Essential roles for GPI-anchored proteins in African trypanosomes revealed using mutants deficient in GPI8.
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.
Lillico
Simon
S
Wellcome Centre for Molecular Parasitology, University of Glasgow, The Anderson College, United Kingdom.
Field
Mark C
MC
Blundell
Pat
P
Coombs
Graham H
GH
Mottram
Jeremy C
JC
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
D015815:Cell Adhesion Molecules; D017261:Glycosylphosphatidylinositols; C491190:PIGK protein, human; D008358:Mannose