Release of GPI-anchored membrane proteins by a cell-associated GPI-specific phospholipase D

EMBO J. 1994 Apr 1;13(7):1741-51.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Antigens, CD / metabolism*
  • Bone Marrow / enzymology
  • Bone Marrow Cells
  • CD55 Antigens
  • DNA, Complementary / genetics
  • Ethanolamine
  • Ethanolamines / metabolism
  • Gene Library
  • Glycosylphosphatidylinositols / metabolism*
  • HeLa Cells / enzymology
  • Heparan Sulfate Proteoglycans
  • Heparitin Sulfate / metabolism*
  • Humans
  • Inositol / metabolism
  • Membrane Glycoproteins / metabolism*
  • Phenanthrolines / pharmacology
  • Phospholipase D / antagonists & inhibitors
  • Phospholipase D / genetics
  • Phospholipase D / metabolism*
  • Proteoglycans / metabolism*
  • RNA, Messenger / genetics

Substances

  • Antigens, CD
  • CD55 Antigens
  • DNA, Complementary
  • Ethanolamines
  • Glycosylphosphatidylinositols
  • Heparan Sulfate Proteoglycans
  • Membrane Glycoproteins
  • Phenanthrolines
  • Proteoglycans
  • RNA, Messenger
  • Inositol
  • Ethanolamine
  • Heparitin Sulfate
  • Phospholipase D
  • glycoprotein phospholipase D
  • 1,10-phenanthroline