Analysis of the role of glycosylation of the human fibronectin receptor.
1-Deoxymannojirimycin (MNJ), an inhibitor of Golgi alpha-mannosidase IA and IB, was used to assess the possible roles of asparagine-linked oligosaccharides in the structure and function of the integrin fibronectin receptor from cultured human fibroblasts. These cells normally attach well to fibronectin substrates and have only mature forms of the fibronectin receptor on their surfaces. MNJ inhibits the intracellular trimming of high mannose oligosaccharides, and cells treated with 0.2 mg/ml MNJ synthesize only immature precursor forms of both the alpha and beta subunits of the fibronectin receptor. The immature receptor polypeptides were found to be nonfunctional by two criteria: 1) cells treated with MNJ attached poorly to fibronectin substrates; and 2) receptor from the treated cells was defective in binding to fibronectin affinity columns. The precursor forms of the fibronectin receptor subunits were found on the surfaces of cells treated with MNJ, demonstrating that processing of receptor carbohydrates to mature forms was not necessary for receptor insertion into the plasma membrane. A monoclonal antibody that specifically bound the alpha subunit of the fibronectin receptor immunoprecipitated both alpha and beta subunit polypeptides from both control cells and cells treated with MNJ. Similarly, a monoclonal antibody that specifically bound only the beta subunit also immunoprecipitated both alpha and beta subunit polypeptides of the receptor from extracts of both control and MNJ-treated cells. These results indicate that receptor assembly can occur in the absence of complete oligosaccharide processing. Thus, oligosaccharide processing to the mature form of the fibronectin receptor is important for its binding function but not for receptor assembly or insertion into the plasma membrane.
Akiyama
S K
SK
Department of Oncology, Howard University Cancer Center, Howard University College of Medicine, Washington, D. C. 20060.
Yamada
S S
SS
Yamada
K M
KM
D017485:1-Deoxynojirimycin; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D005347:Fibroblasts / Q000276:immunology; D005353:Fibronectins / Q000378:metabolism; D005944:Glucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D006020:Glycopeptides / Q000302:isolation & purification; D006031:Glycosylation; D006801:Humans; D007700:Kinetics; D008297:Male; D008715:Methionine / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D012867:Skin / Q000276:immunology
D000900:Anti-Bacterial Agents; D000911:Antibodies, Monoclonal; D005353:Fibronectins; D006020:Glycopeptides; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic; D017485:1-Deoxynojirimycin; D008715:Methionine; D005944:Glucosamine