Downregulation of the Escherichia coli guaB promoter by upstream-bound cyclic AMP receptor protein

Abstract

The Escherichia coli guaB promoter (P-guaB) is responsible for directing transcription of the guaB and guaA genes, which specify the biosynthesis of the nucleotide GMP. P-guaB is subject to growth rate-dependent control (GRDC) and possesses an UP element that is required for this regulation. In addition, PguaB contains a discriminator, three binding sites for the nucleoid-associated protein FIS, and putative binding sites for the regulatory proteins DnaA, PurR, and cyclic AMP receptor protein (CRP). Here we show that the CRP-cyclic AMP (cAMP) complex binds to a site located over 100 bp upstream of the guaB transcription start site, where it serves to downregulate P-guaB. The CRP-mediated repression of P-guaB activity increases in media that support lower growth rates. Inactivation of the crp or cyaA gene or ablation/translocation of the CRP site relieves repression by CRP and results in a loss of GRDC of P-guaB. Thus, GRDC of P-guaB involves a progressive increase in CRP-mediated repression of the promoter as the growth rate decreases. Our results also suggest that the CRP-cAMP complex does not direct GRDC at P-guaB and that at least one other regulatory factor is required for conferring GRDC on this promoter. However, PurR and DnaA are not required for this regulatory mechanism