CadC-mediated activation of the cadBA promoter in Escherichia coli

The transcriptional activator CadC in Escherichia coli, a member of
the ToxR-like proteins, activates transcription of the cadBA operon
encoding the lysine decarboxylase CadA and the lysine-cadaverine
antiporter CadB. cadBA is induced under conditions of acidic
external pH and exogenous lysine; anoxic conditions raise the
expression level up to 10 times. To characterize the binding
mechanism of CadC, procedures for the purification of this
membrane-integrated protein and its reconstitution into
proteoliposomes were established. The binding sites of CadC
upstream of the cadBA promoter region were determined by in vitro
DNaseI protection analysis. Two regions were protected during
DNaseI digestion, one from - 144 to - 112 bp, designated Cad1, and
another one from - 89 to - 59 bp, designated Cad2. Binding of
purified CadC to Cad1 and Cad2 was further characterized by
DNA-binding assays, indicating that CadC was able to bind to both
DNA fragments. Genetic analysis with promoter-lacZ fusions
confirmed that both sites, Cad1 and Cad2, are essential for
activation of cadBA transcription. Moreover, these experiments
revealed that binding of H-NS upstream of the CadC-binding sites is
necessary for repression of cadBA expression at neutral pH and
under aerobic conditions. Based on these results, a model for
transcriptional regulation of the cadBA operon is proposed,
according to which H-NS is involved in the formation of a
repression complex under non-inducing conditions. This complex is
dissolved by binding of CadC to Cad1 under inducing conditions.
Upon binding of CadC to Cad2 cadBA expression is activated.
Copyright (C) 2005 S. Karger AG, Basel.