Characterization of the ETEC Virulence Regulator Rns

ETEC 毒力调节剂 Rns 的表征

• 批准号: 7169876
• 负责人: GEORGE Patrick MUNSON
• 金额: $ 32.09万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-02-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7169876
• 关键词: Adherence; Age-Years; Bacteria; Binding; Binding Sites; Biological; Biological Models; Cessation of life; Child; Chromatin Loop; Complex; Coupled; DNA; DNA Binding; Decompression Sickness; Diarrhea; Disease; Distal; Family; Fimbriae Proteins; Genetic Transcription; Homeostasis; In Vitro; Infant; Infection; Intestinal Mucosa; Lead; Location; Molecular; Phase; Pilum; Prevention; Recruitment Activity; Regulation; Repression; Shigella flexneri; Site; Solutions; System; Transcription Initiation Site; Virulence; attenuation; enteroaggregative Escherichia coli; enterotoxigenic Escherichia coli; genetic analysis; killings; member; monomer; novel; pathogen; prevent; promoter; receptor

DESCRIPTION (provided by the applicant): Worldwide enterotoxigenic E. coli (ETEC) cause an estimated 210 million episodes of diarrheal disease and 380,000 deaths annually. The impact of this pathogen is greatest among infants and children less than 5 years of age. Adherence of the pathogen to the intestinal mucosa of the host is essential for the establishment of an infection. Attachment is usually achieved by pili which bind specific host receptors. The expression of CS1 and CS2 pili is positively regulated at the level of transcription by Rns a member of the AraC/XylS family. There are two Rns binding sites immediately upstream of the CS1 pilin promoter, within the region expected for a typical prokaryotic activator. Rns also positively autoregulates its own expression, however it does so through a highly unusual arrangement of DNA binding sites. Rns requires two DNA binding sites for the activation of its own promoter Prns. One of these is site 1 which is centered 224.5 bp upstream of the transcription start site (TSS), considerably further upstream than expected. Even more unusual is the location of site 3, which is centered 83.5 bp downstream of the TSS. Only a few prokaryotic activators are known to have binding sites downstream of the -10 hexamer. However we have shown that the virulence regulators VirF from Shigella flexneri, CfaR from ETEC, and AggR from enteroaggregative E. coli are also capable of activating Prns and like Rns, each requires the downstream binding site to do so. To date Rns is the only virulence regulator within this group for which an in vitro system has been developed. Thus our in vitro system, coupled with complimentary genetic analysis, affords us an outstanding opportunity to use Rns as a model system for a group of related regulators. These regulators are essential for the virulence of several pathogenic species of bacteria that collectively kill over 2 million people, predominantly children and infants, every year. A detailed understanding of these homologous regulators may eventually lead to new applications for disease treatment and prevention, such as attenuation of bacterial virulence by targeting the function of conserved virulence regulators. Our proposed studies will also provide new and fundamentally important information about transcription and its regulation because of the unique features of Rns autoregulation. These include activation from a downstream binding site and the potential repression of Prns by nonproductive RNAP open (RPo) complexes. Studies in this proposal will determine the molecular mechanism of Rns positive autoregulation and the biological significance of Rns-independent, nonproductive RPo complexes near Prns.

描述（由申请人提供）：全球肠毒素大肠杆菌（ETEC）估计会导致腹泻病的2.1亿次发作和每年380,000例死亡。这种病原体的影响在不到5岁的婴儿和儿童中最大。病原体对宿主肠粘膜的遵守对于建立感染至关重要。通常通过结合特定宿主受体的pili实现附着。 CS1和CS2 PILI的表达在RNS的转录水平上对ARAC/Xyls家族的成员进行了积极调节。在CS1 PILIN启动子的上游，有两个RN结合位点在预期的典型原用核激活剂的区域内。 RN还积极地自动调节了自己的表达，但是它通过高度不寻常的DNA结合位点的排列来做到这一点。 RN需要两个DNA结合位点，以激活其自身的启动子PRN。其中之一是位点1，该站点1以转录起始位点（TSS）上游为中心的224.5 bp，上游的上游比预期的要远大。更不寻常的是站点3的位置，该位置以TSS下游为中心83.5 bp。众所周知，只有少数原核激活剂具有-10六聚体下游的结合位点。但是，我们已经表明，来自Shigella flexneri，ETEC的CFAR和肠球形大肠杆菌的AGGR的毒力调节剂也能够激活PRN，并且像RN一样，每个人都需要下游结合位点才能这样做。迄今为止，RNS是该组中唯一开发了体外系统的毒力调节剂。因此，我们的体外系统，再加上免费的遗传分析，为我们提供了将RNS用作一组相关调节器的模型系统的出色机会。这些调节剂对于每年共同杀死了200万人，主要是儿童和婴儿的几种致病性细菌的毒力至关重要。对这些同源调节剂的详细了解最终可能导致新的疾病治疗和预防应用，例如通过靶向保守的毒力调节剂的功能来衰减细菌毒力。由于RN自动调节的独特特征，我们提出的研究还将提供有关转录及其调节的新的重要信息。这些包括从下游结合位点激活以及通过非生产性RNAP开放（RPO）复合物对PRN的潜在抑制。该提案中的研究将确定RNS阳性自动调节的分子机制以及PRN附近的非依赖性，非生产性RPO复合物的生物学意义。