Differential expression of LacdiNAc sequences (GalNAc (Jl-4GlcNAc-R) in

'To whom correspondence should be addressed The lacdiNAc sequence GalNAcpi->4GlcNAcpi-R occurs in the N- and O-glycans of many glycoproteins in verte-brate and invertebrates. We now report that both human 293 cells and Chinese hamster ovary (CHO) cells contain a UDPGalNAc:GlcNAc pi,4 N-acetylgalactosaminyltrans-ferase (fJl,4GalNAcT) that forms the lacdiNAc sequence. The pi,4GalNAcT in CHO cells is distinct from pi,4 ga-lactosyltransferase in that the latter enzyme, but not the former, binds to a column of immobilized bovine ot-lactal-bumin. To determine whether endogenous glycoproteins in these cells contain lacdiNAc sequences, glycoproteins from 293 cells, CHO and Lec8 CHO cells were desialylated and passed over immobilized Wisteria floribunda agglutinin (WFA), a plant lectin with affinity for terminal GalNAc residues. WFA bound to-120 and-80 kDa glycoproteins in 293 cells and glycans from these glycoproteins contained lacdiNAc sequences. The-120 kDa glycoproteins in 293 cells bound by WFA is a mixture of both the lysosome-associated membrane glycoproteins LAMPs-1 and-2. WFA bound to two glycoproteins of-47 and-78 kDa in Lec8 CHO cells, but these glycoproteins are not LAMPs and they do not contain the lacdiNAc sequence. Instead, they contain multiple GalNAca-Ser/Thr O-glycans that pro-mote binding to WFA. Thus, the pl,4GalNAcT in 293 cells displays a limited specificity in its recognition of acceptors, whereas the pl,4GalNAcT in CHO cells fails to promote synthesis of the cognate lacdiNAc sequence. The presence of the pi,4GalNAcT may not be sufficient for synthesis of lacdiNAc sequences and other factors may contribute to regulate the functionality of the enzyme. Key words: glycoprotein/lacdiNAc/synthesis/expressio