Zhang L, Fan F, Palmer L M, Lonetto M A, Petit C, Voelker L L, St John A, Bankosky B, Rosenberg M, McDevitt D
Anti-Infectives Research, SmithKline Beecham Pharmaceuticals Research and Development, 1250 S. Collegeville Road, Collegeville, PA 19426, USA.
Gene. 2000 Sep 19;255(2):297-305. doi: 10.1016/s0378-1119(00)00325-5.
Selectively regulating gene expression in bacteria has provided an important tool for studying gene function. However, well-regulated gene control systems have been restricted primarily for use in laboratory non-pathogenic strains of bacteria (e.g. Escherichia coli, Bacillus subtilis). The development of analogous systems for use in bacterial pathogens such as Staphylococcus aureus would significantly enhance our ability to examine the contribution of any given gene product to pathogen growth and viability. In this report, we adapt, examine and compare three regulated gene expression systems in S. aureus, which had previously been used in B. subtilis. We demonstrate that all three systems function and exhibit titratable induction, together covering a dynamic range of gene expression of approximately 3000-fold. This dynamic range correlates well with the physiological expression levels of cellular proteins. Importantly, we show that one of these systems, the Spac system, is particularly useful for examining gene essentiality and creating specific conditional lethal phenotypes. Moreover, we find that titration of selective target gene products using this system allows direct demonstration of antibiotic mode of action.
在细菌中选择性地调控基因表达为研究基因功能提供了一个重要工具。然而，调控良好的基因控制系统主要局限于实验室非致病性细菌菌株（如大肠杆菌、枯草芽孢杆菌）的使用。开发适用于诸如金黄色葡萄球菌等细菌病原体的类似系统将显著增强我们检查任何特定基因产物对病原体生长和生存能力贡献的能力。在本报告中，我们对先前在枯草芽孢杆菌中使用过的三种调控基因表达系统进行了改造、检验和比较，并将其应用于金黄色葡萄球菌。我们证明这三种系统均能发挥作用并表现出可滴定诱导，共同覆盖了约3000倍的基因表达动态范围。这个动态范围与细胞蛋白质的生理表达水平密切相关。重要的是，我们表明其中一个系统，即Spac系统，对于检查基因必需性和创建特定的条件致死表型特别有用。此外，我们发现使用该系统滴定选择性靶基因产物能够直接证明抗生素的作用模式。
