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Platelet Biology & its Disorders
Platelet Munc13-4 regulates hemostasis, thrombosis and airway inflammation
Eduardo I. Cardenas,1,2 Keegan Breaux,1 Qi Da,3,4 Jose R. Flores,1
Marco A. Ramos,1 Michael J. Tuvim,1 Alan R. Burns,5 Rolando E. Rumbaut3,4 and Roberto Adachi1
1Department of Pulmonary Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX, USA; 2Tecnologico de Monterrey, Escuela de Ingenieria y Ciencias, Monterrey, Mexico; 3Center for Translational Research on Inflammatory Diseases (CTRID), Michael E. DeBakey Veterans Affairs Medical Center, Houston, TX, USA; 4Department of Medicine, Baylor College of Medicine, Houston, TX, USA and 5College of Optometry, University of Houston, TX, USA
ABSTRACT
Platelet degranulation is crucial for hemostasis and may participate in inflammation. Exocytosis in platelets is mediated by SNARE pro- teins and should be controlled by Munc13 proteins. We found that platelets express Munc13-2 and -4. We assessed platelet granule exocy- tosis in Munc13-2 and -4 global and conditional knockout (KO) mice, and observed that deletion of Munc13-4 ablates dense granule release and indirectly impairs alpha granule exocytosis. We found no exocytic role for Munc13-2 in platelets, not even in the absence of Munc13-4. In vitro, Munc13-4-deficient platelets exhibited defective aggregation at low doses of collagen. In a flow chamber assay, we observed that Munc13-4 acted as a rate-limiting factor in the formation of thrombi. In vivo, we observed a dose-dependency between Munc13-4 expression in platelets and both venous bleeding time and time to arterial thrombosis. Finally, in a model of allergic airway inflammation, we found that platelet-spe- cific Munc13-4 KO mice had a reduction in airway hyper-responsiveness and eosinophilic inflammation. Taken together, our results indicate that Munc13-4-dependent platelet dense granule release plays essential roles in hemostasis, thrombosis and allergic inflammation.
Introduction
A key effector response from platelets is exocytosis of their alpha, dense and lysosomal granules. Alpha granules are the most abundant, and contain soluble molecules and receptors that propagate platelet activation and aggregation.1 Dense granules are secreted at a faster rate and store ADP, an autocrine agonist for platelet activation.2 Lysosomal granules contain membrane-associated proteins and acid- hydrolases, and may contribute to thrombus remodeling.3
During exocytosis, the membrane of a platelet granule fuses with the plasma
membrane. This requires the formation of a SNARE (soluble N-ethylmaleimide-
sensitive factor attachment protein receptor) complex by proteins localized on
both membranes.4 Prior to fusion, granules are brought close to the plasma mem-
brane by tethering and docking processes, but this proximity is not sufficient to
drive fusion, it also requires priming.5 Fundamental to priming is the interaction of
Munc (mammalian homolog of C. elegans uncoordinated gene) 13 with Munc18,
which allows Syntaxin to interact with the other exocytic SNARE proteins.6
Among the four paralogs of Munc13, only Munc13-4 has been studied in mouse
platelets.7 Different groups have agreed that deletion of Munc13-4 inhibits prima-
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Haematologica 2018 Volume 103(7):1235-1244
rily dense granule release,8,9 which may affect alpha granule exocytosis9,10 or inte-
grin α β activation11 depending on experimental variables.12 In addition, global IIb 3
deficiency of Munc13-4 affects hemostasis, probably due to defective platelet exo- cytosis,10,11,13 a difficult conclusion to reach because Munc13-4 is also expressed in other tissues important for hemostasis (e.g. endothelial cells).14 In humans, muta- tions in the gene encoding Munc13-4 cause familial hemophagocytic lymphohisti- ocytosis type 3 (FHL3), an autosomal recessive disorder characterized by defective secretion in cytotoxic T lymphocytes and natural killer cells, multisystemic inflam-
Correspondence:
radachi@mdanderson.org
Received: November 30, 2017. Accepted: April 12, 2018. Pre-published: April 19, 2018.
doi:10.3324/haematol.2017.185637
Check the online version for the most updated information on this article, online supplements, and information on authorship & disclosures: www.haematologica.org/content/103/7/xxx
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