Extensive radioactive labeling of glycolipids in cultured hamster fibroblasts by incubation of whole cells with UDP-[14C]glucose and UDP-[14C]galactose.

NIL 2cl cells, a cloned hamster cell line, are known to have seven classes of neutral glycosphingolipids in addition to hematoside. In this paper, I examined the participation of cell surface glycosyltransferases in the glycolipid synthesis in NIL cells. The cells, harvested from culture bottles by the use of chelating agents of divalent cations, incorporated radioactive glucose and galactose from labeled UDPglucose and UDPgalactose into endogenous glycolipid acceptors to form all eight classes of glycosphingolipids and dolichyl phosphate glucose. The major part of the incorporation occurred by glycosyltransferase reactions from nucleotide sugars, as indicated by a slight inhibition of the incorporation observed in the presence of a large excess of sugars or sugar phosphates. Acid hydrolysis of the glycolipid products showed that 91.4% of the 14C label was in the galactose and glucose moieties of the glycolipids. Exogenously added lactosylceramide stimulated the synthesis of trihexosylceramide in the absence of detergents. Th results presented in this paper suggest that incubation of whole cells with an equilibrium mixture of radioactive UDPglucose and UDPgalactose is a useful radioactive labeling method to determine the glycolipid composition of a cultured cell.