Signal Transduction in the Major Quorum-Sensing Circuit of Vibrio cholerae

描述(申请人提供)：这项研究的目标是了解群体感应背后的分子机制，群体感应是一种细胞-细胞交流过程，允许细菌协调整个种群的基因表达并作为协调的群体发挥作用。群体感应依赖于被称为自体诱导剂的化学信号分子的产生、检测和反应。破坏这些步骤中的任何一个都会使病原菌变得无毒。这里提出的研究将提供对群体感应中信号产生、检测和响应的分子机制的理解，这将使干扰策略的设计能够发展成为对抗传染病的新疗法。霍乱弧菌是霍乱的病原体，它通过一种新的自身诱导剂，称为CAI-1(S)-3-羟基十三碳-4-酮，通过群体感应调节毒力因子的产生和生物膜的形成。CaI-1由CqsA合成酶产生，并由跨膜型组氨酸激酶受体CqsS检测。组氨酸激酶受体广泛存在于细菌中，在细菌的致病过程中发挥着重要作用，但对其跨膜信号转导的分子机制却知之甚少。这在一定程度上是由于严重缺乏组氨酸激酶的化学定义的配体。通过确定CAI-1的生物合成途径和结构，以及研究CAI-1与CqsS的相互作用以及CqsS与一系列相关合成分子的相互作用，我的博士后工作表明，CqsA/CqsS/CAI-1信号通路为研究配体-受体相互作用和组氨酸激酶受体的跨膜信号提供了前所未有的机会。以这项工作为基础，展望未来，我的第一个目标是使用CAI-1/CqsS作为一个模型系统来定义这类重要的受体如何识别外部信号并将其内部传播到细胞内。我的第二个目标集中在CqsA合成酶产生的信号的共同进化和伙伴CqsS受体的信号识别。作为博士后，我展示了每个弧菌CqsA/CqsS对的信号产生和信号识别是匹配的。也就是说，要么特定的CqsA合成多个相关分子，并由同源的CqsS受体检测所有这些分子，要么CqsA酶产生一个特定的分子，同源的CqsS受体对该配体的检测具有极高的选择性。这种共同进化的生物学意义以及指定其严格或缺乏的分子机制尚不清楚，我将研究这一现象。我们对群体感应在霍乱弧菌生理中的作用的了解仅限于毒力因子的产生和生物膜的形成。然而，霍乱弧菌的群体感应调节基因除了与毒力和生物膜形成有关的基因外，还由100多个基因组成。我最近的工作表明，霍乱弧菌突变体无法激活群体感应反应，对低pH值非常敏感。在我的最终目标中，我将研究群体感应依赖的耐酸反应调节的分子机制。长期目标是确定霍乱弧菌群体感应反应在应对环境和宿主变化中的作用。现在已经确定，群体感应被许多细菌物种用来调节有害和有益的特征。群体感应领域的一个长期目标是开发正向群体感应和/或反群体感应小分子来操纵这些行为。这项研究将朝着这一重要目标取得进展。 公共卫生相关性：霍乱弧菌是一种全球重要病原体，每年霍乱的负担估计高达数百万例。霍乱弧菌的毒力依赖于称为群体感应的细胞-细胞通信过程，该过程控制着毒力因子的产生和释放以及生物膜的形成。这里提出的研究将扩大我们对群体感应如何控制这种重要病原菌的毒力的理解，此外，这项研究将促进合成策略的发展。 用于控制霍乱弧菌的毒力，并可能对改善人类健康产生巨大的影响。

DESCRIPTION (provided by applicant): The goal of this research is to understand the molecular mechanisms underlying quorum sensing, a cell-cell communication process that allows bacteria to coordinate population-wide gene expression and function as coordinated groups. Quorum sensing relies on the production, detection, and response to chemical signal molecules called autoinducers. Disruption of any one of these steps renders pathogenic bacteria avirulent. The research proposed here will provide an understanding of the molecular mechanisms underpinning signal production, detection and response in quorum-sensing which will enable the design of interference strategies that can be developed into new therapies to combat infectious diseases. The human pathogen Vibrio cholerae, the causative agent of the disease cholera, regulates virulence factor production and biofilm formation through quorum sensing via a new class of autoinducer, called CAI-1 ((S)-3-hydroxytridecan-4-one). CAI-1 is produced by the CqsA synthase and detected by the transmembrane histidine kinase receptor CqsS. Histidine kinase receptors are ubiquitous in bacteria and play important roles in bacterial pathogenesis, nonetheless, little is known about their molecular mechanism of signal transduction across the membrane. This is, in part, due to a profound lack of chemically defined ligands for histidine kinases. By determining the biosynthetic pathway for and structure of CAI-1, and examining the CAI-1-CqsS interaction as well interactions between CqsS and a series of related, synthetic molecules, my postdoctoral work demonstrated that the CqsA/CqsS/CAI-1 signaling pathway offers an unprecedented opportunity to study ligand-receptor interactions and transmembrane signaling by histidine kinase receptors. With this work as the foundation, going forward, my first aim is to use CAI-1/CqsS as a model system to define how this important class of receptors recognizes and propagates external signals internally into the cell. My second aim focuses on co-evolution of signal production by the CqsA synthases and signal recognition by the partner CqsS receptors. As a postdoc, I showed that signal production and signal recognition in each vibrio CqsA/CqsS pair are matched. That is, either a particular CqsA synthesizes multiple related molecules and the cognate CqsS receptor detects them all, or the CqsA enzyme produces one specific molecule and the cognate CqsS receptor is exquisitely selective for detection of that ligand. The biological significance of this co-evolution and the molecular mechanism that specifies stringency or lack thereof are unclear and I will study this phenomenon. Our understanding of the role quorum sensing plays in V. cholerae physiology is limited to virulence factor production and biofilm formation. However, the V. cholerae quorum-sensing regulon is composed of more than 100 genes in addition to those known to be involved in virulence and biofilm formation. My recent work showed that V. cholerae mutants unable to activate a quorum-sensing response are exquisitely sensitive to low pH. In my final aim, I will study the molecular mechanisms underpinning quorum-sensing dependent regulation of Acid Tolerance Response. The long term goal is to define the roles of V. cholerae quorum-sensing response in coping with the changes in the environment and in the host. It is now well established that quorum sensing is employed by many bacterial species to regulate both harmful and beneficial traits. A long standing goal of the quorum-sensing field is to develop pro-quorum- sensing and/or anti-quorum-sensing small molecules to manipulate these behaviors. This study will make progress toward this important goal. PUBLIC HELATH RELEVANCE: Vibrio cholerae is a globally important pathogen and the burden of cholera is estimated to reach several million cases annually. V. cholerae virulence depends on a cell-cell communication process called quorum sensing that controls the timing of production and release of virulence factors and the formation of biofilms. The investigations proposed here will expand our understanding of how quorum sensing controls virulence in this important pathogen, in addition, this study will facilitate the development of synthetic strategies for controlling V. cholerae virulence and could have enormous ramifications for improving human health.