title: A proximal promoter element required for positive transcriptional control by guanosine tetraphosphate and DksA protein during the stringent response.
abstract: The alarmone guanosine tetraphosphate (ppGpp) acts as both a positive and a negative regulator of gene expression in the presence of DksA, but the underlying mechanisms of this differential control are unclear. Here, using uspA hybrid promoters, we show that an AT-rich discriminator region is crucial for positive control by ppGpp/DksA. The AT-rich discriminator makes the RNA polymerase-promoter complex extremely stable and therefore easily saturated with RNA polymerase. A more efficient transcription is achieved when the RNA polymerase-promoter complex is destabilized with ppGpp/DksA. We found that exchanging the AT-rich discriminator of uspA with the GC-rich rrnB-P1 discriminator made the uspA promoter negatively regulated by ppGpp/DksA both in vivo and in vitro. In addition, the GC-rich discriminator destabilized the RNA polymerase-promoter complex, and the effect of ppGpp/DksA on the kinetic properties of the promoter was reversed. We propose that the transcription initiation rate from promoters with GC-rich discriminators, in contrast to the uspA-promoter, is not limited by the stability of the open complex. The findings are discussed in view of models for both direct and indirect effects of ppGpp/DksA on transcriptional trade-offs.
authors.affiliation: From the Department of Chemistry and Molecular Biology, Gothenburg University, Medicinaregatan 9C, 413 90 Göteborg, Sweden.
authors.forename: Bertil
authors.initials: B
authors.lastname: Gummesson
authors.affiliation: From the Department of Chemistry and Molecular Biology, Gothenburg University, Medicinaregatan 9C, 413 90 Göteborg, Sweden.
authors.forename: Martin
authors.initials: M
authors.lastname: Lovmar
authors.affiliation: From the Department of Chemistry and Molecular Biology, Gothenburg University, Medicinaregatan 9C, 413 90 Göteborg, Sweden. Electronic address: thomas.nystrom@cmb.gu.se.
authors.forename: Thomas
authors.initials: T
authors.lastname: Nyström
mesh_terms: D020856:AT Rich Sequence / Q000235:genetics; D001482:Base Composition / Q000235:genetics; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics; D006159:Guanosine Tetraphosphate / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D014158:Transcription, Genetic
chemical_list: D029968:Escherichia coli Proteins; C090751:dksA protein, E coli; D006159:Guanosine Tetraphosphate; D012321:DNA-Directed RNA Polymerases
keywords: Bacterial Transcription; Discriminator; DksA; Gene Regulation; RNA Polymerase; Ribosomal RNA (rRNA); Stress Response; Stringent Control; ppGpp