Regulation of alternative oxidase activity during phosphate deficiency in bean roots (Phaseolus vulgaris)

Cyanide-resistant respiration was studied in mitochondria isolated from the roots of bean plants (Phaseolus vulgaris L. cv. Zlota Saxa) grown hydroponically up to 16 days on a phosphate-sufficient (+ P, control) or phosphate-deficient (- P) medium. Western blotting indicated that the alternative oxidase (AOX) was present only in its reduced (active) form, both in phosphate-sufficient and phosphate-deficient roots, but in the latter, the amount of AOX protein was greater. Addition of pyruvate to the isolation, washing and reaction media made mitochondria from + P roots cyanide-insensitive, similar to mitochondria from - P roots. The doubled activity of NAD-malic. enzyme (NAD-ME) in - P compared with + P root mitochondria may suggest increased pyruvate production in - P mitochondria. Lower cytochrome c oxidase (COX) activity and no uncoupler effect on respiration indicated limited cytochrome chain activity in - P mitochondria. In - P mitochondria, the oxygen uptake decreased and the level of Q reduction increased from 60 to 80%. With no pyruvate present (AOX not fully activated), inhibition of the cytochrome pathway resulted in an increased level of the ratio of reduced ubiquinone (Qr) to total ubiquinone (Qt) (Qr/Qt) in + P mitochondria, but did not change Qr/Qt in - P mitochondria. When pyruvate was present, the kinetics for AOX were similar in mitochondria from - P and + P roots. It is suggested that AOX participation in - P respiration may provide an acclimation to phosphate deficiency. Stabilization of the ubiquinone reduction level by AOX might prevent the harmful effect of an increased formation of reactive oxygen species.