the effect of temperature (18 °C, 23 °C, and 28 °C) on higher alcohol production using grape juice as the fermentation medium, by analyzing the transcriptomics and metabolomics of S. cerevisiae EC1118-a. By integrating these data, we identified differentially expressed genes (DEGs) associated with higher alcohol metabolism. And established correlations between DEGs and differential metabolites. Furthermore, single-gene knockout experiments of key pathway genes were conducted to elucidate their roles in regulating higher alcohol production. Within the temperature range of 18 °C to 28 °C, the yield of higher alcohols in S. cerevisiae EC1118-a increased with rising temperature. Moreover, temperature significantly influenced metabolic pathways associated with amino acid biosynthesis (ko01230), pyruvate metabolism (ko00620), glycolysis/gluconeogenesis (ko00010), purine metabolism (ko00230), glycine, serine, and threonine metabolism (ko00260). To evaluate their impact on higher alcohol synthesis, we selected 7 genes (FUN14, PET18, PHO84, SER33, SSA1, THI4, and YKR033C) exhibiting significant expression changes. Knockout of those DEGs resulted in reduced levels of 1-propanol (2-methyl), 1-butanol (2-methyl), 1-butanol (3-methyl), and phenethyl alcohol in the EC1118a-PHO84 strain by 14.72%, 8.93%, 11.19%, and 7.66%, respectively. Similarly, in the EC1118a-SER33 strain, these compounds decreased by 12.66%, 18.22%, 6.72%, and 13.37%. In the EC1118a-THI4 strain, levels of 1-butanol (3-methyl) and phenethyl alcohol decreased by 6.65% and 5.00%, respectively. Additionally, fermentation broths from the EC1118a-PHO84 and EC1118a-THI4 knockout strains showed a significant increase in 2,3-butanediol content. Ultimately, this research provides a foundation for developing industrial yeast strains with reduced higher alcohol production through gene knockout technology, thereby contributing to the improvement of wine quality by enhancing its aroma profile and ensuring greater safety in the final product.