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 sigmaS and sigma70 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 sigmaS selectivity. Here, we characterize the first positively acting sigmaS-selective feature in the promoter sequence itself. Using the sigmaS-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 sigmaS. In contrast, sigma70, which features a glutamate at the corresponding position (E458), as well as the sigmaS(K173E) variant, exhibit a preference for a G(-13). C(-13) is highly conserved in sigmaS-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 sigmaS in transcription initiation. Moreover, we propose a consensus sequence for a sigmaS-selective promoter and discuss its emergence and functional properties from an evolutionary point of view.
Institut für Biologie - Mikrobiologie, Freie Universität Berlin, Königin-Luise-Str. 12-16, 14195 Berlin, Germany.
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G
Becker
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Hengge-Aronis
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D016384:Consensus Sequence; D002373:Cyclic AMP Receptor Protein / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D015151:Immunoblotting; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D033902:Periplasmic Binding Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D017433:Protein Structure, Secondary; D012808:Sigma Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D002352:Carrier Proteins; D002373:Cyclic AMP Receptor Protein; D029968:Escherichia coli Proteins; D033902:Periplasmic Binding Proteins; D012808:Sigma Factor; C069567:sigma factor KatF protein, Bacteria; C074783:osmY protein, E coli