Spacing of promoter elements regulates the basal expression of the soxS gene and converts SoxR from a transcriptional activator into a repressor.
SoxR protein of Escherichia coli governs a global response against superoxide-generating agents (such as paraquat) or nitric oxide, and provides broad antibiotic resistance. A redox signal activates SoxR post-translationally to trigger transcription of a second regulatory gene, soxS. Activated and non-activated SoxR bind the soxS promoter with the same high affinity, but only the activated protein stimulates soxS transcription. SoxR acts by an unusual mechanism of positive control: the protein binds the soxS promoter between near-consensus -10 and -35 elements that are separated by an unusually long 19 bp (versus the optimal 17 bp). We have constructed and analyzed site-specific deletions that alter the promoter element spacing. Reducing the spacer length to 16-18 bp dramatically elevated basal expression of soxS in vivo and in vitro, and nearly eliminated additional activation by SoxR in response to paraquat. More strikingly, shortening the spacer converted SoxR from an activator into a repressor regardless of paraquat treatment. Gel mobility-shift assays show that repression by SoxR of the promoters with 17 and 16 bp spacers is due to interference with binding by RNA polymerase. Thus, activated SoxR remodels the unusual configuration of the wild-type soxS promoter into a highly active form, probably by compensating for the suboptimal distance between the -10 and the -35 elements.
Hidalgo
E
E
Department of Molecular and Cellular Toxicology, Harvard School of Public Health, Boston, MA 02115-6021, USA.
Demple
B
B
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D018384:Oxidative Stress / Q000502:physiology; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D017384:Sequence Deletion / Q000235:genetics; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics; D015533:Transcriptional Activation / Q000502:physiology*; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C070724:SoxR protein, Bacteria; C070725:SoxS protein, E coli; D012321:DNA-Directed RNA Polymerases; D001616:beta-Galactosidase