A Defective Vacuolar Proton Pump Enhances Aluminum Tolerance by Reducing Vacuole Sequestration of Organic Acids

Abstract

Plants cope with aluminum (Al) toxicity by secreting organic acids (OAs) into the apoplastic space, which is driven by proton (H⁺) pumps. Here, we show that mutation of vacuolar H⁺-translocating adenosine triphosphatase (H⁺-ATPase) subunit a2 (VHA-a2) and VHA-a3 of the vacuolar H⁺-ATPase enhances Al resistance in Arabidopsis (Arabidopsis thaliana). vha-a2 vha-a3 mutant plants displayed less Al sensitivity with less Al accumulation in roots compared to wild-type plants when grown under excessive Al³⁺ Interestingly, in response to Al³⁺ exposure, plants showed decreased vacuolar H⁺ pump activity and reduced expression of VHA-a2 and VHA-a3, which were accompanied by increased plasma membrane H⁺ pump (PM H⁺-ATPase) activity. Genetic analysis of plants with altered PM H⁺-ATPase activity established a correlation between Al-induced increase in PM H⁺-ATPase activity and enhanced Al resistance in vha-a2 vha-a3 plants. We determined that external OAs, such as malate and citrate whose secretion is driven by PM H⁺-ATPase, increased with PM H⁺-ATPase activity upon Al stress. On the other hand, elevated secretion of malate and citrate in vha-a2 vha-a3 root exudates appeared to be independent of OAs metabolism and tolerance of phosphate starvation but was likely related to impaired vacuolar sequestration. These results suggest that coordination of vacuolar H⁺-ATPase and PM H⁺-ATPase dictates the distribution of OAs into either the vacuolar lumen or the apoplastic space that, in turn, determines Al tolerance capacity in plants.