CYTOCHROME AA3 GENE-REGULATION IN MEMBERS OF THE FAMILY RHIZOBIACEAE - COMPARISON OF COPPER AND OXYGEN EFFECTS IN BRADYRHIZOBIUM-JAPONICUM AND RHIZOBIUM-TROPICI

Dithionite-reduced minus ferricyanide-oxidized difference spectra on m embranes from Rhizobium tropici (formerly Rhizobium leguminosarum bv. phaseoli) incubated at progressively lower O2 concentrations showed on ly a slight concomitant decrease in A603, the alpha-peak of cytochrome aa3. In contrast to previous results on Bradyrhizobium japonicum, R. tropici showed no significant O2-mediated reduction in the level of ei ther coxA transcription or cytochrome aa3 activity (as measured by asc orbate-N,N,N',N'-tetramethyl-p-phenylenediamine [TMPD] oxidase) even i n the cells incubated at 12.5 muM O2. Bean nodule R. tropici bacteroid s contained 65% of the fully aerobic free-living levels of the coxA tr anscript. Primer extension analyses established the transcription init iation site of the R. tropici coxA genes. Sequence analyses of the reg ions upstream of the transcription initiation site revealed no homolog y with previously reported Rhizobiaceae family promoters, including th e coxA promoter of B. japonicum. The R. tropici deduced CoxA sequence itself is highly homologous to the B. japonicum and Paracoccus denitri ficans CoxA sequences. In both B. japonicum and R. tropici, coxA trans cript levels were the same for cells grown with copper (0.02 muM) in t he medium or in medium completely devoid of copper. However, a posttra nscriptional effect of copper deprivation was observed for both bacter ia; difference absorption spectra on membranes from cells grown withou t copper showed that B. japonicum lacked spectroscopically detectable cytochrome aa3, whereas R. tropici retained approximately 50% of norma l cytochrome aa3 levels.