Reprogramming of host cell cholesterol homeostasis by Coxiella burnetii

伯氏柯克斯体对宿主细胞胆固醇稳态的重编程

• 批准号: 9016852
• 负责人: STACEY D GILK
• 金额: $ 23.08万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-12-16 至 2017-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9016852
• 关键词: Acute; Address; Aerosols; Alveolar Macrophages; Bacteria; Biogenesis; Cells; Cholesterol; Cholesterol Homeostasis; Chronic; Complex; Coxiella; Coxiella burnetii; Cytoplasm; Disease; Ectopic Expression; Emerging Communicable Diseases; Endocarditis; Gene Library; Genes; Goals; Growth; Homeostasis; Human; Infection; Integration Host Factors; Libraries; Life; Link; Lipids; Maintenance; Mammalian Cell; Membrane; Microbiology; Modeling; Molecular Profiling; Nutrient; Organelles; Pathogenesis; Pathway interactions; Patients; Pharmacologic Substance; Phenotype; Play; Proteins; Q Fever; Role; Signal Pathway; Signal Transduction; Source; Sterols; System; Testing; Therapeutic Intervention; Type IV Secretion System Pathway; Vacuole; Work; base; flu; inhibitor/antagonist; knock-down; macrophage; monocyte; mutant; novel; pathogen; public health relevance; research study; screening; tissue culture; trafficking; transcriptome; transcriptome sequencing

 DESCRIPTION (provided by applicant): Coxiella burnetii is the causative agent of human Q fever, an emerging infectious disease and a leading cause of culture-negative endocarditis. An obligate intracellular bacterial pathogen spread through aerosols, Coxiella primarily targets alveolar macrophages during natural infection. Once inside the cell, Coxiella manipulates host cell machinery to promote the biogenesis of a specialized compartment called the parasitophorous vacuole (PV). While the PV is central to Coxiella pathogenesis, the mechanisms behind PV biogenesis and maintenance are poorly understood. The Coxiella PV is sterol-rich, and pharmaceutical inhibitors that perturb host cell cholesterol inhibit PV formation and subsequent bacterial growth. Despite increasing evidence that cholesterol plays a critical role in PV formation and Coxiella-host interactions, there is very little information on either the bacterial or host factors involved. The objective of this application is to identify and characterie bacterial-driven changes in host cell cholesterol homeostasis. This proposal will test the hypothesis that Coxiella proteins, secreted into the host cell cytoplasm through the bacteria's specialized Type IV Secretion System (T4SS), manipulate host cholesterol homeostasis as a mechanism to support intracellular bacterial growth and survival. In the first specific aim, host cholesterol homeostasis pathways targeted by Coxiella T4SS effector proteins will be identified through transcriptome analysis of infected alveolar macrophages. This study will identify key host cell genes differentially regulated by Coxiella. Following their identification, the host pathways will be disrupted and the effect on Coxiella survival determined. In the second aim, Coxiella proteins that manipulate host cholesterol will be characterized. This will be accomplished by screening for Coxiella transposon mutants that have lost the ability to alter host cell cholesterol homeostasis. The identified Coxiella proteins will be characterized by analyzing the phenotype of the isolated mutants, as well as functional analysis of the corresponding protein. The proposed work will identify both host and pathogen proteins involved in cholesterol homeostasis during Coxiella infection, opening new avenues for therapeutic intervention.

 描述（由申请人提供）：伯内特柯克斯体是人类 Q 热的病原体，这是一种新出现的传染病，也是培养阴性心内膜炎的主要原因。柯克斯体是通过气溶胶传播的专性细菌细胞内病原体，在自然感染期间主要针对肺泡巨噬细胞。一旦进入细胞，柯克斯体就会操纵宿主细胞机制来促进称为寄生液泡（PV）的特殊隔室的生物发生。作为柯克斯体发病机制的核心，人们对柯克斯体 PV 的生物发生和维持机制知之甚少，而扰乱宿主细胞胆固醇的药物抑制剂会抑制 PV 的形成和随后的细菌生长。 PV 的形成和 Coxiella 与宿主的相互作用，关于这两者的信息很少 该应用的目的是识别和表征细菌驱动的宿主细胞胆固醇稳态变化，该提案将检验柯克斯体蛋白通过细菌专门的 IV 型分泌系统分泌到宿主细胞细胞质中的假设。 （T4SS），宿主胆固醇稳态作为支持细胞内细菌生长和存活的机制在第一个具体目标中，将通过转录组分析来鉴定柯克斯体 T4SS 效应蛋白靶向的宿主胆固醇家庭操纵途径。本研究将鉴定受柯克斯体差异调节的关键宿主细胞基因，并确定其对柯克斯体存活的影响。在第二个目标中，将鉴定操纵宿主胆固醇的柯克斯体蛋白。这将通过筛选失去改变宿主细胞胆固醇稳态能力的柯克斯体转座子突变体来完成，将通过分析分离突变体的表型以及功能分析来表征鉴定的柯克斯体蛋白。拟议的工作将鉴定在柯克斯体感染期间参与胆固醇稳态的宿主和病原体蛋白，为治疗干预开辟新途径。