Vetter Sara M, Schlievert Patrick M
Department of Microbiology, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA.
Biochemistry. 2007 Jun 26;46(25):7343-52. doi: 10.1021/bi700184s. Epub 2007 May 31.
Bacillus anthracis, a bioterrorism threat as well as an agricultural concern, has complex mechanisms for regulation of its major virulence factors. Genome searches identified the putative two-component system that we designated Bacillus anthracis respiratory response (Brr)A-BrrB. A brrA deletion strain was constructed, and real-time reverse transcriptase polymerase chain reaction and Western blot analysis were used to assess the effect of BrrA-BrrB on levels of virulence factors, the regulator atxA, and growth characteristics. When brrA was deleted, the genes for anthrax toxins (lethal factor, protective antigen, and edema factor) where expressed 4-6-log10-fold less than in the parent Sterne strain. The global regulator atxA was downregulated when compared to atxA in the Sterne strain. Thus, the BrrA-BrrB two-component system positively regulates B. anthracis toxin genes as well as the atxA regulator. Aerobic growth was not affected by the DeltabrrA mutation, but colonies showed differences in morphology, the mutant did not sporulate, and the strain lost the ability to synthesize cytochrome aa3. Gel-shift mobility assays demonstrated that BrrA bound to the promoters of genes for both protective antigen and cytochrome aa3, demonstrating that BrrA is a transcription factor. BrrA-BrrB has sequence similarity with the virulence regulator SrrA-SrrB in Staphylococcus aureus and the aerobic/anaerobic regulator, ResD-ResE, in B. subtilis, and appears to share regulatory mechanisms with ResD-ResE.
炭疽芽孢杆菌既是一种生物恐怖主义威胁,也是农业领域关注的对象,它对其主要毒力因子具有复杂的调控机制。通过基因组搜索鉴定出了一个假定的双组分系统,我们将其命名为炭疽芽孢杆菌呼吸反应(Brr)A - BrrB。构建了brrA缺失菌株,并使用实时逆转录聚合酶链反应和蛋白质免疫印迹分析来评估BrrA - BrrB对毒力因子水平、调节因子atxA以及生长特性的影响。当brrA被缺失时,炭疽毒素(致死因子、保护性抗原和水肿因子)的基因表达量比亲本Sterne菌株低4 - 6个对数级。与Sterne菌株中的atxA相比,全局调节因子atxA被下调。因此,BrrA - BrrB双组分系统正向调节炭疽芽孢杆菌毒素基因以及atxA调节因子。有氧生长不受ΔbrrA突变的影响,但菌落形态出现差异,突变体不形成芽孢,并且该菌株失去了合成细胞色素aa3的能力。凝胶迁移率变动分析表明,BrrA与保护性抗原基因和细胞色素aa3基因的启动子结合,表明BrrA是一种转录因子。BrrA - BrrB与金黄色葡萄球菌中的毒力调节因子SrrA - SrrB以及枯草芽孢杆菌中的好氧/厌氧调节因子ResD - ResE具有序列相似性,并且似乎与ResD - ResE共享调节机制。