Oxygen and nitrate-dependent regulation of dmsABC operon expression in Escherichia coli: sites for Fnr and NarL protein interactions.
BACKGROUND
Escherichia coli, can respire anaerobically using dimethyl sulfoxide (DMSO) or trimethylamine-N-oxide (TMAO) as the terminal electron acceptor for anaerobic energy generation. Expression of the dmsABC genes that encode the membrane-associated DMSO/TMAO reductase is positively regulated during anaerobic conditions by the Fnr protein and negatively regulated by the NarL protein when nitrate is present.


RESULTS
The regions of dmsA regulatory DNA required for Fnr and NarL interactions in response to anaerobiosis and nitrate, respectively, were examined. Mutations within the Fnr site that deviated from the wild type sequence, TTGATaccgAACAA, or that removed an entire half-site, either impaired or abolished the anaerobic activation of dmsA-lacZ expression. The region for phosphorylated NarL (NarL-phosphate) binding at the dmsA promoter was identified by DNase I and hydroxyl radical footprinting methods. A large 97 bp region that overlaps the Fnr and RNA polymerase recognition sites was protected by NarL-phosphate but not by the non-phosphorylated form of NarL. Hydroxyl radical footprinting analysis confirmed the NarL-phosphate DNase I protections of both dmsA strands and revealed 8-9 protected sites of 3-5 bp occurring at ten bp intervals that are offset by 3 bp in the 3' direction.


CONCLUSIONS
These findings suggest that multiple molecules of phosphorylated NarL bind along one face of the DNA and may interfere with Fnr and/or RNA polymerase interactions at the dmsA regulatory region. The interplay of these transcription factors insures a hierarchical expression of the dmsABC genes when respiration of the preferred electron acceptors, oxygen and nitrate, is not possible.
Bearson
Shawn M D
SM
Department of Microbiology, Immunology, and Molecular Genetics, and the Molecular Biology Institute, 1602 Molecular Sciences Building, University of California, Los Angeles, CA 90095, USA. robg@microbio.ucla.edu
Albrecht
Jeffrey A
JA
Gunsalus
Robert P
RP
D000332:Aerobiosis / Q000502:physiology; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics / Q000502:physiology; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000502:physiology; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D007506:Iron-Sulfur Proteins / Q000378:metabolism* / Q000502:physiology; D007763:Lac Operon / Q000235:genetics; D008982:Molybdenum / Q000378:metabolism; D009566:Nitrates / Q000502:physiology*; D009876:Operon / Q000235:genetics* / Q000502:physiology; D010088:Oxidoreductases / Q000737:chemistry* / Q000235:genetics*; D010100:Oxygen / Q000502:physiology*; D011401:Promoter Regions, Genetic / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D007506:Iron-Sulfur Proteins; D009566:Nitrates; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C069780:NarL protein, E coli; D008982:Molybdenum; D010088:Oxidoreductases; C045664:dimethyl sulfoxide reductase; D010100:Oxygen