NaCl Induces a Na+/H+ Antiport in Tonoplast Vesicles from Barley Roots

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Abstract
Evidence was found for a Na+/H+ antiport in tonoplast vesicles isolated from barley (Hordeum vulgare L. cv California Mariout 72) roots. The activity of the antiport was observed only in membranes from roots that were grown in NaCl. Measurements of acridine orange fluorescence were used to estimate relative proton influx and efflux from the vesicles. Addition of MgATP to vesicles from a tonoplast-enriched fraction caused the formation of a pH gradient, interior acid, across the vesicle membranes. EDTA was added to inhibit the ATPase, by chelating Mg2+, and the pH gradient gradually dissipated. When 50 millimolar K+ or Na+ was added along with the EDTA to vesicles from control roots, the salts caused a slight increase in the rate of dissipation of the pH gradient, as did the addition of 50 millimolar K+ to vesicles from salt-grown roots. However, when 50 millimolar Na+ was added to vesicles from salt-grown roots it caused a 7-fold increase in the proton efflux. Inclusion of 20 millimolar K+ and 1 micromolar valinomycin in the assay buffer did not affect this rapid Na+/H+ exchange. The Na+/H+ exchange rate for vesicles from salt-grown roots showed saturation kinetics with respect to Na+ concentration, with an apparent Km for Na+ of 9 millimolar. The rate of Na+/H+ exchange with 10 millimolar Na+ was inhibited 97% by 0.1 millimolar dodecyltriethylammonium.