What makes an Escherichia coli promoter sigma(S) dependent? Role of the-13/-14 nucleotide promoter positions and region 2.5 of sigma(S)

The sigma (S) and sigma (70) subunits of Escherichia coli RNA polymerase re cognize very similar promoter sequences. Therefore, many promoters can be a ctivated by both holoenzymes in vitro. The same promoters, however, often e xhibit distinct sigma factor selectivity in vivo. It has been shown that hi gh salt conditions, reduced negative supercoiling and the formation of comp lex nucleoprotein structures in a promoter region can contribute to or even generate sigma (S) selectivity. Here, we characterize the first positively acting sigma (S)-selective feature in the promoter sequence itself. Using the sigma (S)-dependent csiD promoter as a model system, we demonstrate tha t C and T at the -13 and -14 positions, respectively, result in strongest e xpression. We provide allele-specific suppression data indicating that thes e nucleotides are contacted by K173 in region 2.5 of sigma (S). In contrast , sigma (70), which features a glutamate at the corresponding position (E45 8), as well as the sigma (S(K173E)) variant, exhibit a preference for a G(- 13). C(-13) is highly conserved in sigma (S)-dependent promoters, and addit ional data with the osmY promoter demonstrate that the K173/C(-13) interact ion is of general importance. In conclusion, our data demonstrate an import ant role for region 2.5 in sigma (S) in transcription initiation. Moreover, we propose a consensus sequence for a sigma (S)-selective promoter and dis cuss its emergence and functional properties from an evolutionary point of view.