Rab protein functions that impact Coxiella intracellular replication

Rab 蛋白功能影响柯克斯体细胞内复制

• 批准号: 8970333
• 负责人: Craig R. Roy
• 金额: $ 24.98万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-15 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8970333
• 关键词: Affect; Anti-Bacterial Agents; Bacteria; Biogenesis; Biological; Cell physiology; Cells; Complex; Coxiella; Coxiella burnetii; Cytosol; Data; Defect; Development; Disease; Environment; Genes; Goals; Gram-Negative Bacteria; Guanosine Triphosphate Phosphohydrolases; Host Defense; Human; Infection; Integration Host Factors; Lead; Lipids; Lysosomes; Measurable; Membrane Fusion; Molecular; Organelles; Pathway interactions; Play; Process; Property; Proteins; Q Fever; Research Project Grants; Resources; Role; Small Interfering RNA; Staging; System; Testing; Therapeutic; Transmembrane Transport; Transport Process; Type IV Secretion System Pathway; Vacuole; Vesicle Transport Pathway; cohort; genome-wide; human disease; insight; interdisciplinary approach; mutant; novel; novel strategies; pathogen; protein function; public health relevance; rab GTP-Binding Proteins; spatiotemporal

 DESCRIPTION (provided by applicant): Coxiella burnetii is a Gram-negative bacterium and causative agent of the human disease Q fever. This intracellular pathogen has evolved complex mechanisms for evading host cell defenses. One important feature that makes C. burnetii unique is that it survives within the acidic and hydrolytic environment of host lysosomes and modifies this organelle to create a large vacuole that supports replication. To accomplish this, C. burnetii encodes a Dot/Icm type IV secretion system that delivers over 100 effector proteins into the cytosol of the host cell during infection. These effectors modulate host cell functions, which include manipulating host vesicular transport pathways to promote membrane fusion and expansion of the vacuole containing C. burnetii. Host determinants required for biogenesis and fusion of the C. burnetii-containing vacuole (CCV) have not been clearly defined; however, because Rab GTPases coordinate vesicular transport within host cells, and are often modulated by bacterial effector proteins, we hypothesize that Rab proteins are central to the infection strategy used by C. burnetii. This hypothesis is supported by data showing that the silencing of specific Rab proteins in host cells will interfere or enhance C. burnetii replication. Here, we propose studies to determine the spatiotemporal roles for Rab GTPases during C. burnetii infection, and to determine the Rab-directed processes that are required to transform a lysosome-derived vacuole into a unique intracellular compartment that supports C. burnetii replication. We will disrupt the function of specific Rab proteins and determine the stage of CCV maturation that is regulated by the Rab. We will also define the cohort of Rab proteins that localize to the CCV during these specific maturation stages. Together, this will reveal the Rab proteins that are required for C. burnetii transport to a lysosome-derived vacuole, acidification o the vacuole, activation of the Dot/Icm secretion system, and the import of host factors required for bacterial replication. We will also focus on Rab proteins that negatively impact C. burnetii intracellular replication by determining how these Rab proteins affect CCV biogenesis and intracellular survival of C. burnetii and testing whether C. burnetii effector proteins required fo intracellular infection have activities that affect the function of these Rab proteins. These studis will elucidate host membrane transport pathways that are used to create the intracellular environment that supports C. burnetii replication and will identify Rab proteins that have important roles in defending host cells against intracellular pathogens.

 描述（由申请方提供）：贝氏柯克斯体是一种革兰氏阴性细菌，是人类Q热疾病的病原体。这种细胞内病原体已经进化出复杂的机制来逃避宿主细胞的防御。一个重要的特点，使C。Burnetii的独特之处在于它在宿主溶酶体的酸性和水解环境中存活，并修饰该细胞器以产生支持复制的大液泡。为了实现这一点，C。 贝氏体编码Dot/Icm IV型分泌系统，其在感染期间将超过100种效应蛋白递送到宿主细胞的胞质溶胶中。这些效应子调节宿主细胞功能，包括操纵宿主囊泡运输途径以促进膜融合和含C.伯内特氏菌宿主决定因子是该菌生物发生和融合所必需的。然而，由于Rab GTP酶在宿主细胞内协调囊泡运输，并且经常受到细菌效应蛋白的调节，我们假设Rab蛋白是C.伯内特氏菌这一假设得到了数据的支持，这些数据表明，宿主细胞中特定Rab蛋白的沉默将干扰或增强C。贝氏复制在这里，我们提出的研究，以确定时空作用的拉布GTP酶在C。Burnettii感染，并确定将溶酶体衍生的空泡转化为支持C.贝氏复制我们将破坏特定Rab蛋白的功能，并确定由Rab调节的CCV成熟阶段。我们还将定义在这些特定成熟阶段定位于CCV的Rab蛋白的队列。总之，这将揭示C.贝氏体转运至溶酶体衍生的液泡、液泡酸化、Dot/Icm分泌系统的激活以及细菌复制所需的宿主因子的输入。我们还将关注对C产生负面影响的Rab蛋白。通过测定这些Rab蛋白如何影响CCV的生物发生和CCV的细胞内存活，研究了CCV在Burnetii细胞内的复制。burnetii和测试C.胞内感染所需的贝氏体效应蛋白具有影响这些Rab蛋白功能的活性。这些研究将阐明宿主细胞膜转运途径，这些途径用于创造支持C。Burnettii复制，并将鉴定在防御宿主细胞对抗细胞内病原体中具有重要作用的Rab蛋白。