IHF is a protein of the bacterial nucleoid proteins (NAPs for nucleoid-associated proteins) involved in DNA structuring and transcription regulation. In vivo interplay between different NAPs determines selectively the expression rate of many genes. Here, we show that IHF is a trans-acting factor implicated directly in the regulation of the proU promoter of Escherichia coli by binding specifically and solely around the promoter box. proU expression is mainly under the repression effect of another NAP, H-NS. We show that IHF binding to proU organize the promoter DNA local structure in a completely different way than H-NS binding. Thus, we propose that the partial alleviation of H-NS repression is mediated by the promoter structure modification.