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: 9384311
• 负责人: Peter John Bradley
• 金额: $ 37.89万
• 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 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9384311
• 关键词: Animals; Apical; Biology; Biotin; Biotinylation; CRISPR screen; Cell membrane; Cells; Cellular biology; Characteristics; Coccidiosis; Complex; Cone; Cytoskeletal Filaments; Cytoskeleton; Daughter; Disease; Eimeria tenella; Elements; Event; Falciparum Malaria; Family; Human; Immobilization; In Vitro; Infection; Invaded; Knock-out; Knowledge; Lead; Ligase; Mass Spectrum Analysis; Mediating; Medical; Membrane; Methods; Microtubules; Morphology; Motor; Names; Neospora caninum; Organelles; Parasites; Peripheral; Play; Process; Proteins; Proteome; Reporting; Role; Series; Shapes; Streptavidin; Surgical sutures; System; TimeLine; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Vesicle; Virulence; Xenopus oocyte; cell motility; choline transporter; cohort; daughter cell; design; experimental study; fitness; genome-wide; in vivo; insight; member; membrane biogenesis; novel; novel therapeutics; obligate intracellular parasite; parasite invasion; pathogen; protein complex; protein function; scaffold

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. Determining the precise functions of the IMC has been hampered by a limited understanding of its protein constituents, which have predominantly focused on the motor components of the glideosome that powers parasite motility and the family of alveolins that are embedded within the IMC cytoskeletal network underlying the flattened membrane vesicles of the organelle. We have recently overcome this gap in our knowledge using in vivo biotinylation (BioID) to identify over forty new IMC components, more than doubling the known IMC proteome. These new proteins have revealed a surprising level of compartmentalization within the organelle, with distinct components segregating to the cytoskeletal or membrane subregions of the apical cap, IMC body, or the recently discovered IMC sutures, which organize the IMC membranes into their characteristic rectangular plates. Analyses of a few of these proteins have revealed new important roles in controlling parasite shape, cellular division, and invasion. In this proposal, we will leverage our new cohort of IMC proteins to first provide a temporal and spatial understanding of how these components participate in the dynamic process of internal budding known as endodyogeny. This will reveal groups of proteins that function to assemble the IMC at specific steps of endodyogeny and also provide insight into the dynamic process of distinguishing maternal and daughter IMCs during division. We will then explore how the key IMC suture proteins ISC3 and ISC6 regulate proper IMC membrane biogenesis, parasite morphology and replication. Finally, we exploit the recent genome-wide CRISPR screen in Toxoplasma to focus on a subset of our new IMC proteins that are likely to play crucial roles within distinct suborganellar compartments of the organelle. Together, this project will fundamentally transform our understanding of the organization and function of the Toxoplasma IMC. As this organelle is parasite-specific and is not present in its human host, elucidating critical IMC components and their respective functions promises to reveal ideal targets for 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。 由于对IMC蛋白质的了解有限，IMC的精确功能一直受到阻碍 主要集中在为胶质小体提供动力的运动组件上 寄生虫的运动性和嵌入IMC细胞骨架网络的肺泡蛋白家族 细胞器的扁平的膜小泡。我们最近用我们的知识克服了这一鸿沟 体内生物素化(BioID)识别40多个新的IMC成分，是已知IMC的两倍多 蛋白质组。这些新蛋白质揭示了细胞器内令人惊讶的区隔程度， 不同的成分分离到顶帽的细胞骨架或膜亚区，IMC小体， 或者是最近发现的IMC缝合线，它将IMC膜组织成它们的特征 矩形板材。对其中几种蛋白质的分析揭示了新的重要调控作用 寄生虫的形状、细胞分裂和入侵。在这项提案中，我们将利用我们新的IMC蛋白质队列 首先提供对这些组件如何参与动态的时空理解 内部萌发的过程称为内生作用。这将揭示一组蛋白质，它们的功能是 在内生的特定步骤组装IMC，并提供对 在分裂过程中区分母女IMC。然后我们将探索关键的IMC缝合是如何 ISC3和ISC6蛋白调节适当的IMC膜生物发生、寄生虫的形态和复制。 最后，我们利用最近在弓形虫基因组范围内的CRISPR屏幕来关注我们新的 IMC蛋白可能在细胞器不同的亚细胞器隔间中发挥关键作用。 总而言之，这个项目将从根本上改变我们对世界银行组织和职能的理解 弓形虫IMC。由于这种细胞器是寄生虫特有的，不存在于其人类宿主中，因此阐明了关键 IMC组件及其各自的功能有望为小说的设计揭示理想的目标 针对弓形虫和其他尖端复合体寄生虫的治疗。