Exclusion of known protease activated receptors in factor VIIa-induced signal transduction

The protease activity is mandatory for intracellular activities induced by coagulation factor VIIa (FVIIa), and in this way it resembles signal transd uction induced by thrombin and trypsin caused by specific, proteolytic clea vage of protease activated receptors (PARs). The mechanism for FVIIa-induce d signal transduction is, however, not known although a mechanism involving PAR cleavage has been deduced from studies of cytosolic Ca2+ release and p 44/p42 mitogen activated protein kinase (MAPK) activation. In the present w ork we have examined the possibilities that i) FVIIa-induced signal transdu ction involves the activation of one of the four known PARs. or ii) exposur e of cells to FVIIa releases a soluble ligand that is responsible for MAPK activation. For this purpose, we evaluated the effects of FVIIa, thrombin. FXa, trypsin and PAR agonist peptides on the Ca2+ release and MAPK activati on in tissue factor-(TF) transfected baby hamster kidney (BHK[+TF]) cells a nd Madin-Darby canine kidney (MDCK) cells:FVIIa induced a significant MAPK signal in BHK(+TF) cells and in MDCK-I and -II cells whereas no MAPK activa tion was observed with thrombin, FXa or PAR agonist peptides. Thrombin, try psin, PAR-1 and PAR-2 agonist peptides induced a prominent Ca2+ response in both cell types. In contrast the cells did not respond with a detectable C a2+ signal when treated with FVIIa. These results suggest that the intracel lular activity induced by FVIIa is distinctly different from that induced b y trypsin, thrombin and FXa not involving any of the known PARs. Conditione d medium from BHK(+TF) cells treated with FVIIa failed to induce a MAPK res ponse in untreated BHK(+TF) cells when FVIIa was removed by immunoadsorptio n from the medium prior to its transfer to the untreated BHK(+TF) cells. Al though it is not possible entirely to exclude a transient response close to the cell surface, the data suggest that the intracellular response was not induced by an autocrine release of a soluble mediator to the medium.