Arabidopsis thaliana and Saccharomyces cerevisiae NHX1 genes encode amiloride sensitive electroneutral Na+-/H+ exchangers

Sodium at high millimolar levels in the cytoplasm is toxic to plant and yea st cells. Sequestration of Na+ ions into the vacuole is one mechanism to co nfer Na+-tolerance on these organisms. In the present study we provide dire ct evidence that the Arabidopsis thaliana At-NHX1 gene and the yeast NHX1 g ene encode low-affinity electroneutral Na+/H+ exchangers. We took advantage of the ability of heterologously expressed At-NHX1 to functionally complem ent the yeast nhx1-null mutant. Experiments on vacuolar vesicles isolated f rom yeast expressing At-NHX1 or NHX1 provided direct evidence for pH-gradie nt-energized Na+ accumulation into the vacuole. A major difference between NHX1 and At-NHX1 is the presence of a cleavable N-terminal signal peptide ( SP) in the former gene. Fusion of the SP to At-NHX1 resulted in an increase in the magnitude of Na+/H+ exchange, indicating a role for the SP in prote in targeting or regulation. Another distinguishing feature between the plan t and yeast antiporters is their sensitivity to the diuretic compound amilo ride. Whereas At-NHX1 was completely inhibited by amiloride, NHX1 activity was reduced by only 20-40%. These results show that yeast as a heterologous expression system provides a convenient model to analyse structural and re gulatory features of plant Na+/H+ antiporters.