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 mimbresof a significant pool of GPI-anchored proteins which resisted extraction by 1% TX-100 at 4 °C but not at 37 °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 63 kDa component (gpi63) and 23% of a somewhat less abundant component (gpi23) were insoluble in TX-100 at 4 °C. A substantial proportion of the cell's only abundant sphingolipid, ceramideaminoethylphosphonate (CAEP), was also insoluble in 1% TX-100 at 4 °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.
An apparent association between glycosylphosphatidylinositol-anchored proteins and a sphingolipid in Tetrahymena mimbres
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Xiaoqing ZHANG, Guy A. THOMPSON; An apparent association between glycosylphosphatidylinositol-anchored proteins and a sphingolipid in Tetrahymena mimbres. Biochem J 1 April 1997; 323 (1): 197–206. doi: https://doi.org/10.1042/bj3230197
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