酿酒酵母细胞壁和细胞膜应对高渗胁迫机制研究

酿酒酵母是食品发酵最常用的菌种之一,酵母应对高渗胁迫的能力决定食品发酵的成功与否以及食品腐败的危害程度。为研究高糖压力对酿酒酵母的影响,测定了酿酒酵母在应对极端高糖压力(0.6 g/mL)时细胞壁几丁质和葡聚糖含量的变化,采用CFW染料和葡聚糖特异性降解酶对细胞壁的敏感性进行分析,并进一步通过双染料(PI和Hoechst 33342)染色对酿酒酵母细胞膜完整性进行分析,最后通过RNA测序研究了酿酒酵母在极端高糖压力下表达的全局差异基因。结果表明:高糖压力改变了酿酒酵母细胞壁和细胞膜的特性。全局转录测试结果显示,高糖压力通过下调酿酒酵母与细胞壁完整性(CWI)信号传导途径相关的基因ROM1、RLM1、PIR3、YGP1、CWP1等,进行细胞壁损伤的应激反应,高糖压力还通过下调酿酒酵母与细胞膜成分相关的基因(如PDR15和YOR1)造成了细胞膜损伤。最后,比较了酿酒酵母在不同压力因子(0.2 g/mL糖、0.4 g/mL糖、0.6 g/mL糖和柠檬醛压力)下细胞依赖RLM1调控的CWI信号传导途径的差异调节,结果发现,在CWI中起关键作用的RLM1仅在0.6 g/mL糖的极端压力下被下调。

Hyperosmotic Stress Response of Saccharomyces cerevisiae Cell Wall and Cell Membrane

To better understand the physiological and molecular response of S.cerevisiae to hyperosmotic stress,chitin and glucan contents in the cell wall and sensitivity of the cell wall to calcofluor white(CFW)stain and cell wall-degrading enzymes were assayed.Further analysis of the cell membrane integrity of S.cerevisiae was performed by staining with dual dyes(PI and Hoechst 33342).Finally,gene expression profiles of S.cerevisiae under extreme high glucose pressure were conducted by RNA sequencing(RNA-seq).The alterations in membrane integrity and wall integrity in S.cerevisiae were found after high sugar stress.Hyperosmotic stress-treated cells increased sensitivity to CFW stain and cell wall-degrading enzymes demonstrated that hyperosmotic stress disrupted cell wall properties.It was also found that hyperosmotic stress-treated cells increased sensitivity to PI stain,indicating that the membrane was damaged.Global transcript measurements confirmed the membrane damage observations by showing down-regulation of membrane components and changes in fatty acid biosynthesis.High sugar stress did cause a compensatory response to cell wall damage through downregulation of several genes(ROM1,RLM1,PIR3,YGP1 and CWP1)involved with the cell wall integrity(CWI)signaling pathway.In addition,the previous studies were combined to discuss the CWI signaling pathway of yeast response to different stresses.The research results provided a reference for understanding hyperosmotic stress mechanism.