Ji Xue-Xue, Zhang Quan, Yang Bai-Xue, Song Qing-Ran, Sun Zhao-Yong, Xie Cai-Yun, Tang Yue-Qin
College of Architecture and Environment, Sichuan University, Chengdu, 610065, Sichuan, China.
Sichuan Environmental Protection Key Laboratory of Organic Wastes Valorization, Chengdu, 610065, Sichuan, China.
Microb Cell Fact. 2025 Jan 30;24(1):33. doi: 10.1186/s12934-025-02663-7.
Continuous fermentation offers advantages in improving production efficiency and reducing costs, making it highly competitive for industrial ethanol production. A key requirement for Saccharomyces cerevisiae strains used in this process is their tolerance to high ethanol concentrations, which enables them to adapt to continuous fermentation conditions. To explore how yeast cells respond to varying levels of ethanol stress during fermentation, a two-month continuous fermentation was conducted. Cells were collected at different ethanol concentrations (from 60 g/L to 100 g/L) for comparative transcriptomic analysis.
During continuous fermentation, as ethanol concentration increased, the expression of genes associated with cytoplasmic ribosomes, translation, and fatty acid biosynthesis progressively declined, while the expression of genes related to heat shock proteins (HSPs) and ubiquitin-mediated protein degradation gradually increased. Besides, cells exhibited distinct responses to varying ethanol concentrations. At lower ethanol concentrations (nearly 70 g/L), genes involved in mitochondrial ribosomes, oxidative phosphorylation, the tricarboxylic acid (TCA) cycle, antioxidant enzymes, ergosterol synthesis, and glycerol biosynthesis were specifically upregulated compared to those at 60 g/L. This suggests that cells enhanced respiratory energy production, ROS scavenging capacity, and the synthesis of ergosterol and glycerol to counteract stress. At relatively higher ethanol concentrations (nearly 80 g/L), genes involved in respiration and ergosterol synthesis were inhibited, while those associated with glycolysis and glycerol biosynthesis were notably upregulated. This suggests a metabolic shift from respiration towards enhanced glycerol synthesis. Interestingly, the longevity-regulating pathway seemed to play a pivotal role in mediating the cellular adaptations to different ethanol concentrations. Upon reaching an ethanol concentration of 100 g/L, the aforementioned metabolic activities were largely inhibited. Cells primarily focused on enhancing the clearance of denatured proteins to preserve cellular viability.
This study elucidated the mechanisms by which an ethanol-tolerant S. cerevisiae strain adapts to increasing ethanol concentrations during continuous fermentation. The findings suggest that the longevity-regulating pathway may play a critical role in adapting to varying ethanol stress by regulating mitochondrial respiration, glycerol synthesis, ergosterol synthesis, antioxidant enzyme, and HSPs. This work provides a novel and valuable understanding of the mechanisms that govern ethanol tolerance during continuous fermentation.
连续发酵在提高生产效率和降低成本方面具有优势，使其在工业乙醇生产中具有高度竞争力。用于此过程的酿酒酵母菌株的一个关键要求是它们对高乙醇浓度的耐受性，这使它们能够适应连续发酵条件。为了探索酵母细胞在发酵过程中如何应对不同程度的乙醇胁迫，进行了为期两个月的连续发酵。在不同乙醇浓度（从60 g/L到100 g/L）下收集细胞用于比较转录组分析。
在连续发酵过程中，随着乙醇浓度的增加，与细胞质核糖体、翻译和脂肪酸生物合成相关的基因表达逐渐下降，而与热休克蛋白（HSPs）和泛素介导的蛋白质降解相关的基因表达逐渐增加。此外，细胞对不同乙醇浓度表现出不同的反应。在较低乙醇浓度（接近70 g/L）下，与线粒体核糖体、氧化磷酸化、三羧酸（TCA）循环、抗氧化酶、麦角固醇合成和甘油生物合成相关的基因与60 g/L时相比被特异性上调。这表明细胞增强了呼吸能量产生、ROS清除能力以及麦角固醇和甘油的合成以应对胁迫。在相对较高乙醇浓度（接近80 g/L）下，参与呼吸和麦角固醇合成的基因受到抑制，而与糖酵解和甘油生物合成相关的基因显著上调。这表明代谢从呼吸向增强的甘油合成转变。有趣的是，寿命调节途径似乎在介导细胞对不同乙醇浓度的适应中起关键作用。当乙醇浓度达到100 g/L时，上述代谢活动在很大程度上受到抑制。细胞主要专注于增强变性蛋白质的清除以维持细胞活力。
本研究阐明了耐乙醇酿酒酵母菌株在连续发酵过程中适应不断增加的乙醇浓度的机制。研究结果表明，寿命调节途径可能通过调节线粒体呼吸、甘油合成、麦角固醇合成、抗氧化酶和热休克蛋白在适应不同乙醇胁迫中起关键作用。这项工作为连续发酵过程中乙醇耐受性的调控机制提供了新颖且有价值的认识。
