Identification of proteins secreted via Vibrio parahaemolyticus type III secretion system 1
Identification of proteins secreted via Vibrio parahaemolyticus type III secretion system 1
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DOI：
10.1128/iai.74.2.1032-1042.2006
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发表时间：
2006-02-01
影响因子：
3.1
通讯作者：
Honda, T
中科院分区：
文献类型：
--
作者：
Ono, T;Park, KS;Honda, T
副溶血性弧菌是一种革兰氏阴性海洋细菌，是引起食源性胃肠炎或败血症的重要病原菌。RIMD 2210633菌株（血清型03：K6的神奈川现象阳性临床分离株）的最新基因组测序显示，该菌株具有两组编码HI型分泌系统（TTSS）装置的基因簇。第一个簇TTSS1位于大染色体上，第二个簇TTSS2位于小染色体上。之前，我们报道了TTSS 1参与RIMD 2210633菌株对HeLa细胞的细胞毒性。在这里，我们通过使用二维凝胶电泳分析由TTSS 1编码的通过TTSS装置分泌的蛋白质，并鉴定由基因VP1680、VP1686和VPA 450编码的蛋白质。为了研究这些分泌蛋白的作用，我们构建并分析了一系列缺失突变体。流式细胞术分析，使用荧光激活的细胞分选与异硫氰酸荧光素标记的膜联蛋白V表明，TTSSI依赖的细胞死亡是由细胞凋亡。对HeLa细胞的细胞毒性与VP1680编码的一种新鉴定的分泌蛋白有关。腺苷酸环化酶融合蛋白的研究证明，新鉴定的分泌蛋白易位到HeLa细胞。因此，这些似乎是副溶血性弧菌中的TT SS效应蛋白。
Vibrio parahaemolyticus, a gram-negative marine bacterium, is an important pathogen causing food-borne gastroenteritis or septicemia. Recent genome sequencing of the RIMD2210633 strain (a Kanagawa phenomenon-positive clinical isolate of serotype 03:K6) revealed that the strain has two sets of gene clusters that encode the type HI secretion system (TTSS) apparatus. The first cluster, TTSS1, is located on the large chromosome, and the second, TTSS2, is on the small chromosome. Previously, we reported that TTSS1 is involved in the cytotoxicity of the RIMD2210633 strain against HeLa cells. Here, we analyzed proteins secreted via the TTSS apparatus encoded by TTSS1 by using two-dimensional gel electrophoresis and identified the proteins encoded by genes VP1680, VP1686, and VPA450. To investigate the roles of those secreted proteins, we constructed and analyzed a series of deletion mutants. Flow cytometry analysis using fluorescence-activated cell sorting with fluorescein isothiocyanate-labeled annexin V demonstrated that the TTSSI-dependent cell death was by apoptosis. The cytotoxicity to HeLa cells was related to one of the newly identified secreted proteins encoded by VP1680. Adenylate cyclase fusion protein studies proved that the newly identified secreted proteins were translocated into HeLa cells. Thus, these appear to be the TT SS effector proteins in V. parahaemolyticus.