Positive and Negative Transcriptional Regulation of the Escherichia coli Gluconate Regulon Gene gntT by GntR and the Cyclic AMP (cAMP)-cAMP Receptor Protein Complex

Abstract

ABSTRACT The gntT gene of Escherichia coli is specifically induced by gluconate and repressed via catabolite repression. Thus, gluconate is both an inducer and a repressor of gntT expression since gluconate is a catabolite-repressing sugar. In a gntR deletion mutant, the expression of a chromosomal gntT :: lacZ fusion is both high and constitutive, confirming that GntR is the negative regulator of gntT . Indeed, GntR binds to two consensus gnt operator sites; one overlaps the −10 region of the gntT promoter, and the other is centered at +120 with respect to the transcriptional start site. The binding of GntR to these sites was proven in vitro by gel redardation assays and in vivo by site-directed mutagenesis of the binding sites. Binding of GntR to the operators is eliminated by gluconate and also by 6-phosphogluconate at a 10-fold-higher concentration. Interestingly, when gntR deletion strains are grown in the presence of gluconate, there is a twofold decrease in gntT expression which is independent of catabolite repression and binding of GntR to the operator sites. This novel response of gntR mutants to the inducer is termed ultrarepression. Transcription of gntT is activated by binding of the cyclic AMP (cAMP)-cAMP receptor protein (CRP) complex to a CRP binding site positioned at −71 upstream of the gntT transcription start site.