Biosynthesis of murine immunoglobulin D: Heterogeneity of glycosylation

Abstract

AbstractThe biosynthesis of IgD was studied with special reference to aspects of glycosylation in the B1‐8 δ cell line. This cell line was isolated as a spontaneous class switch variant from an IgM‐producing cell line, B1‐8.μ. B1‐8.δ produces both membrane and secretory IgD. The biosynthesis of IgD is unusual in that there is a large degree of heterogeneity in N‐linked glycosylation, most likely involving the number of glycan sidechains that are attached. Similar findings have recently been documented for human IgD. Further modifications of the oligosaccharide side chains follow the pathways established for numerous other glycoproteins. Under conditions where N‐linked glycosylation was inhibited by tunicamycin (TM), the presence of neuraminidase‐sensitive forms of secretory IgD could be shown. Moreover, secretory IgD from TM‐treated cells was susceptible to mild alkaline hydrolysis. Taken together, these findings argue strongly for the presence of O‐linked, sialic acid‐carrying sugars on murine IgD. Both for O‐ and N‐linked carbohydrate side chains, terminal modifications may occur at a different rate and extent for membrane and secretory IgD. For reasons of comparison we also examined some aspects of IgM biosynthesis in the parental cell line and found our results to be in good aggreement with those reported in the literature. Biosynthesis of H‐2K, D antigens was indistinguishable in B1‐8.H μ and B1‐8. δ, suggesting that the unusual features of IgD glycosylation are inherent to this Ig isotype, rather than the consequence of a different complement of glycosyltransfer‐ases or carbohydrate‐modifying enzymes in the Bl‐8.δ cell line. The use of affinity matrices for purification of immunoglobulins allowed us to estimate relative rates for the formation of IgM and IgD molecules capable of binding antigen. The results obtained suggest that Bl‐8 IgM and IgD follow different assembly pathways, IgM most likely through heavy chain dimer formation followed by light chain addition, and IgD through heavy chain‐light chain dimer formation, followed by assembly into full‐sized molecules. Since B1‐8.μ and B1‐8.δ express identical variable regions and light chains, such differences must be attributed to isotypic differences. Inhibition of glycosylation affected neither secretion nor assembly of either IgM or IgD.