Mechanisms of Enteropathogenic E. coli Effects on Polarity and Tight Junctions

肠病性大肠杆菌对极性和紧密连接影响的机制

• 批准号: 8542839
• 负责人: Gail A Hecht
• 金额: $ 31.69万
• 依托单位: LOYOLA UNIVERSITY CHICAGO
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-10 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8542839
• 关键词: Address; Bacterial Proteins; Binding; Biochemical; Biological Assay; Cell Line; Cell Polarity; Cell physiology; Cells; Complex; Coupled; Cyst; Cytoplasm; Data; Developing Countries; Development; Diarrhea; Disease; Dissociation; Endocytosis; Epithelial; Epithelial Cells; Escherichia coli EHEC; Escherichia coli Infections; Escherichia coli Proteins; Event; Genes; Goals; Health; Human; In Vitro; Infant; Infection; Injection of therapeutic agent; Intestinal Content; Intestines; Investigation; Lateral; Light; Maintenance; Maps; Membrane; Microscopic; Modeling; Molecular; Morbidity - disease rate; Morphogenesis; Movement; Mus; Pathogenesis; Physiological; Physiology; Process; Proteins; Public Health; Regulation; Research; Role; Signal Transduction; Sorting - Cell Movement; Structure; Techniques; Therapeutic Intervention; Tight Junctions; Transfection; Translations; Type III Secretion System Pathway; United States; Work; enteropathogenic Escherichia coli; fascinate; foodborne outbreak; holistic approach; in vivo; in vivo Model; intestinal epithelium; migration; mortality; nexin; novel; novel strategies; occludin; overexpression; p21 activated kinase; pathogen; prevent; protein complex; protein protein interaction

DESCRIPTION (provided by applicant): Enteropathogenic E. coli (EPEC) is an important human bacterial pathogen that infects primarily infants in developing countries causing significant morbidity and mortality from diarrhea. EPEC pathogenesis is dependent on the injection of several bacterial effector molecules into host cells by a type III secretion system. EPEC effectors have a multitude of host cell targets. While recent studies focused on single effector proteins have revealed intriguing biochemical data, they have not shed light on the mechanisms by which downstream intestinal physiology is perturbed. Several EPEC effectors contribute to tight junction (TJ) disruption but the mechanisms and cooperative interactions are not known. TJ formation and maintenance in intestinal epithelial cells is dependent on physical and functional interactions between the Par and Crumbs polarity complexes. EPEC effectors may interfere with this process. The long-term goal of this proposal is to enhance our understanding of how multiple EPEC effectors coordinate to perturb intestinal epithelial function, particularly TJ regulation. Recognition of the interdependent relationships between EPEC effectors underscores the need for novel and comprehensive investigative approaches to define the complex mechanisms underlying EPEC pathogenesis. The objective of this proposal is to elucidate the mechanisms by which specific EPEC effectors cooperate to perturb key host intestinal epithelial physiologic endpoints that underlie pathogenesis. The central hypothesis is that EPEC effectors target, in a coordinated fashion, the Par and Crumbs polarity complexes ultimately disrupting TJ structure and barrier function. In vitro infection and transfection models as well as in vivo models will be used to address the hypothesis. Molecular, cellular, microbiological, and physiological approaches will be coupled with state-of-the-art microscopic techniques to achieve the stated goals. The rationale for the proposed research is that the identification of mechanisms by which pathogenic bacterial proteins hijack host cell regulatory controls to alter physiology will guide the development of novel strategies aimed at preserving host functions. The realization that bacterial effectors cooperate highlights the need for expanded efforts toward holistic approaches that examine the interplay between multiple effectors in executing a common goal. The following Specific Aims will address the hypothesis: 1. Define the mechanisms by which Map, EspG, and EspF cooperate to disrupt Par polarity complex. 2. Determine if Map, EspG and EspF disrupt TJ structure/function by targeting cell polarity complexes. 3. Investigate the interdependence of key EPEC effector molecules on disruption of epithelial polarity complexes and TJs using a multicellular and in vivo model of infection.

描述（由申请人提供）：肠病性大肠杆菌（EPEC）是一种重要的人类细菌病原体，主要感染发展中国家的婴儿，导致腹泻的显着发病率和死亡率。 EPEC 发病机制依赖于 III 型分泌系统将多种细菌效应分子注射到宿主细胞中。 EPEC 效应器具有多种宿主细胞靶标。虽然最近针对单一效应蛋白的研究揭示了有趣的生化数据，但它们并没有阐明下游肠道生理学受到干扰的机制。几种 EPEC 效应器会导致紧密连接 (TJ) 破坏，但其机制和协同相互作用尚不清楚。肠上皮细胞中 TJ 的形成和维持取决于 Par 和 Crumbs 极性复合物之间的物理和功能相互作用。 EPEC 效应器可能会干扰此过程。该提案的长期目标是增强我们对多个 EPEC 效应器如何协调扰乱肠上皮功能，特别是 TJ 调节的理解。对 EPEC 效应器之间相互依赖关系的认识强调需要新颖且全面的研究方法来定义 EPEC 发病机制的复杂机制。该提案的目的是阐明特定 EPEC 效应器合作扰乱发病机制背后的关键宿主肠上皮生理终点的机制。核心假设是 EPEC 效应器以协调的方式靶向 Par 和 Crumbs 极性复合物，最终破坏 TJ 结构和屏障功能。体外感染和转染模型 以及体内模型将用于解决该假设。分子、细胞、微生物和生理学方法将与最先进的显微技术相结合，以实现既定目标。拟议研究的基本原理是，鉴定致病细菌蛋白劫持宿主细胞调节控制以改变生理学的机制将指导旨在保护宿主功能的新策略的开发。细菌效应器合作的认识凸显了需要扩大努力以采用整体方法来检查多个效应器在执行共同目标时之间的相互作用。以下具体目标将解决该假设： 1. 定义 Map、EspG 和 EspF 合作破坏 Par 极性复合体的机制。 2. 确定 Map、EspG 和 EspF 是否通过靶向细胞极性复合物破坏 TJ 结构/功能。 3. 使用多细胞和体内感染模型研究关键 EPEC 效应分子对上皮极性复合物和 TJ 破坏的相互依赖性。