Complexity of MAM-6, an epithelial sialomucin associated with carcinomas.
The complexity of epithelial sialomucins was investigated by immunoprecipitation and membrane immunofluorescence, using monoclonal antibodies (MAbs) against MAM-6 and other sialomucins. MAbs against MAM-6 immunoprecipitated from a variety of sources either one or two sialylated glycoproteins with apparent molecular weights of over 400,000 under reducing as well as nonreducing conditions. The electrophoretic mobility of each MAM-6 glycoprotein as isolated from serum, milk, and cell lines of different individuals showed considerable variation. The differences in molecular weight of the MAM-6 glycoproteins were also reflected at the level of MAM-6 precursors which are less heavily glycosylated. Therefore, large differences in apparent molecular weight (150,000 and over) are most likely due to a variable protein backbone. We used this molecular polymorphism to prove that 11 MAbs against different sialomucins, obtained from various investigators, precipitated sialomucins generated from common precursor molecules. The pattern of reactivity of the MAbs with carcinoma cell lines was complex. All but the two MAbs, directed against putative carbohydrate epitopes, immunoprecipitated the precursor molecule from each cell line. However, some of them were unable to immunoprecipitate the mature form of MAM-6 from these cell lines. These results indicate that those epitopes are masked, probably due to cell line- or possibly cell type-dependent variations in glycosylation of the epithelial sialomucin. Even within a single cell line mature molecules with different epitopes were observed. The differential reactivity of the MAbs was confirmed by membrane immunofluorescence. These results show that MAM-6 belongs to a family of epithelial sialomucins with a polymorphic protein backbone and extensive variation in glycosylation.
Hilkens
J
J
Division of Tumor Biology, The Netherlands Cancer Institute (Antoni van Leeuwenhoek Huis), Amsterdam.
Buijs
F
F
Ligtenberg
M
M
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000951:Antigens, Neoplasm / Q000032:analysis*; D001943:Breast Neoplasms / Q000032:analysis; D002460:Cell Line; D005260:Female; D005455:Fluorescent Antibody Technique; D006801:Humans; D007118:Immunoassay; D008562:Membrane Glycoproteins / Q000276:immunology / Q000302:isolation & purification*; D008970:Molecular Weight; D018396:Mucin-1; D009077:Mucins / Q000032:analysis*; D052116:Sialomucins
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000951:Antigens, Neoplasm; D008562:Membrane Glycoproteins; D018396:Mucin-1; D009077:Mucins; D052116:Sialomucins