Control of the ferric citrate transport system of Escherichia coli: Mutations in region 2.1 of the FecI extracytoplasmic-function sigma factor suppress mutations in the FecR transmembrane regulatory protein

Transcription of the ferric citrate transport genes is initiated by binding of ferric citrate to the FecA protein in the outer membrane of Escherichia coli K-12. Bound ferric citrate does not have to be transported but initia tes a signal that is transmitted by FecA across the outer membrane and by F ecR across the cytoplasmic membrane into the cytoplasm, where the FecI extr acytoplasmic-function (ECP) sigma factor becomes active. In this study, we isolated transcription initiation-negative missense mutants in the cytoplas mic region of FecR that were located at four sites, L13Q, W19R, W39R, and W 50R, which are highly conserved in FecR-like open reading frames of the Pse udomonas aeruginosa, Pseudomonas putida, Bordetella pertussis, Bordetella b ronchiseptica, and Caulobacter crescentus genomes. The cytoplasmic portion of the FecR mutant proteins, FecR(1-85), did not interact with wild-type Fe cI, in contrast to wild-type FecR(1-85), which induced FecI-mediated fecB t ransport gene transcription. Two missense mutations in region 2.1 of FecI, S15A and H20E, partially restored induction of ferric citrate transport gen e induction of the fecR mutants by ferric citrate. Region 2.1 of sigma (70) is thought to bind RNA polymerase core enzyme; the residual activity of mu tated FecI in the absence of FecR, however, was not higher than that of wil d-type FecI. In addition, missense mutations in the fecI promoter region re sulted in a twofold increased transcription in fecR wild-type cells and a p artial restoration of fec transport gene transcription in the fecR mutants. The mutations reduced binding of the Fe2+ Fur repressor and as a consequen ce enhanced fecI transcription. The data reveal properties of the FecI ECF factor distinct from those of sigma (70) and further support the novel tran scription initiation model in which the cytoplasmic portion of FecR is impo rtant for FecI activity.