【作者】 李西川；

【导师】 蒋伶活；

【作者基本信息】 天津大学 ， 应用化学， 2009， 博士

【摘要】 酿酒酵母ScRck2p是一种MAP Knase激活的蛋白激酶,被Hog1p磷酸化而激活,响应于细胞对外界高渗透压胁迫和氧化胁迫。在本工作中我们发现酿酒酵母ScRCK2基因的缺失能够导致细胞对TOR抑制剂——雷帕霉素敏感,但ScRCK2在细胞对雷帕霉素敏感过程中的作用并不依赖于其激酶活性。在人体内最常见的机会致病菌-白念珠菌中,我们鉴定了ScRCK2的同源基因CaRCK2,并发现CaRCK2基因的缺失同样导致白念珠菌细胞对雷帕霉素敏感,CaRck2p的这一作用同样不依赖于其激酶活性。此外,我们发现白念珠菌CaHOG1的缺失也导致细胞对雷帕霉素的敏感。这些结果表明在酿酒酵母菌和白念珠菌中,作用于HOG途径下游的RCK2可能调控TOR信号途径的功能。同时,我们发现白念珠菌CaRCK2基因的缺失能够导致细胞对高渗胁迫、氧化胁迫和细胞壁完整性胁迫试剂敏感。比较而言,酿酒酵母ScRCK2基因的缺失只引起细胞对氧化胁迫敏感,而不影响细胞对高渗胁迫和细胞壁完整性胁迫试剂的敏感。但是,酿酒酵母ScRCK2基因却能够弥补白念珠菌CaRCK2基因缺失株对高渗胁迫、氧化胁迫和细胞壁完整性胁迫试剂的敏感性表型。这些结果说明RCK2在细胞内发挥多种功能,但在白念珠菌和酿酒酵母菌中的功能存在分歧。我们还发现,ScRCK2上游激酶基因ScHOG1和ScPBS2的缺失,以及受ScHOG1调控的转录因子基因ScHOT1,ScMSN1,ScMSN2,ScMSN4或ScRLM1的缺失,不影响酵母细胞对雷帕霉素的敏感性。但是,白念珠菌CaHOG1基因的缺失能够导致细胞对雷帕霉素敏感。这些结果表明白念珠菌HOG途径与酿酒酵母HOG途径在细胞功能调控方面存在分歧,它参与细胞对雷帕霉素敏感性的调控。更多还原

【Abstract】 Rck2p is a Hog1p-MAP kinase activated protein kinase and regulates osmotic and oxidative stresses in budding yeast. In this study, we have demonstrated that in both Saccharomycese cerevisiae and, the most medically important human fungal pathogen, Candida albicans deletion of RCK2 causes cells sensitive to rapamycin, the inhibitor of TOR (target of rapamycin) protein kinase controlling cell growth. In addition, the kinase activity of Rck2p seems to be not required for this rapamycin sensitive function in both eukaryotic microorganisms. We also have found that CaRCK2 and its kinase activity are required for cell growth in C. albicans. Furthermore, we have found that Candida cells deleted for CaRCK2 display an increased sensitivity to high osmotic stress, oxidative stress and agents disrupting cell wall integrity, whereas yeast cells deleted for ScRCK2 only show an increased sensitivity to oxidative stress. Our results indicate that RCK2 plays divergent roles in C. albicans and budding yeast cells.We also found that deletion of HOG1 and PBS2 as well as genes encoding HOG1-regulated transcription factors does not affect the rapamycin sensitivity in S. cerevisiae, whereas disruption of CaHOG1 causes cells sensitive to rapamycin in C. albicans. These results suggest that the HOG pathway also plays divergent roles in C. albicans and budding yeast cells.更多还原