High V‐PPase activity is beneficial under high salt loads, but detrimental without salinity.

Summary: The membrane‐bound proton‐pumping pyrophosphatase (V‐PPase), together with the V‐type H⁺‐ATPase, generates the proton motive force that drives vacuolar membrane solute transport. Transgenic plants constitutively overexpressing V‐PPases were shown to have improved salinity tolerance, but the relative impact of increasing PP_i hydrolysis and proton‐pumping functions has yet to be dissected. For a better understanding of the molecular processes underlying V‐PPase‐dependent salt tolerance, we transiently overexpressed the pyrophosphate‐driven proton pump (NbVHP) in Nicotiana benthamiana leaves and studied its functional properties in relation to salt treatment by primarily using patch‐clamp, impalement electrodes and pH imaging. NbVHP overexpression led to higher vacuolar proton currents and vacuolar acidification. After 3 d in salt‐untreated conditions, V‐PPase‐overexpressing leaves showed a drop in photosynthetic capacity, plasma membrane depolarization and eventual leaf necrosis. Salt, however, rescued NbVHP‐hyperactive cells from cell death. Furthermore, a salt‐induced rise in V‐PPase but not of V‐ATPase pump currents was detected in nontransformed plants. The results indicate that under normal growth conditions, plants need to regulate the V‐PPase pump activity to avoid hyperactivity and its negative feedback on cell viability. Nonetheless, V‐PPase proton pump function becomes increasingly important under salt stress for generating the pH gradient necessary for vacuolar proton‐coupled Na⁺ sequestration. [ABSTRACT FROM AUTHOR]