Cryptic messages: is noncoagulant tissue factor reserved for cell signaling?

T issue factor (TF) is the principal initiator of blood clotting in response to vascular injury and in thrombotic disease (1). TF, an integral membrane protein, is normally excluded from the vascular compartment. Coagulation is initiated when TF is exposed to zymogen coagulation proteases in plasma upon vascular damage. TF first binds factor VII, supporting both its activation to VIIa and activation of factor X (Fig. 1). Factor Xa acts with its cellular cofactor Va to generate thrombin (IIa), which drives both fibrin deposition and platelet aggregation. Thrombin elicits signaling in cells through protease-activated receptors (PARs), a family of G proteincoupled receptors, which are activated proteolytically by unmasking of an Nterminal tethered ligand (Fig. 1) (2). In addition to hemostasis, TF expression and coagulant protease activation have been implicated in inflammation, vascular development, and cancer progression. Signaling by coagulant proteases is an important consequence of TF exposure in these settings. Factors VIIa and Xa have been shown to cleave and activate PAR2, the only PAR not activated by thrombin, and may act in concert with thrombin to elicit cellular responses to coagulation activation (Fig. 1). VIIa requires its cofactor TF to facilitate PAR2 cleavage and activation. The molecular determinants that specify TF function as a cofactor for cellular signaling versus coagulation are not known. In a recent issue of PNAS, Ahamed et al. (3) provide fascinating insight suggesting that coagulant and signaling TF may, in fact, be two different structural entities, that signaling TF may be the long-elusive encrypted TF, and that disulfide isomerization facilitates a dynamic and reversible switch between these two distinct functional TF species. TF encryption has been widely studied in cell culture and may represent a physiological mechanism for controlling expression of cellular coagulant activity in vivo. Recent reports suggest the existence of circulating TF important for clot propagation that only becomes active in the context of a growing thrombus (4). Cells that constitutively express TF also display latent forms of this protein. Although there is usually good correlation between cell surface TF expression and binding and allosteric activation of VIIa, the ability of this complex to activate X is variable and usually attributable to only a fraction of the TF:VIIa complexes formed (5, 6). The TF population unable to promote coagulation has been dubbed ‘‘encrypted.’’ When cells are damaged, lysed by cycles of freeze–thawing, or treated with certain cell-activating agents such as calcium ionophores, coagulant potential is dramatically increased. One prevailing model for induction of TF activity was the exposure of phosphatidylserine on the outer leaflet of the plasma membrane, because negatively charged lipids are limiting and required for optimal TF:VIIa activity toward factor X. TF dimerization and or compartmentalization in lipid rafts such as caveolae might also limit enzymatic activity. The studies by Ahamed et al. (3) now suggest a new mechanism for modulating TF coagulant activity involving conformational changes induced by disulfide bond cleavage. Disulfide bond rearrangements can function as an allosteric mechanism to regulate protein function, as demonstrated for several cell surface proteins including CD4 and integrins (7). Protein disulfide isomerase (PDI) is capable of catalyzing all of the reactions involved in disulfide bond formation, cleavage, and isomerization (rearrangements) (8). PDI facilitates folding of nascent proteins in the endoplasmic reticulum but also resides on the surface of mammalian cells and platelets. PDI has been shown to cleave disulfide bonds in the extracellular domains of certain receptors to allosterically regulate protein activity. Interestingly, PDI readily cleaves rare disulfide bonds in which high strain is introduced by linking strands in the same or parallel -sheets (7). TF disulfide-bonded Cys186 and Cys209 link two parallel -sheets, are required for coagulation (9), and were predicted to function in allosteric modulation of TF function (7, 10). In a series of elegant experiments, Ahamed et al. now demonstrate that PDImediated cleavage of the Cys186-Cys209 disulfide indeed disables coagulant activity. Under conditions that favored TF:VIIa signaling and disabled coagulant