Regulation of hemolysin genes in Bacillus

描述（由申请方提供）：炭疽芽孢杆菌（炭疽的病原体）的毒力因子研究主要集中在质粒编码的荚膜以及致死和水肿毒素。很少有关于孔形成溶血素的研究报道。研究有限的一个主要原因是B。炭疽通常被认为是一种非溶血性病原体，尽管溶血素基因存在于基因组中。然而，最近的研究表明，B。炭疽菌在一定条件下能产生溶血素。另一种非溶血性芽孢杆菌也携带溶血素基因。该研究计划的长期目标是调查溶血素是否在B的致病性中发挥作用。炭疽病，如成功地从巨噬细胞逃逸到细胞外环境。拟议的研究旨在了解溶血素等毒力因子是如何因环境变化和调节基因的单一突变而被诱导的。“潜伏”毒力基因的激活可能对各种细菌感染产生强烈影响。该提案的具体目标是：1）识别激活？B中的溶血素基因。通过基因组作图和DNA序列分析，2)为了表征？-通过调节突变和通过氧限制使用与溶血素基因的启动子的转录lacZ融合来修饰溶血素基因。3)研究硝酸盐对？-溶血素基因在B。枯草芽孢杆菌中的需氧/厌氧表达和在B.炭疽病硝酸盐依赖性诱导需要硝酸还原酶活性的假设将通过使用B来检验。枯草杆菌硝酸还原酶突变体和测定好氧B的硝酸还原酶活性。炭疽菌培养物B。编码CRP家族推定转录调节因子的炭疽基因将被引入B中。枯草杆菌，以确定是否推定的调节能够激活呼吸硝酸还原酶操纵子在有氧条件下的表达，从而导致有氧诱导？溶血素在B中。枯草杆菌。

DESCRIPTION (provided by applicant): Studies of virulence factors of Bacillus anthracis, the causative agent of anthrax, have mainly focused on plasmid-encoded capsule as well as lethal and edema toxins. Very few studies have been reported on pore-forming hemolysins. A primary reason for the limited studies is that B. anthracis is generally accepted as a nonhemolytic pathogen, although hemolysin genes are present in the genome. However, recent studies have shown that B. anthracis can produce hemolysin under certain conditions. Another nonhemolytic sporeformer Bacillus subtilis also carries putative hemolysin genes. The long-term objectives of the research plan are to investigate whether hemolysin plays a role in pathogenicity of B. anthracis such as successful escape from macrophage into extracellular milieu. The proposed studies will aim at understanding how virulence factors such as hemolysins are induced in response to environmental changes and as a result of a single mutation in regulatory genes. The activation of 'latent' virulence genes could have a strong impact on various bacterial infections. The specific aims of the proposal are: 1) To identify regulatory mutation(s) that activate ?-hemolysin gene(s) in B. subtilis by genome mapping and DNA sequence analysis. 2) To characterize the Induction mechanism of the ?-hemolysin gene by the regulatory mutation and by oxygen limitation using a transcriptional lacZ fusion to the promoter of the hemolysin gene. 3) To investigate the stimulatory effect of nitrate on anaerobic expression of ?-hemolysin gene in B. subtilis and on aerobic/anaerobic expression in B. anthracis. The hypothesis that nitrate-dependent induction requires nitrate reductase activity will be examined by using a B. subtilis nitrate reductase mutant and measuring nitrate reductase activity of aerobic B. anthracis cultures. B. anthracis gene(s) encoding putative transcriptional regulator(s) of the CRP-family will be introduced into B. subtilis to determine whether the putative regulator is able to activate expression of the respiratory nitrate reductase operon under aerobic conditions, and thus leading to the aerobic induction of ?-hemolysin in B. subtilis.