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 analysis of affinity chromatography showed differential interactions between laminin-binding beta-1 integrins (two beta-1 polypeptides of 105 and 120 kDa) and fibronectin and collagen-binding beta-1 integrins (elution 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 study this difference we also submitted material eluted from WGA-Sepharose columns to IEF but now only the acidic beta-1 isoform was found. Extracts of B16-F10 treated with neuraminidase showed only the basic beta-1 isoform, suggesting that terminal sialic acid residues may be responsible for this acidic pattern, an interpretation supported by the fact that MAA (Maackia ammurensis agglutinin) reacts only with the acidic isoform. Differential glycosylation of beta-1 integrin isoforms in B16-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 agglutinin) and MAA; thus this heterogeneity of beta-1 chains is essentially 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.
Veiga
S S
SS
Ludwig Institute for Cancer Research, São Paulo, Brazil.
Chammas
R
R
Cella
N
N
Brentani
R R
RR
D000818:Animals; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005353:Fibronectins / Q000378:metabolism; D027801:Galanthus; D006031:Glycosylation; D019012:Integrin beta1; D016023:Integrins / Q000302:isolation & purification / Q000378:metabolism*; D007525:Isoelectric Focusing; D007797:Laminin / Q000378:metabolism; D008546:Melanoma, Experimental / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008970:Molecular Weight; D009439:Neuraminidase; D011499:Protein Processing, Post-Translational; D012794:Sialic Acids / Q000032:analysis
D005353:Fibronectins; D019012:Integrin beta1; D016023:Integrins; D007797:Laminin; D012794:Sialic Acids; D009439:Neuraminidase