Oxygen-dependent transcriptional regulation of cytochrome aa3 in Bradyrhizobium japonicum

Abstract

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 02 concentrations showed a concomitant decrease in theA603, 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 02 level. Dot blots of total RNA isolated from cells grown at various 02 levels were probed with a fragment of the coxA gene from B. japonicum; a sixfold reduction in transcription from the highest (250 ,M) to the lowest (12.5 ,uM) 02 concentration was observed. Bacteroids had even less coxA message, approximately 19%O that in the 12.5 ,uM 02-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.

A high-affinity cbb3-type cytochrome oxidase terminates the symbiosis-specific respiratory chain of Bradyrhizobium japonicum

“…A K m of 56 nM was determined for such an alternative oxidase (or oxidase mixture). This value is about eightfold higher than that of the cbb 3 -type oxidase and falls within (5,14), the values of 55 to 62 nM measured with aerobically grown coxA ϩ strains probably reflect its K m for O 2 , even more so as the V max for the coxA mutant drops by almost 80% compared with that for the wild type (Table 2). In this context it should be noted that the derived K m for O 2 of the aerobically expressed B. japonicum oxidases accords well with the recently reported K m values of the low-affinity cytochrome boЈ from E. coli (46 nM) (11) and the mitochondrial cytochrome aa 3 from soybean (50 to 147 nM) (26,27).…”

“…In such mutants, we consistently measured 20% residual oxidase activity with bacteroids or with cells grown microaerobically or anaerobically (30), but we do not know the relative contribution of each of the three alternative oxidases to this activity. It has been suggested that the level of cytochrome aa 3 is down-regulated in symbiosis (14,23), and the physiological conditions for maximal expression of the CoxMNOP oxidase and the CoxX oxidase ( Fig. 1) are not known at all.…”

Enzymatic Characterization and In Vivo Function of Five Terminal Oxidases in Pseudomonas aeruginosa

“…However, it was unexpected that the quadruple mutant strain QXAa was unable to grow aerobically (Fig. 1A), because aa 3 -type oxidases play major roles in aerobic growth in many bacterial species (23)(24)(25)(26). Transcription of the cox genes encoding aa 3 is repressed by the two-component regulator RoxSR and is dependent on the stationary-phase sigma factor RpoS (21).…”

“…aa 3 is closely related to mitochondrial terminal oxidases. aa 3 -type oxidases typically have low affinity for oxygen and function as main enzymes under high-oxygen conditions in many bacterial species (23)(24)(25)(26). However, in P. aeruginosa, aa 3 is expressed at very low levels in nutrient-rich medium under high-oxygen conditions and is upregulated only in response to nutrient starvation (21).…”

Transcriptional regulation of delta-aminolevulinic acid dehydratase synthesis by oxygen in Bradyrhizobium japonicum and evidence for developmental control of the hemB gene

“…Cytochrome aa 3 is a terminal oxidase necessary for normal aerobic respiration; its expression is diminished or absent in microaerobically cultured cells and in symbiotic bacteroids, and the message level of a gene encoding a subunit of the oxidase is under O 2 control (17). The persistence of cytochrome aa 3 in O 2 -restricted cells of mutant strain 7360 (Fig.…”