In vivo and in vitro effects of integration host factor at the DmpR-regulated sigma(54)-dependent Po promoter

Transcription from the Pseudomonas CF600-derived sigma (54)-dependent promo ter Po is controlled by the aromatic-responsive activator DmpR. Here we exa mine the mechanism(s) by which integration host factor (IHF) stimulates Dmp R-activated transcriptional output of the Po promoter both in vivo and in v itro. In vivo, the Po promoter exhibits characteristics that typify many si gma (54)-dependent promoters, namely, a phasing-dependent tolerance with re spect to the distance from the regulator binding sites to the distally loca ted RNA polymerase binding site, and a strong dependence on IHF for optimal promoter output. IHF is shown to affect transcription via structural reper cussions mediated through binding to a single DNA signature located between the regulator and RNA polymerase binding sites. In vitro, using DNA templa tes that lack the regulator binding sites and thus bypass It role of THF in facilitating physical interaction between the regulator and the transcript ional apparatus, IHF still mediates a DNA binding-dependent stimulation of Po transcription. This stimulatory effect is shown to be independent of pre viously described mechanisms for the effects of IHF at sigma (54) promoters such as aiding binding of the regulator or recruitment of sigma (54)-RNA p olymerase via UP element-like DNA. The effect of IHF could be traced to pro motion and/or stabilization of open complexes within the nucleoprotein comp lex that may involve an A+T-rich region of the IHF binding site and promote r-upstream DNA. mechanistic implications are discussed in the context of a model in which IHF binding results in transduction of DNA instability from an A+T-rich region to the melt region of the promoter.