Fatty acid related regulation of enteric infectious disease

肠道传染病的脂肪酸相关调节

• 批准号: 8030148
• 负责人: Fredrick Jon Kull
• 金额: $ 23.7万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-30 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8030148
• 关键词: 5 year old; Accounting; Affect; Age-Years; Amino Acids; Bacteria; Bacterial Infections; Bile fluid; Binding; Biochemical; Biological Assay; Cessation of life; Characteristics; Charge; Child; Cholera; Cholera Toxin; Code; Codon Nucleotides; Communicable Diseases; Country; DNA; DNA Binding; DNA Binding Domain; Development; Diarrhea; Disease; Dysentery; Elderly; Electrophoretic Mobility Shift Assay; Enteral; Enterobacteriaceae; Escherichia coli; Family member; Fatty Acids; Food; Gene Expression; Gene Expression Regulation; Genes; Genetic; Homology Modeling; Individual; Infection; Ingestion; Laboratories; LacZ Genes; Lead; Ligand Binding; Ligands; Lysine; Measures; Mediating; Methods; Modeling; Molecular Conformation; Mutate; Organism; Pathology; Pilum; Play; Positioning Attribute; Process; Production; Proteins; Regulation; Reporter; Resolution; Role; Salmonella; Salmonella enteritidis; Salmonella typhi; Salmonella typhimurium; Sequence Alignment; Shigella flexneri; Side; Signal Transduction; Site-Directed Mutagenesis; Structure; System; Techniques; Testing; Therapeutic; Toxin; Traveler' s diarrhea; Typhoid Fever; United States; Vibrio cholerae; Virulence; Virulence Factors; Water; World Health Organization; Yersinia enterocolitica; base; enteropathogenic Escherichia coli; enterotoxigenic Escherichia coli; gel mobility shift assay; interest; member; palmitoleic acid; pathogenic bacteria; prevent; promoter; prophylactic; response; therapeutic development; transcription factor

DESCRIPTION (provided by applicant): According to the World Health Organization, diarrhea caused by enteric infections affects some four billion individuals annually throughout the world, resulting in an estimated four million to six million deaths, many in children under five years of age. In the United States, diarrhea is the second most common infectious illness, and in some countries diarrheal diseases account for up to one third of all deaths. In many cases, enteric disease is caused by the ingestion of pathogenic bacteria via contaminated food or water. Once ingested by the host, environmental signals in the gut initiate a virulence gene cascade leading to the production of virulence factors. The expression of these virulence factors, many of which are toxins, effector molecules, and colonization factors, results in a wide variety of debilitating and often deadly diarrheal diseases including travelers' diarrhea, dysentery, and cholera. In cholera the primary virulence factors are the toxin- coregulated pilus (TCP), which is required for the organism to colonize the host, and cholera toxin (CT), which is responsible for the pathology of the disease. The expression of both virulence factors is regulated by ToxT, the paramount regulator of virulence gene expression in Vibrio cholerae. ToxT, a member of the AraC/XylS (A/X) superfamily of regulators, of which there are numerous members throughout the bacterial world, is inhibited by fatty acids present in bile. In the structure of ToxT, a palmitoleic acid bridges the regulatory and DNA binding domains of the transcription factor, locking it in an inactive conformation. Two positively charged lysine side chains, one from each domain, play a critical role in fatty acid binding and, thereby, in the mechanism of virulence gene regulation. The studies proposed in this application will test the hypothesis that many other enteric bacteria use a similar, if not identical, mechanism to regulate pathogenic activity. The approach will utilize secondary structure prediction and homology modeling to identify candidates from the A/X protein superfamily containing positively charged amino acid residues at positions homologous to those found in ToxT. The effect of fatty acids on virulence gene production in the various candidate organisms will be measured utilizing a lacZ fusion system and confirmed using electrophoretic mobility shift assays (EMSAs), an NMR-based ligand binding assay, and site directed mutagenesis. Preliminary analysis has indicated that a number of pathogenic bacteria, including several types of Escherichia coli, Salmonella typhi, Salmonella typhimurium, Shigella flexneri, and Yersinia enterocolitica, contain A/X regulators with homologous positively charged residues and ligand binding pockets. The studies proposed here will confirm if these bacteria use a common, fatty acid mediated mechanism to regulate pathogenic activity, thereby laying the groundwork for development of therapeutics that could be broadly applied to treat enteric bacterial infections causing diseases such as travelers' diarrhea, dysentery, and typhoid fever. PUBLIC HEALTH RELEVANCE: Enteric infectious diarrheal diseases affect more than 4 billion individuals yearly throughout the world, resulting in an estimated 4 million to 6 million deaths, primarily among the elderly and children less than 5 years of age. This proposal investigates how pathogenic bacteria use fatty acids to regulate virulence activity. Understanding this process could lead to the development of therapeutics to treat or prevent enteric bacterial infections including travelers' diarrhea, salmonella, dysentery, and typhoid fever.

