Characterization of a [formula omitted] symport associated with plasma membrane vesicles of maize roots using 36CIO 3− as a radiotracer analog

Abstract

The mechanism of NO 3 − transport was examined in isolated plasma membrane vesicles from maize ( Zea mays L., hybrid B73 × LH 51) roots using 36ClO 3 − as a radiotracer analog for NO 3 −. When an acid-exterior ΔpH was imposed across the vesicle membrane, uptake of 36ClO 3 − was stimulated and the time course of radiolabel uptake displayed an overshoot phenomenon characteristic of the coupling of one solute gradient to the movement of another solute. Evidence supporting ΔpH as the driving force for 36ClO 3 − uptake included a dependence of the overshoot peak and initial rate of 36ClO 3 − uptake on the magnitude of the imposed ΔpH, the occurrence of ΔpH-driven 36ClO 3 − uptake in the presence of KSCN/valinomycin, and the ability of an imposed ΔpH to drive 36ClO 3 − uptake when radiolabel was equilibrated across the membrane. When ΔpH-driven 36ClO 3 − transport was examined in the presence of NO 3 − radiolabel uptake was inhibited in a competitive manner. This was consistent with the carrier having the capacity to use either ClO 3 − or NO 3 − and supports the use of this radiotracer as an analog for NO 3 − in transport studies. When ΔpH-driven 36ClO 3 − uptake was examined as a function of 36ClO 3 − concentration and ΔpH, saturation kinetics were observed and the magnitude of the imposed ΔpH affected the K m but not the V max for 36ClO 3 − uptake. This suggested an ordered binding mechanism where 36ClO 3 − would bind to the protonated form of the carrier prior to translocation. Radiolabeled 36ClO 3 − uptake was inhibited by treatment of the vesicles with phenylglyoxal, suggesting the involvement of arginine moieties in the process of transport. Taken together, these results support the presence of a H + NO 3 − symport carrier at the plasma membrane which could be involved in mediating energy-dependent NO 3 − uptake into plant cells.