Pagán-Ramos E, Song J, McFalone M, Mudd M H, Deretic V
Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0620, USA.
J Bacteriol. 1998 Sep;180(18):4856-64. doi: 10.1128/JB.180.18.4856-4864.1998.
Oxidative stress response in pathogenic mycobacteria is believed to be of significance for host-pathogen interactions at various stages of infection. It also plays a role in determining the intrinsic susceptibility to isoniazid in mycobacterial species. In this work, we characterized the oxyR-ahpC and furA-katG loci in the nontuberculous pathogen Mycobacterium marinum. In contrast to Mycobacterium smegmatis and like Mycobacterium tuberculosis and Mycobacterium leprae, M. marinum was shown to possess a closely linked and divergently oriented equivalents of the regulator of peroxide stress response oxyR and its subordinate gene ahpC, encoding a homolog of alkyl hydroperoxide reductase. Purified mycobacterial OxyR was found to bind to the oxyR-ahpC promoter region from M. marinum and additional mycobacterial species. Mobility shift DNA binding analyses using OxyR binding sites from several mycobacteria and a panel of in vitro-generated mutants validated the proposed consensus mycobacterial recognition sequence. M. marinum AhpC levels detected by immunoblotting, were increased upon treatment with H2O2, in keeping with the presence of a functional OxyR and its binding site within the promoter region of ahpC. In contrast, OxyR did not bind to the sequences upstream of the katG structural gene, and katG expression did not follow the pattern seen with ahpC. Instead, a new open reading frame encoding a homolog of the ferric uptake regulator Fur was identified immediately upstream of katG in M. marinum. The furA-katG linkage and arrangement are ubiquitous in mycobacteria, suggesting the presence of additional regulators of oxidative stress response and potentially explaining the observed differences in ahpC and katG expression. Collectively, these findings broaden our understanding of oxidative stress response in mycobacteria. They also suggest that M. marinum will be useful as a model system for studying the role of oxidative stress response in mycobacterial physiology, intracellular survival, and other host-pathogen interactions associated with mycobacterial diseases.
致病性分枝杆菌中的氧化应激反应被认为在感染的各个阶段对于宿主 - 病原体相互作用具有重要意义。它在决定分枝杆菌物种对异烟肼的内在敏感性方面也发挥作用。在这项工作中，我们对非结核病原体海分枝杆菌中的oxyR - ahpC和furA - katG基因座进行了表征。与耻垢分枝杆菌不同，与结核分枝杆菌和麻风分枝杆菌相似，海分枝杆菌被证明拥有紧密相连且方向相反的过氧化物应激反应调节因子oxyR及其下属基因ahpC的等效物，ahpC编码烷基过氧化氢还原酶的同源物。发现纯化的分枝杆菌OxyR可与海分枝杆菌及其他分枝杆菌物种的oxyR - ahpC启动子区域结合。使用来自几种分枝杆菌的OxyR结合位点和一组体外产生的突变体进行的迁移率变动DNA结合分析验证了所提出的分枝杆菌共有识别序列。通过免疫印迹检测到，经H2O2处理后，海分枝杆菌AhpC水平升高，这与功能性OxyR及其在ahpC启动子区域内的结合位点的存在一致。相比之下，OxyR不与katG结构基因上游的序列结合，并且katG的表达并不遵循ahpC的模式。相反，在海分枝杆菌中，在katG的紧邻上游发现了一个新的编码铁摄取调节因子Fur同源物的开放阅读框。furA - katG的连锁和排列在分枝杆菌中普遍存在，这表明存在氧化应激反应的其他调节因子，并可能解释了在ahpC和katG表达中观察到的差异。总体而言，这些发现拓宽了我们对分枝杆菌中氧化应激反应的理解。它们还表明，海分枝杆菌将作为一个模型系统，用于研究氧化应激反应在分枝杆菌生理学、细胞内存活以及与分枝杆菌疾病相关的其他宿主 - 病原体相互作用中的作用。
