Oxygen-dependent transcriptional regulation of cytochrome aa3 in Bradyrhizobium japonicum.

Cytochrome aa3 is one of two terminal oxidases expressed in free-living Bradyrhizobium japonicum but not symbiotically in bacteroids. Difference spectra (dithionite reduced minus ferricyanide oxidized) for membranes from cells incubated with progressively lower O2 concentrations showed a concomitant decrease in the A603, the absorption peak characteristic of cytochrome aa3. The level of N,N,N',N'-tetramethyl-p-phenylenediamine oxidase activity, a measure of cytochrome aa3 activity, was also found to depend on the O2 level. Dot blots of total RNA isolated from cells grown at various O2 levels were probed with a fragment of the coxA gene from B. japonicum; a sixfold reduction in transcription from the highest (250 microM) to the lowest (12.5 microM) O2 concentration was observed. Bacteroids had even less coxA message, approximately 19% that in the 12.5 microM O2-incubated cells. Primer extension analysis established the transcription initiation site of the coxA gene at 72 bases upstream of the putative translational start codon. Sequence analysis of the region upstream of the transcription initiation site revealed no homology with previously reported B. japonicum promoters.