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: 10550156
• 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/10550156
• 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; Collaborations; Complement; Complex; Cone; Cryo-electron tomography; Cytoskeletal Filaments; Cytoskeletal Modeling; Cytoskeleton; Daughter; Disease; Eimeria tenella; Elements; Funding; Grant; Human; Invaded; Life Style; Malaria; Mediating; Medical; Membrane; Microtubules; Molecular; Motor; Names; Neospora caninum; Organelles; Parasites; Pattern; Peripheral; Plasmodium falciparum; Play; Process; Proteins; Proteome; Reporting; Role; Series; Shapes; Structure; Surgical sutures; System; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Vesicle; Work; 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; segregation; 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.

项目摘要 弓形虫Gondii和相关的Apicomplexan寄生虫包含一个专门的细胞器 膜复合物（IMC）在宿主细胞侵袭和子细胞形成中起重要作用。 IMC 由扁平的膜囊泡和支撑的细胞骨架网隔板组成。 寄生虫外围的膜和细胞微管。最近的体内生物素化（生物学） 研究揭示了IMC细胞器内的隔室化水平令人惊讶的水平，有不同的组 将蛋白隔离到锥形顶端帽，身体或基底络合物的蛋白质的蛋白质 和膜子组门。来自我们小组和其他人的分析已经暴露了新的基本功能 IMC的of，包括顶盖复合物AC9/ac10/erk7在安装蛋白固醇中的作用，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女儿芽蛋白IMC32，这是必不可少的 复制。在此续约中，我们将扩展这些研究，以确定这些研究如何和其他关键 IMC组件能够执行其功能。首先，我们将确定新型顶盖蛋白的作用 是AC9/AC10/ERK7复合物的推定相互作用者，并评估它们在顶端cap函数和 圆锥体组件。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起作用，并使用这些蛋白质进一步探索早期女儿 芽蛋白质组。最后，我们还将利用我们最近开发的光电反应性非天然氨基酸系统 作为冷冻电子层析成像，以确定如何组织寄生虫细胞骨架的临界肺泡素 充当细胞器的脚手架。该项目一起将促进对 弓形虫IMC的架构和功能。因为这个细胞器是寄生虫特异性的，并且不存在于其 人类宿主，确切地确定这些基本IMC组件的功能如何有望实现设计 针对T. gondii和其他Apicomplexan寄生虫的新型疗法。