Functional characterization of Chlamydia trachomatis inclusion membrane proteins and their role in subversion of host vesicular trafficking

沙眼衣原体包涵膜蛋白的功能特征及其在破坏宿主囊泡运输中的作用

• 批准号: 10363758
• 负责人: Mary Weber
• 金额: $ 44.45万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10363758
• 关键词: Address; Animals; Antibiotics; Bacteria; Bacterial Infections; Bacterial Proteins; Binding; Blindness; Cell Culture Techniques; Cells; Ceramides; Chlamydia; Chlamydia Infections; Chlamydia trachomatis; Complex; Data; Defect; Development; Disease; Ectopic Pregnancy; Endosomes; Etiology; Genetic; Goals; Golgi Apparatus; Growth; Impairment; Incidence; Infection; Infertility; Integration Host Factors; Iron; Knowledge; Label; Lipids; Mediating; Membrane; Membrane Proteins; Metabolic; Microtubules; Molecular; Nutrient; Organelles; Paper; Pathogenesis; Pathway interactions; Pelvic Inflammatory Disease; Positioning Attribute; Prevalence; Process; Proteins; Recurrence; Recycling; Reporting; Role; SNAP receptor; Sexual Transmission; Sexually Transmitted Diseases; Source; Techniques; Therapeutic Intervention; Trachoma; Transferrin; Vacuole; Vesicle; Work; base; genetic manipulation; global health; insight; new therapeutic target; novel therapeutic intervention; pathogen; rab GTP-Binding Proteins; recruit; trafficking; trans-Golgi Network; vesicle transport

Project Summary Chlamydia trachomatis (C.t.) is the leading cause of non-congenital blindness and the most prevalent sexually transmitted bacterial infection in the world and thus, has a major impact on global health. C.t. replicates in a membrane-bound vacuole, termed the inclusion, that is extensively modified early in infection through the incorporation of over 50 type III secreted effector proteins termed inclusion membrane proteins (Incs). From within the confines of the inclusion, the bacterium must engage select host organelles to obtain key nutrients such as lipids and iron for bacterial replication and inclusion expansion. Despite the importance of nutrient acquisition to chlamydial infection, a major gap in our knowledge exists regarding how C.t. acquires these essential substrates from the host cell. Recent work from our group shows that the Inc protein CT229 binds and recruits Rab GTPases and specific Rab effectors to the inclusion. Using cutting-edge genetics, we showed that the absence of CT229 results in defects in intracellular replication and inclusion development. We hypothesize that CT229-Rab interactions direct post-Golgi vesicles and recycling endosomes to the inclusion for the acquisition of essential substrates needed for proliferation and inclusion expansion. In Aim 1, we will determine how CT229-Rab interactions promote lipid acquisition by hijacking host vesicular trafficking pathways and iron acquisition by promoting the recruitment of transferrin-positive vesicles to the inclusion. We will evaluate the lipid composition of the bacterial and inclusion membranes and evaluate the importance of lipid acquisition for incorporation into the inclusion and bacterial membranes. We will also answer a major long-standing question in the field by determining whether transferrin indeed serves as a major source of iron for chlamydia and furthermore mechanistically probe the mechanisms by which chlamydia may extract iron from this pathway. It is unlikely that CT229 alone controls vesicle hijacking from start to finish, therefore in Aim 2 we will identify the complex of host and bacterial proteins that are formed at the inclusion during CT229-Rab interactions. Using cellular, genetic, and molecular techniques we will evaluate the impact of Inc-Inc interactions on Rab-dependent vesicle transport to the inclusion. Detailed characterization of the bacterial and host proteins that co-opt host vesicular trafficking during C.t. infection will provide a holistic view of how intracellular pathogens coordinate the capture and fusion of host vesicles that are necessary for bacterial infection.

项目摘要 衣原体沙眼（C.T.）是非占否盲目的主要原因，也是最普遍的性行为 世界上传播的细菌感染对全球健康有重大影响。 C.T.在A中复制 膜结合的液泡称为包容性，在感染早期通过 掺入50多种III型分泌效应子蛋白称为纳入膜蛋白（INCS）。从 在包含的范围内，细菌必须接合选择的宿主细胞器以获取关键营养素 例如脂质和铁，用于细菌复制和纳入膨胀。尽管营养很重要 获得衣原体感染，我们的知识中存在有关如何C.T.的一个主要差距获取这些 来自宿主单元的必需底物。我们小组的最新工作表明，INC蛋白CT229结合并 招募RAB GTPases和特定的RAB效应子来包含。使用尖端的遗传学，我们表明 CT229的缺失会导致细胞内复制和包容性发育的缺陷。我们假设 CT229-RAB相互作用将高尔基囊泡和回收内体直接到包含 获取增殖和包容性扩展所需的必需底物。在AIM 1中，我们将确定 CT229-RAB相互作用如何通过劫持宿主的囊泡贩运途径和铁来促进脂质的获取 通过促进转铁蛋白阳性囊泡的募集来获取。我们将评估脂质 细菌和包含膜的组成，并评估脂质采集的重要性 掺入包含和细菌膜中。我们还将回答一个主要的长期问题 通过确定转铁蛋白是否确实是衣原体铁的主要来源和 此外，机械探测衣原体可以从该途径中提取铁的机制。这是 仅CT229就不太可能控制囊泡从头到尾劫持，因此在AIM 2中，我们将确定 宿主和细菌蛋白的复合物在CT229-RAB相互作用期间形成。使用 细胞，遗传和分子技术我们将评估Inc-Inc相互作用对RAB依赖性的影响 囊泡转运到包容性。对宿主的细菌和宿主蛋白的详细表征 C.T.期间的囊泡贩运感染将为细胞内病原体如何坐标提供整体视图 捕获和融合细菌感染所需的宿主囊泡。