Microdomains of GPI-anchored proteins in living cells revealed by crosslinking.
There is some discussion as to whether glycosyl-phosphatidylinositol(GPI)-anchored proteins occur in microdomains in the cell membrane. These putative microdomains have been implicated in processes such as sorting in polarized cells and signal transduction. Complexes enriched in GPI-anchored proteins, cholesterol and glycosphingolipids have been isolated from cell membranes by using non-ionic detergents: these complexes were thought to represent a clustered arrangement of GPI-anchored proteins. However, results obtained when clustering of GPI-anchored proteins induced by antibodies or by detergents was prevented support the idea of a dispersed surface distribution of GPI-anchored proteins at steady state. Here we use chemical crosslinking to show that membrane microdomains of a GPI-anchored protein exist at the surface in living cells. This clustering is specific for the GPI-anchored form, as two transmembrane forms bearing the same ectodomain do not form oligomers. Depletion of membrane cholesterol causes the clustering of GPI-anchored proteins to break up, whereas treatment of cells with detergent substantially increases the size of the complexes. We find that in living cells these GPI-anchored proteins reside in microdomains consisting of at least 15 molecules, which are much smaller than those seen after detergent extraction.
Friedrichson
T
T
Department of Cell Biology, Max Delbrück Centre for Molecular Medicine, Berlin-Buch, Germany.
Kurzchalia
T V
TV
D000818:Animals; D018958:CD55 Antigens / Q000737:chemistry*; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D003001:Cloning, Molecular; D006224:Cricetinae; D003432:Cross-Linking Reagents; D003902:Detergents; D017261:Glycosylphosphatidylinositols / Q000737:chemistry*; D013006:Growth Hormone / Q000737:chemistry*; D051381:Rats
D018958:CD55 Antigens; D003432:Cross-Linking Reagents; D003902:Detergents; D017261:Glycosylphosphatidylinositols; D013006:Growth Hormone