Epac1-deficient mice have bleeding phenotype and thrombocytes with decreased GPIbβ expression

Epac1 (Exchange protein directly activated by cAMP 1) limits fluid loss from the circulation by tightening the endothelial barrier. We show here that Epac1−/− mice, but not Epac2−/− mice, have prolonged bleeding time, suggesting that Epac1 may limit fluid loss also by restraining bleeding. The Epac1−/− mice had deficient in vitro secondary hemostasis. Quantitative comprehensive proteomics analysis revealed that Epac1−/− mouse platelets (thrombocytes) had unbalanced expression of key components of the glycoprotein Ib-IX-V (GPIb-IX-V) complex, with decrease of GP1bβ and no change of GP1bα. This complex is critical for platelet adhesion under arterial shear conditions. Furthermore, Epac1−/− mice have reduced levels of plasma coagulation factors and fibrinogen, increased size of circulating platelets, increased megakaryocytes (the GP1bβ level was decreased also in Epac1−/− bone marrow) and higher abundance of reticulated platelets. Viscoelastic measurement of clotting function revealed Epac1−/− mice with a dysfunction in the clotting process, which corresponds to reduced plasma levels of coagulation factors like factor XIII and fibrinogen. We propose that the observed platelet phenotype is due to deficient Epac1 activity during megakaryopoiesis and thrombopoiesis, and that the defects in blood clotting for Epac1−/− is connected to secondary hemostasis

Multiple roles for the actin cytoskeleton during regulated exocytosis

Regulated exocytosis is the main mechanism utilized by specialized secretory cells to deliver molecules to the cell surface by virtue of membranous containers (i.e. secretory vesicles). The process involves a series of highly coordinated and sequential steps, which include the biogenesis of the vesicles, their delivery to the cell periphery, their fusion with the plasma membrane and the release of their content into the extracellular space. Each of these steps is regulated by the actin cytoskeleton. In this review, we summarize the current knowledge regarding the involvement of actin and its associated molecules during each of the exocytic steps in vertebrates, and suggest that the overall role of the actin cytoskeleton during regulated exocytosis is linked to the architecture and the physiology of the secretory cells under examination. Specifically, in neurons, neuroendocrine, endocrine, and hematopoietic cells, which contain small secretory vesicles that undergo rapid exocytosis (on the order of milliseconds), the actin cytoskeleton plays a role in pre-fusion events, where it acts primarily as a functional barrier and facilitates docking. In exocrine and other secretory cells, which contain large secretory vesicles that undergo slow exocytosis (seconds to minutes), the actin cytoskeleton plays a role in post-fusion events, where it regulates the dynamics of the fusion pore, facilitates the integration of the vesicles into the plasma membrane, provides structural support, and promotes the expulsion of large cargo molecules