描述（由申请人提供）：根据世界卫生组织的说法，由肠道感染引起的腹泻每年在全球范围内影响约40亿个人，导致估计有400万至600万人死亡，其中许多儿童五岁以下。在美国，腹泻是第二常见的传染病，在某些国家，腹泻疾病占所有死亡的三分之一。在许多情况下，肠道疾病是由通过污染的食物或水摄入致病细菌引起的。一旦被宿主摄入，肠道中的环境信号会引发毒力基因级联反应，从而导致毒力因子产生。这些毒力因子的表达，其中许多是毒素，效应分子和定植因子，导致多种使人衰弱的腹泻疾病，包括旅行者的腹泻，痢疾和霍乱。在霍乱中，主要的毒力因子是生物体定居宿主所必需的毒素核心菌毛（TCP），以及负责疾病病理学的霍乱毒素（CT）。两种毒力因子的表达均由Toxt（Toxt）调节，Toxt是蛇形霍乱中毒力基因表达的最高调节剂。 TOXT是调节剂的ARAC/Xyls（A/X）超家族的成员，在整个细菌世界中有许多成员受到胆汁中存在的脂肪酸的抑制。在TOXT的结构中，棕榈酸桥接了转录因子的调节和DNA结合结构域，将其锁定在不活跃的构象中。两个正电荷的赖氨酸侧链，一个来自每个结构域，在脂肪酸结合中起关键作用，从而在毒力基因调节的机理中起着关键作用。 在本应用中提出的研究将检验以下假设：许多其他肠细菌使用类似（即使不是完全相同的机制）来调节致病活性。该方法将利用二级结构预测和同源模型来鉴定来自A/X蛋白超家族的候选物，这些蛋白质包含与TOXT中发现的位置同源的位置的正电荷氨基酸残基。通过LACZ融合系统，将测量脂肪酸对各种候选生物中毒力基因产生的影响，并使用电泳迁移率转移测定法（EMSA），一种基于NMR的配体结合测定法和位点定向诱变。初步分析表明，许多致病性细菌，包括几种类型的大肠杆菌，鼠伤寒沙门氏菌，鼠伤寒沙门氏菌，shigella flexneri和Yersinia肠结肠膜炎，含有A/X的调节剂，具有同源的带电的带电的带电残基和粘合式口袋。此处提出的研究将证实这些细菌是否使用常见的脂肪酸介导的机制来调节致病活性，从而为开发治疗剂的基础奠定了基础，可以广泛应用来治疗肠细菌感染，从而导致旅行者的腹泻，孕妇和疾病。 公共卫生相关性：肠道感染性腹泻疾病每年影响超过40亿人，导致估计400万至600万人死亡，主要是在老年人和不到5岁的儿童中。该提案研究了致病细菌如何使用脂肪酸调节毒力活性。了解这一过程可能会导致治疗或预防肠道细菌感染（包括旅行者腹泻，沙门氏菌，痢疾和伤寒）的疗法发展。