Cellular glycosylphosphatidylinositol-specific phospholipase D regulates urokinase receptor shedding and cell surface expression.
The glycosylphosphatidylinositol (GPI)-anchored, multifunctional receptor for the serine proteinase, urokinase plasminogen activator (uPAR, CD87), regulates plasminogen activation and cell migration, adhesion, and proliferation. uPAR occurs in functionally distinct, membrane-anchored and soluble isoforms (s-uPAR) in vitro and in vivo. Recent evidence indicates that s-uPAR present in the circulation of cancer patients correlates with tumor malignancy and represents a valuable prognostic marker in certain types of cancer. We have therefore analyzed the mechanism of uPAR shedding in vitro. We present evidence that uPAR is actively released from ovarian cancer cells since the rate of receptor shedding did not correlate with uPAR expression. While s-uPAR was derived from the cell surface, it lacked the hydrophobic portion of the GPI moiety indicating anchor cleavage. We show that uPAR release is catalyzed by cellular GPI-specific phospholipase D (GPI-PLD), an enzyme cleaving the GPI anchor of the receptor. Thus, recombinant GPI-PLD expression increased receptor release up to fourfold. Conversely, a 40% reduction in GPI-PLD activity by GPI-PLD antisense mRNA expression inhibited uPAR release by more than 60%. We found that GPI-PLD also regulated uPAR expression, possibly by releasing a GPI-anchored growth factor. Our data suggest that cellular GPI-PLD might be involved in the generation of circulating prognostic markers in cancer and possibly regulate the function of GPI-anchored proteins by generating functionally distinct, soluble counterparts.
Frauenklinik der Technischen Universität München, Munich, Germany.
O G
OG
Wilhelm
S
S
Wilhelm
G M
GM
Escott
V
V
Lutz
V
V
Magdolen
M
M
Schmitt
D B
DB
Rifkin
E L
EL
Wilson
H
H
Graeff
G
G
Brunner
D016375:Antisense Elements (Genetics); D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D017261:Glycosylphosphatidylinositols / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D008565:Membrane Proteins / Q000378:metabolism; D010051:Ovarian Neoplasms; D010739:Phospholipase D / Q000235:genetics / Q000378:metabolism*; D010960:Plasminogen Activators / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000201:enzymology
D016375:Antisense Elements (Genetics); D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins; D010739:Phospholipase D; D010960:Plasminogen Activators