Abnormal surface expression of sialoglycans on B lymphocyte cell lines from patients with carbohydrate deficient glycoprotein syndrome I A (CDGS I A).

The carbohydrate-deficient glycoprotein syndromes (CDGS) are genetic, multisystemic diseases characterized by deficiencies in the glycosylation of many secretory glycoproteins, lysosomal enzymes, and possibly cell surface glycoproteins resulting in central nervous system abnormalities and frequent early death by infection. Here we examined whether membranous glycoconjugates of lymphocytes are affected by this disorder. For this, we analyzed cell surface-expressed sialoglycans of Epstein Barr virus (EBV)-transformed B cell lines derived from peripheral B lymphocytes of several patients with CDGS I A. These CDG-LCL (lymphoblastoid cell lines) expressed differentiation markers comparable to those of other EBV-transformed B cell lines. No apparent defects in the gross glycosylation process of defined complex glycosylated proteins such as the surface-expressed major histocompatibility complex class I glycoprotein or secreted immunoglobulin (IgM) were identified. However, using a novel flow cytometric enzyme assay to measure cell surface alpha2,6 sialylation on live cells we found that CDG-LCL express less alpha2,6 sialylated glycans in comparison to other EBV-transformed B cell lines. Also, CDG-LCL bound less of the B lymphocyte lectin CD22, specific for alpha2,6 sialylated lactosamines and known to modulate B cell receptor mediated signaling, as demonstrated by using a soluble CD22-immunoglobulin fusion protein in flow cytometry. CDG-LCL showed stronger surface staining with the monoclonal antibody 1B2 which detects a distinct group of surface-expressed lactosaminyl epitopes. After pretreatment with neuraminidase of Newcastle disease virus (NDVN) it became apparent that in CDG-LCL a significantly larger portion of the 1B2 epitopes was sialylated in alpha2,3 linkage as compared to other B cell lines. Intracellular alpha2,6 sialyltransferase activity as well as polymerase chain reaction products specific for four different sialyltransferases did not significantly differ in CDG-LCL as compared to other EBV-B cell lines. Differences in sialylation may be caused by the respective oligosaccharide core structures available for alpha2,6 or alpha2,3 sialylation in CDG-LCL. Therefore, lymphocytes derived from CDGS patients have distinct deviations in their surface-expressed lactosaminoglycan structures which may affect functions as exemplified by reduced interactions of CD22 with its ligands.

Organisation of the murine 5-HT3 receptor gene and assignment to human chromosome 11.

We have isolated the murine gene encoding the 5-HT3 receptor (5-HT3R), a member of the ligand-gated ion channels, that mediates a variety of physiological effects in central and peripheral neurons. DNA sequence analysis of the 5-HT3R gene revealed its organisation in 9 exons distributed over approximately 12 kbp of DNA. Alternative use of exon 9 splice acceptor sites generated two 5-HT3R variants. The 5-HT3R gene, whose structure is closely related to neuronal and muscle AChR alpha genes, as demonstrated by four common splice junctions, was localised on human chromosome 11.