Characterization of the ETEC Virulence Regulator Rns

ETEC 毒力调节剂 Rns 的表征

• 批准号: 6924167
• 负责人: GEORGE Patrick MUNSON
• 金额: $ 33.85万
• 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/6924167
• 关键词: Characterization; ETEC; Virulence; Regulator; Rns

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.

性状（由申请方提供）：全球产肠球菌E.大肠杆菌（ETEC）每年估计造成2.1亿例腹泻病和38万例死亡。这种病原体对婴儿和5岁以下儿童的影响最大。病原体粘附于宿主的肠粘膜对于感染的建立是必不可少的。附着通常通过结合特异性宿主受体的皮利来实现。CS 1和CS2皮利的表达在转录水平上受到AraC/XylS家族成员Rns的正调控。在CS 1菌毛蛋白启动子的上游有两个Rns结合位点，在典型的原核激活剂预期的区域内。Rns也积极地自动调节其自身的表达，但它是通过一种非常不寻常的DNA结合位点排列来实现的。Rns需要两个DNA结合位点来激活其自身的启动子Prns。其中之一是位于转录起始位点（TSS）上游224.5 bp处的位点1，比预期的要远得多。更不寻常的是位点3的位置，其位于TSS下游83.5 bp处。已知只有少数原核激活剂在-10六聚体下游具有结合位点。然而，我们已经表明来自福氏志贺菌的毒力调节因子VirF、来自ETEC的CfaR和来自肠聚集性E.大肠杆菌也能够激活Prns，并且与Rns一样，每个都需要下游结合位点才能做到这一点。到目前为止，Rns是该组中唯一的毒力调节剂，已开发出体外系统。因此，我们的体外系统，加上免费的遗传分析，为我们提供了一个很好的机会，使用RNS作为一组相关监管机构的模型系统。这些调节剂对几种致病性细菌的毒力至关重要，这些细菌每年总共杀死200多万人，主要是儿童和婴儿。对这些同源调节子的详细了解可能最终导致疾病治疗和预防的新应用，例如通过靶向保守毒力调节子的功能来减弱细菌毒力。由于Rns自身调节的独特功能，我们提出的研究也将提供关于转录及其调节的新的和根本上重要的信息。这些包括从下游结合位点的激活和非生产性RNAP开放（RPo）复合物对Prns的潜在抑制。在这个建议中的研究将确定Rns积极的自动调节的分子机制和Rns独立的，非生产性RPo复合物附近的Prns的生物学意义。