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）：细菌痢疾的病因志贺氏菌Flexneri使用其III型分泌系统（TTSS）作为导管，通过该导管，效应蛋白从细菌到宿主细胞质的效应蛋白都可以颠覆正常的宿主功能。尽管已经阐明了有关TTSS功能的许多有关TTSS功能的信息，但最初的主机互动的详细信息尚未很好地理解。已经表明，III型分泌设备（TTSA）的整体成分与胆固醇（CHOL）相互作用，并且最近的证据表明含有CHOL和鞘磷脂（SM）的脂质体诱导TTS。这些数据和其他数据表明，脂质筏微区域在志贺氏菌接触，稳定相互作用以及对宿主细胞的效应子注入中的作用，但是，没有持续的尝试直接探索病原体与脂质筏状结构域之间的联系。我的假设是，人类细胞中的脂质筏子像类似的结构域提供了一个位点，志贺氏菌TTS可以锚定在细胞的表面上，同时提供诱导TTS的信号并允许将效应蛋白注射到宿主细胞的细胞细胞质中。为了更好地理解脂质筏在志贺氏菌的染色中的作用，本研究的具体目的是：1）确定宿主细胞LIPD筏成分和膜相对于S. flexneri的初始附着和进入宿主细胞的作用； 2）确定脂质膜动力学对活志贺氏菌与模型膜之间相互作用的影响。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的生化研究，与培养细胞相互作用的实时成像，以及开发改善大分子组件成像的方法。在完成这些目标时，我将首先对这种病原体的微环境进行首次全面检查，因为它首先遇到了宿主细胞以及脂质对此过程的影响。这将洞悉TTSS的ROE，作为宿主细胞表面上的锚以及宿主 - 病原体串扰所需的信号平台。从这些研究中获得的信息将提供有关Shigella TTS的致病机制的基本信息，这可能与其他TTS有关。这最终可能导致开发实用化合物或通过表达TTSS病原体感染感染的方法。