Asparagine-linked oligosaccharides in murine tumor cells: Comparison of a WGA-resistant (WGA(r)) nonmetastatic mutant and a related WGA-sensitive (WGA(s)) metastatic line

Abstract

MDW40, a wheat germ agglutinin-resistant (WGA(r)) mutant of the highly metastatic tumor cell line called MDAY-D2, is restricted to local growth at the subcutaneous site of inoculation. The WGA(r) tumor cells acquire metastatic ability by fusing spontaneously with a normal host cell followed by chromosome segregation, a process accompanied by reversion of the WGA(r) phenotype (i.e., WGA(s)). Since lectin-resistant mutant cell lines often have oligosaccharide alterations that may affect membrane function and consequently metastatic capacity, we compared the major Asn-linked glycopeptides in WGA(r) and WGA(s) cell lines. [2-3H]mannose-labeled glycopeptides were separated into four fractions on a DEAE-cellulose column and then further fractionated on a concanavalin A-Sepharose column. Glycopeptide structures were determined by: (a) sequential exoglycosidase digestion followed by chromatography on lectin/agarose and Bio-Gel P-4 columns and (b) proton nuclear magnetic resonance analysis. The metastatic WGA(s) cells had a sialylated poly-N-acetyllactosamine-containing glycopeptide which was absent in the nonmetastatic mutant cell line. Unique to the mutant was a neutral triantennary class of glycopeptide lacking sialic acid and galactose; the WGA(r) lesion therefore appeared to be a premature truncation of the antennae of the poly-N-acetyllactosamine-containing glycopeptide found in the WGA(s) cells. High mannose glycopeptides containing five to nine mannose residues constituted a major class in both WGA(r) and WGA(s) cells. Lysates of both wild-type and mutant cells had similar levels of galactosyltransferase activity capable of adding galactose to the N-acetylglucosamine-terminated glycopeptide isolated from mutant cells; the basis of the WGA(r) lesion remains to be determined.