The Escherichia coli NarX/NarL two-component response-regulator system regulates gene expression in response to nitrate ions and the NarL protein is a global transcription factor, which activates transcript initiation at many target promoters. One such target, the E. coli ogt promoter, which controls the expression of an O6-alkylguanine-DNA-alkyltransferase, is dependent on NarL binding to two DNA targets centred at positions -44.5 and -77.5 upstream from the transcript start. Here, we describe ogt promoter derivatives that can be activated solely by NarL binding either at position -44.5 or position -77.5. We show that NarL can also activate the ogt promoter when located at position -67.5. We present data to argue that NarL-dependent activation of transcript initiation at the ogt promoter results from a direct interaction between NarL and a determinant in the C-terminal domain of the RNA polymerase α subunit. Footprinting experiments show that, at the -44.5 promoter, NarL and the C-terminal domain of the RNA polymerase α subunit bind to opposite faces of promoter DNA, suggesting an unusual mechanism of transcription activation. Our work suggests new organisations for activator-dependent transcription at promoters and future applications for biotechnology.
Bibliographical noteThis is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY). Open access for this article was enabled by the participation of University of Birmingham in an all-inclusive Read & Publish pilot with Portland Press and the Biochemical Society under a transformative agreement with JISC.
- DNA-Binding Proteins/genetics
- DNA-Directed RNA Polymerases/genetics
- Escherichia coli K12/genetics
- Escherichia coli Proteins/genetics
- Gene Expression Regulation, Bacterial/drug effects
- Promoter Regions, Genetic
- Transcription Initiation, Genetic