Deencryption of cellular tissue factor is independent of its cytoplasmic domain

Tissue factor (TF) is a transmembrane molecule that, when exposed to plasma , is the key initiator of coagulation. Cellular TF activity is normally "en crypted", but treating cells with calcium ionophore (i.e., ionomycin or A23 187) increases ("deencrypts") TF activity without increasing TF mRNA or ant igen expression. Deencryption results from both plasma membrane phosphatidy lserine (PS)-dependent and -independent mechanisms; however, the nature of the PS-independent component is unclear. Since deencryption has been sugges ted to result from release of TF dimers on the cell surface, and since TF's cytoplasmic domain binds to actin-binding protein 280 and interacts with t he cytoskeleton, we hypothesized that interactions with the cytoskeleton, t hrough the cytoplasmic domain, play a role in mediating encryption/deencryp tion, We examined TF deencryption and the role of the cytoplasmic domain in the PS-independent component using baby hamster kidney (BHK) cells express ing full length TF (BHK-TF) or TF lacking its cytoplasmic domain (BHK-descy t) (Sorensen et al. (1999) J. Biol. Chem. 274, 21349). Both BHK-TF and BHK- descyt cells exhibited a dose-dependent, 1.5- to 10-fold increase in TF act ivity upon treatment with calcium ionophore, and this increase in activity was only partially blocked by annexin V. These results indicate that deencr yption is not restricted to cells which naturally express TF and that the P S-independent component of deencryption is intact on cells transfected with either full length or truncated TF. Our results clearly indicate that deen cryption is not dependent on an intact cytoplasmic domain in transfected BH K cells. (C) 2000 Academic Press.