Glycosylation of the epidermal growth factor receptor and its relationship to membrane transport and ligand binding.
Epidermal growth factor (EGF) receptor biosynthesis was examined in an oral squamous cell carcinoma line, NA, which overproduces the receptor to an even greater extent than the widely studied A431 cells. The EGF receptor of NA cells synthesized in the presence of tunicamycin had an apparent molecular weight of 130,000. The nascent protein in untreated cells was cotranslationally glycosylated to Mr 160,000 and further processed to Mr 170,000. The endo-beta-N-acetylglucosaminidase H (Endo H) digestion analysis revealed the presence of high mannose type oligosaccharide on the Mr 170,000 mature receptor. We extended the analysis by correlating the biosynthesis with the acquisition of binding activity. The unglycosylated Mr 130,000 receptor and the Mr 160,000 receptor seen after pulse-labeling had no EGF binding activity, whereas the Mr 160,000 receptor seen after chase-incubation and the Mr 170,000 receptor had binding activity. Thus, not only glycosylation but also some oligosaccharide processing is apparently necessary for the EGF binding. Treatment with processing inhibitors, such as monensin, swainsonine and 1-deoxynojirimycin, affected neither receptor transport to the plasma membrane nor binding activity. Inhibition by 1-deoxynojirimycin is thought to be incomplete since the surface receptor in treated cells had the same molecular weight as that in control cells. An Mr 160,000 receptor without binding activity accumulated in the intracellular fraction in the presence of brefeldin A, an inhibitor of intracellular transport. Thus, the EGF binding activity is thought to be acquired after the brefeldin A-sensitive process but prior to the swainsonine-sensitive mannose removal in NA cells.
Gamou
S
S
Department of Molecular Biology, Keio University School of Medicine, Tokyo.
Shimizu
N
N
D000118:Acetylglucosaminidase / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D066246:ErbB Receptors / Q000096:biosynthesis / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007700:Kinetics; D008024:Ligands; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D010779:Photofluorography; D011233:Precipitin Tests; D014407:Tumor Cells, Cultured / Q000378:metabolism
D008024:Ligands; D066246:ErbB Receptors; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase