Regulation of hemolysin genes in Bacillus

芽孢杆菌溶血素基因的调控

• 批准号: 7073689
• 负责人: MICHIKO M NAKANO
• 金额: $ 7.66万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-15 至 2008-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7073689
• 关键词: Bacillus; gene mutation; genes; hemolysin

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.

描述（由申请人提供）：对炭疽病病原体炭疽杆菌毒力因子的研究主要集中在质粒编码的荚膜以及致死毒素和水肿毒素上。关于成孔溶血素的研究报道很少。研究有限的主要原因是炭疽芽孢杆菌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，炭疽杆菌在一定条件下可以产生溶血素。另一种非溶血性芽孢杆菌枯草芽孢杆菌也携带假定的溶血素基因。该研究计划的长期目标是调查溶血素是否在炭疽杆菌的致病性中发挥作用，例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响应环境变化以及调节基因的单一突变而诱导的。 “潜在”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) 通过基因组作图和 DNA 序列分析来识别激活枯草芽孢杆菌中 β-溶血素基因的调节突变。 2)通过使用与溶血素基因启动子的转录lacZ融合来表征通过调节突变和氧限制的α-溶血素基因的诱导机制。 3)研究硝酸盐对枯草芽孢杆菌α-溶血素基因无氧表达和炭疽芽孢杆菌需氧/厌氧表达的刺激作用。硝酸盐依赖性诱导需要硝酸盐还原酶活性的假设将通过使用枯草芽孢杆菌硝酸盐还原酶突变体并测量需氧炭疽芽孢杆菌培养物的硝酸盐还原酶活性来检验。将编码 CRP 家族推定转录调节因子的炭疽芽孢杆菌基因引入枯草芽孢杆菌中，以确定推定调节因子是否能够在有氧条件下激活呼吸硝酸还原酶操纵子的表达，从而导致枯草芽孢杆菌中 β-溶血素的有氧诱导。