title: The GPI anchor of cell-surface proteins is synthesized on the cytoplasmic face of the endoplasmic reticulum.
abstract: Glycosylphosphatidylinositol (GPI) membrane protein anchors are synthesized from sugar nucleotides and phospholipids in the ER and transferred to newly synthesized proteins destined for the cell surface. The topology of GPI synthesis in the ER was investigated using sealed trypanosome microsomes and the membrane-impermeant probes phosphatidylinositol-specific phospholipase C, Con A, and proteinase K. All the GPI biosynthetic intermediates examined were found to be located on the external face of the microsomal vesicles suggesting that the principal steps of GPI assembly occur in the cytoplasmic leaflet of the ER. Protease protection experiments showed that newly GPI-modified trypanosome variant surface glycoprotein was primarily oriented towards the ER lumen, consistent with eventual expression at the cell surface. The unusual topographical arrangement of the GPI assembly pathway suggests that a biosynthetic intermediate, possibly the phosphoethanolamine-containing anchor precursor, must be translocated across the ER membrane bilayer in the process of constructing a GPI anchor.
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison 53706.
authors.forename: J
authors.initials: J
authors.lastname: Vidugiriene
authors.forename: A K
authors.initials: AK
authors.lastname: Menon
mesh_terms: D000818:Animals; D003208:Concanavalin A / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D019286:Endopeptidase K; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000096:biosynthesis*; D008051:Lipid Bilayers / Q000378:metabolism; D008358:Mannose / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000378:metabolism* / Q000648:ultrastructure; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000378:metabolism
chemical_list: D017261:Glycosylphosphatidylinositols; D008051:Lipid Bilayers; D014643:Variant Surface Glycoproteins, Trypanosoma; D003208:Concanavalin A; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D012697:Serine Endopeptidases; D019286:Endopeptidase K; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D008358:Mannose