Distinct interactions among GPI-anchored, transmembrane and membrane associated intracellular proteins, and sphingolipids in lymphocyte and endothelial cell plasma membranes.
Glycosylphosphatidylinositol (GPI)-anchored glycoproteins are enriched in sphingolipid-rich plasma membrane domains, which are often isolated as low-density membrane complexes. This association is believed to arise from the interactions between the GPI-acyl chains and sphingolipids, but is not fully understood. In this study, we compared the physical properties of GPI-anchored glycoproteins from a non-polarized (murine T-lymphocyte) and a polarized (human endothelial) cell by equilibrium density gradient centrifugation after extraction by detergents under identical conditions. Unlike those on epithelial cells, the GPI-anchored proteins of lymphocytes (Thy-1 and the heat stable antigen CD24) were enriched in the floating fractions after extraction over a wide range of octylglucoside concentrations. In contrast, the floatability of endothelial GPI-anchored CD59 was markedly diminished, not only by octylglucoside, but also by increasing concentrations of Triton X-100. Distribution of cholera toxin binding ganglioside GM1 in the sucrose gradient fractions closely followed that of the GPI-anchored proteins in both lymphocytes and endothelial cells under most extraction conditions. Analysis of the intracellular acylated molecules revealed that a significant amount of p56(lck) was always associated with the floating GPI-rich fractions of lymphocytes when extracted by Triton X-100 or octylglucoside at 4 degrees C, while the behaviour of endothelial cell caveolin was comparable to that of CD59. The transmembrane glycoproteins CD45 in lymphocytes and MHC class I antigen in endothelial cells interacted weakly with GPI domains, whereas endothelial CD44 and lymphocyte CD26 displayed a strong association. These results show that: (1) the physical properties of different GPI-anchored proteins may vary significantly; and (2) transmembrane and acylated intracellular proteins could be associated with GPI domains to a variable extent. These differences probably reflect cell type-specific interactions of GPI anchors with the sphingolipid framework of plasma membranes, as well as extracellular interactions of GPI-anchored glycoproteins with neighbouring cell surface molecules.
Ilangumaran
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Department of Pathology, Centre Médical Universitaire, Geneva, Switzerland.
Briol
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Hoessli
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D000818:Animals; D002462:Cell Membrane / Q000737:chemistry*; D016764:Cell Polarity; D002499:Centrifugation, Density Gradient; D004730:Endothelium, Vascular / Q000737:chemistry* / Q000166:cytology; D005677:G(M1) Ganglioside / Q000032:analysis; D017261:Glycosylphosphatidylinositols / Q000032:analysis*; D006801:Humans; D008214:Lymphocytes / Q000737:chemistry* / Q000166:cytology; D008565:Membrane Proteins / Q000032:analysis*; D051381:Rats; D013107:Sphingolipids / Q000032:analysis*; D013347:Subcellular Fractions / Q000737:chemistry
D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D013107:Sphingolipids; D005677:G(M1) Ganglioside