Control of virulence in Vibrio cholerae by fatty acids

通过脂肪酸控制霍乱弧菌的毒力

• 批准号: 9174511
• 负责人: Fredrick Jon Kull
• 金额: $ 37.13万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9174511
• 关键词: 5' Untranslated Regions; Acids; Address; Adult; Affect; Bacterial Infections; Beryllium; Bicarbonates; Bile fluid; Binding; Cessation of life; ChIP-seq; Child; Cholera; Cholera Toxin; Communication; Complex; DNA; DNA Binding; DNA Binding Domain; Data; Development; Dimerization; Disease; Enteral; Environment; Epidemic; Factor V; Family; Fatty Acids; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Goals; Gram-Negative Bacterial Infections; Hour; Human; Hybrids; Laboratories; LacZ Genes; Lead; Length; Ligands; Light; Link; Mediating; Mission; Modeling; Molecular; Mutation; N-terminal; Nucleotides; Pathogenesis; Pathway interactions; Persons; Pharmaceutical Preparations; Pilum; Play; Prevention; Process; Proteins; RNA; Regulation; Regulatory Element; Research; Rhamnose; Ribosomes; Role; Signal Transduction; Stimulus; Stomach; Structure; Testing; Thermometers; Toxin; Transcriptional Activation; Translations; United States National Institutes of Health; Unsaturated Fatty Acids; Untranslated Regions; Vaccines; Vibrio cholerae; Vibrio cholerae O1; Virulence; Virulence Factors; Virulent; Work; base; drug discovery; fatty acid metabolism; in vivo; insight; killings; member; monomer; mutant; novel therapeutic intervention; pandemic disease; pathogenic bacteria; prevent; promoter; research study; small molecule; stem; structural biology; transcriptome sequencing

Vibrio cholerae O1 causes the fatal epidemic disease cholera. The ability of V. cholerae to cause disease in humans is dependent upon two primary virulence factors, the toxin-coregulated pilus (TCP), a critical colonization factor, and cholera toxin (CT). The expression of these factors is controlled by a highly regulated transcriptional cascade that serves as a paradigm for the regulation of bacterial virulence. Expression of the cascade is initiated at the tcpPH promoter by a cooperative interaction between the regulators AphA and AphB. TcpPH and ToxRS are homologous pairs of transmembrane regulators that then cooperate to activate expression from the toxT promoter. ToxT, an AraC-type regulator, directly activates the expression of TCP and CT. Transcriptional activation of the virulence cascade is strongly dependent upon a variety of stimuli from the external environment. The long-term goals of this proposal are to understand the molecular basis of virulence gene regulation in V. cholerae so as to facilitate the development of new strategies to control its infectivity. Through a collaborative effort involving laboratories with expertise in structural biology, virulence gene regulation and pathogenesis, we have found that exogenous unsaturated fatty acids (UFAs), which are components of human bile, are capable of binding to ToxT and impairing its ability to activate virulence gene expression. UFAs bind into a ligand pocket in the N-terminal domain of ToxT and inhibit its dimerization as well as its ability to bind to DNA. Bicarbonate, which neutralizes the acid that comes from the stomach, has been shown to function as a second in vivo signal that, in contrast to UFAs, stimulates ToxT and enhances its DNA binding through an unknown mechanism. We have recently identified a new link between fatty acids (FAs) and virulence gene expression in the current pandemic strain of V. cholerae with the discovery that the master regulator of FA metabolism, FadR, influences the translation of ToxT by an unknown mechanism. This proposal will build upon the ToxT structural and functional data, as well as our recent studies involving FadR, in order to elucidate several key mechanisms involved in regulating the expression of the virulence cascade. In Aim 1, we propose to elucidate the allosteric mechanisms controlling the dimerization of ToxT and to investigate the opposing effects of UFAs and bicarbonate on this process. In Aim 2, we propose to elucidate the mechanism by which FadR influences the translation of ToxT in V. cholerae. These studies will contribute significantly toward our understanding of how virulence gene expression is regulated in V. cholerae and will likely provide new avenues for antivirulence drug discovery.

霍乱弧菌 O1 引起致命的流行病霍乱。霍乱弧菌引起的能力 人类疾病依赖于两种主要毒力因子，即毒素共调节菌毛 (TCP)、 关键定植因子和霍乱毒素 (CT)。这些因子的表达受高度控制 调节转录级联，作为细菌毒力调节的范例。 级联的表达是通过 tcpPH 启动子之间的协作相互作用启动的。 调节剂 AphA 和 AphB。 TcpPH 和 ToxRS 是同源的跨膜调节因子对，然后 协同激活 toxT 启动子的表达。 ToxT 是一种 AraC 型调节剂，可直接激活 TCP和CT的表达。毒力级联的转录激活强烈依赖于 来自外部环境的各种刺激。该提案的长期目标是了解 霍乱弧菌毒力基因调控的分子基础，以促进新药物的开发 控制其传染性的策略。通过具有专业知识的实验室的共同努力 通过结构生物学、毒力基因调控和发病机制等方面的研究，我们发现外源不饱和 脂肪酸 (UFA) 是人类胆汁的组成部分，能够与 ToxT 结合并削弱其 激活毒力基因表达的能力。 UFA 结合到 ToxT N 端结构域的配体口袋中 并抑制其二聚化及其与 DNA 结合的能力。碳酸氢盐，可中和酸 来自胃部，已被证明可作为第二体内信号，与 UFA 相比， 刺激 ToxT 并通过未知机制增强其 DNA 结合。我们最近确定了一个 当前流行的霍乱弧菌菌株中脂肪酸（FA）和毒力基因表达之间的新联系 发现 FA 代谢的主调节因子 FadR 通过一种方式影响 ToxT 的翻译 未知的机制。该提案将建立在 ToxT 结构和功能数据以及我们的 最近涉及 FadR 的研究，以阐明参与调节的几个关键机制 毒力级联的表达。在目标 1 中，我们建议阐明控制的变构机制 ToxT 的二聚化并研究 UFA 和碳酸氢盐对此过程的相反影响。在 目标 2，我们建议阐明 FadR 影响 V 中 ToxT 翻译的机制。 霍乱。这些研究将极大地有助于我们了解毒力基因如何 霍乱弧菌的表达受到调节，可能为抗毒药物的发现提供新的途径。