The Arabidopsis Na+/H+ exchanger AtNHX1 catalyzes low affinity Na+ and K+ transport in reconstituted liposomes

In saline environments, plants accumulate Na+ in vacuoles through the activity of tonoplast Na+/H+ antiporters. The first gene for a putative plant vacuolar Na+/H+ antiporter, AtNHX1, was isolated from Arabidopsis and shown to increase plant tolerance to NaCl. However, AtNHX1 mRNA was up-regulated by Na+ or K+ salts in plants and substituted for the homologous protein of yeast to restore tolerance to several toxic cations. To study the ion selectivity of the AtNHX1 protein, we have purified a histidine-tagged version of the protein from yeast microsomes by Ni2+ affinity chromatography, reconstituted the protein into lipid vesicles, and measured cation-dependent H+ exchange with the fluorescent pH indicator pyranine. The protein catalyzed Na+ and K+ transport with similar affinity in the presence of a pH gradient. Li+ and Cs+ ions were also transported with lower affinity. Ion exchange by AtNHX1 was inhibited 70% by the amiloride analog ethylisopropylamiloride. Our data indicate a role for intracellular antiporters in organelle pH control and osmoregulation.