PIKfyve-dependent phosphoinositide dynamics in megakaryocyte/platelet granule integrity and platelet functions
Auteurs : Manuella Caux, Rana Mansour, Jean-Marie Xuereb, Gaëtan Chicanne, Julien Viaud1, Alicia Vauclard, Frédéric Boal, Bernard Payrastre, Hélène Tronchère, Sonia Severin
Arterioscler Thromb Vasc Biol. 2022 Jun 16:101161ATVBAHA122317559 , doi: 10.1161/ATVBAHA.122.317559
Background: Secretory granules are key elements for platelet functions. Their biogenesis and integrity are regulated by fine-tuned mechanisms that need to be fully characterized. Here, we investigated the role of the phosphoinositide 5-kinase PIKfyve and its lipid products, phosphatidylinositol 5 monophosphate (PtdIns5P) and phosphatidylinositol (3,5) bisphosphate (PtdIns(3,5)P2) in granule homeostasis in megakaryocytes and platelets.
Methods: For that, we invalidated PIKfyve by pharmacological inhibition or gene silencing in megakaryocytic cell models (human MEG-01 cell line, human imMKCLs, mouse primary megakaryocytes) and in human platelets.
Results: We unveiled that PIKfyve expression and its lipid product levels increased with megakaryocytic maturation. In megakaryocytes, PtdIns5P and PtdIns(3,5)P2 were found in alpha and dense granule membranes with higher levels in dense granules. Pharmacological inhibition or knock-down of PIKfyve in megakaryocytes decreased PtdIns5P and PtdIns(3,5)P2 synthesis and induced a vacuolar phenotype with a loss of alpha and dense granule identity. Permeant PtdIns5P and PtdIns(3,5)P2 and the cation channel TRPML1 and TPC2 activation were able to accelerate alpha and dense granule integrity recovery following release of PIKfyve pharmacological inhibition. In platelets, PIKfyve inhibition specifically impaired the integrity of dense granules culminating in defects in their secretion, platelet aggregation and thrombus formation.
Conclusions: These data demonstrated that PIKfyve and its lipid products PtdIns5P and PtdIns(3,5)P2 control granule integrity both in megakaryocytes and platelets.
KEYWORDS: phosphoinositides, granules, megakaryocytes, blood platelets