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Distinct Modes of Promoter Recognition by Two Iron Starvation σ Factors with Overlapping Promoter Specificities

ECF σ factors regulate subsets of bacterial genes in response to environmental stress signals by directing RNA polymerase to promoter sequences known as the −35 and −10 elements. In this work, we identify the −10 and −35 elements that are recognized by the ECF σ factor OrbS. Furthermore, we demonstrate that efficient promoter utilization by this σ factor also requires a polyadenine tract located downstream of the −35 region. We propose that the unique architecture of A-tract DNA imposes conformational features on the −35 element that facilitates efficient recognition by OrbS. Our results show that sequences located between the core promoter elements can make major contributions to promoter recognition by some ECF σ factors. ABSTRACT OrbS and PvdS are extracytoplasmic function (ECF) σ factors that regulate transcription of operons required for the biosynthesis of the siderophores ornibactin and pyoverdine in the Burkholderia cepacia complex and Pseudomonas spp., respectively. Here we show that promoter recognition by OrbS requires specific tetrameric −35 and −10 element sequences that are strikingly similar to those of the consensus PvdS-dependent promoter. However, whereas Pseudomonas aeruginosa PvdS can serve OrbS-dependent promoters, OrbS cannot utilize PvdS-dependent promoters. To identify features present at OrbS-dependent promoters that facilitate recognition by OrbS, we carried out a detailed analysis of the nucleotide sequence requirements for promoter recognition by both OrbS and PvdS. This revealed that DNA sequence features located outside the sigma binding elements are required for efficient promoter utilization by OrbS. In particular, the presence of an A-tract extending downstream from the −35 element at OrbS-dependent promoters was shown to be an important contributor to OrbS specificity. Our observations demonstrate that the nature of the spacer sequence can have a major impact on promoter recognition by some ECF σ factors through modulation of the local DNA architecture. IMPORTANCE ECF σ factors regulate subsets of bacterial genes in response to environmental stress signals by directing RNA polymerase to promoter sequences known as the −35 and −10 elements. In this work, we identify the −10 and −35 elements that are recognized by the ECF σ factor OrbS. Furthermore, we demonstrate that efficient promoter utilization by this σ factor also requires a polyadenine tract located downstream of the −35 region. We propose that the unique architecture of A-tract DNA imposes conformational features on the −35 element that facilitates efficient recognition by OrbS. Our results show that sequences located between the core promoter elements can make major contributions to promoter recognition by some ECF σ factors.