Translocation of newly synthesized gangliosides to the cell surface.

A new method was developed to follow the translocation of gangliosides from their site of synthesis within the cell to the plasma membrane. Cultured mouse neuroblastoma N18 and rat glioma C6 cells were labeled for increasing times with D- [1-3H]galactose and then subjected to mild oxidation with NaIO4. Under the conditions chosen, oxidation was essentially restricted to cell-surface sialic acid residues, which were converted to derivatives with an aldehyde function. The labeled gangliosides were isolated from the cells and reacted with dinitrophenylhydrazine to form dinitrophenyl (DNP) derivatives of the oxidized gangliosides. The DNP-gangliosides then were separated from their unmodified counterparts by thin-layer chromatography. Thus, the rate of labeling of surface gangliosides was distinguished from the rate of labeling of total gangliosides. Our results indicated that the transfer of gangliosides from the site of synthesis to the cell surface required approximately 20 min and that newly synthesized gangliosides appeared to be transported to the plasma membrane at a constant rate. No essential differences were found in the rates of translocation of different ganglioside species by N18 cells or between gangliosides of N18 and C6 cells.