SEQUENCE AND DNA STRUCTURAL DETERMINANTS OF N4 VIRION RNA POLYMERASE-PROMOTER RECOGNITION

Coliphage N4-coded, virion-encapsidated RNA polymerase (vRNAP) is able to bind to and transcribe promoter-containing double-stranded DNAs wh en the template is supercoiled and Escherichia coli single-stranded DN A-binding protein (Eco SSB) is present. We report that vRNAP-promoter recognition and activity on these templates require specific sequences and a hairpin structure on the template strand. Hairpin extrusion,ind uced by Mg(PI) and physiological superhelical density, is essential to provide the correct DNA structure for polymerase recognition, as muta nt promoters that do not form hairpins show reduced in vitro activity. Therefore, a supercoil-induced DNA structural transition regulates N4 VRNAP transcription. Eco SSB activates transcription at physiological superhelical densities by stabilizing the template-strand hairpin. Sp ecific sequences at the promoters are conserved to provide proper cont acts for VRNAP, to support hairpin extrusion, or both. We propose a mo del for in vivo utilization of the VRNAP promoters, and discuss the ro les of DNA supercoiling and Eco SSB in promoter activation.