In this study we have identified the first plant K+/H+ exchanger, LeNHX2 from tomato (Lycopersicon esculentum Mill. cv. Moneymaker), which is a member of the intracellular NHX exchanger protein family. The LeNHX2 protein, belonging to a subfamily of plant NHX proteins closely related to the yeast NHX1 protein, is abundant in roots and stems and is induced in leaves by short term salt or abscisic acid treatment. LeNHX2 complements the salt- and hygromycin-sensitive phenotype caused by NHX1 gene disruption in yeast, but affects accumulation of K+ and not Na+ in intracellular compartments. The LeNHX2 protein co-localizes with Prevacuolar and Golgi markers in a linear sucrose gradient in both yeast and plants. A histidine-tagged version of this protein could be purified and was shown to catalyze K+/H+ exchange but only minor Na+/H+ exchange in vitro. These data indicate that proper functioning of the endomembrane system relies on the regulation of K+ and H+ homeostasis by K+/H+ exchangers.

Wild halophytic tomato has long been considered as an ideal gene donor for improving salt tolerance in tomato cultivars. Extensive research has been focused on physiological and quantitative trait locus (QTL) characterization of wild tomato species in comparison with cultivated tomato. However, the global gene expression modification of wild tomato in response to salt stress is not well known. A wild tomato genotype, Solanum pimpinellifolium 'PI365967' is significantly more salt tolerant than the cultivar, Solanum lycopersicum 'Moneymaker', as evidenced by its higher survival rate and lower growth inhibition at the vegetative stage. The Affymetrix Tomato Genome Array containing 9,200 probe sets was used to compare the transcriptome of PI365967 and Moneymaker. After treatment with 200 mM NaCl for 5 h, PI365967 showed relatively fewer responsive genes compared with Moneymaker. The salt overly sensitive (SOS) pathway was found to be more active in PI365967 than in Moneymaker, coinciding with relatively less accumulation of Na(+) in shoots of PI365967. A gene encoding salicylic acid-binding protein 2 (SABP2) was induced by salinity only in PI365967, suggesting a possible role for salicylic acid signaling in the salt response of PI365967. The fact that two genes encoding lactoylglutathione lyase were salt inducible only in PI365967, together with much higher basal expression of several glutathione S-transferase genes, suggested a more effective detoxification system in PI365967. The specific down-regulation in PI365967 of a putative high-affinity nitrate transporter, known as a repressor of lateral root initiation, may explain the better root growth of this genotype during salt stress.

In general, wild tomato species are more salt tolerant than cultivated species, a trait that is related to enhanced Na(+) accumulation in aerial parts in the wild species, but the molecular basis for these differences is not known. Plant NHX proteins have been suggested to be important for salt tolerance by promoting accumulation of Na(+) or K(+) inside vacuoles. Therefore, differences in expression or activity of NHX proteins in tomato could be at the basis of the enhanced salt tolerance in wild tomato species. To test this hypothesis, we studied the expression level of four NHX genes in the salt sensitive cultivated species Solanum lycopersicum L. cv. Volgogradskij and the salt tolerant wild species Solanum pimpinelifolium L in response to salt stress. First, we determined that in the absence of salt stress, the RNA abundance of LeNHX2, 3 and 4 was comparable in both species, while more LeNHX1 RNA was detected in the tolerant species. LeNHX2 and LeNHX3 showed comparable expression levels and were present in all tissues, while LeNHX4 was expressed above all in stem and fruit tissues. Next, we confirmed that the wild species was more tolerant and accumulated more Na(+) in aerial parts of the plant. This correlated with the observation that salt stress induced especially the LeNHX3 and LeNHX4 isoforms in the tolerant species. These results support a role of NHX genes as determinants of salt tolerance in tomato, inducing enhanced Na(+) accumulation observed in the wild species when grown in the presence of NaCl.