Electrochemical investigation of active malic acid transport at the tonoplast into the vacuoles of the CAM plantKalanchoë daigremontiana

Abstract

The membrane potential of cells in leaf slices of the CAM plantKalanchoe daigremontiana Hamet et Perrier in the light and in the dark is −200 mV on the average; it is reversibly depolarized by the metabolic inhibitors FCCP (5×10−6 m) and CN− (5×10−3 m); it shows the light-dependent transient oscillations ubiquitously observed in green cells; it is independent of the amount of malic acid accumulated in the cells (in a tested range between 30 and 140mm); and it is considerably hyperpolarized by the fungal toxin fusicoccin (30×10−6 m). Fusicoccin inhibits nocturnal malic acid accumulation in intact isolated phyllodia of the CAM plantKalanchoe tubiflora (Harv.) Hamet but does not affect remobilization of malic acid during the day. Electrochemical gradients for the various ions resulting from dissociation of malic acid, i.e., H+, Hmal− and mal2−, were calculated using the Nernst equation. With a very wide range of assumptions on cytoplasmic pH and malate concentration results of calculations suggest uphill transport of H+ and Hmal− from the cytoplasm into the vacuole, while mal2− might be passively distributed at the tonoplast. On the basis of the present data the most likely mechanism of active malic acid accumulation in the vacuoles of CAM plants appears to be an active H+ transport at the tonoplast coupled with passive movement of mal2− possibly mediated by a translocator (“catalyzed diffusion”), with subsequent formation of Hmal− (2 H++mal2−→H++Hmal−) at vacuolar pH's.