副溶血弧菌VI型分泌系统的表型功能及基因调控研究

副溶血弧菌表达两种 VI 型分泌系统（Type VI secretion system, T6SS），分别称作 T6SS1 和 T6SS2。有关副溶血弧菌 T6SS1 和 T6SS2 的具体功能及其基因调控机制，均不甚明确。在本研究中，将分析 T6SS1 和 T6SS2 在小鼠致死活性、细胞毒性、细胞粘附、抑菌活性等方面的功能表型效应；再通过引物延伸实验、LacZ 报告基因融合实验、蛋白表达与纯化、凝胶阻滞实验、DNase I 足迹实验，探究密度感应（Quorum sensing, QS）系统核心调控子 AhpA 和 OpaR 调控 T6SS1 和 T6SS2 基因转录的精细分子机制；最后将表型和分子实验数据结合起来，确证 T6SS1 和 T6SS2 的功能差异性，并确证由于 QS 调控引起的 T6SS1 和 T6SS2的菌密度依赖性的差异表达。

Vibrio parahaemolyticus expresses two different type VI secretion systmes (T6SSs) called T6SS1 and T6SS2. The phenotpes and gene regulation of T6SS1 and T6SS2 of V. parahaemolyticus are still poorly understood. In this study, the phenotpic effects including lethality against mice, cytotoxicity, cell adhesion, and antimicrobial activity will be systematically characterized for T6SS1 and T6SS2 of pandemic V. parahaemolyticus; the primer extension, LacZ fusion reporter, protein expression and purification, gel mobility shift, and DNase I footprinting assays will be then carried out to characterize the trancscriptional regulation of T6SS1 and T6SS2 genes by the quorum-sensing (QS) master regulators OpaR and AphA; the final combined phenotpic and melecular dataset will promote us to gain the deeper understanding of distinct phenotpes and QS-mediated cell dencsity-dependent reciprocal regulation of T6SS1 and T6SS2 expression in V. parahaemolyticus.