VAMP Usage In Regulated Platelet Secretion

Abstract

Platelets play important roles in hemostasis and thrombosis through secretion of their granular content. The releasate mediates activation of other platelets, clot formation, wound healing, angiogenesis and inflammation. Granule‐plasma membrane fusion requires Soluble N‐ethylmaleimide Sensitive Factor Attachment Protein Receptors (SNAREs) proteins from the granule (v‐SNAREs) and plasma membrane (t‐SNARE). The SNARE proteins form a trans‐bilayer complex that is essential for fusion. We showed that syntaxin‐11 and SNAP‐23 combine to form the functionally relevant t‐SNARE heterodimer in platelets. VAMP‐8 functions as the primary v‐SNARE though residual release occurs in VAMP‐8−/− platelets. This residual release is inhibited by treatment of permeabilized VAMP‐8−/− platelets with tetanus toxin, implying a secondary role for VAMP‐2 or ‐3. To address this differential usage of platelet v‐SNAREs, we have created a mouse strain lacking all brevin family VAMPs and one that lacks VAMP‐2 and ‐3. By crossing a RC::PFtox strain, containing a genomic insertion encoding the catalytic subunit of tetanus toxin which is controlled by a Cre‐recombinase‐excisable element with a platelet/megakaryocyte‐specific PF4‐promotor‐driven Cre strain, we created platelets that lacked VAMP‐2 and ‐3. These mice were crossed to VAMP‐8−/− mice to create VAMP‐2,‐3,‐8 null platelets. Secretion assays measuring the release of cargo from platelets derived from these strains are being used to evaluate the differential roles of these v‐SNAREs in platelet secretion. Upon completion of this study we will have a better understanding of molecular nature of platelet secretion. This work was supported by Grants HL56652 and HL091893 from the National Institutes of Health.