Gene Regulation by ToxR, TcpP and ToxT in V. Cholerae

描述(申请人提供)：霍乱弧菌中ToxR、TCPP和ToxT的基因调控霍乱弧菌、霍乱毒素和毒素共调控菌毛(分别为CT和TCPP)的主要毒力因子分别由一系列调节剂控制，最终导致ToxT被两种膜定位激活剂ToxR和TCPP激活。ToxT是霍乱毒素和毒素共同调节的菌毛基因表达的直接激活剂，也是其他基因的表达，这些基因在霍乱弧菌的生物学和致病性中的作用尚不清楚。膜激活剂ToxR和TCPP各自依赖于同源效应蛋白ToxS和TcpH的活性，尽管这种依赖的机制对于每一对蛋白质可能是不同的。先前资金阶段的一个主要发现是，TCPP受制于被TcpH拮抗的膜内蛋白分解。这一过程需要Yael蛋白酶作为从细胞中消除TCPP的两种酶中的第二种。TCPP是激活toxT基因所必需的，NIH最近一次支持这项工作的另一项重大发现是鉴定了ToxT直接转录控制下的一个小RNA分子(Tara)。基于大量的初步数据和已发表的文献，目前的建议有三个目标：1.全面分析TCPP和TcpH的相互作用以及调控细胞内TCPP水平的膜内蛋白分解途径，包括鉴定仍未知的位点I酶；2.筛选调控途径中ToxR/TCPP臂的小分子抑制剂，并表征这些抑制剂的靶点和抑制剂的作用机制；3，Tara的作用机制。 与公共卫生相关：霍乱弧菌是一种导致人类腹泻疾病霍乱的细菌。拟议的研究旨在揭示霍乱弧菌用来调节其毒力特征的新知识机制。其中一个目标是识别干扰导致疾病的调控途径关键步骤的小分子。这些分子可能代表了针对毒力的新型药物。

DESCRIPTION (provided by applicant): Gene regulation by ToxR, TcpP and ToxT in V. cholerae The major virulence factors of Vibrio cholerae, cholera toxin and toxin-coregulated pilus (CT and TCP, respectively) are controlled by a cascade of regulators that ultimately leads to activation of ToxT by two membrane-localized activators, ToxR and TcpP. ToxT is the direct activator of cholera toxin and toxin-coregulated pilus gene expression, as well as expression of other genes whose roles in the biology and pathogenicity of V. cholerae are less well defined. The membrane activators, ToxR and TcpP, each depend for their activity on a cognate effector protein, ToxS and TcpH, although the mechanism of this dependence may be different for each pair of proteins. A major discovery of the prior funding period is that TcpP is subjected to regulate intramembrane proteolysis that is antagonized by TcpH. This process requires the YaeL protease as the second of two proteases that eliminate TcpP from the cell. TcpP is necessary to activate the toxT gene, and another major discovery from the most recent period of NIH support for this work was identification of a small RNA molecule (tarA) under direct transcription control of ToxT. Based on significant preliminary data and published manuscripts, the current proposal has three aims: 1. Comprehensive analysis of TcpP and TcpH interaction and the regulated intramembrane proteolysis pathway that governs TcpP levels in the cell, including the identification of the still unknown site I protease; 2. Screens for small molecule inhibitors of the ToxR/TcpP arm of the regulatory pathway and characterization of the targets of those inhibitors as well as the mechanism of inhibitor; 3, the tarA mechanism of action. PUBLIC HEALTH RELEVANCE: Vibrio cholerae is a bacterium that causes the human diarrheal disease cholera. The proposed studies are designed to uncover new knowledge mechanisms used by V. cholerae to regulate its virulence traits. One of the aims is intended to identify small molecules that interfere with key steps in the regulation pathway leading to disease. Such molecules might represent new classes of drugs that target virulence.