Phase variation of virulence factors in Clostridium difficile

艰难梭菌毒力因子的相位变化

• 批准号: 10231056
• 负责人: RITA TAMAYO
• 金额: $ 41.55万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-20 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10231056
• 关键词: 5' Untranslated Regions; Adherence; Affect; Anabolism; Animal Disease Models; Animal Model; Antibiotics; Area; Attenuated; Bacteria; Binding Sites; Biochemistry; Cells; Centers for Disease Control and Prevention (U.S.); Cessation of life; Clostridium difficile; Code; Cytotoxin; DNA; DNA Sequence; Data; Development; Diarrhea; Disease; Down-Regulation; Epithelial Cells; Flagella; Gene Expression; Gene Expression Regulation; Genes; Genetic Recombination; Genetic Transcription; Goals; Health Care Costs; Human; Immune; In Vitro; Infection; Inflammatory Response; Intestines; Investigation; Knowledge; Laboratories; Link; Mediating; Messenger RNA; Mission; Modeling; Molecular; Molecular Biology; Operon; Pathogenesis; Pathogenicity; Phase; Physiological; Physiological Processes; Physiology; Preventive; Production; Pseudomembranous Colitis; Public Health; Publishing; Regulation; Regulatory Element; Reporting; Research; Resources; Rho Factor; Role; Sigma Factor; Site; Swimming; Symptoms; Testing; Therapeutic; Toxin; Transcript; Transcription Initiation; United States National Institutes of Health; Variant; Virulence; Virulence Factors; bacterial genetics; base; cell motility; combat; gene product; gut colonization; host colonization; in vivo; innovation; intestinal epithelium; mutant; pathogen; pathogenic bacteria; premature; recombinase; rho; transcription termination

PROJECT SUMMARY Clostridium difficile is a major public health threat, causing disease ranging from mild diarrhea to pseudomembranous colitis. C. difficile disease symptoms are largely mediated by the secreted cytotoxins, TcdA and TcdB. Many aspects of the pathogenicity of this bacterium remains poorly understood, including how C. difficile regulates the production of the toxins and other virulence factors. Toxin gene expression is linked to expression of flagellar genes via the flagellar sigma factor sigma-D, which is encoded in the flgB operon. Thus, factors that regulate the expression of the flgB operon will impact toxin production in addition to flagellar motility. Our long-term goal is to determine how C. difficile coordinately regulates flagellum and toxin gene expression, as these mechanisms are likely fundamental to pathogenesis. We recently showed that flagellar gene expression is subject to phase variation through site-specific DNA recombination of an invertible sequence upstream of the flgB operon. Notably, inversion of the DNA sequence, which we term the “flagellar switch”, also impacts the production of toxins essential for virulence. We identified RecV as the DNA recombinase that mediates inversion of the flagellar switch, and we have begun to define the mechanism by which the orientation of the flagellar switch sequence controls gene expression. Our central hypothesis is that phase variation of flagellum and toxin production is critical to C. difficile pathogenesis, and consequently interfering with the ability of C. difficile to phase vary will attenuate one or more aspects of virulence. While flagella and/or motility may be important for some aspects of infection, downregulation of flagella is likely vital for C. difficile to evade host immune recognition. Toxin production may similarly be beneficial or detrimental to C. difficile during different stages of infection. The objective of this proposal is to define the molecular mechanisms underlying phase variation and to evaluate the impact of phase variation on host colonization and virulence. This proposal is innovative because it represents an entirely new area of investigation toward understanding C. difficile disease: phase variable expression of flagella and toxins. Completion of the proposed studies will provide much needed mechanistic information on how C. difficile controls virulence factor production during infection, and may expose potential targets for inhibition of intestinal colonization and toxin production to facilitate efforts to combat this increasingly problematic pathogen.

项目摘要 艰难梭菌是主要的公共卫生威胁，导致疾病从轻度腹泻到 假膜结肠炎。艰难疾病症状的症状在很大程度上是由分泌的细胞毒素介导的， TCDA和TCDB。该细菌致病性的许多方面仍然很了解，包括如何 艰难梭菌调节毒素和其他病毒因素的产生。毒素基因表达与 通过在FLGB歌剧中编码的Flagllar Sigma因子Sigma-D的旗杆基因表达。那， 调节FLGB歌剧表达的因素除鞭毛外还会影响毒素的产生 运动。我们的长期目标是确定艰难梭菌如何协调调节鞭毛和毒素基因 表达，因为这些机制可能是发病机理的基础。我们最近证明了鞭毛 基因表达通过位点特异性DNA重组的相变的相位变化。 FLGB歌剧的上游序列。值得注意的是，DNA序列的反转，我们将其称为“鞭毛 开关”，还影响病毒必不可少的毒素的产生。我们确定RECV为DNA 介导鞭毛开关反转的重组酶，我们已经开始通过 鞭毛开关序列的方向控制基因表达。我们的中心假设是 鞭毛和毒素产生的相位变化对于艰难梭菌发病机理至关重要，因此 干扰艰难梭菌变化的能力会减弱病毒的一个或多个方面。尽管 鞭毛和/或运动对于感染的某些方面可能很重要，鞭毛的下调可能至关重要 艰难梭菌逃避宿主的免疫识别。毒素的产生可能同样有益或有害 艰难梭菌在不同的感染阶段。该提议的目的是定义分子 相位变化的机制，并评估相位变异对宿主定植的影响 病毒。该提议具有创新性，因为它代表了一个全新的投资领域 了解艰难梭菌疾病：鞭毛和毒素的相变表达。拟议的完成 研究将提供有关艰难梭菌如何控制病毒因子的急需的机械信息 感染过程中的产生，并可能暴露于抑制肠道定植和毒素的潜在靶标 生产以促进努力打击越来越有问题的病原体。