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The activity of the ς⁵⁴-promoter Pu of Pseudomonas putida was examined in vitro with a DNA template lacking upstream activating sequences, such that RNA polymerase can be activated by the enhancer-binding protein XylR only from solution. Although the transcription activation pathway in this system lacked the step of integration host factor (IHF)-mediated looping of the XylR·DNA complex toward the prebound RNA polymerase, IHF still stimulated promoter activity. The positive effect of IHF became evident not only with XylR from solution, but also with other ς⁵⁴-dependent activators such as NtrC and NifA. Furthermore, an equivalent outcome was shown for the nonspecific DNA-binding protein HU. This stimulation of transcription in the absence of the enhancer was traced to the recruitment of RNA polymerase (i.e. increased efficiency of formation of closed complexes) brought about by IHF or HU binding. Thus …