Sodium transport by endocytic vesicles in cultured Arabidopsis thaliana (L.) Heynh. cells

Abstract

The involvement of endocytosis in Na+ internalization by suspension-cultured Arabidopsis thaliana (L.) Heynh. cells under salt stress was investigated. Using epifluorescence and confocal laser-scanning microscopy in co-localization experiments with a marker of the endocytic structures, N-(3-triethylammoniumpropyl)-4-(6-(4-(diethylamino) phenyl) hexatrienyl) pyridinium dibromide (FM 4-64), and a membrane-impermeable Na+ indicator, Asante Natrium Green-2 tetramethylammonium salt (ANG-2 TMA), intracellular Na+ ion uptake via endocytosis, but not through the plasma-membrane ion channels or transporters, was detected. The punctate fluorescence of the two markers was co-localized in the microvacuoles (MVs) with sizes up to 5 μm. The study of cell ultrastructure by transmission electron microscopy revealed the fusion of MVs into larger ones during an incubation of the cells in medium containing 100 mM NaCl. Wortmannin, an inhibitor of multivesicular body/late endosome/pre-vacuolar compartment fusion with vacuoles, substantially lowered intracellular Na+ content. Apparently, endocytic Na+ uptake and subsequent fusion of Na+-containing membrane structures provided direct transport of external Na+ ions into the vacuoles, bypassing the cytosol.