Release of GPI-anchored membrane proteins by a cell-associated GPI-specific phospholipase D.
Although many glycosylphosphatidylinositol (GPI)-anchored proteins have been observed as soluble forms, the mechanisms by which they are released from the cell surface have not been demonstrated. We show here that a cell-associated GPI-specific phospholipase D (GPI-PLD) releases the GPI-anchored, complement regulatory protein decay-accelerating factor (DAF) from HeLa cells, as well as the basic fibroblast growth factor-binding heparan sulfate proteoglycan from bone marrow stromal cells. DAF found in the HeLa cell culture supernatants contained both [3H]ethanolamine and [3H]inositol, but not [3H]palmitic acid, whereas the soluble heparan sulfate proteoglycan present in bone marrow stromal cell culture supernatants contained [3H]ethanolamine. 125I-labeled GPI-DAF incorporated into the plasma membranes of these two cell types was released in a soluble form lacking the fatty acid GPI-anchor component. GPI-PLD activity was detected in lysates of both HeLa and bone marrow stromal cells. Treatment of HeLa cells with 1,10-phenanthroline, an inhibitor of GPI-PLD, reduced the release of [3H]ethanolamine-DAF by 70%. The hydrolysis of these GPI-anchored molecules is likely to be mediated by an endogenous GPI-PLD because [3H]ethanolamine DAF is constitutively released from HeLa cells maintained in serum-free medium. Furthermore, using PCR, a GPI-PLD mRNA has been identified in cDNA libraries prepared from both cell types. These studies are the first demonstration of the physiologically relevant release of GPI-anchored proteins from cells by a GPI-PLD.
Metz
C N
CN
Department of Pathology, New York University Medical Center, NY 10016.
Brunner
G
G
Choi-Muira
N H
NH
Nguyen
H
H
Gabrilove
J
J
Caras
I W
IW
Altszuler
N
N
Rifkin
D B
DB
Wilson
E L
EL
Davitz
M A
MA
D015703:Antigens, CD / Q000378:metabolism*; D001853:Bone Marrow / Q000201:enzymology; D001854:Bone Marrow Cells; D018958:CD55 Antigens; D018076:DNA, Complementary / Q000235:genetics; D019856:Ethanolamine; D004983:Ethanolamines / Q000378:metabolism; D015723:Gene Library; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006367:HeLa Cells / Q000201:enzymology; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000378:metabolism*; D006801:Humans; D007294:Inositol / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D010618:Phenanthrolines / Q000494:pharmacology; D010739:Phospholipase D / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011509:Proteoglycans / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics
D015703:Antigens, CD; D018958:CD55 Antigens; D018076:DNA, Complementary; D019856:Ethanolamine; D004983:Ethanolamines; D017261:Glycosylphosphatidylinositols; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D007294:Inositol; D008562:Membrane Glycoproteins; D010618:Phenanthrolines; D010739:Phospholipase D; D011509:Proteoglycans; D012333:RNA, Messenger; C065017:glycoprotein phospholipase D; C025205:1,10-phenanthroline