基于交互保护的氯化钠胁迫提高库德毕赤酵母高温耐性机制

Application of thermotolerant yeasts has extensive prospects in fermented food industries. The improvement in thermotolerance of yeasts can enhance fermentation efficiency and reduce fermentation cost. In this study, the multi-stress-tolerant Pichia kudriavzevii is chosen as the research object to study the cross-protection between the sodium chloride stress and high temperature stress. Proteome and transcriptome association analysis is used to study the effect of sodium chloride stress on the gene expression in P. kudriavzevii under high temperature stress. The key differentially expressed genes which can parse the protective effect of sodium chloride on the thermotolerance are selected. The expression levels and biological functions of the key genes are verified by SYBR Green qRT-PCR, RNAi and gene overexpression. Combined with the results of metabolic changes in P. kudriavzevii under different stress treatments, it is expected to illuminate the mechanisms of the improvement in thermotolerance of P. kudriavzevii under sodium chloride stress at multi-level and multi-angle. The results of this study can provide valuable references for improving the yeast thermotolerance, the genetic engineering of thermotolerant yeasts and the research of cross-protection in other tolerant microorganisms, which has a certain role in promoting the industrialized application of P. kudriavzevii and the development of fermented food industries.

耐高温酵母菌在发酵食品行业中有广阔的应用前景，高温耐性的改善可有效提高酵母菌的发酵效率，降低发酵成本。本研究以多耐性库德毕赤酵母为研究对象，系统研究氯化钠胁迫对库德毕赤酵母高温胁迫的交互保护作用，利用转录组和蛋白质组联合分析技术解析氯化钠胁迫对库德毕赤酵母高温胁迫基因表达的影响，筛选氯化钠胁迫对库德毕赤酵母高温胁迫保护的关键基因，并通过qRT-PCR、RNAi、基因过表达等技术对关键基因的表达量和生物学功能进行验证，结合不同胁迫处理下库德毕赤酵母应激代谢变化结果，以期从多层次、多角度阐释氯化钠胁迫提高库德毕赤酵母高温耐性的分子机制，其研究成果不仅为酵母菌高温耐性的提高提供新的思路，也为耐高温酵母菌的基因工程改造提供技术基础，同时还为其他耐性微生物的交互保护研究提供参考，对于库德毕赤酵母的工业化应用和发酵食品行业的发展有一定的推动作用。

耐高温酵母菌在发酵食品行业中有广阔的应用前景，高温耐性的改善可有效提高酵母菌的发酵效率，降低发酵成本。本研究以多耐性库德毕赤酵母为研究对象，系统研究氯化钠胁迫对库德毕赤酵母高温胁迫的交互保护作用和作用机制。通过对生物量、细胞形态和乙醇生产能力观察发现，盐胁迫对高温乙醇生产具有明显的交互保护作用。在45℃条件下，其生物量和乙醇产量分别提高了2.6倍和3.9倍。代谢网络图显示，盐胁迫显著改善了碳水化合物代谢中的关键酶和中间体，有助于乙醇、ATP、氨基酸、核苷酸和不饱和脂肪酸的合成，以及随后的细胞内代谢。海藻糖、甘油、热休克蛋白和麦角甾醇有助于维持细胞的正常功能。盐胁迫显著抑制了热诱导的ROS爆发、氧化损伤和细胞死亡，这主要是与GSH和GST的大量合成有关。乙醇产量的提高不仅是与耐热性的提高，也与盐胁迫下醇脱氢酶的上调和乙醛脱氢酶的下调密切相关。qRT-PCR和RNA-Seq结果具有显著正相关性，表明RNA-Seq结果的准确性。将HSP12重组到库德毕赤酵母染色体上，能显著提高其高温耐性和高温产乙醇能力。本研究结果不仅为酵母菌高温耐性的提高提供新的思路，也为耐高温酵母菌的基因工程改造提供技术基础，同时还为其他耐性微生物的交互保护研究提供参考，对于库德毕赤酵母的工业化应用和发酵食品行业的发展有一定的推动作用。