Interplay between global regulators of Escherichia coli: effect of RpoS, Lrp and H-NS on transcription of the gene osmC.

The transcription of the osmC gene of Escherichia coli is regulated as a function of the phase of growth. It is induced during the decelerating phase, before entry into stationary phase. osmC expression is directed by two overlapping promoters, osmCp1 and osmCp2. osmCp2 is mainly transcribed by E-sigma(s), the RNA polymerase using the sigma(s) (RpoS) sigma factor, and is responsible for the growth phase regulation. Transcription from osmCp1 is independent of sigma(s). The leucine-responsive protein (Lrp) has been shown to bind the osmC promoter region in band shift experiments. In vivo analysis using osmC-lacZ transcriptional fusions demonstrated that Lrp affects the expression of both promoters. It represses the transcription of osmCp1 and activates the transcription of osmCp2 by E-sigma(s). An absence of Lrp results in an increase in the amount of RpoS during exponential growth in minimal medium. The nucleoid-associated protein H-NS also represses osmC transcription from both promoters. However, this happens through different mechanisms. The effect on osmCp2 is probably mediated by the increase in sigma(s) concentration in the cytoplasm of hns- mutants, while the effect on osmCp1 is independent of sigma(s). No binding of H-NS to the promoter region DNA could be detected, indicating that the effect on osmCp1 could also be indirect.

Growth-phase-dependent expression of the osmotically inducible gene osmC of Escherichia coli K-12.

The transcription of the osmotically inducible gene osmC of Escherichia coli is initiated by two overlapping promoters, osmCp1 and osmCp2. The existence of these two promoters was confirmed by site-directed mutagenesis. osmC transcription is regulated by the growth phase. In a medium of low osmotic pressure, expression of osmC is induced at the onset of decelerating phase and continues during the beginning of stationary phase. At elevated osmotic pressure, the induction occurs somewhat earlier during growth. Both promoters are repressed during early exponential phase. osmCp2 is induced during entry into stationary phase. Transcription from osmCp1, which is approximately 10-fold lower than that of osmCp2 in rich medium, starts during the mid-log phase and stops in early stationary phase. In the absence of sigmaS, the stationary-phase sigma factor encoded by rpoS, osmCp2 expression is much reduced while expression of osmCp1 is unaffected. As a consequence, the regulation of osmC as a function of growth is at least partially independent of sigmaS.

Characterization of the osmotically inducible gene osmE of Escherichia coli K-12.

osmE, an osmotically inducible gene of Escherichia coli, was physically mapped on the bacterial chromosome, cloned and sequenced. osmE appeared to encode a 12,021 Da protein of unknown function, with a lipoprotein-type signal sequence at the amino-terminus. The osmE reading frame was confirmed by sequencing the junction of an osmE-phoA gene fusion. osmE was demonstrated to be transcribed as a single cistron. A phi [osmEp-lac] operon fusion was constructed, and analysis of its expression demonstrated that osmE osmotic regulation probably occurs at the transcriptional level. The osmE promoter was identified by both S1 nuclease and primer extension mapping of the 5' end of the osmE mRNA, by deletion analysis and by identification of a point mutation reducing its activity. Sequence information sufficient for expression and osmotic regulation is present on a DNA fragment extending from positions -37 to +52 with respect to the osmE transcription start. Uninduced expression of the osmE-lac fusion was increased in the presence of mutations in the hns and himA genes. The osmE promoter overlaps a promoter for a gene transcribed in the opposite direction, efg. Transcription from the efg promoter is only weakly affected by osmotic pressure and is independent of the presence of an intact OsmE protein.