Uhl M A, Miller J F
Department of Microbiology and Immunology, School of Medicine, University of California, Los Angeles 90024, USA.
J Bacteriol. 1995 Nov;177(22):6477-85. doi: 10.1128/jb.177.22.6477-6485.1995.
BvgA and BvgS, which regulate virulence gene expression in Bordetella pertussis, are members of the two-component signal transduction family. The effects of growth conditions on the ability of BvgAS to activate transcription of fhaB (encoding filamentous hemagglutinin) and ptxA (encoding the S1 subunit of pertussis toxin) were assessed in Escherichia coli by using chromosomal fhaB-lacZYA and ptxA-lacZYA fusions. Although it had previously been reported that a ptxA-lacZYA transcriptional fusion was not activated by bvgAS in E. coli (J. F. Miller, C. R. Roy, and S. Falkow, J. Bacteriol. 171:6345-6348, 1989), we now present evidence that ptxA is activated by bvgAS in E. coli in a manner that is highly dependent on the growth conditions. Higher levels of beta-galactosidase were produced by ptxA-lacZYA in the presence of bvgAS during growth in Stainer-Scholte medium or M9 minimal salts medium with glucose than in Luria-Bertani medium. In contrast, the level of fhaB-lacZYA expression was high during growth in all media. Addition of modulating stimuli which inhibit BvgAS function eliminated expression of ptxA-lacZYA. Levels of beta-galactosidase expressed from the ptx-lacZYA fusion correlated with growth rate and with the final optical density at 600 nm, suggesting that the lower growth rate in M9-glucose and Stainer-Scholte media was responsible for greater accumulation of beta-galactosidase than was seen in Luria-Bertani medium. Overproduction of BvgA was not sufficient for activation of ptxA expression but was sufficient for fhaB expression. However, overproduction of a constitutive BvgA allele (bvgA-Cl) or overproduction of BvgA in the presence of BvgS was able to activate ptxA. Our results demonstrate Bvg-dependent activation of a ptxA-lacZYA fusion in E. coli and indicate that bvg is the only Bordetella locus required for ptxA activation in this heterologous system.
在百日咳博德特氏菌中调节毒力基因表达的BvgA和BvgS是双组分信号转导家族的成员。通过使用染色体fhaB - lacZYA和ptxA - lacZYA融合体，在大肠杆菌中评估了生长条件对BvgAS激活fhaB（编码丝状血凝素）和ptxA（编码百日咳毒素S1亚基）转录能力的影响。尽管先前有报道称ptxA - lacZYA转录融合体在大肠杆菌中不被bvgAS激活（J. F. Miller、C. R. Roy和S. Falkow，《细菌学杂志》171:6345 - 6348，1989年），但我们现在提供证据表明ptxA在大肠杆菌中被bvgAS激活，其激活方式高度依赖于生长条件。在Stainer - Scholte培养基或含葡萄糖的M9基本盐培养基中生长时，ptxA - lacZYA在bvgAS存在下产生的β - 半乳糖苷酶水平高于在Luria - Bertani培养基中。相反，fhaB - lacZYA的表达水平在所有培养基中生长期间都很高。添加抑制BvgAS功能的调节刺激物可消除ptxA - lacZYA的表达。从ptx - lacZYA融合体表达的β - 半乳糖苷酶水平与生长速率以及600nm处的最终光密度相关，这表明在M9 - 葡萄糖和Stainer - Scholte培养基中较低的生长速率导致β - 半乳糖苷酶的积累比在Luria - Bertani培养基中更多。过量表达BvgA不足以激活ptxA的表达，但足以激活fhaB的表达。然而，组成型BvgA等位基因（bvgA - Cl）的过量表达或在BvgS存在下BvgA的过量表达能够激活ptxA。我们的结果证明了大肠杆菌中ptxA - lacZYA融合体的Bvg依赖性激活，并表明bvg是该异源系统中激活ptxA所需的唯一博德特氏菌基因座。
