title: Cell lysis induces redistribution of the GPI-anchored variant surface glycoprotein on both faces of the plasma membrane of Trypanosoma brucei.
abstract: African trypanosomes are coated by 10 million copies of a single variant specific glycoprotein (VSG) which are anchored in the plasma membrane by glycosylphosphatidylinositol (GPI). A GPI-specific phospholipase C (GPI-PLC) triggers fast VSG release upon cell lysis but in vivo it is safely controlled and topologically concealed from its substrate by being intracellular. One enigmatic aspect of GPI-PLC action therefore consists of how it could gain access to the VSG in the exoplasmic leaflet of the membrane. The data presented herewith disclose an unexpected possible solution for this puzzle: upon cell rupture the VSG invades the cytoplasmic face of the plasma membrane which thus becomes double coated. This unusual VSG rearrangement was stable in ruptured plasma membrane from GPI-PLC null mutant trypanosomes but transiently preceded VSG release in wild-type parasites. The formation of double coat membrane (DCM) was independent of the presence or activation of GPI-PLC, occurred both at 4 degrees C and 30 degrees C and was unaffected by the classical inhibitor of VSG release, p-choromercuryphenylsulfonic acid (PCM). DCMs conserved the same coat thickness and association with subpellicular microtubules as in intact cells and were prone to form vesicles following gradual detachment of the latter. Our data also demonstrate that: (i) GPI-PLC expressed by one trypanosome only targets its own plasma membrane, being unable to release VSG of another parasite; (ii) DCMs concomitantly formed from trypanosomes expressing different VSGs do not intermix, an indication that DCM might be refractory to membrane fusion.
authors.affiliation: Laboratory of Molecular Parasitology, Free University of Brussels, rue des Chevaux, B-1640 Rhode-St-Genèse, Belgium. mlcalmei@alize. ulb.ac.be.
authors.forename: M L
authors.initials: ML
authors.lastname: Cardoso De Almeida
authors.forename: M
authors.initials: M
authors.lastname: Geuskens
authors.forename: E
authors.initials: E
authors.lastname: Pays
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000502:physiology* / Q000648:ultrastructure; D003593:Cytoplasm / Q000502:physiology; D017353:Gene Deletion; D054800:Glycosylphosphatidylinositol Diacylglycerol-Lyase; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D007038:Hypotonic Solutions; D008870:Microtubules / Q000502:physiology / Q000648:ultrastructure; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D010738:Type C Phospholipases / Q000378:metabolism*; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*
chemical_list: D017261:Glycosylphosphatidylinositols; D007038:Hypotonic Solutions; D014643:Variant Surface Glycoproteins, Trypanosoma; D010738:Type C Phospholipases; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054800:Glycosylphosphatidylinositol Diacylglycerol-Lyase