The LysR-type regulator AtzR binding site: DNA sequences involved in activation, repression and cyanuric acid-dependent repositioning

Abstract

The LysR-type transcriptional regulator (LTTR) AtzR of Pseudomonas sp. strain ADP activates the cyanuric acid-utilization atzDEF operon in response to low nitrogen availability and the presence of cyanuric acid. AtzR also represses expression of its own gene, atzR, transcribed divergently from atzDEF. Here we identify and functionally characterize the cis-acting sequences at the atzR-atzDEF divergent promoter region required for AtzR-dependent regulation. AtzR binds a single site overlapping both the PatzR and PatzDEF promoters and induces a DNA bend immediately upstream from PatzDEF. Interaction of AtzR with the inducer cyanuric acid shortens the protein-DNA interaction region and relaxes the DNA bend. The AtzR binding site contains a strong binding determinant, the repression binding site (RBS), centred at position -65 relative to the atzDEF transcriptional start, containing the LTTR binding consensus motif. Integrity of the RBS is essential for high-affinity AtzR binding, activation and autorepression. A second, weaker binding determinant, the activation binding site (ABS), is present between the RBS and PatzDEF. Deletion of the ABS only provokes a modest decrease in AtzR affinity for the promoter region in vitro, but abolishes repression of PatzR in vivo. Involvement of the ABS in autorepression has not been previously reported.