Requirement of FXIII transglutaminase for fibrin translocation to DRM raft fraction of thrombin-stimulated platelets. (A) Inhibition of fibrin translocation to DRM raft fraction by transglutaminase inhibitor cystamine. Immunoblotting under nonreduced condition by antifibrinogen antibody of releasates (upper panel), DRM raft fraction (middle panel), and nonraft fraction (lower panel) in resting (lane 1) and thrombin-stimulated human platelets in the absence (lane 2) or presence (lane 3) of cystamine. Total amount of fibrin in releasate, raft, and nonraft fraction of thrombin-stimulated human platelets in the presence of cystamine (lane 3) is comparable to that of fibrinogen in nonraft fraction of resting platelets (lane 1). (B) Transglutaminase-dependent association of fibrinogen γ-chain fusion protein with DRM raft fraction in thrombin-stimulated platelets. Immunoblotting of DRM raft fraction (lanes 1, 2, and 3) and nonraft fraction (lanes 4, 5, and 6) in resting (lanes 1 and 4) and thrombin-stimulated platelets in the absence (lanes 2 and 5) or presence (lanes 3 and 6) of cystamine. (C) FXIII-dependent association of fibrinogen γ-chain fusion protein with DRM raft fraction in thrombin-stimulated platelets. Immunoblotting by anti-His tag antibody of DRM raft fraction (lanes 1, 2, and 3) and nonraft fraction (lanes 4, 5, and 6) using fibrinogen γ-chain fusion protein (lanes 1 and 4), fibrinogen γ-chain fusion protein mutants of FXIII-crosslinking site Q398Q399 (lanes 2 and 5), or K406 (lanes 3 and 6). (D) Impaired fibrin translocation to DRM raft fraction of thrombin-stimulated platelets in FXIIIA subunit-deficient mice. Immunoblotting under nonreduced condition by antifibrinogen antibody of DRM raft fraction (lanes 2 and 4) and nonraft fraction (lanes 1 and 3) in resting platelets (lanes 1 and 2) and thrombin-stimulated platelets (lanes 3 and 4) of wild-type mice (upper panel) and FXIIIA-deficient mice (lower panel).