Exposporium determinants of Bacillus anthracis

描述(申请人提供)：炭疽杆菌是一种形成内孢子的土壤微生物，是一种重要的人畜共患病病原体。作为潜在的生物恐怖主义制剂，它引起了极大的关注。孢子是这种病原体的感染形式。孢子进入人类宿主的途径决定了由此产生的疾病的性质，是皮肤病、胃肠道病还是肺病。胃肠道和肺部炭疽病与高死亡率有关。炭疽杆菌的孢子有一个外层，称为外孢子体。这个气球状的外层包含孢子表面分子，这些分子构成了与宿主天然免疫系统最初相互作用的场所。外孢子体上覆盖着一层毛发状的绒毛层，主要由胶原样糖蛋白BCLA组成。这种外孢子菌蛋白与吞噬细胞上的受体结合，促进孢子的吸收，从而启动感染过程。虽然它在感染的初始阶段起着重要的作用，但人们对外孢子层的组成、组装过程和功能知之甚少。我们对芽胞形成过程的许多了解都是通过对枯草芽孢杆菌的生化和遗传学研究来了解的，枯草杆菌是一种模式细菌，其孢子缺乏外孢子体。 我们建议用遗传学和生物化学的方法来研究炭疽杆菌的外孢子菌组成。在我的实验室开发的转座子突变系统已经产生了一系列潜在的外孢子体、间隙和外孢子皮突变体。这些突变体将在这个项目中被描述出来。更多的外孢子蛋白将通过蛋白质组学方法和在产孢子细胞中与已知外孢子菌蛋白形成复合体的蛋白质鉴定来鉴定。除了鉴定外孢子菌基因及其产物外，鉴定孢子形成细胞中较大复合体中存在的蛋白质将使我们能够开始确定哪些蛋白质在孢子形成过程中相互作用，我们可以开始绘制外孢子菌组装过程中蛋白质的顺序图。该项目将有助于更好地了解炭疽杆菌的内孢子成熟，并将确定可能被证明对未来疫苗或治疗开发有用的孢子靶标。此外，包括蜡状芽孢杆菌、产气荚膜梭菌和艰难梭菌在内的许多形成内孢子的人类病原体都具有外孢子，对炭疽杆菌外孢子的更详细的了解可能会为这些重要的人类病原体提供孢子生物学方面的见解。 与公共卫生相关：炭疽芽孢杆菌是一种芽胞形成细菌，可引起炭疽病，是一种重要的潜在生物制剂。孢子是细菌的感染形式，也是炭疽感染过程中最初宿主相互作用的中心。该项目将利用遗传和生化方法来表征孢子的最外层--外孢子体。这项工作将提供可能对了解这种重要病原体的生物学很重要的信息，并可能确定针对这种微生物的疫苗、疗法或诊断的未来发展目标。

DESCRIPTION (provided by applicant); Bacillus anthracis is an endospore-forming, soil microorganism that is an important zoonotic pathogen. It is of significant concern as a potential bioterrorism agent. The spore is the infective form of this pathogen. The route by which the spore gains access to a human host defines the nature of the resulting disease, cutaneous, gastrointestinal, or pulmonary. Gastrointestinal and pulmonary anthrax are associated with high mortality rates. The B. anthracis spore possesses an outer layer referred to as the exosporium. This balloon-like outer layer contains the spore surface molecules that constitute the site of initial interactions with the host innate immune system. The exosporium is covered with a hair-like nap layer made up principally by the collagen-like glycoprotein BclA. This exosporium protein binds to receptors on phagocytic cells to promote uptake of the spores and which initiates the infectious process. Although it plays an important role in the initial stages of infection, very little is known regarding the composition, assembly process, and function of the exosporium layer. Much of what we know regarding the sporulation process was learned through biochemical and genetic studies of Bacillus subtilis, a model bacterium whose spores lack an exosporium. We propose to study exosporium composition in B. anthracis with a genetic and biochemical focused approach. A transposon mutagenesis system developed in my laboratory has produced a collection of potential exosporium, interspace, and outer spore coat mutants. These mutants will be characterized in this project. Additional outer spore proteins will be identified by a proteomic approach and by identification of proteins which form complexes with known exosporium proteins in sporulating cells. In addition to identifying exosporium genes and their products, identification of proteins present in larger complexes in sporulating cells will permit u to begin to determine which proteins interact in the sporulation process and we can begin to map the order of proteins in the exosporium assembly process. This project will lead to a better understanding of endospore maturation in B. anthracis and will identify spore targets that may prove useful for future vaccine or therapeutic development. In addition, a number of endospore-forming human pathogens possess exosporia, including Bacillus cereus, Clostridium perfringens, and Clostridium difficile and a more detailed understanding of the exosporium of B. anthracis may provide insights on spore biology for these other important human pathogens. PUBLIC HEALTH RELEVANCE: Bacillus anthracis is a spore-forming bacterium that causes anthrax and is important as a potential biothreat agent. The spore is the infectious form of the bacterium and is the center of the initial host interactions in the anthrax infectious process. Thi project will make use of genetic and biochemical approaches to characterize the exosporium, the outermost layer of the spore. The work will provide information that may be important in understanding the biology of this important pathogen and may identify targets for the future development of vaccines, therapeutics, or diagnostics against this organism.