Activity of tonoplast proton pumps and Na+/H+ exchange in potato cell cultures is modulated by salt

The efficient exclusion of excess Na from the cytoplasm and vacuolar Na+ accumulation are the main mechanisms for the adaptation of plants to salt stress. This is typically carried out by transmembrane transport proteins that exclude Na+ from the cytosol in exchange for H+, a secondary transport process which is energy-dependent and driven by the proton-motive force generated by plasma-membrane and tonoplast proton pumps. Tonoplast enriched-vesicles from control and 150 mM NaCl-tolerant calli lines were used as a model system to study the activity of V-H+-PPase and V-H+-ATPase and the involvement of Na+ compartmentalization into the vacuole as a mechanism of salt tolerance in Solanum tuberosum. Both ATP- and pyrophosphate (PPi)-dependent H+-transport were higher in tonoplast vesicles from the salt-tolerant line than in vesicles from control cells. Western blotting of tonoplast proteins confirmed that changes in V-H+-PPase activity are correlated with increased protein amount. Conversely, immunodetection of the A-subunit of V-H+-ATPase revealed that a mechanism of post-translational regulation is probably involved. Na+-dependent dissipation of a pre-established pH gradient was used to measure Na+/H+ exchange in tonoplast vesicles. The initial rates of proton efflux followed Michaelis-Menten kinetics and the V-max of proton dissipation was 2-fold higher in NaCl-tolerant calli when compared to the control. H+-coupled exchange was specific for Na+ and Li+ and not for K+. The increase of both the pH gradient across the tonoplast and the Na+/H+ antiport activity in response to salt strongly suggests that Na+ sequestration into the vacuole contributes to salt tolerance in potato.