title: Regions 1.2 and 3.2 of the RNA Polymerase σ Subunit Promote DNA Melting and Attenuate Action of the Antibiotic Lipiarmycin.
abstract: Initiation of RNA synthesis by bacterial RNA polymerase (RNAP) requires melting of promoter DNA, which is nucleated by the σ subunit during formation of the "open" promoter complex (RPo). The antibiotic lipiarmycin (Lpm) inhibits promoter melting by blocking access of the template DNA strand to the RNAP active-site cleft. Here we show that Escherichia coli RNAP holoenzymes containing either housekeeping σ(70), with a deletion in the region 3.2, or the stationary phase σ(S) subunits exhibited hypersensitivity to Lpm and increased cold sensitivity of RPo formation. Similar effects were produced by mutation located ~60 Å away from the Lpm binding site within σ(70) region 1.2, controlling -10 promoter element recognition. Our data suggested that template strand single-stranded DNA competes with Lpm for binding to RNAP and that σ(70) regions 1.2 and 3.2 attenuate Lpm action by promoting DNA duplex opening.
authors.affiliation: CNRS - FRE3689 Université de Montpellier, 1919 Route de Mende, 34293 Montpellier, France.
authors.forename: Zakia
authors.initials: Z
authors.lastname: Morichaud
authors.affiliation: CNRS - FRE3689 Université de Montpellier, 1919 Route de Mende, 34293 Montpellier, France.
authors.forename: Laurent
authors.initials: L
authors.lastname: Chaloin
authors.affiliation: CNRS - FRE3689 Université de Montpellier, 1919 Route de Mende, 34293 Montpellier, France. Electronic address: konstantin.brodolin@inserm.fr.
authors.forename: Konstantin
authors.initials: K
authors.lastname: Brodolin
mesh_terms: D000617:Aminoglycosides / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D000077732:Fidaxomicin; D009691:Nucleic Acid Denaturation; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000378:metabolism*
chemical_list: D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D012808:Sigma Factor; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases; D000077732:Fidaxomicin
keywords: TFIIB B-reader; fidaxomicin; promoter melting; rpoS; switch region