Activation of RpoS-dependent proP P2 transcription by the Fis protein in vitro.
The proP gene, encoding a transporter of the osmoprotecting compounds proline and glycine betaine, is expressed from two promoters. Transcription of the P2 promoter occurs at a transient period in late exponential phase and is dependent upon Fis and the RpoS (sigma38) sigma factor. Here we characterize Fis-mediated activation of the P2 promoter in vitro. We find that this promoter displays unusually high specificity for sigma38. Fis strongly activates P2 when bound to site I centered at -41 within the promoter region. There is a complex relationship involving DNA supercoiling and potassium glutamate concentration on Fis activation, but most efficient transcription occurs under high salt conditions when the superhelical density is above -0.03. The major stimulatory effect of DNA supercoiling occurs between superhelical densities of 0 to -0.02 suggesting that, while supercoiling is mechanistically important, it may not be a physiologically relevant controlling factor. However, the stimulation of transcription by high potassium glutamate concentrations may contribute to the osmotic inducibility of the P2 promoter. We show that Fis and E sigma38 bind cooperatively on supercoiled DNA to form a stable complex at P2 that involves promoter melting. Fis also binds to a second site within the proP regulatory region. While binding to this site appears to play no role in Fis activation of the P2 promoter, it functions as a repressor of transcription initiating from the P1 promoter by either sigma70 or sigma38.
Xu
J
J
Department of Biological Chemistry, UCLA School of Medicine, Los Angeles, CA 90095-1737, USA.
Johnson
R C
RC
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism*; D004278:DNA, Superhelical / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; D005971:Glutamates / Q000494:pharmacology; D034982:Integration Host Factors; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009994:Osmolar Concentration; D011401:Promoter Regions, Genetic / Q000235:genetics; D012808:Sigma Factor / Q000378:metabolism*; D027981:Symporters; D015533:Transcriptional Activation / Q000502:physiology*
D001426:Bacterial Proteins; D002352:Carrier Proteins; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; D005971:Glutamates; D034982:Integration Host Factors; C079244:ProP protein, E coli; D012808:Sigma Factor; D027981:Symporters; C069567:sigma factor KatF protein, Bacteria