The platelet release reaction plays a critical role in thrombosis and contributes to the events that follow hemostasis. Previous studies have shown that platelet secretion is mediated by Soluble NSF Attachment Protein Receptor (SNARE) proteins from granule and plasma membranes. The SNAREs form transmembrane complexes that mediate membrane fusion and granule cargo release. Although VAMP-8 (v-SNARE) and SNAP-23 (a t-SNARE class) are important for platelet secretion, the identity of the functional syntaxin (another t-SNARE class) has been controversial. Previous studies using anti-syntaxin Abs in permeabilized platelets have suggested roles for both syntaxin-2 and syntaxin-4. In the present study, we tested these conclusions using platelets from syntaxin-knockout mouse strains and from a Familial Hemophagocytic Lymphohistiocytosis type 4 (FHL4) patient. Platelets from syntaxin-2 and syntaxin-4 single- or double-knockout mice had no secretion defect. Platelets from a FHL4 patient deficient in syntaxin-11 had a robust defect in agonist-induced secretion although their morphology, activation, and cargo levels appeared normal. Semiquantitative Western blotting showed that syntaxin-11 is the more abundant syntaxin in both human and murine platelets. Coimmunoprecipitation experiments showed that syntaxin-11 can form SNARE complexes with both VAMP-8 and SNAP-23. The results of the present study indicate that syntaxin-11, but not syntaxin-2 or syntaxin-4, is required for platelet exocytosis.
"The SNARE binding partners, and functions, of syntaxin 11 may therefore be cell type specific. Notably, syntaxin 11 also binds to SNAP23 in platelets, in which both proteins are required for granule secretion . FHL-4 mutant proteins, in which there is a partial or complete deletion of the SNARE domain, had reduced binding to SNAP23. "
[Show abstract][Hide abstract] ABSTRACT: Natural killer (NK) cell secretory lysosome exocytosis and cytotoxicity are impaired in familial hemophagocytic lymphohistiocytosis type 4 (FHL-4), a disorder caused by mutations in the gene encoding the SNARE protein syntaxin 11. We show that syntaxin 11 binds to SNAP23 in NK cells and that this interaction is reduced by FHL-4 truncation and frameshift mutation proteins that delete all or part of the SNARE domain of syntaxin 11. In contrast the FHL-4 mutant proteins bound to the Sec-1/Munc18-like (SM) protein Munc18-2. We demonstrate that the C-terminal cysteine rich region of syntaxin 11, which is deleted in the FHL-4 mutants, is S-acylated. This posttranslational modification is required for the membrane association of syntaxin 11 and for its polarization to the immunological synapse in NK cells conjugated to target cells. Moreover, we show that Munc18-2 is recruited by syntaxin 11 to intracellular membranes in resting NK cells and to the immunological synapse in activated NK cells. This recruitment of Munc18-2 is abolished by deletion of the C-terminal cysteine rich region of syntaxin 11. These results suggest a pivotal role for S-acylation in the function of syntaxin 11 in NK cells.
PLoS ONE 06/2014; 9(6):e98900. DOI:10.1371/journal.pone.0098900 · 3.23 Impact Factor
"). Similarly, the recent report that STX11 is crucial for secretion overturned the previously accepted model implicating STX2 and/or STX4 (Ye et al, 2012). It is possible that tight spatial and temporal regulation of secretion is achieved by a number of different SNARE complexes interacting and mediating fusion at different rates and/or residing on granules located in different parts of the cell. "
[Show abstract][Hide abstract] ABSTRACT: Upon activation by extracellular matrix components or soluble agonists, platelets release in excess of 300 active molecules from intracellular granules. Those factors can both activate further platelets and mediate a range of responses in other cells. The complex microenvironment of a growing thrombus, as well as platelets' roles in both physiological and pathological processes, require platelet secretion to be highly spatially and temporally regulated to ensure appropriate responses to a range of stimuli. However, how this regulation is achieved remains incompletely understood. In this review we outline the importance of regulated secretion in thrombosis as well as in 'novel' scenarios beyond haemostasis and give a detailed summary of what is known about the molecular mechanisms of platelet exocytosis. We also discuss a number of theories of how different cargoes could be released in a tightly orchestrated manner, allowing complex interactions between platelets and their environment.
British Journal of Haematology 11/2013; 165(2). DOI:10.1111/bjh.12682 · 4.71 Impact Factor
"In a new development, defective platelet secretion (dense granule, a-granule and lysosomal) despite normal granule cargo has been shown in familial haemophagocytic lymphohistiocytosis (FHL) types 3, 4 and 5, potentially lethal disorders of immune dysregulation caused respectively by defects in Munc (mammalian uncoordinated) 13–4 (UNC13D), syntaxin-11 (STX11) and Munc18b (STXBP2) coding genes (Sandrock et al, 2010; Al Hawas et al, 2012; Ye et al, 2012). Munc18b appears to be a partner of syntaxin-11 in platelet exocytosis. "
[Show abstract][Hide abstract] ABSTRACT: Genetic defects of platelets constitute rare diseases that include bleeding syndromes of autosomal dominant, recessive or X-linked inheritance. They affect platelet production, resulting in a low circulating platelet count and changes in platelet morphology, platelet function, or a combination of both with altered megakaryopoiesis and a defective platelet response. As a result, blood platelets fail to fulfil their haemostatic function. Most studied of the platelet function disorders are deficiencies of glycoprotein mediators of adhesion and aggregation while defects of primary receptors for stimuli include the P2Y12 ADP receptor. Studies on inherited defects of (i) secretion from storage organelles (dense granules, α-granules), (ii) the platelet cytoskeleton and (iii) the generation of pro-coagulant activity have identified genes indirectly controlling the functional response. Signalling pathway defects leading to agonist-specific modifications of platelet aggregation are the current target of exome-sequencing strategies. We now review recent advances in the molecular characterization of platelet function defects.
British Journal of Haematology 11/2013; 165(2). DOI:10.1111/bjh.12662 · 4.71 Impact Factor
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