GLYCOSYLATION OF BETA-1 INTEGRINS IN B16-F10 MOUSE MELANOMA-CELLS AS DETERMINANT OF DIFFERENTIAL BINDING AND ACQUISITION OF BIOLOGICAL-ACTIVITY

Studying B16-F10 cells we could identify beta-1 integrins as laminin, fibronectin and collagen receptors. Gradient ionic strength elution an alysis of affinity chromatography showed differential interactions bet ween laminin-binding beta-1 integrins (two beta-1 polypeptides of 105 and 120 kDa) and fibronectin and collagen-binding beta-1 integrins (el ution of one major beta-1 polypeptide of 120 kDa) and their respective ligands. To evaluate this diversity we submitted B16-F10 extracts to IEF and SDS-PAGE and found that one beta-1 integrin formed acidic and larger isoforms, while another formed basic and smaller isoforms. To s tudy this difference we also submitted material eluted from WGA-Sephar ose columns to IEF but now only the acidic beta-1 isoform was found. E xtracts of B16-F10 treated with neuraminidase showed only the basic be ta-1 isoform, suggesting that terminal sialic acid residues may be res ponsible for this acidic pattern, an interpretation supported by the f act that MAA (Maackia ammurensis agglutinin) reads only with the acidi c isoform. Differential glycosylation of beta-1 integrin isoforms in B 16-F10 was also demonstrated since the smaller laminin-binding beta-1 integrin isoform reacted only with GNA (Galanthus nivalis agglutinin), whereas the mature larger form reacted with DSA (Datura stramonium ag glutinin) and MAA; thus this heterogeneity of beta-1 chains is essenti ally due to variable glycosylation. Autoradiography and immunoblotting analysis of material separated by 2-dimensional electrophoresis show that only the processed forms of beta-1 integrins are expressed at the cell surface. (C) 1995 Wiley-Liss, Inc.