創傷弧菌（Vibrio vulnificus）一種嗜塩性的海洋致病菌，暴露在海水之中或是吃食了不潔的海鮮會對人體致病，包括了傷口感染、敗血症和急性下痢。之前的實驗中，我們實驗室對創傷弧菌建立了一個Tn5突變株的library，並對其篩選對血清有高度感受性的變異株。在眾多的突變株中利用基因構築（cloning）的技術定序出一段基因(命名為polR)。其轉譯出的蛋白質具有對σ54-depentent response regulators有相當高的相似度，其相鄰的一段基因命名為polS，其對histidine kinase sensor proteins也有相當高的相似度。
　　在本實驗中，由我們的磷酸化實驗結果顯示，PolS與PolR這兩個蛋白質會自我磷酸化（auto-phosphorylation）而形成一組訊息傳遞系統，並利用site-directed mutagenesis來確認。在與細胞做共同培養的實驗中發現到創傷弧菌polR突變株會對嗜中性球，單核球細胞，抗菌蛋白及巨噬細胞相較於野生株具有較低的抗性，這也意味著，polR基因的缺失對創傷弧菌的人體感染及抵抗能力有很大的影響。在polS及polR promoter 的預測中，發現到polS前端具有Sigma54 factor recognition的序列，而在promoter fusion 實驗指出polS promoter 會在stationary phase受到polymyxin B及單核球細胞的誘導而表現。由EMSA的結果看出的確Sigma54蛋白質會與polS前端的序列有結合，可能表示了此一訊息傳遞系統是由Sigma54 factor所調控，因此利用EMSA實驗發現了Sigma54 factor會與polS promoter結果，其結果指向Sigma54可能是調節polS 上游的轉錄因子，因此進行了sigma54基因的剔除。利用homologous recombination技術構築了sigma54的突變株，其游動性與生物膜的形成及黏附能力都較野生株為弱。，而進一步實驗指出，polR突變株與sigma54突變株利用對老鼠皮下注射可明顯觀測出對創傷弧菌致死率的下降，代表了由Sigma54所調控的polS/R two-component system在老鼠感染及致死能力扮演了一個重要的角色。
　　在本篇文章中，我們發現了polS/polR 這兩個基因會形成一組調節組織感染初期的訊息傳遞系統（two-component signal transductional system）。當創傷弧菌感染人體初期，人體白球（PMN等）或抗菌蛋白的不足以殺死創傷弧菌卻又能誘導此Sigma54所調控的訊息傳遞系統，而活化了PolS進而磷酸化PolR，PolR又會去調控活化下游某些致病基因，造成宿主的感染。

　　Vibrio vulnificus is a halophilic, marine pathogen that has been associated with primary septicemia and serious wound infections, often acquired by eating shellfish, and associated with exposure to seawater. In our previous studies, we constructed a library of transposon (Tn5) mutants of V. vulnificus and screened these mutants for hypersensitivity to human serum. One of the isolated serum-susceptible mutants had a mutation in the gene encoding product (PolR) with homology to the σ54-depentent response regulators of two-component signal transductional system (TCSTS). We also found an ORF, which is located immediately upstream of polR, and this gene (polS)’s product showed similarity to typical histidine kinase sensor proteins. A polR isogenic mutant exhibited more sensitive than the wild-type strain to killing by serum and polymyxin B. In this study, we found that the polR isogenic mutant was significantly more sensitive than the wild-type strain to opsonophagocytic killing by human PMNs or monocytes in vitro. Our data suggest that purified truncated C-terminal domain (244 a.a. residues) of PolS protein could phosphotransfer the intact PolR in vitro. With site-direct mutagenic protein PolR-D53A, we also confirmed that PolS could phosphotransfer to PolR. Promoter fusion experiments indicated that polS promoter could be markedly induced during exposure of bacteria to polymyxin B and monocytes, especially when bacteria cells were in stationary phase. Electrophoresis mobility shift assays revealed that purified protein of σ54 especially binds to the upstream region of polS. We also constructed a sigma54 isogeneic mutant by homologous recombination. The sigma54 isogeneic mutant, like polR isogenic mutant, showed more sensitive than parental strain to killing by PMNs, monocytes, macrophages or polymyxin B. In addition, both sigma54 mutant and polR mutant exhibited a reduced motility, biofilm formation ability and adhesion ability as compared to the parental strain. The mice of BABL/cJ strain inoculated with polR mutant or sigma54 mutant demonstrated virulence attenuation when the strains were administered subcutaneously. These results indicate that the polS and polR are cognate regulatory components and the expression of polS may be especially induced by σ54 when Vibrio vulnificus is within the infected tissue. Under Sigma54 factor regulation, the polS/polR components appear to play a role in mediating resistance to host innate defense during early events in a natural infection.

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