Identification with monoclonal antibody 140.240 of a structural variant of melanotransferrin shed by human melanoma cell lines in vitro.
Shedding by cultured human melanoma cells of a well-characterised cell- surface glycoprotein antigen known as "melanotransferrin" was studied with two monoclonal antibodies, 140.240 and 96.5. By means of [35S]-cysteine metabolically-labelled melanoma cells and immunoprecipitation studies, identification was made, by 140.240 in the spent media of two of six melanoma cell lines, of a new molecule of 100-kDa, aside from the 88-kDa molecule. Only the 88-kDa shed molecule was detected in the remaining four melanoma cell lines with both antibodies. None of nine clonal sublines derived from the two melanoma cell lines were found to shed the 100-kDa or 88-kDa molecule exclusively. Both shed antigens were released spontaneously to the medium from the live melanoma cells rather than as a result of cell death and lysis, since there was no obvious cell death or debris in the spent medium nor in the monolayer cells detected at the time of spent medium collection. Digestion of the isolated 100-kDa and 88-kDa shed molecules with N-glycanase followed by SDS-polyacrylamide gel electrophoresis resulted in the appearance of a single band of the 77-kDa molecule, which is deduced to be the polypeptide precursor of the cell-associated 87-kDa antigen. It is concluded that some melanoma lines shed the variant 100-kDa molecule, in addition to the 88-kDa molecule, and that both shed molecules and their cellular counterpart 87-kDa differ in their degrees of glycosylation.
Liao
S K
SK
Tumor Immunology Laboratory, Graduate Institute of Clinical Medicine, Chang Gung College of Medicine and Technology, Kweishan, Taoyuan, Taiwan, Republic of China.
D000581:Amidohydrolases / Q000378:metabolism; D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm / Q000032:analysis*; D003545:Cysteine / Q000378:metabolism; D006801:Humans; D008545:Melanoma / Q000737:chemistry* / Q000276:immunology; D058950:Melanoma-Specific Antigens; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000032:analysis* / Q000378:metabolism; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D013462:Sulfur Radioisotopes; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins; D013462:Sulfur Radioisotopes; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D003545:Cysteine