Isolation of a temperature-sensitive FM3A mutant deficient in asparagine-linked glycosylation by selecting for resistance to tritiated mannose suicide.
Protein glycosylation mutants in the mouse mammary carcinoma cell line FM3A were selected for ability to withstand exposure to [2-3H]mannose at 39 degrees C. G258 , one of the mutant cells isolated, has been characterized. G258 cells were temperature-sensitive for cell growth. Moreover, G258 cells showed temperature sensitivity for [3H]mannose incorporation into the TCA-insoluble fraction. To study the biochemical basis of the defect in glycoprotein biosynthesis, the formation of lipid-linked saccharides was examined. The results showed that the formation of lipid-linked oligosaccharides was severely inhibited in G258 cells at 39 degrees C. At 33 degrees C, G258 cells synthesized Glc3Man9GlcNAc2-PP-Dol, the fully assembled lipid-linked oligosaccharides, but at 39 degrees C, G258 cells were able to synthesize merely the smaller lipid-linked oligosaccharides (approximately up to Man3GlcNAc2 -PP-Dol), but were unable to synthesize the larger lipid-linked oligosaccharides.
D000818:Animals; D002455:Cell Division; D002460:Cell Line; D005260:Female; D006017:Glycolipids / Q000032:analysis; D006023:Glycoproteins / Q000096:biosynthesis*; D008325:Mammary Neoplasms, Experimental; D051379:Mice; D009154:Mutation; D011105:Polyisoprenyl Phosphate Sugars / Q000096:biosynthesis*; D013696:Temperature
D006017:Glycolipids; D006023:Glycoproteins; D011105:Polyisoprenyl Phosphate Sugars