Membrane topology and transient acylation of Toxoplasma gondii glycosylphosphatidylinositols

Jurgen Kimmel, Terry K Smith, Nahid Azzouz, Peter Gerold, Frank Seeber, Klaus Lingelbach, Jean-Francois Dubremetz, Ralph T. Schwarz

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

Using hypotonically permeabilized Toxoplasma gondii tachyzoites, we investigated the topology of the free glycosylphosphatidylinositols (GPIs) within the endoplasmic reticulum (ER) membrane. The morphology and permeability of parasites were checked by electron microscopy and release of a cytosolic protein. The membrane integrity of organelles (ER and rhoptries) was checked by protease protection assays. In initial experiments, GPI biosynthetic intermediates were labeled with UDP-[6-H-3]GlcNAc in permeabilized parasites, and the transmembrane distribution of the radiolabeled lipids was probed with phosphatidylinositol-specific phospholipase C (PI-PLC). A new early intermediate with an acyl modification on the inositoll was identified, indicating that inositol acylation also occurs in T. gondii. A significant portion of the early GPI intermediates (GlcN-PI and GlcNAc-PI) could be hydrolyzed following PI-PLC treatment, indicating that these glycolipids are predominantly present in the cytoplasmic leaflet of the ER. Permeabilized T. gondii parasites labeled with either GDP-[2-H-3]mannose or UDP-[6-H-3]glucose showed that the more mannosylated and side chain (Glc-GalNAc) -modified GPI intermediates are also preferentially localized in the cytoplasmic leaflet of the ER.

Original languageEnglish
Pages (from-to)1420-1429
Number of pages10
JournalEukaryotic Cell
Volume5
Issue number8
DOIs
Publication statusPublished - Aug 2006

Keywords

  • GLYCOSYL-PHOSPHATIDYLINOSITOL ANCHOR
  • VARIANT SURFACE GLYCOPROTEIN
  • HOST-CELL INVASION
  • ENDOPLASMIC-RETICULUM
  • TRYPANOSOMA-BRUCEI
  • PLASMODIUM-FALCIPARUM
  • BIOSYNTHETIC-PATHWAY
  • AFRICAN TRYPANOSOMES
  • INOSITOL ACYLATION
  • GPI BIOSYNTHESIS

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