Functional Analysis of Novel Components of the Toxoplasma Inner Membrane Complex

弓形虫内膜复合物新成分的功能分析

基本信息

批准号：
10444432
负责人：
Peter John Bradley
金额：
$ 44.88万
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：
2017
资助国家：
美国
起止时间：
2017-08-01 至 2026-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10444432
关键词：
Ablation Acylation Affect Alveolar Alveolus Animals Apical Apicomplexa Architecture Binding Biotinylation C-terminal Cell membrane Cells Cellular biology Coccidiosis Coiled-Coil Domain Complement Complex Cone Cryo-electron tomography Cytoskeletal Filaments Cytoskeleton Daughter Disease Eimeria tenella Elements Falciparum Malaria Funding Grant Human Invaded Lead Life Style Mediating Medical Membrane Microtubules Molecular Motor Names Neospora caninum Organelles Parasites Pattern Peripheral Play Process Proteins Proteome Reporting Role Series Structure Surgical sutures System Toxoplasma Toxoplasma gondii Toxoplasmosis Vesicle Work base cell motility crosslink daughter cell design human pathogen in vivo knock-down knockout gene member new therapeutic target novel novel therapeutics obligate intracellular parasite pathogen protein complex protein function scaffold unnatural amino acids

项目摘要

PROJECT SUMMARY Toxoplasma gondii and related apicomplexan parasites contain a specialized organelle called the inner membrane complex (IMC) that plays essential roles in host cell invasion and daughter cell formation. The IMC consists of flattened membrane vesicles and a supporting cytoskeletal meshwork that is flanked by the plasma membrane and subpellicular microtubules at the periphery of the parasite. Recent in vivo biotinylation (BioID) studies have revealed a surprising level of compartmentalization within the IMC organelle, with distinct groups of proteins segregating to the cone-shaped apical cap, the body, or to the basal complex of both the cytoskeletal and membrane subcompartments. Analyses from our group and others have exposed new essential functions of the IMC including the role of the apical cap complex AC9/AC10/ERK7 in mounting the conoid which is essential for organelle secretion and invasion as well as the conserved early daughter bud protein IMC32 that is essential for replication. In this renewal, we will expand on these studies to determine precisely how these and other critical IMC components are able to carry out their functions. First, we will determine the role of novel apical cap proteins that are putative interactors of the AC9/AC10/ERK7 complex and assess their role in apical cap function and conoid assembly. We will then explore how IMC32 collaborates with the newly discovered partner IMC48 to function at the earliest stages of parasite division and use these proteins to further explore the early daughter bud proteome. Finally, we will exploit our recently developed photoreactive unnatural amino acid system as well as cryo-electron tomography to determine how critical alveolins of the parasite cytoskeleton are organized to serve as scaffolds for the organelle. Together, this project will promote a much deeper understanding of the architecture and function of the Toxoplasma IMC. As this organelle is parasite-specific and not present in its human host, determining precisely how these essential IMC components function promises to enable the design of novel therapies against T. gondii and other apicomplexan parasites.

项目概要弓形虫和相关的顶复门寄生虫含有一种特殊的细胞器，称为内部细胞器。膜复合物（IMC）在宿主细胞入侵和子细胞形成中发挥重要作用。法团校际委员会由扁平膜囊泡和两侧是血浆的支撑细胞骨架网络组成寄生虫周围的膜和膜下微管。最近的体内生物素化（BioID）研究揭示了 IMC 细胞器内令人惊讶的划分水平，具有不同的组蛋白质分离到锥形顶盖、主体或细胞骨架的基底复合体和膜子室。我们小组和其他人的分析揭示了新的基本功能 IMC 的功能，包括顶盖复合体 AC9/AC10/ERK7 在安装圆锥体中的作用，这一点至关重要对于细胞器分泌和侵袭以及保守的早期子芽蛋白 IMC32 至关重要用于复制。在本次更新中，我们将扩展这些研究，以确定这些研究和其他关键研究如何准确地 IMC 组件能够执行其功能。首先，我们将确定新型顶帽蛋白的作用是 AC9/AC10/ERK7 复合体的假定相互作用因子，并评估它们在心尖帽功能中的作用，圆锥体装配。然后我们将探讨 IMC32 如何与新发现的合作伙伴 IMC48 合作，在寄生虫分裂的最早阶段发挥作用，并利用这些蛋白质进一步探索早期子体芽蛋白质组。最后，我们还将利用我们最近开发的光反应性非天然氨基酸系统作为冷冻电子断层扫描来确定寄生虫细胞骨架的关键肺泡是如何组织的作为细胞器的支架。总之，这个项目将促进人们更深入地了解弓形虫 IMC 的结构和功能。由于这种细胞器是寄生虫特异性的并且不存在于其体内人类宿主，精确确定这些重要的 IMC 组件如何发挥作用，以实现设计针对弓形虫和其他顶复门寄生虫的新疗法。