The Role of Toxoplasma Rhoptries in Host Cell Infection

弓形虫在宿主细胞感染中的作用

基本信息

批准号：
8792359
负责人：
Peter John Bradley
金额：
$ 37.79万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-12-15 至 2016-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8792359
关键词：
Animals Apicomplexa Architecture Bacterial Adhesins Binding Biological Models Cell membrane Cells Cellular biology Cessation of life Chemicals Coccidia Coccidiosis Complex Coupled Development Disease Eimeria tenella Elements Event Face Falciparum Malaria Funding Grant Human Immunocompromised Host In Vitro Individual Infection Integral Membrane Protein Invaded Knock-out Link Macromolecular Complexes Malaria Maps Mass Spectrum Analysis Mediating Medical Membrane Methods Molecular Motor Names Neck Neospora caninum Organelles Organism Parasites Pathway interactions Plasmodium falciparum Play Population Process Proteins Role Shapes Site Structure Surface Therapeutic Intervention Tight Junctions Toxoplasma Toxoplasma gondii Toxoplasmosis Vacuole Virulence Work Yeasts crosslink design driving force gel electrophoresis human MST1R protein in vivo insight member molecular sieving neonate novel novel therapeutics obligate intracellular parasite parasite invasion pathogen protein complex rhoptry scaffold stoichiometry tool yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): Toxoplasma gondii is an obligate intracellular parasite that causes severe disease in immunocompromised individuals and congenitally infected neonates. Toxoplasma also serves as a model system for the study of related apicomplexan parasites including Plasmodium falciparum, the causative agent of malaria. Intracellular survival of these organisms is critically dependent on the ability of the parasite to actively invade their host cell, establish a replication-permissive vacuole, and avoid host cell defenses. Apicomplexan parasites share a unique mechanism for invasion that involves the formation of a tight junction between the invading parasite and the host cell called the moving junction (MJ). The MJ is believed to form a stable anchor for the parasite to invade the host cell and also serve as a molecular sieve that modifies the nascent vacuole to render it non-fusogenic with the host endocytic pathway. The Toxoplasma MJ consists of micronemal AMA1 on the parasite's surface connected to a macromolecular complex of rhoptry neck proteins (RONs 2/4/5/8) that are injected into the host cell. While RON2 spans the host membrane and establishes the link to AMA1, the remaining members of the complex are surprisingly on the cytoplasmic face of the host membrane and how they function in invasion is largely unknown. We have disrupted the coccidial-specific component RON8 and shown that while not essential, this protein plays an important role in parasite invasion in vitro and in virulence in vivo. As the remaining members are conserved in the Apicomplexa and believed to be essential, this indicates that apicomplexans contain a conserved core complex that is required for invasion, which is enhanced in the coccidia via RON8. This is supported by our recent development of a conditional knockout for RON5, which shows that this protein plays a critical role in assembly of the complex and that the MJ complex is indeed essential for invasion. Our objectives in this first renewal application are to conduct an in depth functional analysis of the MJ complex and determine its architecture. Specifically, we will first focus on RON8 to determine how this component enhances invasion and links the complex to the host cell. We will then exploit the conditional knockout of RON5 to study its role in the organization and function of the MJ complex. Lastly, we will explore the architecture of the complex by determining its stoichiometry and identifying key interactions of its component proteins. These studies will open completely new insight into the mechanism by which apicomplexan parasites use this novel invasion machine to infect their mammalian hosts and cause disease.

描述（由申请人提供）：弓形虫Gondii是一种强制性细胞内寄生虫，可在免疫功能低下的个体和先天感染的新生儿中引起严重疾病。弓形虫还可以作为研究相关的丙糖寄生虫（包括恶性疟原虫）的模型系统，疟疾是疟疾的致病药物。这些生物的细胞内存活至关重要取决于寄生虫积极入侵其宿主细胞，建立复制 - 腐蚀性液泡并避免宿主细胞防御能力的能力。 Apicomplexan寄生虫具有一种独特的入侵机制，涉及入侵寄生虫与称为移动结（MJ）的宿主细胞之间形成紧密的连接。据信，MJ形成了寄生虫侵入宿主细胞的稳定锚，并作为一种分子筛，可修饰新生的液泡，以使其与宿主内吞途径相关。弓形虫MJ由寄生虫的表面上的微生物AMA1组成，该表面与注入宿主细胞的大型颈部蛋白（Rons 2/4/5/8）的大分子复合物（RONS 2/4/5/8）组成。尽管RON2跨越宿主膜并建立了与AMA1的联系，但该综合体的其余成员出奇地位于宿主膜的细胞质面上，以及它们在入侵中的作用在很大程度上是未知的。我们已经破坏了球球菌特异性成分RON8，并表明，尽管不是必不可少的，但该蛋白质在体外和体内毒力中起着重要作用。由于其余成员在apicomplexa中是保守的，并且认为这是必不可少的，因此这表明Apicomplexans包含一种保守的核心复合物，这是入侵所需的，这可以通过RON8在球球菌中增强。我们最近开发了RON5的有条件敲除，这表明该蛋白在复合物的组装中起着至关重要的作用，并且MJ复合物确实对于侵袭至关重要。我们的目标在第一个更新应用将对MJ复合物进行深入的功能分析，并确定其架构。具体而言，我们将首先关注RON8，以确定该组件如何增强侵袭并将复合物与宿主细胞联系起来。然后，我们将利用RON5的条件敲除以研究其在MJ复合物的组织和功能中的作用。最后，我们将通过确定其化学计量并识别其组分蛋白的关键相互作用来探索该复合物的体系结构。这些研究将对Apicomplexan寄生虫使用这种新型入侵机感染其哺乳动物宿主并引起疾病的机制开辟全新的见解。