PLATELETS AND THROMBOPOIESIS I k B kinase phosphorylation of SNAP-23 controls platelet secretion

Several hundred molecules are released from activated platelets; these components are stored in at least 3 classes of platelet granules. The small molecules (ie, adenosine 59-diphosphate, polyphosphate, serotonin) in dense granules are important for hemostasis, given the bleeding associated with dense granule biogenesis defects (ie, Hermansky-Pudlak Syndrome). Release of polypeptides from a-granules has a more heterogeneous impact. Patients lacking a-granules (ie, Gray Platelet Syndrome) have diverse bleeding phenotypes, ranging from severe to mild. a-Granule cargo contribute to the sequelae of thrombosis (eg, tissue repair, angiogenesis, and inflammation). The role of lysosome release remains unclear. Secreting these components at a vascular damage site allows platelets to control the microenvironment at the lesion. Given this central role, understanding platelet secretion should identify potential therapeutic targets for modulating spurious thrombosis and ameliorating platelet-based bleeding disorders. Exocytosis is mediated by membrane proteins called Soluble N-ethylmaleimide sensitive factor attachment protein receptors (SNAREs). In platelets, vesicle-associated membrane protein (VAMP)8/endobrevin (a vesicle, or v-SNARE) is required for each granule type. Two target membrane SNAREs (t-SNAREs) are present: the syntaxin class (syntaxin-11 being functionally relevant) and the SNAP-23/25 class (SNAP-23 being functionally relevant). The SNAREs form a trans-bilayer complex, which facilitates fusion of the granule and plasma membranes for cargo release. Although SNAREs are essential for fusion, how, when, and where they assemble into fusigenic complexes represent key secretion control points. Thus, SNARE complex assembly is determined by numerous SNARE regulators (ie, Munc18s and Munc13s) and posttranslational modifications (ie, acylation and phosphorylation). Reports of SNARE phosphorylation focus on t-SNAREs and an array of kinases (ie, protein kinase A [PKA], IkB kinase [IKK], casein kinase II, GSK3b, and protein kinase C [PKC] isoforms). In platelets, PKC phosphorylates SNAP-23 and syntaxin-4-modulating SNARE complex formation. In neurons, SNAP-25 phosphorylation on Ser187 by PKC promotes syntaxin-1 binding. Hepp et al demonstrated phosphorylation of SNAP-23, on Ser95 and Ser120, in mast cells and in thrombin-activated platelets. In mast cells, SNAP-23 is phosphorylated by IKK, which is important for secretion. Conversely, PKA phosphorylation of SNAP-25 or syntaxin disrupts interactions. These reports illustrate the varied modes by which kinases control SNAREs and thus secretion. Liu et al showed that IkB is phosphorylated in activated platelets; whereas, Malaver et al showed that inhibiting IKK affects platelet function. These data suggest a role for IKK in platelets, but a mechanistic understanding is lacking. In this article, we asked whether IKK regulates SNAP-23, and thus secretion from platelets and whether IKK could be targeted for anti-thrombotic