Tissue factor pathway inhibitor-γ is an active alternatively spliced form of tissue factor pathway inhibitor present in mice but not in humans

Abstract

Background: Tissue factor pathway inhibitor (TFPI) is a potent inhibitor of tissue factor procoagulant activity produced as two alternatively spliced isoforms, TFPIα and TFPIβ, which differ in domain structure and mechanism for cell surface association. 3′ Rapid amplification of cDNA ends was used to search for new TFPI isoforms. TFPIγ, a new alternatively spliced form of TFPI, was identified and characterized. Methods: The tissue expression, cell surface association and anticoagulant activity of TFPIγ were characterized and compared to those of TFPIα and TFPIβ through studies of mouse and human tissues and expression of recombinant proteins in Chinese hamster ovary (CHO) cells. Results: TFPIγ is produced by alternative splicing using the same 5′-splice donor site as TFPIβ and a 3′-splice acceptor site 276 nucleotides beyond the stop codon of TFPIβ in exon 8. The resulting protein has the first two Kunitz domains connected to an 18 amino acid C-terminal region specific to TFPIγ. TFPIγ mRNA is differentially produced in mouse tissues but is not encoded within the human TFPI gene. When expressed in CHO cells, TFPIγ is secreted into conditioned media and effectively inhibits tissue factor procoagulant activity. Conclusions: TFPIγ is a third alternatively spliced form of TFPI that is widely expressed in mouse tissues but not made by human tissues. It contains the first two Kunitz domains and is a secreted, rather than a cell surface-associated, protein. It is a functional anticoagulant and may partially explain the resistance of mice to coagulopathy in tissue factor-mediated models of disease. © 2008 International Society on Thrombosis and Haemostasis.