Aerobic activation of transcription of the anaerobically inducible Escherichia coli focA‐pfl operon by fumarate nitrate regulator

Expression of the anaerobically inducible focA‐pfl operon in Escherichia coli was activated nearly sevenfold relative to wild‐type under aerobic growth conditions by increasing the dosage of the fnr gene on a pBR322‐based plasmid (pCH21). No effect on anaerobic expression levels was observed, suggesting that operon expression under these conditions is maximal. Examination of the complex transcript pattern of the focA‐pfl operon confirmed that in strains bearing pCH21 all transcripts, with the exception of the promoter 7 transcript, were up‐regulated aerobically. Western analysis of strains bearing pCH21 revealed that the fumarate nitrate regulator (FNR) level was increased approximately ninefold relative to the level in strains bearing a single copy of the fnr gene aerobically, but was only overproduced threefold anaerobically. Analysis of an fnr‐lacZ fusion indicated that fnr expression was more strongly negatively autoregulated in anaerobic cells compared with aerobic cells when pCH21 was present. Taken together, these findings suggest that high‐level overproduction of FNR is prevented anaerobically by active FNR repressing expression of the fnr gene. Furthermore, transcription from promoter 7 of the focA‐pfl operon, which depends on both ArcA‐P and FNR, cannot be activated aerobically by overproduction of FNR alone, while promoter 6, which is less dependent on ArcA‐P, can be activated under these conditions.