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NsrR targets in the Escherichia coli genome: new insights into DNA sequence requirements for binding and a role for NsrR in the regulation of motility

The Escherichia coli NsrR protein is a nitric oxide‐sensitive repressor of transcription. The NsrR‐binding site is predicted to comprise two copies of an 11 bp motif arranged as an inverted repeat with 1 bp spacing. By mutagenesis we confirmed that both 11 bp motifs are required for maximal NsrR repression of the ytfE promoter. We used chromatin immunoprecipitation and microarray analysis (ChIP‐chip) to show that NsrR binds to 62 sites close to the 5′ ends of genes. Analysis of the ChIP‐chip data suggested that a single 11 bp motif (with the consensus sequence AANATGCATTT) can function as an NsrR‐binding site in vivo. NsrR binds to sites in the promoter regions of the fliAZY, fliLMNOPQR and mqsR‐ygiT transcription units, which encode proteins involved in motility and biofilm development. Reporter fusion assays confirmed that NsrR negatively regulates the fliA and fliL promoters. A mutation in the predicted 11 bp NsrR‐binding site in the fliA promoter impaired repression by NsrR and prevented detectable binding in vivo. Assays on soft‐agar confirmed that NsrR is a negative regulator of motility in E. coli K12 and in a uropathogenic strain; surface attachment assays revealed decreased levels of attached growth in the absence of NsrR.