title: Role of glycosylation in expression of functional diphtheria toxin receptors.
abstract: We have previously demonstrated, by using a detergent-solubilized system, the existence of specific diphtheria toxin-binding glycoproteins on the surface of toxin-sensitive cells. We have now tested the effect of tunicamycin treatment on the sensitivity of cells in culture to diphtheria toxin and have investigated the toxin sensitivity of mutant cells with known defects in glycosylation of asparagine-linked glycoproteins. Treatment of CHO-K1 cells with tunicamycin, which blocks the synthesis of both high-mannose-type and complex-type oligosaccharide chains of asparagine-linked glycoproteins, resulted in a 50- to 100-fold decrease in sensitivity to diphtheria toxin. In contrast, CHO-K1 mutants, defective in the synthesis of either high-mannose-type or complex-type oligosaccharides, showed no difference in toxin sensitivity compared with that of their parental cell lines. When we used an acid shock system, which is believed to result in receptor-dependent direct toxin penetration at the cell surface, the toxin sensitivity of tunicamycin-treated cells was not restored to that of untreated cells, suggesting that tunicamycin treatment results in a decrease in functional toxin receptors. Direct binding studies with 125I-labeled toxin demonstrated that this decrease in functional receptors is due to a decrease in the affinity of the receptors rather than to a change in the number of receptors. Taken together, these data are consistent with the interpretation that the diphtheria toxin receptor is a glycoprotein and suggest that the toxin binds neither to carbohydrate residues unique to the high-mannose-type oligosaccharides nor to those unique to the complex-type oligosaccharides. Furthermore, these data are consistent with the hypothesis that diphtheria toxin binds to the peptide backbone of the glycoprotein receptor.
mesh_terms: D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D003412:Cricetulus; D004167:Diphtheria Toxin / Q000378:metabolism* / Q000633:toxicity; D005260:Female; D006023:Glycoproteins / Q000096:biosynthesis / Q000378:metabolism*; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D007700:Kinetics; D009154:Mutation; D009844:Oligosaccharides / Q000378:metabolism; D010053:Ovary; D011956:Receptors, Cell Surface; D011950:Receptors, Cholinergic / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D017026:Swainsonine; D014415:Tunicamycin / Q000494:pharmacology
chemical_list: D000470:Alkaloids; D004167:Diphtheria Toxin; D006023:Glycoproteins; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D009844:Oligosaccharides; D011956:Receptors, Cell Surface; D011950:Receptors, Cholinergic; D014415:Tunicamycin; D017026:Swainsonine