Bacteria have branched aerobic respiratory chains that terminate at different terminal oxidases. These terminal oxidases have varying properties such as their affinity for oxygen, transcriptional regulation and proton pumping ability. The focus of this study was a quinol oxidase encoded by cyoABCD. Although this oxidase (Cyo) is widespread among bacteria, not much is known about its role in the cell, particularly in bacteria that contain both cytochrome c oxidases and quinol oxidases. Using Rhizobium etli CFN42 as a model organism, a cyo mutant was analysed for its ability to grow in batch cultures at high (21 % O2) and low (1 and 0.1 % O2) ambient oxygen concentrations. In comparison with other oxidase mutants, the cyo mutant had a significantly longer lag phase under low-oxygen conditions. Using a cyo :: lacZ transcriptional fusion, it was shown that cyo expression in the wild type peaks between 1 and 2.5 % O2. In addition, it was shown with quantitative reverse transcriptase PCR that cyoB is upregulated approximately fivefold in 1 % O2 compared with fully aerobic (21 % O2) conditions. Analysis of the cyo mutant during symbiosis with Phaseolous vulgaris indicated that Cyo is utilized during early development of the symbiosis. Although it is commonly thought that Cyo is utilized only at higher oxygen concentrations, the results from this study indicate that Cyo is important for adaptation to and sustained growth under low oxygen.

Rhizobium etli aerobically respires with several terminal oxidases. The quinol oxidase (Cyo) encoded by cyoABCD is needed for efficient adaptation to low oxygen conditions and cyo transcription is upregulated at low oxygen. This study sought to determine how transcription of the cyo operon is regulated. The 59 sequence upstream of cyo was analysed in silico and revealed putative binding sites for ActR of the ActSR two-component regulatory system. The expression of cyo was decreased in an actSR mutant regardless of the oxygen condition. As ActSR is known to be important for growth under low pH in another rhizobial species, the effect of growth medium pH on cyo expression was tested. As the pH of the media was incrementally decreased, cyo expression gradually increased in the WT, eventually reaching ,10-fold higher levels at low pH (4.8) compared with neutral pH (7.0) conditions. This upregulation of cyo under decreasing pH conditions was eliminated in the actSR mutant. Both the actSR and cyo mutants had severe growth defects at low pH (4.8). Lastly, the actSR and cyo mutants had severe growth defects when grown in media treated with an iron chelator. Under these conditions, cyo was upregulated in the WT, whereas cyo was not induced in the actSR mutant. Altogether, the results indicated cyo expression is largely dependent on the ActSR two-component system. This study also demonstrated additional physiological roles for Cyo in R. etli CFN42, in which it is the preferred oxidase for growth under acidic and low iron conditions.