Suppression of innate immunity by an ADP-ribosyltransferase type III effector

ADP-核糖基转移酶 III 型效应子对先天免疫的抑制

基本信息

批准号：
7538341
负责人：
JAMES Robert ALFANO
金额：
$ 36.33万
依托单位：
UNIVERSITY OF NEBRASKA LINCOLN
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-12-15 至 2012-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7538341
关键词：
ADP Ribose Transferases ADP ribosylation Active Sites Affect Affinity Animals Arabidopsis Area Bacterial Infections Bacterial Toxins Binding Biochemical Biological Biological Assay Chloroplasts Cholera Toxin Data Disease Eukaryota Eukaryotic Cell Genes Glycine Goals Gram-Negative Bacteria Immune response Immune system Immunity Immunologic Surveillance In Vitro Infection Link Literature Mammals Mass Spectrum Analysis Metabolism Microbe Mission Modeling Molecular Mono(ADP-Ribose) Transferases Natural Immunity Outcome Output Pathogenesis Pattern recognition receptor Plant Components Plant Proteins Plants Play Positioning Attribute Predisposition Protein Secretion Proteins Proteomics Pseudomonas syringae Qualifying RNA RNA-Binding Proteins Reporting Research Research Personnel Resources Role System Tomatoes Toxin Type III Epithelial Receptor Cell Type III Secretion System Pathway United States National Institutes of Health Virulence Work base cost efficient disorder control experience improved innovation interest microorganism mutant novel pathogen plant glycine-rich RNA-binding protein protein function research study response

项目摘要

DESCRIPTION (provided by applicant): The eukaryotic innate immune system represents an important barrier that pathogens need to circumvent in order to cause disease. Several components of this system are conserved in eukaryotes. Recently, bacterial pathogen effectors that are injected into host cells by type III protein secretion systems (TTSSs) have been shown to be capable of suppressing innate immunity in eukaryotes. The bacterial plant pathogen Pseudomonas syringae is dependent on a TTSS to cause disease on plants. The P. s. pv. tomato DC3000 effector gene hopU1 resembles ADP ribosyltransferases (ADP-RTs) genes. These genes encode some of the best understood toxins in bacterial pathogens of animals (e. g., cholera toxin). Preliminary data within this proposal show that HopU1 is an active ADP-RT and that it ADP-riboslylates several plant proteins. Mass spectrometry determined that chloroplast and glycine-rich RNA-binding proteins acted as in vitro substrates for HopU1. These are novel substrates for ADP-RTs. Moreover, HopU1 has the ability to suppress several responses of the plant innate immune system in a manner that is dependent on its ADP-RT active site. An Arabidopsis mutant lacking one HopU1 substrate, AtGRP7, displayed enhanced susceptibility to P. syringae suggesting that it is a component of innate immunity. AtGRP7 is a glycine-rich RNA-binding protein, which suggests this pathogen targets proteins involved in RNA metabolism to suppress innate immunity. The central hypothesis of the proposed experiments is that AtGRP7 and perhaps other targets of the HopU1 ADP-RT type III effector are components of innate immunity. Several of the experiments seek to elucidate the role AtGRP7 plays in innate immunity using biochemical and molecular biological approaches. The P. syringae-Arabidopsis pathosystem is an excellent model to study the innate immune system because of the resources available, the similarities between innate immune systems between eukaryotes, and the cost efficient research. These experiments will contribute to a fundamental understanding of the molecular mechanism of bacterial pathogenesis and innate immunity. The Specific Aims are the following: (1) Determine the molecular consequence of ADP- ribosylation on the function of AtGRP7 and elucidate the role this protein plays in innate immunity; (2) Identify additional substrates of HopU1 and verify their involvement in innate immunity; (3) Analyze the affect that HopU1 has on host-microbe interactions. Project Narrative: Identifying the eukaryotic targets for the P. syringae HopU1 ADP-ribosyltransferase will contribute to our understanding of bacterial pathogenesis and will likely reveal important components of the innate immune system. One HopU1 target belongs to a large group of proteins called glycine-rich RNA binding proteins, which are not well understood, and this research will likely increase our understanding of these proteins. Because there are considerable similarities between the innate immune systems in plants and mammals we expect that our findings will be relevant to the mission of the NIH and be broadly interesting to researchers studying molecular mechanisms of bacterial pathogenesis and innate immunity.

描述（由申请人提供）：真核先天免疫系统代表了病原体需要绕过才能引起疾病的重要屏障。该系统的几个组件在真核生物中是保守的。最近，通过 III 型蛋白分泌系统 (TTSS) 注入宿主细胞的细菌病原体效应子已被证明能够抑制真核生物的先天免疫。细菌性植物病原体丁香假单胞菌依赖于 TTSS 来引起植物疾病。 P.s。光伏。番茄 DC3000 效应基因 hopU1 类似于 ADP 核糖基转移酶 (ADP-RT) 基因。这些基因编码动物细菌病原体中一些最容易理解的毒素（例如霍乱毒素）。该提案中的初步数据表明，HopU1 是一种活性 ADP-RT，并且它可以 ADP 核糖基化多种植物蛋白。质谱分析确定叶绿体和富含甘氨酸的 RNA 结合蛋白是 HopU1 的体外底物。这些是 ADP-RT 的新型底物。此外，HopU1 能够以依赖于其 ADP-RT 活性位点的方式抑制植物先天免疫系统的多种反应。缺乏HopU1底物AtGRP7的拟南芥突变体表现出对丁香假单胞菌的敏感性增强，表明它是先天免疫的一个组成部分。 AtGRP7 是一种富含甘氨酸的 RNA 结合蛋白，这表明该病原体以参与 RNA 代谢的蛋白质为目标，抑制先天免疫。所提出的实验的中心假设是 AtGRP7 以及可能的 HopU1 ADP-RT III 型效应子的其他靶标是先天免疫的组成部分。一些实验试图利用生化和分子生物学方法阐明 AtGRP7 在先天免疫中的作用。由于可用资源、真核生物之间先天免疫系统之间的相似性以及具有成本效益的研究，丁香假单胞菌-拟南芥病理系统是研究先天免疫系统的绝佳模型。这些实验将有助于对细菌发病机制和先天免疫的分子机制的基本理解。具体目标如下：（1）确定ADP-核糖基化对AtGRP7功能的分子影响，并阐明该蛋白在先天免疫中的作用； (2) 鉴定HopU1的其他底物并验证它们在先天免疫中的参与； (3)分析HopU1对宿主-微生物相互作用的影响。项目叙述：确定丁香假单胞菌 HopU1 ADP-核糖基转移酶的真核靶点将有助于我们了解细菌发病机制，并可能揭示先天免疫系统的重要组成部分。一个 HopU1 靶标属于一大类称为富含甘氨酸的 RNA 结合蛋白的蛋白质，目前人们对这些蛋白质的了解还不是很清楚，而这项研究可能会增加我们对这些蛋白质的了解。由于植物和哺乳动物的先天免疫系统之间存在相当大的相似性，我们预计我们的发现将与 NIH 的使命相关，并对研究细菌发病机制和先天免疫的分子机制的研究人员产生广泛的兴趣。