Membrane trafficking in soybean protoplasts

Abstract

Membrane trafficking in plant cells was investigated by examining soybean (Glycine max L.) cell suspension cultures. The convergence of the endocytic pathway and the lysosomal and secretory pathways was examined by incubating protoplasts in an electron-dense, adsorptive-endocytosis marker, cationized ferritin (CF), and histochemically labeling for acid phosphatase (AcPase) activity. Both labels were ultrastructurally co-localized in endocytic vesicles, the Golgi apparatus, multivesicular bodies (MVB), and the large central vacuole. Protoplasts were also incubated in CF and immunolabeled for glucosidase II (GSII), a glycosylation-trimming enzyme. CF and GSII were co-localized along the plasma membrane, in endocytic vesicles, the partially-coated reticulum, the Golgi apparatus, and MVB. The ability of cells to sort proteins and polysaccharides to the correct destination was investigated by incubating protoplasts in monensin, an ionophore, and CF and histochemically labeling for AcPase activity. Monensin inhibited the localization of AcPase in multivesicular bodies, indicating that there may be a monensin-sensitive sorting mechanism in plants Monensin did not inhibit the internalization of CF. Lucifer yellow (LY) internalization was examined for its usefulness as a fluid-phase marker. Monensin did not inhibit by internalization. Probenecid, an anion transport inhibitor, changed the localization of LY, suggesting that some BY is internalized by an anion transport mechanism. This would rule out LY as an appropriate fluid-phase marker in plant cells.