Department of Biosciences and Biomedical Research Laboratory, George Mason University, 10650 Pyramid Place, Manassas, Virginia 20110, USA.
Cell Microbiol. 2012 Aug;14(8):1219-30. doi: 10.1111/j.1462-5822.2012.01791.x. Epub 2012 Apr 17.
To achieve widespread dissemination in the host, Bacillus anthracis cells regulate their attachment to host endothelium during infection. Previous studies identified BslA (Bacillus anthracis S-layer Protein A), a virulence factor of B. anthracis, as necessary and sufficient for adhesion of vegetative cells to human endothelial cells. While some factors have been identified, bacteria-specific contributions to BslA mediated adhesion remain unclear. Using the attenuated vaccine Sterne 7702 strain of B. anthracis, we tested the hypothesis that InhA (immune inhibitor A), a B. anthracis protease, regulates BslA levels affecting the bacteria's ability to bind to endothelium. To test this, a combination of inhA mutant and complementation analysis in adhesion and invasion assays, Western blot and InhA inhibitor assays were employed. Results show InhA downregulates BslA activity reducing B. anthracis adhesion and invasion in human brain endothelial cells. BslA protein levels in ΔinhA bacteria were significantly higher than wild-type and complemented strains showing InhA levels and BslA expression are inversely related. BslA was sensitive to purified InhA degradation in a concentration- and time-dependent manner. Taken together these data support the role of InhA regulation of BslA-mediated vegetative cell adhesion and invasion.
为了在宿主中广泛传播，炭疽杆菌细胞在感染过程中调节其与宿主内皮细胞的附着。先前的研究确定了 BslA（炭疽杆菌 S 层蛋白 A），一种炭疽杆菌的毒力因子，是营养细胞与人类内皮细胞附着所必需和充分的。虽然已经确定了一些因素，但细菌特异性对 BslA 介导的附着的贡献仍不清楚。我们使用减毒疫苗 Sterne 7702 株炭疽杆菌来检验假设，即 InhA（免疫抑制剂 A），一种炭疽杆菌蛋白酶，调节 BslA 水平，影响细菌与内皮细胞结合的能力。为了验证这一点，我们在粘附和侵袭实验中结合了 inhA 突变体和互补分析、Western blot 和 InhA 抑制剂实验。结果表明，InhA 下调了 BslA 活性，降低了炭疽杆菌在人脑血管内皮细胞中的粘附和侵袭能力。ΔinhA 细菌中的 BslA 蛋白水平明显高于野生型和互补株，表明 InhA 水平与 BslA 表达呈负相关。Bsla 对纯化的 InhA 的降解敏感，呈浓度和时间依赖性。这些数据共同支持了 InhA 调节 BslA 介导的营养细胞粘附和侵袭的作用。
