Vibrio cholerae c-diGMP signaling: Motile to biofilm transition and transmission

项目总结 霍乱弧菌导致霍乱，霍乱是世界范围内的一个重要公共卫生问题。霍乱弧菌的能力 引起流行病与其在水生栖息地的传播和生存以及对人类的传播有关。 主持人。病原体形成生物膜(即基质封闭的、表面相关的群落)的能力对 它在流行之间的水生生境中存活，有利于在 流行病。信号核酸环二聚鸟苷一磷酸(c-diGMP)是广泛保守的 在细菌中，是生物膜形成的关键调节因子。了解c-diGMP如何控制生物膜的形成， 哪些环境信号调节c-diGMP水平和生物膜的形成，以及c-diGMP的后果 霍乱弧菌信号在感染性、传播性和传播性方面是有限的。这些信息差距将 通过专注于两个具体目标来解决。1)分析控制生物被膜的c-diGMP信号通路 2)分析c-diGMP信号通路的激活及其在V。 霍乱弧菌感染周期。在第一个目的下，c-diGMP调控的分子机制(S) IVA型MSHA菌毛的生产，初始表面附着的主要成分将被确定 使用结构和生化方法。C-diGMP下游信号通路启动于 表面附着将通过使用基于显微镜的群落跟踪方法进行分析 运动、分裂和第二信使信号水平。特定密钥c-diGMP信令的机制 控制表面附着和生物膜基质产生的蛋白质将通过组合使用来确定 遗传和生化方法。在第二个目标下，密钥c-diGMP的激活机制 信号蛋白将通过结构和配体结合研究来确定。C-diGMP信号转导途径的作用 在体内生物膜的形成中，霍乱弧菌的传播和扩散也将被调查，使用状态- 最先进的成像工具和新型c-diGMP传感器。拟议的工作将极大地促进对 C-diGMP信号如何运作，确定影响c-diGMP产生和降解的输入，以及 揭示c-diGMP信号的生物学后果。这项拟议的工作有望实现分子/机械论。 洞察力将使我们能够设计出控制c-di-GMP信号转导通路的方法，控制运动性和 生物被膜的形成，最终为霍乱弧菌传播抑制剂的开发提供靶标。

PROJECT SUMMARY Vibrio cholerae causes the disease cholera, an important public health problem worldwide. V. cholerae’s ability to cause epidemics is tied to its dissemination and survival in aquatic habitats and its transmission to the human host. The pathogen’s ability to form biofilms (i.e., matrix-enclosed, surface-associated communities) is crucial for its survival in aquatic habitats between epidemics and is advantageous for host-to-host transmission during epidemics. The signaling nucleotide cyclic dimeric guanosine monophosphate (c-diGMP) is broadly conserved in bacteria and is a key regulator of biofilm formation. Understanding of how c-diGMP controls biofilm formation, which environmental signals modulate c-diGMP levels and biofilm formation, and consequences of c-diGMP signaling in V. cholerae infectivity, transmission and dissemination, is limited. These information gaps will addressed by focusing on two specific aims. 1) Analyze c-diGMP signaling pathways that control biofilm formation dynamics; and 2) Analyze activation of c-diGMP signaling pathways and their consequences in V. cholerae infection cycle. Under the first aim, the molecular mechanism(s) through which c-diGMP controls production of the type IVa MSHA pilus, the primary component of initial surface attachment will be determined using structural and biochemical approaches. The down-stream c-diGMP signaling pathways initiated upon surface attachment will be analyzed by employing microscopy-based community tracking methods to measure motility, division, and second messenger signal levels. The mechanism by which specific key c-diGMP signaling proteins act to control surface attachment and biofilm matrix production will be determined using combination of genetic and biochemical approaches. Under the second aim, the mechanism of activation of key c-diGMP signaling proteins will be determined using structural and ligand binding studies. The role of c-diGMP signaling in in vivo biofilm formation, in V. cholerae transmission and dissemination will also be investigated, using state- of-the-art imaging tools and novel c-diGMP sensors. The proposed work will greatly advance the understanding of how c-diGMP signaling operates, identify the inputs that influence c-diGMP production and degradation, and unveil the biological consequences of c-diGMP signaling. The proposed work promises molecular/mechanistic insights that will allow us to devise ways to control c-di-GMP signal transduction pathways governing motility and biofilm formation, ultimately providing targets for the development of inhibitors of V. cholerae transmission.