Platelet α-granule cargo release is fundamental to both hemostasis and thrombosis. Granule matrix hydration is a key regulated step in this process, yet its mechanism is poorly understood. In endothelial cells, there is evidence for 2 modes of cargo release: a jack-in-the-box mechanism of hydration-dependent protein phase transitions and an actin-driven granule constriction/extrusion mechanism. The third alternative considered is a prefusion, channel-mediated granule swelling, analogous to the membrane “ballooning” seen in procoagulant platelets. Using thrombin-stimulated platelets from a set of secretion-deficient, soluble N-ethylmaleimide factor attachment protein receptor (SNARE) mutant mice and various ultrastructural approaches, we tested predictions of these mechanisms to distinguish which best explains the α-granule release process. We found that the granule decondensation/hydration required for cargo expulsion was (1) blocked in fusion-protein-deficient platelets; (2) characterized by a fusion-dependent transition in granule size in contrast to a preswollen intermediate; (3) determined spatially with α-granules located close to the plasma membrane (PM) decondensing more readily; (4) propagated from the site of granule fusion; and (5) traced, in 3-dimensional space, to individual granule fusion events at the PM or less commonly at the canalicular system. In sum, the properties of α-granule decondensation/matrix hydration strongly indicate that α-granule cargo expulsion is likely by a jack-in-the-box mechanism rather than by gradual channel-regulated water influx or by a granule-constriction mechanism. These experiments, in providing a structural and mechanistic basis for cargo expulsion, should be informative in understanding the α-granule release reaction in the context of hemostasis and thrombosis.

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Published in Blood Advances, v. 2, no. 21, p. 2947-2958.

This research was originally published in Blood Advances. Irina D. Pokrovskaya, Smita Joshi, Michael Tobin, Rohan Desai, Maria A. Aronova, Jeffrey A. Kamykowski, Guofeng Zhang, Sidney W. Whiteheart, Richard D. Leapman and Brian Storrie. SNARE-dependent membrane fusion initiates α-granule matrix decondensation in mouse platelets. Blood Adv. 2018;2:2947-2958. © the American Society of Hematology.

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The Storrie laboratory was supported in part by grants from the National Heart, Lung, and Blood Institute, National Institutes of Health (R01 HL119393 and R56 HL119393). The Whiteheart laboratory was supported in part by the National Heart, Lung, and Blood Institute, National Institutes of Health (grants HL56652 and HL138179), the American Heart Association (grant AHA16GRNT27620001), a Veterans Affairs Merit Award (S.W.W.), and an American Heart Association predoctoral grant (AHA15PRE25550020) (S.J.). The Leapman laboratory was supported by the intramural program at National Institute of Biomedical Imaging and Bioengineering at the National Institutes of Health (Bethesda, MD).

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