Endocytosis of a glycosylphosphatidylinositol-anchored protein via clathrin-coated vesicles, sorting by default in endosomes, and exocytosis via RAB11-positive carriers

Recently, proteins linked to glycosylphosphatidylinositol (GPI) residues have received considerable attention both for their association with lipid microdomains and for their specific transport between cellular membranes. Basic features of trafficking of GPI-anchored proteins or glycolipids may be explored in flagellated protozoan parasites, which offer the advantage that their surface is dominated by these components. In Trypanosoma brucei, the GPI-anchored variant surface glycoprotein (VSG) is efficiently sorted at multiple intracellular levels, leading to a 50-fold higher membrane concentration at the cell surface compared with the endoplasmic reticulum. We have studied the membrane and VSG flow at an invagination of the plasma membrane, the flagellar pocket, the sole region for endo-and exocytosis in this organism. VSG enters trypanosomes in large clathrin-coated vesicles (135 nm in diameter), which deliver their cargo to endosomes. In the lumen of cisternal endosomes, VSG is concentrated by default, because a distinct class of small clathrin-coated vesicles (50-60 nm in diameter) budding from the cisternae is depleted in VSG. TbRAB11-positive cisternal endosomes, containing VSG, fragment by an unknown process giving rise to intensely TbRAB11- as well as VSG-positive, disk-like carriers (154 nm in diameter, 34 nm in thickness), which are shown to fuse with the flagellar pocket membrane, thereby recycling VSG back to the cell surface.