Functional analysis of the Francisella capBCA gene locus

弗朗西斯菌 capBCA 基因位点的功能分析

• 批准号: 7712743
• 负责人: JING-REN ZHANG
• 金额: $ 23.55万
• 依托单位: ALBANY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7712743
• 关键词: Amino Acids; Anthrax disease; Antigens; Bacillus anthracis; Bacteria; Bacterial Genes; Biology; Bioterrorism; Categories; Cells; Data; Disease; Francisella; Francisella tularensis; Genes; Glutamic Acid; Goals; Gram-Negative Bacteria; Growth; Human; Infection; Knowledge; Licensing; Molecular; Molecular Target; Mutagenesis; Mutation; Pathogenesis; Polymerase; Polymers; Property; Proteins; Research Design; Structure; Surface; System; Testing; Therapeutic; Ticks; Tularemia; Vaccines; Virulence; Virulence Factors; base; capsule; fitness; genome-wide; in vivo; macrophage; mortality; pathogen; public health relevance; tool

DESCRIPTION (provided by applicant): Francisella tularensis, the causative agent of tularemia, is a Gram-negative facultative intracellular bacterium. Because of its extreme infectivity, high virulence, and ease of dissemination, F. tularensis is considered as a category A potential agent of bioterrorism. The virulence mechanisms of F. tularensis are poorly understood, and there are no licensed vaccines against tularemia. Our long-term goals are: (i) to understand at the molecular level the pathogenic mechanisms of F. tularensis; and (ii) identify molecular targets for effective vaccines and other therapeutics against F. tularensis infections in humans. This application is based on our recent genome-wide search for virulence determinants in F. tularensis strain LVS by signature-tagged mutagenesis (STM). This study identified 95 F. tularensis genes that are required for the in vivo survival of this pathogen. Among these are three adjacent genes (capB, capC, and capA) in a gene locus. The capBCA locus closely resembles the biosynthetic gene locus of the poly-3-glutamic acid (PGA) capsule in Bacillus anthracis, the anthrax agent. The B. anthracis capsule is a major virulence factor. However, PGA has not been described in any Gram-negative bacteria or any intracellular bacterial pathogen. Based on our exciting preliminary findings, we hypothesize that the F. tularensis capBCA locus promotes the virulence of F. tularensis by producing PGA. The Specific Aims are: (i) to characterize the F. tularensis capBCA locus, (ii) to determine PGA polymerase activity of the capBCA locus, and (iii) to determine how capB enhances F. tularensis intracellular growth and virulence. Our expertise and existing tools have provided us the unique capability to accomplish the goal of this application. We will be the first to study a potential PGA biosynthetic system in Gram-negative bacteria. The data of the proposed studies will provide valuable information concerning the significance of PGA in biology and pathogenesis of Gram-negative bacteria including F. tularensis. The PGA may be an attractive vaccine target for tularemia if it proves to be surface-exposed like its counterparts in the Gram-positive pathogens. PUBLIC HEALTH RELEVANCE: The bacterium Francisella tularensis is the causative agent of tularemia and also a bioweapon agent due to its extreme infectivity and ability to cause mortality. There are no licensed vaccines against tularemia. F. tularensis can infect a wide range of host from ticks to humans. However, the mechanisms for this extreme versatility are unknown. This project will study three bacterial genes that are potentially responsible for synthesis of an unconventional amino acid polymer called poly-glutamic acid (PGA). As the sole constituent of the capsule (coating structure) of Bacillus anthracis, causative agent of anthrax, PGA protects the bacterium from host clearance during infection of mammalian hosts. We will pursue our hypothesis that F. tularensis also produces PGA to enhance its adaptation when the bacterium is located in different microenvironments. The information will significantly enhance our knowledge in disease pathogenesis of F. tularensis. PGA can be used to develop vaccines against F. tularensis infection if it is placed at the surface of F. tularensis as its counterpart in B. anthracis

描述（由申请人提供）：土拉热弗朗西斯菌是土拉热病的病原体，是一种革兰氏阴性兼性细胞内细菌。由于其极高的传染性、高毒力和易于传播的特点，图拉拉弗朗西斯被认为是 A 类潜在的生物恐怖分子。人们对土拉菌的毒力机制知之甚少，并且没有针对土拉菌病的许可疫苗。我们的长期目标是：（i）在分子水平上了解土拉弗拉菌的致病机制； (ii) 确定针对人类土拉弗朗西斯虫感染的有效疫苗和其他疗法的分子靶标。该应用基于我们最近通过特征标记诱变 (STM) 对 F. tularensis 菌株 LVS 中的毒力决定簇进行全基因组搜索。这项研究鉴定了 95 个土拉弗拉菌基因，这些基因是该病原体在体内生存所必需的。其中有一个基因座中的三个相邻基因（capB、capC 和 capA）。 capBCA 基因座与炭疽杆菌（炭疽病原）中聚 3-谷氨酸 (PGA) 荚膜的生物合成基因座非常相似。炭疽芽孢杆菌荚膜是主要毒力因子。然而，尚未在任何革兰氏阴性细菌或任何细胞内细菌病原体中描述过 PGA。基于我们令人兴奋的初步发现，我们假设土拉弗朗西斯 capBCA 基因座通过产生 PGA 来促进土拉弗朗西斯的毒力。具体目标是：(i) 表征土拉杆菌 capBCA 基因座，(ii) 确定 capBCA 基因座的 PGA 聚合酶活性，以及​​ (iii) 确定 capB 如何增强土拉杆菌 capBCA 细胞内生长和毒力。我们的专业知识和现有工具为我们提供了实现此应用程序目标的独特能力。我们将成为第一个研究革兰氏阴性细菌中潜在的 PGA 生物合成系统的人。拟议研究的数据将为 PGA 在包括土拉杆菌在内的革兰氏阴性菌的生物学和发病机制中的重要性提供有价值的信息。如果 PGA 被证明像革兰氏阳性病原体中的对应物一样是表面暴露的，那么它可能是兔热病的一个有吸引力的疫苗靶点。公共卫生相关性：土拉弗朗西斯菌是兔热病的病原体，由于其极强的传染性和导致死亡的能力，它也是一种生物武器。目前还没有针对兔热病的许可疫苗。土拉热蜱可以感染从蜱虫到人类的多种宿主。然而，这种极端多功能性的机制尚不清楚。该项目将研究三种细菌基因，它们可能负责合成一种称为聚谷氨酸（PGA）的非常规氨基酸聚合物。作为炭疽杆菌（炭疽病的病原体）荚膜（涂层结构）的唯一成分，PGA 可以保护细菌在哺乳动物宿主感染期间免遭宿主清除。我们将继续我们的假设，即当细菌位于不同的微环境中时，土拉弗朗西斯菌也会产生 PGA 以增强其适应能力。这些信息将显着增强我们对土拉弗拉菌疾病发病机制的了解。如果将 PGA 放置在土拉杆菌表面（与炭疽杆菌中的对应物一样），则可用于开发针对土拉杆菌感染的疫苗