Abstract 197: p300-Based Acetylation Signaling Regulates Cytoskeletal Reorganization and Procoagulant Phenotype of Platelets

Abstract

Introduction: Reversible acetylation signaling catalyzed by acetyltransferases and deacetylases regulates multiple cellular events. We recently showed a role for deacetylation signaling in platelet function via the actions of the deacetylase enzyme HDAC6. Here we investigate the role of acetylation mediated by the p300 class of acetyltransferase proteins in platelet function. Methods: To test the hypothesis that p300 acetylation signaling controls cytoskeletal remodeling and the exposure of phosphatidylserine (PS) in platelets, we used cell biological and hematological methods, including enzymatic assays, microscopy, cell sorting, aggregometry and in vitro assays of thrombus formation. Results: Platelets express high levels of p300 protein. Upon stimulation with the glycoprotein VI (GPVI) agonist collagen-related peptide (CRP), total platelet acetyltransferase enzyme activity increases. Pre-treatment of platelets with pharmacological inhibitors of p300 blocks agonist-induced acetyltransferase activity and prevents the acetylation of p300 substrate proteins. Inhibition of p300 blocked the deacetylation of HDAC6 and tubulin and disrupted the platelet tubulin cytoskeleton. p300 inhibition also increased basal platelet PS exposure via a caspase-dependent apoptotic pathway. On a functional level, p300 inhibition prevented platelet shape change, aggregation and spreading while increasing the procoagulant activity of platelets in whole blood. Conclusions: These results suggest that p300-based acetylation signaling regulates separable aspects of platelet function. p300 plays a role in the regulation of the platelet cytoskeleton as well as PS exposure to effect platelet spreading, aggregation, thrombus formation as well as platelet procoagulant activities. Future studies of the platelet acetylome will aim to identify p300 target substrates and specify roles for p300 in in the context of platelet activation, thrombus formation and pathologies of the cardiovascular system. Acknowledgements: This work is funded by the American Heart Association (13POST13730003 to J.E.A. and 13EIA12630000 to O.J.T.M.) and the National Institutes of Health (R01HL101972 to O.J.T.M.).