Regions 1.2 and 3.2 of the RNA Polymerase σ Subunit Promote DNA Melting and Attenuate Action of the Antibiotic Lipiarmycin

Initiation of RNA synthesis by bacterial RNA polymerase (RNAP) requires melting of promoter DNA, which is nucleated by the a 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 sigma(70), with a deletion in the region 3.2, or the stationary phase sigma(s) subunits exhibited hypersensitivity to Lpm and increased cold sensitivity of RPo formation. Similar effects were produced by mutation located similar to 60 angstrom away from the Lpm binding site within sigma(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 sigma(70) regions 1.2 and 3.2 attenuate Lpm action by promoting DNA duplex opening. (C) 2015 Elsevier Ltd. All rights reserved.