Topology of O-linked N-acetylglucosamine in murine lymphocytes.
A unique form of protein glycosylation in which N-acetylglucosamine monosaccharides are O-glycosidically linked to serine or threonine residues (O-GlcNAc) was initially reported in studies that used purified bovine milk galactosyltransferase to exogenously probe living populations of murine lymphocytes. However, in this same study, detergent latency experiments surprisingly indicated that unlike other known forms of protein glycosylation, O-GlcNAc-modified proteins occur predominantly intracellularly. Since we now know that as little as 5% lysis could have accounted for the putative cell surface O-GlcNAc seen in these earlier studies, and also in the light of recent data on the subcellular localization of the O-GlcNAc glycosyltransferase(s), we decided to critically reexamine the topology and polypeptide distribution of O-GlcNAc in primary cultures of murine lymphocytes that were prepared using improved cell selection techniques that do not involve complement-mediated lysis for cell enrichment. We have also examined two well-characterized T cell hybridoma lines. Under these highly stringent conditions of cell viability, we were unable to detect O-GlcNAc bearing proteins on the cell surfaces of any of these cell types. Also, O-GlcNAc was found on a similar subset of proteins in all of the various lymphocyte cell types. These data suggest that O-GLcNAc is highly restricted to the cytoplasmic/nucleoplasmic compartment of the cell and is found on a similar subset of nuclear and cytoplasmic proteins in functionally different types of lymphocytes.
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
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Kearse
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Hart
D000117:Acetylglucosamine / Q000737:chemistry*; D000818:Animals; D002460:Cell Line; D003600:Cytosol / Q000737:chemistry*; D006031:Glycosylation; D006825:Hybridomas / Q000737:chemistry; D008214:Lymphocytes / Q000737:chemistry* / Q000201:enzymology; D008565:Membrane Proteins / Q000737:chemistry; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D013154:Spleen; D013950:Thymus Gland
D008565:Membrane Proteins; D000117:Acetylglucosamine