Tropomyosin 1 promotes platelet adhesion and clot contraction to inhibit thrombosis

Abstract

Abstract Background: Failures in thrombosis prevention and treatment demand novel therapeutic approaches. We hypothesized that novel genetic determinants of thrombosis may be hiding in plain sight in human genetics data sets for relevant traits. Genome-wide associations studies (GWAS) have linked the Tropomyosin 1 (Tpm1) gene locus to quantitative platelet trait variation. Tpm1 encodes an actin-binding protein that stabilizes actin filaments and influences cell adhesion and actomyosin contractility in many cells, but it had no known role in platelet function. We hypothesized that Tpm1 functions in platelet adhesion and hemostasis. Methods: We used murine models to test the role of Tpm1 in vivo. We created conditional Tpm1 knockout (Tpm1KO) models using Cdh5Cre or VavCre to ablate Tpm1 in embryonic hemogenic endothelium or nascent hematopoietic stem and progenitor cells (HSPCs), respectively. These models helped bypass cardiac dysmorphology and embryonic lethality associated with Tpm1 deficiency. All experiments were based on murine in vivo or ex vivo samples. Results: Both models ablated Tpm1 in adult peripheral blood by qPCR and produced healthy Tpm1KO mice in normal Mendelian ratios. Cdh5Cre Tpm1fl/fl mice increased formation of definitive hemogenic endothelial cells in embryos (immediate precursors), consistent with our prior findings. However, postnatal HSPCs and adult blood counts remained unchanged. This suggested separate roles for Tpm1 in the embryonic and adult blood systems. Genetic colocalization analysis confirmed a platelet trait-specific GWAS locus at the Tpm1 locus, suggesting platelet-specific function. Alleles that decrease TPM1 expression increase platelet counts. We hypothesized that Tpm1 deficiency might diminish adhesion based on actin perturbations. Static adhesion assays showed reduced adhesion to fibronectin and fibrinogen and led to longer in vivo platelet half-life in healthy adult mice by almost 50%. Platelet-fibrin binding mediates blood clot contraction, which prevents vascular occlusion during thrombosis. Tpm1KO delayed clot contraction in assays of whole blood and platelet rich plasma. Perturbed clot contraction can worsen vascular occlusion. In a ferric chloride-induced thrombosis model, Tpm1KO showed more rapid occlusion and larger clots forming within 5-10 minutes whereas littermate controls took over 30 minutes. Conclusions: Our findings reveal a new role for Tpm1 and cytoskeletal regulation in platelet adhesion and clot contraction. Loss of Tpm1 perturbs clot contraction, worsening vascular occlusion and thrombosis. This mechanism represents a new target for modifying thrombotic risk and related complications.