Coxiella secreted proteins mediating inter-organelle membrane contact sites

柯克斯体分泌的蛋白质介导细胞器间膜接触位点

• 批准号: 10575434
• 负责人: STACEY D GILK
• 金额: $ 23.03万
• 依托单位: UNIVERSITY OF NEBRASKA 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/10575434
• 关键词: Acute; Address; Antibiotic Therapy; Antibiotics; Bacteria; Bacterial Infections; Bacterial Proteins; Binding; Binding Proteins; Bioinformatics; Biological Assay; Cell physiology; Cells; Cessation of life; Cholesterol; Cholesterol Homeostasis; Chronic; Complement; Coxiella; Coxiella burnetii; Data; Development; Disease Outbreaks; Ectopic Expression; Endocarditis; Endoplasmic Reticulum; Etiology; Eukaryotic Cell; Family; Family member; Fatigue; Fluorescence; Funding; Future; Genetic Complementation Test; Growth; Health; Homeostasis; Human; Immune response; Individual; Infection; Lipids; Location; Macrophage; Maintenance; Measures; Mediating; Membrane; Modeling; Molecular; Molecular Structure; Molecular Target; Netherlands; Organelles; Pathogenesis; Patient Noncompliance; Phagolysosome; Play; Protein Family; Protein Secretion; Proteins; Publishing; Q Fever; Regimen; Research; Role; Site; Symptoms; System; Testing; Therapeutic Intervention; Vacuole; Virulence Factors; chronic infection; experimental study; flu; insight; lipid metabolism; member; mutant; novel; pathogen; therapeutic target; yeast two hybrid system

PROJECT SUMMARY/ABSTRACT Coxiella burnetii is an obligate intracellular bacterium and the etiological agent of Q fever. Inside the host cell, the bacterium survives in a phagolysosome-like vacuole called the Coxiella Containing Vacuole (CCV). Coxiella is uniquely sensitive to host cholesterol levels, where elevated cholesterol, especially in the CCV and endolysosomal system, leads to bacterial death. While Coxiella directly regulates host cholesterol homeostasis through effector proteins secreted by the Coxiella Type 4B Secretory System (T4BSS), the T4BSS effector proteins responsible for manipulating host cell cholesterol have not yet been identified. Using a bioinformatic approach, we identified a new family of Coxiella T4BSS effector proteins which contain a eukaryotic FFAT motif. In eukaryotic cells, FFAT proteins bind to the VAP protein family on the endoplasmic reticulum (ER) and mediate membrane contact sites (MCS) between cell organelles and ER. These inter-organelle MCS play a critical role in regulating lipid homeostasis. The proposed experiments will test our hypothesis that Coxiella FFAT-containing T4BSS effector proteins mediate MCS between the ER and other host organelles in order to manipulate host cholesterol metabolism. Aim 1 will establish which members of the Coxiella FFAT protein family bind to VAP proteins, and determine where these proteins localize in the host cell. Aim 2 will test whether these proteins are essential for Coxiella infection, as well as whether they function in modulating host cholesterol. Completion of these studies will not only reveal a new strategy utilized by Coxiella to modulate host lipid metabolism, but significantly add to our knowledge on how pathogen-secreted proteins manipulate the host cell.

项目概要/摘要 伯内特柯克斯体是一种专性细胞内细菌，也是 Q 热的病原体。在宿主细胞内， 该细菌在称为含有柯克斯体的液泡（CCV）的吞噬溶酶体样液泡中生存。柯克斯体 对宿主胆固醇水平特别敏感，其中胆固醇升高，尤其是 CCV 和 内溶酶体系统，导致细菌死亡。而柯克斯体直接调节宿主胆固醇稳态 通过 Coxiella 4B 型分泌系统 (T4BSS) 分泌的效应蛋白，T4BSS 效应子 负责操纵宿主细胞胆固醇的蛋白质尚未确定。使用生物信息学 通过这种方法，我们鉴定了一个新的 Coxiella T4BSS 效应蛋白家族，其中包含真核 FFAT 基序。 在真核细胞中，FFAT 蛋白与内质网 (ER) 上的 VAP 蛋白家族结合并介导 细胞器和内质网之间的膜接触位点（MCS）。这些细胞器间 MCS 发挥着关键作用 调节脂质稳态。所提出的实验将检验我们的假设，即含有 FFAT 的 Coxiella T4BSS 效应蛋白介导 ER 和其他宿主细胞器之间的 MCS，以操纵宿主 胆固醇代谢。目标 1 将确定 Coxiella FFAT 蛋白家族的哪些成员与 VAP 结合 蛋白质，并确定这些蛋白质在宿主细胞中的定位位置。目标 2 将测试这些蛋白质是否 对于柯克斯体感染至关重要，以及它们是否具有调节宿主胆固醇的功能。完成 这些研究不仅揭示了柯克斯体用来调节宿主脂质代谢的新策略，而且 显着增加了我们对病原体分泌蛋白如何操纵宿主细胞的了解。