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：
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
项目类别：
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

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. 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。由于对其蛋白质的了解有限，确定 IMC 的精确功能受到了阻碍成分，主要集中在为滑翔体提供动力的运动成分上寄生虫运动性和嵌入 IMC 细胞骨架网络中的肺泡蛋白家族细胞器的扁平膜囊泡。我们最近克服了这一知识差距，利用体内生物素化 (BioID) 可识别 40 多种新的 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室化水平，具有分离到顶盖、IMC体的细胞骨架或膜子区域的不同成分，或者最近发现的 IMC 缝合线，它将 IMC 膜组织成其特征矩形板。对其中一些蛋白质的分析揭示了在控制中新的重要作用寄生虫的形状、细胞分裂和入侵。在本提案中，我们将利用我们的新 IMC 蛋白群首先提供对这些组件如何参与动态的时间和空间理解内部出芽的过程称为内生作用。这将揭示发挥作用的蛋白质组在内生作用的特定步骤组装 IMC，并提供对内生作用动态过程的深入了解在分裂过程中区分母体和女儿的 IMC。接下来我们将探讨关键的IMC如何缝合蛋白质 ISC3 和 ISC6 调节适当的 IMC 膜生物发生、寄生虫形态和复制。最后，我们利用最近在弓形虫中进行的全基因组 CRISPR 筛选，重点关注我们新发现的一个子集。 IMC 蛋白可能在细胞器的不同亚细胞器区室中发挥关键作用。总之，这个项目将从根本上改变我们对组织和职能的理解。弓形虫 IMC。由于这种细胞器是寄生虫特异性的，并且不存在于其人类宿主中，因此阐明了关键的 IMC 组件及其各自的功能有望揭示新型设计的理想目标针对弓形虫和其他顶复门寄生虫的疗法。