GLYCOSYLATION PATTERN AND PROCESSING OF ENVELOPE GENE-PRODUCTS ENCODED BY GLYCOSYLATION MUTANTS OF FRIEND SPLEEN FOCUS-FORMING VIRUS

The protein encoded by the envelope gene of Friend spleen focus-formin g virus is responsible for the acute leukaemogenicity of this virus. I n order to correlate glycosylation and intracellular processing of thi s protein with viral pathogenicity, envelope gene products of pathogen ic and apathogenic glycosylation mutants were expressed in Rat-I cells and metabolically labelled with [6-H-3]glucosamine. Following immunop recipitation, primary and secondary gene products (gp55, gp65) were se parated by preparative polyacrylamide gel electrophoresis. Oligosaccha rides were released from tryptic glycopeptides by treatment with endo- beta-N-acetyl-glucosaminidase H (gp55), peptide-N4-(N-acetyl-beta-gluc osaminyl)asparagine amidase F (gp65) or by reductive beta-elimination. Resulting glycans were characterized by cochromatography with authent ic oligosaccharide standards using different HPLC systems and digestio n with exoglycosidases. The results revealed that the primary envelope gene products of pathogenic glycosylation mutants were, in part, furt her processed in Rat-1 cells similar to wild-type glycoprotein, result ing in polypeptides carrying complex-type N-glycans as well as partial ly sialylated O-linked oligosaccharides. In contrast, corresponding gl ycoproteins encoded by apathogenic mutants were found to remain at the level of the primary translation product exclusively comprising high- mannose-type N-glycans. Hence, intracellular maturation of the envelop e gene products in this model cell line seems to correlate with the in vivo pathogenicity of the glycosylation mutants studied.