Saponin-induced release of cell-surface-anchored Thy-1 by serum glycosylphosphatidylinositol-specific phospholipase D.
A glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) was purified from human serum and used for studies on the release of GPI-anchored Thy-1 glycoprotein from mouse T lymphoma cells Y191. Previous studies have shown that whereas GPI-PLD is highly active against detergent-solubilized GPI-anchored proteins, it is normally unable to release GPI-containing proteins anchored in a lipid bilayer. Confirming these findings, the addition of GPI-PLD to intact Y191 cells did not result in cleavage of Thy-1. However, pretreatment of cells with saponin, a cholesterol-sequestering agent, rendered Thy-1 susceptible to hydrolysis. Very little solubilization of GPI-containing Thy-1 occurred under these conditions. From experiments with reconstituted liposomes it was inferred that the effect of saponin on cells was to aid in the presentation of Thy-1 to GPI-PLD. Furthermore, it was concluded that cholesterol-saponin complexes formed in the membrane were not alone responsible for the effect. Rather, additional molecules in the plasma membrane are possibly involved in the presentation of Thy-1 on saponin-treated cells. This finding may have implications for a physiological role of circulating GPI-PLD in the regulation of GPI-anchored proteins on cells.
Bergman
A S
AS
Department of Medical Biochemistry and Biophysics, University of Umeå, Sweden.
Carlsson
S R
SR
D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism*; D001769:Blood; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006801:Humans; D007163:Immunosorbent Techniques; D008081:Liposomes / Q000378:metabolism; D008223:Lymphoma / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D010739:Phospholipase D / Q000737:chemistry / Q000378:metabolism*; D012503:Saponins / Q000494:pharmacology*; D017421:Sequence Analysis; D012995:Solubility; D018800:Thy-1 Antigens; D014407:Tumor Cells, Cultured
D000954:Antigens, Surface; D017261:Glycosylphosphatidylinositols; D008081:Liposomes; D008562:Membrane Glycoproteins; D012503:Saponins; D018800:Thy-1 Antigens; D010739:Phospholipase D; C065017:glycoprotein phospholipase D