Enzymatic basis for a lectin-resistant phenotype: increase in a fucosyltransferase in mouse melanoma cells.

Abstract

In the search for the biochemical basis of the control of glycosylation of cell surface carbohydrates, revertant clones were isolated from previously characterized wheat germ agglutinin-resistant clones of B16 mouse melanoma cells by selection for resistance to Lotus tetragonolobus lectin or to ricin. Comparison of the wheat germ agglutinin-resistant clones with the parent and revertant clones indicated that this phenotype was correlated with an increased sensitivity to the Lotus lectin, a 60-to 70-fold increase in a1 -* 3 fucosyltransferase activity and a decreased sialic acid content of the N-glycosidic chains of glycoproteins . The results suggest a novel type of control mechanism for lectin resistance, an increase in a glycosyltransferase activity . The presence of a1 ---> 3 bound fucose on N-acetylglucosamine residues would interfere with the addition of sialic acid by a2--> 3 linkages to galactose residues in the carbohydrate units, and this change could explain the resistance to wheat germ agglutinin and the increased sensitivity to the Lotus lectin . A change in a regulatory gene for the fucosyltransferase as a possible primary cause for the changed phenotype is discussed.The availability of lectin-resistant cell lines with altered carbohydrate moieties of cell surface glycoproteins and glycolipids has provided a powerful tool for the study of the biosynthesis and function of the complex carbohydrates in animal cells (1-4). Some of the variant cell lines appear to have a block at some level of the biosynthetic pathway of protein-bound oligosaccharides, but the enzymatic basis for the changed phenotype is usually not known . For the study of the biological role of glycoproteins and glycolipids, it would be important to know how the expression of cell surface carbohydrates is regulated.Many lectin-resistant variant cell lines differ from the parent lines with respect to more than one property (5-7). Because the occurrence of more than one genetic or epigenetic change even in single-step isolates cannot be ruled out, it is not known whether all the changes observed could be ascribed to one primary change. Other evidence should therefore be obtained by the isolation of independent variants or revenants (7,8) . The isolation of independent variants has so far been shown only for the lectin-resistant cells with a block in a specific Nacetylglucosaminyltransferase (9, 10), whereas the isolation of revenants has usually not been successful (1, 7) . We report here the isolation of variants and revenants that could be obtained repeatedly as well as their enzymic defect .Previous work from this laboratory has described a wheat THE JOURNAL OF CELL BIOLOGY " VOLUME 92 FEBRUARY 1982 277-282 © The Rockefeller University Press -0021-9525/82/02/0277/06 $1 .00 germ agglutinin (WGA)-resistant mouse melanoma cell line with many altered properties, including changes in cell adhesion and metastasis (11) . Analysis of the protein-bound carbohydrates revealed a structural change involving increased fucose-content and de...

Wheatgerm agglutinin-mediated toxicity in pancreatic cancer cells

“…When WGA was used to generate lectin-resistant clones of cultured cancer cells, most resulting clones showed a remarkable loss of metastatic capabilities (Stanley et al, 1980;Kerbel et al, 1982;Ishikawa et al, 1988;Ishikawa and Kerbel, 1989;Kim et al, 1993). The abnormal truncation of asparagine-linked cell surface carbohydrate with loss of sialylated poly-N-acetyllactosamine complexes and the resulting increase in adherence to laminin, fibronectin and type-4 collagen appears to account for this loss of metastatic potential in WGA-resistant clones (Finne et al, 1980(Finne et al, , 1982Dennis, 1985Dennis, , 1986a. Therefore, the determination of the presence of sialic acid binding sites on cancer cell surface carbohydrate may carry important functional information.…”

Cellular sialic acid level and phenotypic expression in B16 melanoma cells: Comparison of spontaneous variations and bromodeoxyuridine- and theophylline-induced changes.

“…Higher adhesiveness of amelanotic and BrdU-treated melanotic cells could be explained by their lower sialic acid levels, as with the case of lectin-resistant cells (2,25). Galactosyl residues exposed as a result of desialylation (detected as lower sialic acid levels) and lectin-like molecules recognizing the exposed sugar moieties (9, 10) might be therein involved.…”

“…In melanoma cells, involvement of sialic acid residues has been well documented in determining metastatic potential (13). A lectin (WGA)-resistant phenotype and the related alteration of adhesive properties have also been shown to be due to a decrease in cell surface sialic acid content (2,25). However, little has been known of the functional significance of sialic acid relating to other aspects of melanoma cell phenotypes.…”

Protein Glycosylation and Cancer

“…Transfection of a glioma cell line, U373 MG cells with u2,6SA-T reduced invasion [82]. However, a2,6-sialylation in H-RAS-transformed rat fibroblasts correlates positively with invasive potential [ 831, and loss of sialylation in B16 melanoma mutants due to over-expression of al,3Fuc-T results in loss of metastatic potential [38]. Similarly, metastatic properties of B16 melanoma cells were reduced in cells transfected with either al,2-Fuc-T or al,3Gal-T [ 841.…”