I. Ott et al., Reversible regulation of tissue factor-induced coagulation by glycosyl phosphatidylinositol-anchored tissue factor pathway inhibitor, ART THROM V, 20(3), 2000, pp. 874-882
Endothelial and tumor cells synthesize tissue factor pathway inhibitor (TFP
I-1), which regulates tissue factor (TF) function by TF . VIIa . Xa . TFPI-
1 quaternary complex formation (where VIIa and Xa are coagulation factors)
and by translocation of these complexes into glycospbingolipid-rich microdo
mains of the cell membrane. Recombinant TFPI-1 added exogenously to cells i
s targeted to a degradation pathway. This study analyzes whether quaternary
complex formation with endogenous TFPI-1 results also in internalization a
nd degradation. We demonstrate that endogenous TFPI-1 and recombinant TFPI-
1 differ in their distribution on the cell surface, Recombinant TFPT-1 is f
ound in phospholipid- and glycosphingolipid-rich membrane domains, whereas
endogenous TFPI-1 preferentially localizes to glycosphingolipid-rich microd
omains. On quaternary complex formation, endogenous TFPT-1 remains protease
sensitive and accessible for antibodies on intact cells, demonstrating tha
t it is not appreciably internalized Rather, regulation of TP by TFPI-1 is
restored within 12 hours, consistent with dissociation of quaternary comple
xes on the cell surface. Endogenous TFPI-1 can be released from the cell su
rface by phospholipase treatment, indicating that TFPI-1 either is a glycos
yl phosphatidylinositol (GPI)-anchored protein or binds to a GPI-linked rec
eptor. We demonstrate that expression of a recombinant GPI-anchored form of
TFPI-1 targets TF . VIIa complexes to glycosphingolipid-rich membrane frac
tions. Thus, GPI anchoring of TFPI-1 is sufficient for regulation of TF . V
IIa complex function by a pathway of reversible inhibition rather than inte
rnalization and degradation.