Superimposed Levels of Regulation of the 4-Hydroxyphenylacetate Catabolic Pathway in Escherichia coli

Abstract

The regulation of the Pg promoter, which controls the expression of the meta operon of the 4-hydroxyphenylacetic acid (4-HPA) catabolic pathway of Escherichia coli W, has been examined through in vivo and in vitro experiments. By using Pg-lacZ fusions we have demonstrated that Pg is a promoter only inducible in the stationary phase when cells are grown on glucose as the sole carbon and energy source. This strict catabolite repression control is mediated by the cAMP receptor protein (CRP). This event does not require the presence of the specific HpaR repressor or the 4-HPA permease (HpaX), excluding the involvement of a typical inducer exclusion mechanism. However, the acetic acid excreted in the stationary phase by the cells growing in glucose acts as an overflow metabolite, which can provide the energy to produce cAMP and to adapt the cells rapidly to the utilization of a new less preferred carbon source such as the aromatic compounds. Although Pg is not a final sigma(38)-dependent promoter, it is activated by the global regulator integration host factor (IHF) in the stationary phase of growth. Gel retardation assays have demonstrated that both CRP and IHF simultaneously bind to the Pg upstream region. DNase I footprint experiments showed that cAMP-CRP and IHF binding sites are centered at -61.5 and -103, respectively, with respect to the transcription start site +1 of the Pg promoter.