Department of Histology, Microbiology and Medical Biotechnologies, University of Padova, Via Gabelli, 63 35100 Padova, Italy.
Nucleic Acids Res. 2010 Jul;38(12):e134. doi: 10.1093/nar/gkq235. Epub 2010 Apr 20.
Tightly regulated gene expression systems represent invaluable tools for studying gene function and for the validation of drug targets in bacteria. While several regulated bacterial promoters have been characterized, few of them have been successfully used in mycobacteria. In this article we describe the development of a novel repressible promoter system effective in both fast- and slow-growing mycobacteria based on two chromosomally encoded repressors, dependent on tetracycline (TetR) and pristinamycin (Pip), respectively. This uniqueness results in high versatility and stringency. Using this method we were able to obtain an ftsZ conditional mutant in Mycobacterium smegmatis and a fadD32 conditional mutant in Mycobacterium tuberculosis, confirming their essentiality for bacterial growth in vitro. This repressible promoter system could also be exploited to regulate gene expression during M. tuberculosis intracellular growth.
基因表达调控系统是研究基因功能和验证细菌药物靶点的宝贵工具。虽然已经有几种调控细菌启动子被描述过，但在分枝杆菌中成功应用的却很少。本文描述了一种新型的可诱导抑制启动子系统，它基于两种染色体编码的抑制剂，分别是依赖于四环素（TetR）和普那霉素（Pip）的 repressor，可在快速生长和缓慢生长的分枝杆菌中有效工作。这种独特性使得该系统具有很高的多功能性和严格性。使用这种方法，我们能够在耻垢分枝杆菌中获得 ftsZ 条件突变体，在结核分枝杆菌中获得 fadD32 条件突变体，这证实了它们对细菌体外生长的必要性。这种可诱导抑制启动子系统也可以被用来在结核分枝杆菌的细胞内生长过程中调节基因表达。
