CONTROL OF PROTECTIVE ANTIGEN SYNTHESIS BY B ANTHRACIS

要成为成功的病原体，细菌需要感知 宿主环境，处理环境更改，并进行 适当的适应。在许多细菌中，表达不同的 毒力因子由共同的调控因子协调控制 系统。病原菌炭疽芽孢杆菌的毒力表达 炭疽病，代表了一个独特的例子，监管机构对 环境信号。已知的炭疽杆菌的毒力因子 产生三方外毒素和D-谷氨酰多肽胶囊 只有在二氧化碳水平升高的情况下才能在体外生长 含重碳酸盐的介质。小苏打对毒素和细胞毒性的影响 胶囊的生产是特定的，而不仅仅是因为它的缓冲 细菌生长过程中的能力。这个二氧化碳/碳酸氢盐信号是 一种入侵哺乳动物宿主的病原体的生理学意义 纸巾。这个项目的长期目标是阐明 炭疽杆菌毒力基因表达的分子基础。毒素和 囊膜合成可能受反式作用因子的共同调节 对二氧化碳/碳酸氢盐的信号。 这里描述的实验将探索PAG的调节，PAG是一种 免疫原性毒素蛋白，保护性抗原。这篇文章的抄本 基因对重碳酸盐的反应增加了10到40倍。炭疽杆菌 PX01是刺激PAg转录所必需的。我们会 通过以下途径确定影响PAg表达的反式作用调控基因 转座子介导的插入诱变及pX01的筛选 激活PAG报告基因的克隆文库。A监管机构 已发现pX01上PAG上游至少3 kb的基因座 作为保护性抗原合成所必需的。这个轨迹和其他轨迹 在这里提出的实验中确定的调控基因座将是 其特征是定义结构限制、排序、确定开放 阅读框架，并构建零突变体。的影响。 二氧化碳/碳酸氢盐信号对调控基因功能和表达的影响 接受检查。将详细分析PAG启动子：上游 对二氧化碳依赖转录很重要的序列将被识别出来。 这些实验将阐明炭疽杆菌信号的成分- 负责识别和传输信号的转导系统 CO2/小苏打信号与毒力因子的可能全局调控 由这种有机体合成的。这项研究的结果将提高人们的认识 寄主-寄生虫关系和信号转导，并提供 与炭疽病发病机制相关的信息。

To be successful pathogens, bacteria require mechanisms for sensing the host environment, processing environmental changes, and making appropriate adaptations. In many bacteria, expression of disparate virulence factors is coordinately controlled by a common regulatory system. Virulence expression by Bacillus anthracis, the causative agent of anthrax, represents a unique example of a regulatory response to an environmental signal. The known virulence factors of B. anthracis, a tripartite exotoxin and a D-glutamyl polypeptide capsule, are produced in vitro only during growth under elevated levels of carbon dioxide in media containing bicarbonate. The effect of bicarbonate on toxin and capsule production is specific, and not simply due to its buffering capacity during bacterial growth. This CO2/bicarbonate signal is of physiological significance for a pathogen which invades mammalian host tissues. the long-term goal of this project is to elucidate the molecular basis for virulence gene expression in B. anthracis. Toxin and capsule synthesis may be co-regulated by trans-acting factors in response to the CO2/bicarbonate signal. Experiments described here will probe the regulation of pag, the gene for the immunogenic toxin protein, protective antigen. Transcription of this gene increases 10 to 40-fold in response to bicarbonate. B. anthracis plasmid pX01 is required for stimulation of pag transcription. We will identify trans-acting regulatory genes that affect expression of pag by using transposon-mediated insertional mutagenesis and by screening a pX01 library for clones which activate a pag reporter gene. A regulatory locus at least 3 kb upstream from pag on pX01 has already been implicated as necessary for protective antigen synthesis. This locus, and other regulatory loci identified in experiments proposed here, will be characterized by defining structural limits, sequencing, identifying open reading frames, and constructing null mutants. Effects of the CO2/bicarbonate signal on regulatory gene function and expression will be examined. The pag promoter will be analyzed in detail: upstream sequences important for CO2-dependent transcription will be identified. These experiments will elucidate components of the B. anthracis signal- transduction system responsible for recognition and transmission of the CO2/bicarbonate signal and possibly global control of virulence factor synthesis by this organism. Results of this study will enhance awareness of host-parasite relationships and signal transduction and provide information relevant to the pathogenesis of anthrax disease.