Gene Expression and Manipulation of Coxiella Burnetii

简介-贝氏柯克斯体是Q热的病原体，是一种潜在的生物武器和选择剂， 已知最具传染性的病原体之一然而，关于这一义务的报告很少。 细胞内因子分子发病机制或发育周期，以及缺乏定点系统 基因操纵严重阻碍了研究进展。为了解决信息的缺乏问题， 为了改进目前的分子生物学技术，我们建议：1）检测表达谱 小细胞变异（SCV）和大细胞变异（LCV）的发育周期，以确定基因和 使Coxiella能够在环境中长时间存活（SCV阶段）， 随后在宿主细胞吞噬溶酶体的不适宜的范围内生长（LCV阶段）; 2）分析和 比较体外与体内培养的柯克斯体以及I期与II期的表达谱 生物体，以确定潜在的毒力决定因素，以及3）开发定点诱变系统 对于这种生物体，允许常规遗传操作和未来开发减毒 疫苗株。这些数据有望为柯克斯体的生命周期和环境保护提供有价值的信息。 各自的基因和蛋白质参与，并应导致破坏细胞发育的策略， 控制Q热。最后，柯克斯体的常规遗传操作系统是一个重要的分子生物学系统。 生物学工具，用于检查可疑的毒力基因，并在体内执行分子科赫假设。 项目互动-项目3（Harmsen）将与项目1（本研究）互动，因为它涉及活体培养 小鼠模型中的柯克斯体。项目4（Pascual）和项目2（Jutila）将受益于项目1的数据 阶段特异性基因产物; SCV特异性基因/抗原被假设参与 传播/早期感染和LCV特异性基因/抗原参与细胞内复制。

Introduction- Coxiella burnetii is the etiologic agent of Q fever, a potential bioweapon and select agent, and one of the most infectious pathogens known. However, there are few reports concerning this obligate intracellular agent's molecular pathogenesis or developmental cycle, and lack of a system for site-directed genetic manipulation has severely hampered research progress. To address this dearth of information and to improve the current state of molecular biology technology, we propose to: 1) examine the expression profiles of small cell variants (SCVs) and large cell variants (LCVs) of the developmental cycle to identify genes and proteins that allow Coxiella to survive for extended periods of time in the environment (SCV stage) and to subsequently grow in the inhospitable confines of the host cell phagolysosome (LCV stage); 2) analyze and compare expression profiles of in vitro versus in vivo-cultivated Coxiella and phase I versus phase II organisms to identify potential virulence determinants, and 3) develop a system of site-directed mutagenesis for this organism to allow for the routine genetic manipulation and future development of an attenuated vaccine strain. These data are expected to provide valuable information on the life cycle of Coxiella and the respective genes and proteins involved, and should lead to strategies for disrupting cellular development to control Q fever. Finally, a system for routine genetic manipulation of Coxiella is an essential molecular biology tool for examining suspected virulence genes and performing molecular Koch's postulates in vivo. Project Interactions- Project 3 (Harmsen) will interact with Project 1 (this study) as it relates to in vivocultivated Coxiella from the mouse model. Projects 4 (Pascual) and 2 (Jutila) will benefit from Project 1 data on stage-specific gene products; SCV-specific genes / antigens are hypothesized to be involved in transmission / early infection and LCV-specific genes / antigens are involved in intracellular replication.