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副溶血弧菌Ⅲ型分泌系统vscH基因生物学功能分析OA北大核心

Characterization and functional analysis of the vscH gene in the Vibrio parahaemolyticus type Ⅲ secretion system

中文摘要英文摘要

副溶血弧菌是一种重要的食源性病原菌.Ⅲ型分泌系统(type Ⅲ secretion system,T3SS)作为该菌的主要毒力因子之一,在致病过程中发挥重要作用.vscH位于T3SS1基因座中,功能未知.为探究vscH基因的生物学功能,通过同源重组的方法在副溶血弧菌POR-1菌株的基础上构建vscH缺失株ΔvscH及回补株CΔvscH,比较各菌株在生物被膜形成、细胞毒性等生物学特性上的差异;应用Western blot检测T3SS1效应蛋白的分泌量及其在HeLa细胞中易位量;通过GST-Pull Down试验鉴定与VscH相互作用的T3SS1相关蛋白.结果:与POR-1菌株相比,缺失株ΔvscH在生长性能上无明显差异,但生物被膜形成能力下降,同时对HeLa细胞和CaCo-2细胞的毒性作用明显下降;vscH基因的缺失使效应蛋白VopR、VopS和胞外针结构蛋白VscI的分泌显著减少,同时使VopR和VopS在HeLa细胞内易位量显著下降;GST-Pull Down试验发现VscH与一种位于T3SS1的ATP酶VscN存在相互作用.本研究表明,vscH基因影响副溶血弧菌生物被膜形成,参与T3SS1介导的毒性作用,同时对T3SS1效应蛋白的分泌和易位发挥重要功能.

Vibrio parahaemolyticus is an important food-borne pathogen,and type Ⅲ secretion system 1(T3SS1)plays a major role in its pathogenic process.vscH gene is correlated with T3SS1,and its effects on the pathogenesis of Vibrio parahaemolyticus have not been clarified.To investigate the biological functions of the vscH gene in Vibrio parahaemolyticus.vscH gene deletion strains ΔvscH and vscH gene compensa-tion strain CΔvscH were constructed here based on POR-1 strains using the homologous recombination technology.The biological characteris-tics,such as biofilm formation and cytotoxicity,of the different strains were compared.The secretion of effector proteins and their transloca-tion in HeLa cells were detected by Western blot.The potential proteins interacting with vscH were identified by GST-Pull Down.The results showed that vscH deletion did not affect the growth of the mutant,compared with its parent strain;but it inhibited biofilm formation and re-duced toxicity to HeLa cells and CaCo-2 cells.Meanwhile,vscH deletion greatly reduced the secretion of effector proteins VopR and VopS,the extracellular needle structure protein Vsci,and it markedly suppressed the translocation of VopR and VopS in the HeLa cells.Moreover,there was a potential interaction between vscH and vscN with an ATPase located at T3SS1.The present study indicated that the vscH gene had effect on the biofilm formation of Vibrio parahaemolyticus,participated in T3SS1-mediated cytotoxicity,and was essential for the secretion and translocation of T3SS1 effector proteins.

田珅;李婉君;曹钰莹;许立慧;任建鸾;汤芳;戴建君;薛峰

南京农业大学动物健康与食品安全国际合作联合实验室,江苏南京 210095南京农业大学动物健康与食品安全国际合作联合实验室,江苏南京 210095||中国药科大学生命科学与技术学院,江苏南京 210009

畜牧业

副溶血弧菌Ⅲ型分泌系统vscH分泌易位

Vibrio parahaemolyticustype Ⅲ secretion systemvscHsecretiontranslocation

《畜牧与兽医》 2025 (001)

57-65 / 9

国家自然科学基金项目(31871893)

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