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

• 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
• 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

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.

项目总结 弓形虫和相关的顶端复合体寄生虫含有一种特殊的细胞器，称为内细胞器 膜复合体(IMC)，在宿主细胞入侵和子代细胞形成中起重要作用。国际管理信息中心 由扁平的膜小泡和支撑的细胞骨架网组成，两侧是血浆 寄生虫周围的膜和膜下微管。最新体内生物素化(BioID) 研究表明，IMC细胞器内的区隔程度令人惊讶，具有不同的群组 分离到锥形顶帽、身体或两个细胞骨架的基底层复合体的蛋白质 和膜小室。我们小组和其他人的分析揭示了新的基本功能 包括顶帽复合体AC9/AC10/ERK7在安装圆锥体中的作用，这是必不可少的 对于细胞器的分泌和入侵以及保守的早期子芽蛋白IMC32是必不可少的 用于复制。在这次更新中，我们将对这些研究进行扩展，以准确地确定这些和其他关键 IMC组件能够执行其功能。首先，我们将确定新的顶帽蛋白的作用。 它们可能是AC9/AC10/ERK7复合体的相互作用因子，并评估它们在心尖帽功能和 圆锥面组件。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女儿 芽蛋白质组。最后，我们还将开发我们最近开发的光反应非天然氨基酸系统。 作为冷冻电子断层扫描，以确定寄生虫细胞骨架的关键肺泡蛋白是如何组织起来的 作为细胞器的脚手架。总而言之，这个项目将促进对 弓形虫整合细胞的结构和功能。因为这个细胞器是寄生虫特有的，不存在于它的 以人为本，准确地确定这些基本的IMC组件如何工作，有望实现设计 针对弓形虫和其他尖端复合体寄生虫的新疗法。