Elucidating the role of the Toxoplasma residual body in cytoskeleton turnover

阐明弓形虫残体在细胞骨架更新中的作用

• 批准号: 10591825
• 负责人: Michael Lloyd Reese
• 金额: $ 28.7万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-11-04 至 2024-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10591825
• 关键词: Actins; Address; Animals; Apical; Auxins; Biochemical; Biochemistry; Biology; Biophysics; Biotinylation; Cell Cycle; Cell membrane; Cell physiology; Cells; Complement; Complex; Complication; Coupled; Cryptosporidiosis; Cytokinesis; Cytoplasm; Cytoskeleton; Data; Daughter; Dependence; Disease; Enzymes; Fluorescence Microscopy; Foundations; Future; Garbage; Giant Cells; Goals; Growth; Human; Immunocompromised Host; In Vitro; Infection; Invaded; Knock-out; Length; Life; Lytic Phase; Malaria; Mass Spectrum Analysis; Mediating; Microtubules; Modeling; Mothers; Names; Organelles; Parasites; Pathogenesis; Phenotype; Planet Earth; Population; Protein Region; Proteins; Reaction; Recombinants; Research; Residual state; Ring Finger Domain; Role; Signal Transduction; Stimulus; Structure; Testing; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Ubiquitination; Vacuole; cell type; cost; daughter cell; experimental study; fitness; human disease; immunosuppressed; innovation; live cell imaging; loss of function; model organism; multicatalytic endopeptidase complex; mutant; pathogen; premature; prevent; programs; protein degradation; reconstitution; response; targeted treatment; tool; ubiquitin ligase; ubiquitin-protein ligase; virtual

PROJECT SUMMARY/ABSTRACT Toxoplasma gondii has the remarkable ability to infect virtually any cell type of almost all warm-blooded animals and is arguably the most successful parasite on earth, having infected an estimated one-third of humans globally. While initial infection typically resolves without complication, the parasite is able to persist for the life of its host, and can re-emerge in the immunocompromised and immunosuppressed to cause fatal disease. Toxoplasma, like other apicomplexan parasites, has an unusual cell cycle in which daughter cells are fully assembled within a mother before cytokinesis. Because of its unusual features and regulatory checkpoints distinct from mammalian hosts, the parasite cell cycle is an ideal target for therapy In the final steps of Toxoplasma division, the mother cell organelles and cytoskeleton are degraded to make room for the daughter cells. Because the daughter cells must insert their own cytoskeleton, including the host cell invasion machinery, into the plasma membrane, this is a critical point in parasite assembly. Once divided, the daughter cells remain connected through a poorly understood organelle called the residual body. While the residual body was first described over 60 years ago, little is know about it. We have found that maternal cytoskeleton is actively stripped from the plasma membrane and delivered to the residual body, where it is then degraded. We have identified a ubiquitin E3 ligase that localizes to the residual body, and is essential to the turnover of maternal cytoskeleton. Moreover, we have shown that mistargeting of the E3 outside of the residual body results in premature degradation of the daughter cells' invasion machinery, blocking the infective cycle. We have therefore identified the sequestration of protein turnover as a major cellular function for the residual body. The goal of the proposed studies is to (i) understand the functional role of the residual body in Toxoplasma cytokinesis and the turnover of maternal materials, and (ii) to delineate the components of the ubiquitination cascade that targets the parasite cytoskeleton. We will determine what other organelles and structures are delivered to the residual body for turnover and identify the proteins that sequester E3 ligase activity and cytoskeleton degradation within the residual body. Furthermore, we will build the components required for the creation of an innovative reconstitution of the E3 ligase activity that will enable future studies towards generating a mechanistic model of maternal cytoskeleton turnover.

项目摘要/摘要 弓形虫几乎可以感染几乎所有温血动物的任何细胞类型。 可以说是地球上最成功的寄生虫，已经感染了大约三分之一的 全球的人类。虽然最初的感染通常不会出现并发症，但寄生虫能够持续存在。 为其宿主的生命，并可再次出现在免疫功能受损和免疫抑制而导致致命 疾病。弓形虫和其他顶端复合体寄生虫一样，有一个不寻常的细胞周期，在这个周期中，子代细胞 在细胞质分裂前在母体内完全组装。因为它的不同寻常的特征和监管 有别于哺乳动物宿主的检查点，寄生虫的细胞周期是治疗的理想靶点 在弓形虫分裂的最后阶段，母细胞细胞器和细胞骨架被降解为 为子代细胞腾出空间。因为子代细胞必须插入自己的细胞骨架，包括 宿主细胞入侵机械，进入质膜，这是寄生虫组装的关键点。 一旦分裂，子细胞仍然通过一种鲜为人知的细胞器连接在一起，这种细胞器被称为 残存身体。虽然遗体最早是在60多年前被描述的，但人们对它知之甚少。我们有 发现母体细胞骨架主动地从质膜上剥离并输送到 残余体，在那里它被降解。我们已经确定了一个泛素E3连接酶，它定位于 残留体，对母体细胞骨架的周转是必不可少的。此外，我们已经表明， 错误地将E3定位在残留体之外会导致子代细胞的过早降解 入侵机器，阻断感染循环。因此，我们确定了蛋白质的封存 周转是残余体的一项主要细胞功能。 拟议研究的目标是(I)了解残余体在 弓形虫胞质分裂和母体物质的周转，以及(Ii)描述弓形虫胞质分裂的成分 以寄生虫细胞骨架为目标的泛素化级联反应。我们将确定还有哪些细胞器和 结构被运送到残余体进行周转，并识别隔离E3连接酶的蛋白质 残留体内的活性和细胞骨架的降解。此外，我们还将构建组件 创建E3连接酶活动的创新重组所需的，从而使未来 关于建立母体细胞骨架周转的机制模型的研究。