title: Granule-mediated release of sphingosine-1-phosphate by activated platelets.
abstract: Sphingosine-1-phosphate (S1P) is an intracellularly generated bioactive lipid essential for development, vascular integrity, and immunity. These functions are mediated by S1P-selective cell surface G-protein coupled receptors. S1P signaling therefore requires extracellular release of this lipid. Several cell types release S1P and evidence for both plasma membrane transporter-mediated and vesicle-dependent secretion has been presented. Platelets are an important source of S1P and can release it in response to agonists generated at sites of vascular injury. S1P release from agonist-stimulated platelets was measured in the presence of a carrier molecule (albumin) using HPLC-MS/MS. The kinetics and agonist-dependence of S1P release were similar to that of other granule cargo e.g. platelet factor IV (PF4). Agonist-stimulated S1P release was defective in platelets from Unc13d(Jinx) (Munc13-4 null) mice demonstrating a critical role for regulated membrane fusion in this process. Consistent with this observation, platelets efficiently converted fluorescent NBD-sphingosine to its phosphorylated derivative which accumulated in granules. Fractionation of platelet organelles revealed the presence of S1P in both the plasma membrane and in α-granules. Resting platelets contained a second pool of constitutively releasable S1P that was more rapidly labeled by exogenously added sphingosine. Our studies indicate that platelets contain two pools of S1P that are released extracellularly: a readily-exchangeable, metabolically active pool of S1P, perhaps in the plasma membrane, and a granular pool that requires platelet activation and regulated exocytosis for release.
authors.affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, Lexington, KY 40536, USA.
authors.forename: Deepa
authors.initials: D
authors.lastname: Jonnalagadda
authors.affiliation: The Gill Heart Institute and Division of Cardiovascular Medicine, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Manjula
authors.initials: M
authors.lastname: Sunkara
authors.affiliation: The Gill Heart Institute and Division of Cardiovascular Medicine, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Morris
authors.affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, Lexington, KY 40536, USA. Electronic address: whitehe@uky.edu.
authors.forename: Sidney W
authors.initials: SW
authors.lastname: Whiteheart
keywords: Munc13-4; Platelet; Platelet activation; Secretion; Sphingosine