What makes an Escherichia coli promoter σS dependent?: Role of the-13/-14 nucleotide promoter positions and region 2.5 of σS

The sigma (S) and sigma (70) subunits of Escherichia coli RNA polymerase recognize very similar promoter sequences. Therefore, many promoters can be activated by both holoenzymes in vitro. The same promoters, however, often exhibit distinct sigma factor selectivity in vivo. It has been shown that high salt conditions, reduced negative supercoiling and the formation of complex 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 that C and T at the -13 and -14 positions, respectively, result in strongest expression. We provide allele-specific suppression data indicating that these nucleotides are contacted by K173 in region 2.5 of sigma (S). In contrast, sigma (70), which features a glutamate at the corresponding position (E458), 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 additional data with the osmY promoter demonstrate that the K173/C(-13) interaction is of general importance. In conclusion, our data demonstrate an important role for region 2.5 in sigma (S) in transcription initiation. Moreover, we propose a consensus sequence for a sigma (S)-selective promoter and discuss its emergence and functional properties from an evolutionary point of view.