title: Essential roles for GPI-anchored proteins in African trypanosomes revealed using mutants deficient in GPI8.
abstract: The survival of Trypanosoma brucei, the causative agent of Sleeping Sickness and Nagana, is facilitated by the expression of a dense surface coat of glycosylphosphatidylinositol (GPI)-anchored proteins in both its mammalian and tsetse fly hosts. We have characterized T. brucei GPI8, the gene encoding the catalytic subunit of the GPI:protein transamidase complex that adds preformed GPI anchors onto nascent polypeptides. Deletion of GPI8 (to give Deltagpi8) resulted in the absence of GPI-anchored proteins from the cell surface of procyclic form trypanosomes and accumulation of a pool of non-protein-linked GPI molecules, some of which are surface located. Procyclic Deltagpi8, while viable in culture, were unable to establish infections in the tsetse midgut, confirming that GPI-anchored proteins are essential for insect-parasite interactions. Applying specific inducible GPI8 RNAi with bloodstream form parasites resulted in accumulation of unanchored variant surface glycoprotein and cell death with a defined multinuclear, multikinetoplast, and multiflagellar phenotype indicative of a block in cytokinesis. These data show that GPI-anchored proteins are essential for the viability of bloodstream form trypanosomes even in the absence of immune challenge and imply that GPI8 is important for proper cell cycle progression.
authors.affiliation: Wellcome Centre for Molecular Parasitology, University of Glasgow, The Anderson College, United Kingdom.
authors.forename: Simon
authors.initials: S
authors.lastname: Lillico
authors.forename: Mark C
authors.initials: MC
authors.lastname: Field
authors.forename: Pat
authors.initials: P
authors.lastname: Blundell
authors.forename: Graham H
authors.initials: GH
authors.lastname: Coombs
authors.forename: Jeremy C
authors.initials: JC
authors.lastname: Mottram
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002240:Carbohydrate Sequence; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle / Q000502:physiology; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D008358:Mannose / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D034622:RNA Interference; D016415:Sequence Alignment; D014346:Trypanosoma brucei brucei / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D014353:Trypanosomiasis, African; D014370:Tsetse Flies / Q000378:metabolism / Q000469:parasitology
chemical_list: D015815:Cell Adhesion Molecules; D017261:Glycosylphosphatidylinositols; C491190:PIGK protein, human; D008358:Mannose