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Lipid raft effects on Shigella type III secretion system-based interactions

脂筏对志贺氏菌 III 型分泌系统相互作用的影响

基本信息

批准号：
8686737
负责人：
Nicholas E Dickenson
金额：
$ 10.8万
依托单位：
UTAH STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-08-01 至 2016-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8686737
关键词：
Adherence Anti-Infective Agents Antibiotics Antigens Atomic Force Microscopy Bacillary Dysentery Biochemical Cell Communication Cell membrane Cell model Cell physiology Cell surface Cells Characteristics Cholesterol Complex Cultured Cells Cytoplasm Data Dependence Detergents Development Exhibits Film Fluorescence Fluorescence Microscopy Human Image Infection prevention Injection of therapeutic agent Intestines Investigation Lead Life Link Lipids Liposomes Liquid substance Membrane Membrane Lipids Membrane Microdomains Methods Microscopy Molecular Monitor Needles Phase Plasmids Play Process Proteins Real-Time Systems Recombinants Recruitment Activity Research Research Design Resistance Role Series Shigella Shigella Infections Shigella flexneri Signal Transduction Site Sphingolipids Sphingomyelins Structure Syringes System Techniques Testing Time Transmission Electron Microscopy Type III Secretion System Pathway Virulence Virulence Factors Work base bile salts improved insight macromolecular assembly membrane model novel pathogen prevent public health relevance sensor uptake

项目摘要

DESCRIPTION (provided by applicant): Shigella flexneri, the causative agent of bacillary dysentery, uses its type III secretion system (TTSS) as a conduit through which effector proteins are shuttled from bacterial to host cell cytoplasm where they subvert normal host functions. While much about TTSS function has been worked out, details on initial host interaction are not well understood. It has been shown that integral components of the type III secretion apparatus (TTSA) interact with cholesterol (Chol) and recent evidence indicates that liposomes containing Chol and sphingomyelin (Sm) induce TTS. These and other data suggest a role for lipid raft microdomains in Shigella contact, stable interaction, and effector injection into host cells, however, there has been no sustained attempts to directly explore the link between the pathogen and lipid raft-like domains. It is my hypothesis that lipid raft- like domains in human cells provide a site at which the Shigella TTSS can anchor to the surface of the cell while simultaneously supplying the signal that induces TTS and allows the injection of effector proteins into the host cell's cytoplasm. To better understand the role of lipid rafts in the virulece of Shigella, the specific aims of this investigation are to: 1) determine the role of host cell lipd raft components and membrane phases on S. flexneri initial attachment to and entry into host cells; and 2) determine the effects of lipid membrane dynamics on the interaction between live Shigella and model membranes. These studies will use a diverse array of techniques and approaches including: biochemical study of purified proteins using model membrane systems, real-time imaging of Shigella interaction with cultured cells, and developing methods for improving imaging of macromolecular assemblies. In completing these aims, I will provide the first comprehensive examination of this pathogen's microenvironment as it first encounters a host cell and the influence that lipids have on this process. This will provide insight into the roe of the TTSS as both an anchor on the host cell surface and as a signaling platform that is needed for host-pathogen crosstalk. The information obtained from these studies will provide basic information on the pathogenic mechanisms of Shigella TTS with likely implications with regard to other TTSS. This could eventually lead to the development of practical compounds or methods for preventing infections by TTSS-expressing human pathogen.

描述（由申请人提供）：福氏志贺菌是细菌性痢疾的病原体，其使用III型分泌系统（TTSS）作为通道，通过该通道，效应蛋白从细菌穿梭至宿主细胞质，在那里它们破坏正常的宿主功能。虽然关于TTSS的功能已经研究了很多，但关于初始宿主相互作用的细节还没有很好的理解。已经表明，III型分泌器（TTSA）的组成部分与胆固醇（Chol）相互作用，最近的证据表明，含有Chol和鞘磷脂（Sm）的脂质体诱导TTS。这些和其他数据表明脂筏微结构域在志贺氏菌接触、稳定相互作用和效应物注射到宿主细胞中的作用，然而，没有持续的尝试直接探索病原体和脂筏样结构域之间的联系。我的假设是人细胞中的脂筏样结构域提供了志贺氏菌TTSS可以锚到细胞表面的位点，同时提供诱导TTS的信号并允许效应蛋白注射到宿主细胞的细胞质中。为了更好地了解脂筏在志贺菌死亡过程中的作用，本研究的具体目的是：1）确定宿主细胞脂筏组分和膜相在志贺菌死亡过程中的作用;福氏菌初始附着和进入宿主细胞;和2）确定脂质膜动力学对活志贺氏菌和模型膜之间的相互作用的影响。这些研究将使用多种技术和方法，包括：使用模型膜系统对纯化蛋白质进行生物化学研究，志贺氏菌与培养细胞相互作用的实时成像，以及开发改善大分子组装成像的方法。在完成这些目标，我将提供这种病原体的微环境的第一次全面检查，因为它第一次遇到宿主细胞和脂质对这一过程的影响。这将提供对TTSS作为宿主细胞表面上的锚和作为宿主-病原体串扰所需的信号传导平台的作用的深入了解。从这些研究中获得的信息将提供有关志贺菌TTS致病机制的基本信息，并可能对其他TTSS产生影响。这可能最终导致开发用于预防表达TTSS的人类病原体感染的实用化合物或方法。