title: Autoactivation of the marRAB multiple antibiotic resistance operon by the MarA transcriptional activator in Escherichia coli.
abstract: Transcriptional activation of the promoters of the mar/soxRS regulons by the sequence-related but independently inducible MarA and SoxS proteins renders Escherichia coli resistant to a broad spectrum of antibiotics and superoxide generators. Here, the effects of MarA and SoxS on transcription of the marRAB promoter itself were assayed in vitro by using a minimal transcription system and in vivo by assaying beta-galactosidase synthesized from marR::lacZ fusions. Purified MarA and MalE-SoxS proteins stimulated mar transcription about 6- and 15-fold, respectively, when the RNA polymerase/DNA ratio was 1. Purified MarA bound as a monomer to a 16-bp "marbox" located 69 to 54 nucleotides upstream of a putative RNA initiation site. Deletion of the marbox reduced MarA-mar binding 100-fold, abolished the stimulatory effects of MarA and SoxS on transcription in vitro, and reduced marR::lacZ synthesis about 4-fold in vivo. Deletion of upstream DNA adjoining the marbox reduced MarA binding efficiency 30-fold and transcriptional activation 2- to 3-fold, providing evidence for an accessory marbox. Although MarA and the mar operon repressor, MarR, bound to independent sites, they competed for promoter DNA in band shift experiments. Assays of marR::lacZ transcriptional fusions in marRAB deletion or soxRS deletion strains showed that the superoxide generator paraquat stimulates mar transcription via soxRS and that salicylate stimulates mar transcription both by antagonizing MarR and by a MarR-independent mechanism. Thus, transcription of the marRAB operon is autorepressed by MarR and autoactivated by MarA at a site that also can be activated by SoxS.
authors.affiliation: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland 20892-0560, USA. rgmartin@helix.nih.gov
authors.forename: R G
authors.initials: RG
authors.lastname: Martin
authors.forename: K W
authors.initials: KW
authors.lastname: Jair
authors.forename: R E
authors.initials: RE
authors.lastname: Wolf
authors.forename: J L
authors.initials: JL
authors.lastname: Rosner
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins; D004352:Drug Resistance, Microbial / Q000235:genetics; D018432:Drug Resistance, Multiple / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009876:Operon; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics; D012459:Salicylates / Q000494:pharmacology*; D020156:Salicylic Acid; D017384:Sequence Deletion; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C080002:MarA protein, E coli; C080004:MarR protein, E coli; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D012459:Salicylates; D015534:Trans-Activators; D014157:Transcription Factors; C080003:multiple antibiotic resistance protein B; C070725:SoxS protein, E coli; D020156:Salicylic Acid