title: An apparent association between glycosylphosphatidylinositol-anchored proteins and a sphingolipid in Tetrahymena mimbres.
abstract: Sphingolipids are thought to stabilize glycosylphosphatidylinositol (GPI)-anchored protein-rich membrane domains of yeast and polarized higher animal cells during the processing and targeting of these proteins to the plasma membrane. A widely used criterion for identifying the stable sphingolipid- and GPI-anchored protein-enriched membrane domains is the resistance of these lipid-modified proteins to solubilization by the detergent Triton X-100 (TX-100) at low temperature. Surprisingly, there have been no reports of sphingolipid/GPI-anchored protein association in protozoans, despite the fact that these cells contain considerably higher levels of GPI-anchored proteins than does any other organism. We report here the presence in Tetrahymena mimbres of a significant pool of GPI-anchored proteins which resisted extraction by 1% TX-100 at 4 degrees C but not at 37 degrees C. Of the total cellular complement of GPI-anchored proteins, which together accounted for more than 2% of whole-cell protein and were especially enriched in surface membranes, 10% of the major 63kDa component (gpi63) and 23% of a somewhat less abundant component (gpi23) were insoluble in TX-100 at 4 degrees C. A substantial proportion of the cell's only abundant sphingolipid, ceramideaminoethylphosphonate (CAEP), was also insoluble in 1% TX-100 at 4 degrees C. Radiolabelling studies involving [3H]leucine incorporation into proteins and [3H]palmitic acid incorporation into lipids revealed that the TX-100-resistant gpi63, gpi23 and CAEP molecules were all metabolically distinct from their TX-100-soluble counterparts in other compartments of the cell. The presence of detergent-resistant sphingolipid/GPI-anchored protein domains in non-polarized ciliate and trypanosomatid cells was probably obscured in previous studies by the profusion of accompanying detergent-soluble molecules.
authors.affiliation: Department of Botany, University of Texas at Austin, Austin, TX 78713, USA.
authors.forename: X
authors.initials: X
authors.lastname: Zhang
authors.forename: G A
authors.initials: GA
authors.lastname: Thompson
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000737:chemistry; D002478:Cells, Cultured; D002518:Ceramides / Q000737:chemistry; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D017261:Glycosylphosphatidylinositols / Q000032:analysis*; D017830:Octoxynol; D011092:Polyethylene Glycols; D011506:Proteins / Q000737:chemistry*; D013107:Sphingolipids / Q000737:chemistry*; D013696:Temperature; D013768:Tetrahymena / Q000737:chemistry*
chemical_list: D002518:Ceramides; D003902:Detergents; D017261:Glycosylphosphatidylinositols; D011506:Proteins; D013107:Sphingolipids; D011092:Polyethylene Glycols; D017830:Octoxynol; C010615:Nonidet P-40