Role of RpoS in stress response of Vibrio parahaemolyticus and cold response genes selection

• Main Authors: Zih-Li Wang, 王自立
• Other Authors: Mei-Shiuan Yu
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/73903319958636857427

碩士 === 慈濟大學 === 微免暨分子醫學研究所 === 97 === Vibrio parahaemolyticus is a food-borne pathogen that inhabits in brackish seawater throughout the world. For microorganism, to survive in a variety of diverse and harsh stresses during their life cycle, it’s important to regulate gene expression responding to the continuous changing of surrounding environment. Switching of sigma factor, a subunit of RNA polymerase is a common way for microorganisms to regulate the gene expression pattern. It is believed that RpoS (σS) participates in general stress responses in many bacteria. To understand the role of V. parahaemolyticus RpoS in stress response, we constructed a rpoS mutant and compared the survival and gene expression of the mutant and wild type under selected stresses. We found that there was no significant difference between wild type and mutant in growth performance at 37oC. In stationary-phase cells, rpoS mutants were more sensitive to oxidative, starvation, and acid stress conditions, but both of wild type and mutant showed similar survival characteristics under cold treatment. It indicated that RpoS might play an important role in stationary-phase V. parahaemolyticus in regulating stress responses. Based on catalase zymogram, three divergent patterns (Kat A, B, C) were found in the wild type, but the catalase activity of group C was faintly expressed in the rpoS mutant. Then the expression of catalase genes were also monitored by RT-PCR. In rpoS mutant, both katG genes (VPA0453 and VPA0768) expression was decreased after 4 hr, but katE genes (VPA0305 and VPA1418) were continuously expressed at 8 hr. These results showed that expression of katG and kat E was affected by RpoS. Moreover, several cold-sensitive strains were selected from mini-Tn10 mutant library. The flanking sequence of transposon insertion site was amplified by arbitrary primed PCR and sequenced. A novel gene (VP2094), encodes a molybdenum cofactor biosynthesis protein, that are related to cold resistance in V. parahaemolyticus was identified. In summary, RpoS might involved in the response of oxidative stress, starvation, acid stress. Both of katG and katE was also regulated by RpoS.