Nature of glycosylphosphatidylinositols produced by mouse embryonic stem cells.
Incorporation of [(3)H]inositol into mouse embryonic stem cells of the CCE cell line leads to the labelling of the three common phosphoinositides, phosphatidylinositol, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate, and a fourth unknown lipid (lipid X). Incubation with [(3)H]glucosamine results in the labelling of lipid X and at least one other lipid that co-migrates with phosphatidylinositol (lipid Y), indicating that both of these lipids are putative glycosylphosphatidylinositols. In this study, the incorporation of other possible glycosylphosphatidylinositol precursors, ethanolamine, mannose and galactose, into lipids X and Y was examined. Galactose was incorporated into lipids X and Y, and ethanolamine and mannose into lipid Y only. Inhibitors of glycosylphosphatidylinositol biosynthesis pathways, mannosamine and 2-fluoro-2-deoxyglucose, both significantly inhibited ethanolamine incorporation into lipid Y. A high glucose concentration (25 mmol l(-1)) abolished the action of both inhibitors. Phospholipase C treatment of embryonic stem cells that had been labelled in culture with [(3)H]ethanolamine caused a large release of ethanolamine label into the incubation medium and markedly decreased the amount of ethanolamine-labelled lipid Y remaining in the cell membranes. These effects were almost totally abolished by incubation with mannosamine before ethanolamine labelling. These studies strongly indicate that lipid Y is a member of the protein anchor class of glycosylphosphatidylinositol, whereas lipid X is a member of the signal transduction inositol phosphoglycan class of glycosylphosphatidylinositol.
Quinlan
L R
LR
Department of Physiology, National University of Ireland, Galway, University Road, Galway, Ireland.
Kane
M T
MT
D000818:Animals; D001345:Autoradiography; D002460:Cell Line; D002855:Chromatography, Thin Layer; D019856:Ethanolamine / Q000378:metabolism; D005260:Female; D019788:Fluorodeoxyglucose F18 / Q000494:pharmacology; D005690:Galactose / Q000378:metabolism; D005944:Glucosamine / Q000378:metabolism; D006017:Glycolipids / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000096:biosynthesis* / Q000378:metabolism; D006595:Hexosamines / Q000494:pharmacology; D007294:Inositol / Q000378:metabolism; D008358:Mannose / Q000378:metabolism; D051379:Mice; D010716:Phosphatidylinositols / Q000378:metabolism; D013234:Stem Cells / Q000378:metabolism*; D010738:Type C Phospholipases / Q000494:pharmacology
D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D006595:Hexosamines; D010716:Phosphatidylinositols; D019788:Fluorodeoxyglucose F18; C008322:mannosamine; D007294:Inositol; D019856:Ethanolamine; C044282:lipid Y; D010738:Type C Phospholipases; D005944:Glucosamine; D008358:Mannose; D005690:Galactose