Effect of some electron donors and acceptors on redox capacity and simultaneous net H⁺/K⁺ fluxes by aeroponic sunflower seedling roots: Evidence for a CN⁻-resistant redox chain accessible to nonpermeative redox compounds

Summary

Excised roots from aeroponic axenically 48 h dark-grown sunflower (Helianthus annuus L.) seedlings showed redox activities, being able to oxidize/reduce all the exogenously added electron donors/acceptors, that affected the H⁺/K⁺ net fluxes simultaneously measured in the medium. Trials were performed with in vivo and CN⁻-poisoned roots; these showed null⁺/K⁺ net flux activity but still oxidized/reduced all the e⁻ donors/acceptors tested except NADH. NADH enhanced the rate of H⁺ efflux by in vivo roots, otherwise not changing any of the normal flux kinetic characteristics, suggesting that NADH donates e⁻ and H⁺ to the exocellular NADH oxidoreductase activity of a CN⁻-sensitive redox chain in the plasmalemma of the root cells. K⁺ influx was not affected, probably because the NADH concentration was not very high. The e⁻ donor HFC(hexacyanoferrate)(II) activated the H⁺ efflux in a very different way: maximum H⁺ efflux rate was maintained, but both the maximum rate plateau and the optimal pH range were extended, and hence the total H⁺ efflux was significantly enhanced. At the same time, the K⁺ influx was doubled. The different H⁺-efflux kinetics, together with the small but significant HCF(II) oxidation by CN⁻-poisoned roots, were taken as evidence that, besides the CN⁻-sensitive redox chain, an alternative CN⁻-resistant redox chain in the plasmalemma was involved in HCF(II) oxidation. The effect of the oxidized form HCF(III) on H⁺ and K⁺ fluxes was the opposite to that described for HCF(II), but the other H⁺ efflux kinetic characteristics were similar (the maximum rate plateau was extended so that total H⁺ efflux equaled that of the controls). It is proposed that HCF(III) accepts e⁻ only from the alternative CN⁻-resistant redox chain. We could not measure the effect of HCI(hexachloroiridate)(IV) on H+ efflux, as the pH electrodes alone quickly reduced the compound. HCI(IV) promoted a rapid transitory K⁺ efflux, followed by recovery of K⁺ influx. The HCI(IV) reduction by in vivo or CN⁻-poisoned roots was extremely rapid, following similar kinetics. Thus, only the CN⁻-resistant redox chain was involved in both cases. The redox chain inhibitor cis-platinum(II) annulled ion fluxes in the presence of both NADH and HCF(III), and later even inverted them (a small H⁺ influx down the gradient would induce K⁺ efflux). Cis-platinum(II) did not affect HCF(III) reduction by in vivo roots, and only slightly depressed that by CN⁻-poisoned roots. Overall, the effects of the exogenously added e⁻ donors/acceptors tested were consistent with the existence of a CN⁻-resistant redox chain in the plasmalemma of the root cells which would donate/accept e⁻ even when the H⁺ and K⁺ fluxes were annulled by CN⁻ or even inverted by cis-platinum(II) treatments. Thus, in the plasmalemma of in vivo roots this chain would compete for electrons with the normal CN⁻-sensitive one, as in plant mitochondria. The effects on the K⁺ flux were consistent with the current hypothesis that this contributes to counteracting the changes in membrane potential caused by redox activities and the H⁺ flux induced by the different redox compounds tested.