A non-enzymatic C-type lectin from bullhead shark Heterodontus japonicus skin amplifies coagulation via common-pathway downstream of factor Xa.

The rapid arrest of bleeding after injury is essential for survival, and blood coagulation is the central mechanism of hemostasis. We previously reported that a C-type lectin (HjCL) present in the skin mucus of the bullhead shark (Heterodontus japonicus) exhibits procoagulant activity; however, its underlying molecular mechanism remains unclear. In this study, we evaluated the effects of HjCL on pathway-specific clotting reactions using prothrombin time (PT) and activated partial thromboplastin time (aPTT) assays. HjCL accelerated aPTT reagent-triggered clotting, as indicated by the shortened time to reach 50% of maximal fibrin formation (t50). In contrast, no procoagulant effect was observed in the reactions initiated by the PT reagent. We performed synthetic substrate assays for the factors Xa and thrombin to further assess the activation of common-pathway enzymes at the plasma level. HjCL enhanced thrombin generation in the aPTT assay but did not increase factor Xa generation. Together, these results suggest that HjCL does not act as an initiator of coagulation but rather promotes coagulation at a stage downstream of factor Xa formation, consistent with its role in the amplification phase of the common pathway. Our findings suggest a previously unrecognized mechanism by which a non-enzymatic protein modulates blood coagulation in cartilaginous fish, providing new insights into hemostatic regulation in early-diverging vertebrates.