Stable expression of a cDNA encoding a human beta 1 --> 3galactosyltransferase responsible for lacto-series type 1 core chain synthesis in non-expressing cells: variation in the nature of cell surface antigens expressed.
Transient expression of a human colonic adenocarcinoma Colo 205 cell derived cDNA in cell lines which ordinarily express only neolacto-series glycolipids has resulted in the expression of a beta 1 --> 3galactosyltransferase gene responsible for synthesis of glycolipids based upon the lacto-series type 1 core chain. Calcium phosphate transfected cells were panned on anti-IgM coated plates after initial treatment with a combination of monoclonal antibodies specific for type 1 chain terminal structures (TE-3) and a very broadly specific antibody reactive with multiple type 1 chain derivatives (TE-2). Adherent cells after panning were capable of efficiently transferring Gal in beta 1 --> 3-linkage to the acceptor glycolipid Lc3. Using these reagents, clones of stably transfected human colonic adenocarcinoma HCT-15 cells were produced and isolated. Parental HCT-15 cells do not express type 1 chain based antigens. The nature of the type 1 chain based antigens produced in each of these clones was analyzed by solid phase antibody binding assays. Three types of behavior were observed. Formation of type 1 terminal structures that were either exclusively sialylated or fucosylated, or a mixture of sialylated and fucosylated determinants occurred. In contrast, no difference in type 2 antigen expression between any clone and the parental cells was observed. These data suggest that coordination of subsequent reactions capable of modifying type 1 chain structures is not the same in all clones. The relationship of these results to aspects of cellular regulation of carbohydrate biosynthesis is discussed.
Sherwood
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AL
Pacific Northwest Research Foundation, Seattle, Washington 98122.
Greene
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TG
Holmes
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EH
D000230:Adenocarcinoma; D000918:Antibody Specificity; D000954:Antigens, Surface / Q000096:biosynthesis* / Q000276:immunology; D002240:Carbohydrate Sequence; D002855:Chromatography, Thin Layer; D003110:Colonic Neoplasms; D004247:DNA; D005434:Flow Cytometry; D005700:Galactosyltransferases / Q000235:genetics* / Q000378:metabolism; D005732:Gangliosides / Q000378:metabolism; D006017:Glycolipids / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology; D006801:Humans; D008969:Molecular Sequence Data; D011863:Radioimmunoassay; D014162:Transfection; D014407:Tumor Cells, Cultured
D000954:Antigens, Surface; D005732:Gangliosides; D006017:Glycolipids; D004247:DNA; D005700:Galactosyltransferases