Expression of human alpha-L-fucosyltransferase gene homologs in monkey kidney COS cells and modification of potential fucosyltransferase acceptor substrates by an endogenous glycosidase

Previous investigations on the monkey kidney COS cell line demonstrated the weak expression of fucosylated cell surface antigens and presence of endog enous fucosyltransferase activities in cell extracts, RT-PCR analyses have now revealed expression of five homologs of human fucosyltransferase genes, FUT1, FUT4, FUT5, FUT7, and FUT8, in COS cell mRNA, The enzyme in COS cell extracts acting on unsialylated Type 2 structures is closely similar in it s properties to the alpha 1,3-fucosyltransferase encoded by human FUT4 gene and does not resemble the product of the FUT5 gene. Although FUT1 is expre ssed in the COS cell mRNA, it has not been possible to demonstrate alpha 1, 2-fucosyltransferase activity in cell extracts but the presence of Le(y) an d blood-group A antigenic determinants on the cell surface imply the format ion of H-precursor structures at some stage. The most strongly expressed fu cosyltransferase in the COS cells is the alpha 1,6-enzyme transferring fuco se to the innermost N-acetylglucosamine unit in N-glycan chains; this enzym e is similar in its properties to the product of the human FUT8 gene. The e nzymes resembling the human FUT4 and FUT8 gene products both had pH optima of 7.0 and were resistant to 10 mM NEM, The incorporation of fucose into as ialo-fetuin was optimal at 5.5 and was inhibited by 10 mM NEM. This result initially suggested the presence of a third fucosyltransferase expressed in the COS cells but we have now shown that triantennary N-glycans with termi nal nonreducing galactose units, similar to those present in asialo-fetuin, are modified by a weak endogenous beta-galactosidase in the COS cell extra cts and thereby rendered suitable substrates for the alpha 1,6-fucosyltrans ferase.