Abstract
Skeletal muscle myosin (SkM) has been shown to possess procoagulant activity; however, the mechanisms of this coagulation-enhancing activity involving plasma coagulation pathways and factors are incompletely understood. Here, we discovered direct interactions between immobilized SkM and coagulation factor XI (FXI) using biolayer interferometry (Kd = 0.2 nM). In contrast, we show that prekallikrein, a FXI homolog, did not bind to SkM, reflecting the specificity of SkM for FXI binding. We also found that the anti-FXI monoclonal antibody, mAb 1A6, which recognizes the Apple (A) 3 domain of FXI, potently inhibited binding of FXI to immobilized SkM, implying that SkM binds FXI A3 domain. In addition, we show that SkM enhanced FXI activation by thrombin in a concentration-dependent manner. We further used recombinant FXI chimeric proteins in which each of the four A domains of the heavy chain (designated A1 through A4) was individually replaced with the corresponding A domain from prekallikrein to investigate SkM-mediated enhancement of thrombin-induced FXI activation. These results indicated that activation of two FXI chimeras with substitutions of either the A3 domains or A4 domains was not enhanced by SkM, whereas substitution of the A2 domain did not reduce the thrombin-induced activation compared with wildtype FXI. These data strongly suggest that functional interaction sites on FXI for SkM involve the A3 and A4 domains. Thus, this study is the first to reveal and support the novel intrinsic blood coagulation pathway concept that the procoagulant mechanisms of SkM include FXI binding and enhancement of FXI activation by thrombin.
Highlights
Surface-bound skeletal muscle myosin (SkM) promotes thrombus formation when exposed to fresh flowing human blood [1, 2], and clinical studies suggested that various plasma SkM isoforms and phenotypes are linked to pulmonary embolism and thrombin generation in plasma [1]
Since bleeding in muscle is often observed in coagulation factor deficiency [12], hemostasis is a critical event in muscle damage
SkM and cardiac muscle myosin promote hemostasis and thrombotic events in murine model studies [13, 14], and they enhance prothrombin activation by clotting factor Xa in combination with factor V [1, 2, 15], an activity termed prothrombinase activity, which is associated with procoagulant anionic phospholipids that are linked to these two muscle myosins [16]
Summary
Shravan Morla1,‡ , Hiroshi Deguchi1,‡, Jevgenia Zilberman-Rudenko1,2 , András Gruber, Owen J. Griffin1,4,* From the 1Department of Molecular Medicine, The Scripps Research Institute, La Jolla, California, USA; 2Departments of Biomedical Engineering and Medicine, School of Medicine, Oregon Health & Science University, Portland, Oregon, USA; 3Departments of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA; 4Department of Medicine, University of California, San Diego, California, USA
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