ThevirFpromoter inShigella: more than just a curved DNA stretch

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Abstract
Summary: In the human enteropathogenShigellatranscription ofvirF, the primary regulator of the invasion functions, is strictly temperature‐dependent and is antagonistically mediated by H‐NS and FIS, which bind to specific sites on thevirFpromoter. Here we report on the relevance of DNA geometry to the themoregulation ofvirFand demonstrate that thevirFpromoter hosts a major DNA bend halfway between two H‐NS sites. The bent region has been mutagenizedin vitroto mimic temperature‐induced changes of DNA curvature. Functional analysis of curvature mutants and of promoter constructs in which the two H‐NS sites are phased‐out by a half–helix turn reveals that modifying the spatial relationships between these sites severely affects the interaction of H‐NS with thevirFpromoter, as well as itsin vivoandin vitrotemperature‐dependent activity. The role of promoter curvature as thermosensor is also compatible with the present observation that, with increasing temperature, thevirFbending centre moves downstream at a rate having its maximum around the transition temperature, abruptly unmasking a binding site for the transcriptional activator FIS.