SORTING OF NEWLY SYNTHESIZED GALACTOSPHINGOLIPIDS TO THE 2 SURFACE DOMAINS OF EPITHELIAL-CELLS

The high concentration of glycosphingolipids on the apical surface of epithelial cells may be generated by selective transport from their si te of synthesis to the cell surface. Previously, we showed that canine kidney MDCK and human intestinal Caco-2 cells converted a ceramide ca rrying the short fluorescent fatty acid C-6-NBD to glucosylceramide (G lcCer) and sphingomyelin (SM), and that GlcCer was preferentially tran sported to the apical surface as compared to SM. Here, we address the point that not all glycosphin-golipid classes are apically enriched in epithelia. We show that a ceramide containing the 2-hydroxy fatty aci d C6OH was preferentially converted by MDCK and Caco-2 cells to galact osylceramide (Ga(2)Cer) and its derivatives galabiosylceramide (Ga(2)C er) and sulfatide (SGalCer) as compared to SM and GlcCer-all endogenou s lipid classes of these cells. Transport to the apical and basolatera l cell surface was monitored by a BSA-depletion assay. In MDCK cells, GalCer reached the cell surface with a two- to sixfold lower apical/ba solateral polarity than GlcCer. Remarkably, in Caco-2 cells GalCer and GlcCer displayed the same apical/basolateral polarity, but it was six fold lower for lipids with a C6OH chain than for C-6-NBD lipids. There fore, the sorting of a sphingolipid appears to depend on lipid structu re and cell type. We propose that the different ratios of gluco- and g alactosphingolipid synthesis in the various epithelial tissues govern lipid sorting in the membrane of the trans Golgi network by dictating the composition of the domains from where vesicles bud to the apical a nd basolateral cell surface.