The Eph Tyrosine Kinase Receptors EphB2 and EphA2 Are Novel Proteolytic Substrates of Tissue Factor/Coagulation Factor VIIa |
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Authors: | Oskar Eriksson Margareta Ramstr?m Katarina H?rnaeus Jonas Bergquist Dariush Mokhtari Agneta Siegbahn |
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Institution: | From the ‡Department of Medical Sciences, Clinical Chemistry, Uppsala University Hospital, SE-751 85 Uppsala.;the §Department of Chemistry, Analytical Chemistry, BMC, SE-751 24 Uppsala, and ;¶Science for Life Laboratory, Uppsala University, Sweden |
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Abstract: | Tissue factor (TF) binds the serine protease factor VIIa (FVIIa) to form a proteolytically active complex that can trigger coagulation or activate cell signaling. Here we addressed the involvement of tyrosine kinase receptors (RTKs) in TF/FVIIa signaling by antibody array analysis and subsequently found that EphB2 and EphA2 of the Eph RTK family were cleaved in their ectodomains by TF/FVIIa. We used N-terminal Edman sequencing and LC-MS/MS analysis to characterize the cleaved Eph isoforms and identified a key arginine residue at the cleavage site, in agreement with the tryptic serine protease activity of FVIIa. Protease-activated receptor 2 (PAR2) signaling and downstream coagulation activity was non-essential in this context, in further support of a direct cleavage by TF/FVIIa. EphB2 was cleaved by FVIIa concentrations in the subnanomolar range in a number of TF expressing cell types, indicating that the active cellular pool of TF was involved. FVIIa caused potentiation of cell repulsion by the EphB2 ligand ephrin-B1, demonstrating a novel proteolytical event to control Eph-mediated cell segregation. These results define Eph RTKs as novel proteolytical targets of TF/FVIIa and provide new insights into how TF/FVIIa regulates cellular functions independently of PAR2. |
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Keywords: | Coagulation Factor Mass Spectrometry (MS) Receptor Tyrosine Kinase Serine Protease Tissue Factor |
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