EFFECTS OF MONENSIN ON GLYCOSPHINGOLIPID METABOLISM IN CULTURED HUMAN PROXIMAL TUBULAR CELLS

Effects of monensin, a monovalent cationic ionophore which disrupts Golgi apparatus and its related functions, on glycosphingolipid (GSL) metabolism were investigated in cultured human proximal tubular (PT) cells. Monensin (10(-6)M) stimulated [H-3]Gal incorporation into GlcCer, GalCer and LacCer by 8.5-fold and 15-fold, respectively, in PT cells as compared to control. In contrast, [H-3]Gal incorporation into GbOse(3)Cer and GM(3) remained unchanged and that into GbOse(4)Cer was decreased 2-fold as compared to control. GSL measured by HPLC revealed that in cells incubated with monensin, GlcCer, GalCer and LacCer levels were increased 1.6-fold and 7-fold, respectively, whereas GbOse(3)Cer and GbOse(4)Cer levels were decreased several folds. Cells incubated with monensin contained 2.5- to 3-fold higher activity of alpha-galactosidase, beta-galactosidase and beta-glucosidase than control, whereas the activity of UDP-gal: glucosylceramide galactosyltransferase (beta-GalT-2) was 8-fold lower than control cells. Cells incubated with monensin took up and degraded one-half as much I-125-LDL as that of control cells. In control cells, exogenously derived [H-3]LacCer on LDL was rapidly taken up and catabolized to monoglycosylceramide, or it was used for the endogenous synthesis of globotriosylceramide (trihexosylceramide), globotetraosylceramide (tetrahexosylceramide) and a ganglioside, GM(3). In contrast, cells incubated with monensin accumulated most of the [H-3]LacCer-LDL. Exogenously derived [H-3]LacCer on LDL was catabolized to GlcCer, but was not utilized, for the synthesis of globotriosylceramide, globotetraosylceramide and GM(3) in cells incubated with monensin. We conclude that monensin-induced increase in the cellular levels of LacCer in human PT cells is not due to increased endogenous synthesis, but due to increased catabolism of GbOse(3)Cer and storage of exogenous LacCer.