Qiu J, Helmann J D
Department of Microbiology, Cornell University, Ithaca, New York 14853-8101, USA.
J Bacteriol. 2001 Mar;183(6):1921-7. doi: 10.1128/JB.183.6.1921-1927.2001.
Transcriptional selectivity derives, in large part, from the sequence-specific DNA-binding properties of the sigma subunit of RNA polymerase. There are 17 sigma factors in Bacillus subtilis which, in general, recognize distinct sets of promoters. However, some sigma factors have overlapping promoter selectivity. We hypothesize that the overlap between the regulons activated by the sigma(X) and sigma(W) factors can be explained by overlapping specificity for the -10 region: sigma(X) recognizes -10 elements with the sequence CGAC and sigma(W) recognizes CGTA, while both can potentially recognize CGTC. To test this model, we mutated the sigma(X)-specific autoregulatory site (P(X)), containing the -10 element CGAC, to either CGTC or GCTA. Conversely, the sigma(W) autoregulatory site (P(W)) was altered from CGTA to CGTC or CGAC. Transcriptional analyses, both in vitro and in vivo, indicate that changes to the -10 element are sufficient to switch a promoter from the sigma(X) to the sigma(W) regulon or, conversely, from the sigma(W) to the sigma(X) regulon, but context effects clearly play an important role in determining promoter strength. It seems likely that these subtle differences in promoter selectivity derive from amino acid differences in conserved region 2 of sigma, which contacts the -10 element. However, we were unable to alter promoter selectivity by replacements of two candidate recognition residues in sigma(W).
转录选择性在很大程度上源于RNA聚合酶σ亚基的序列特异性DNA结合特性。枯草芽孢杆菌中有17种σ因子,它们通常识别不同的启动子集合。然而,一些σ因子具有重叠的启动子选择性。我们推测,由σ(X)和σ(W)因子激活的调控子之间的重叠可以通过对-10区域的重叠特异性来解释:σ(X)识别序列为CGAC的-10元件,σ(W)识别CGTA,而两者都可能识别CGTC。为了验证这个模型,我们将含有-10元件CGAC的σ(X)特异性自调控位点(P(X))突变为CGTC或GCTA。相反,σ(W)自调控位点(P(W))从CGTA改变为CGTC或CGAC。体外和体内的转录分析表明,-10元件的变化足以使启动子从σ(X)调控子切换到σ(W)调控子,或者相反,从σ(W)调控子切换到σ(X)调控子,但上下文效应在决定启动子强度方面显然起着重要作用。启动子选择性的这些细微差异似乎源于σ的保守区域2中与-10元件接触的氨基酸差异。然而,我们无法通过替换σ(W)中的两个候选识别残基来改变启动子选择性。