Protease-activated receptor 4 variant p.Tyr157Cys reduces platelet functional responses and alters receptor trafficking
Norman, Jane E. and Cunningham, Margaret R. and Jones, Matthew L. and Walker, Mary E. and Westbury, Sarah K. and Sessions, Richard B. and Mundell, Stuart J. and Mumford, Andrew D. (2016) Protease-activated receptor 4 variant p.Tyr157Cys reduces platelet functional responses and alters receptor trafficking. Arteriosclerosis, Thrombosis, and Vascular Biology, 36 (5). pp. 952-960. ISSN 1079-5642 (https://doi.org/10.1161/ATVBAHA.115.307102)
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Abstract
OBJECTIVE—: Protease-activated receptor 4 (PAR4) is a key regulator of platelet reactivity and is encoded by F2RL3, which has abundant rare missense variants. We aimed to provide proof of principle that rare F2LR3 variants potentially affect on platelet reactivity and responsiveness to PAR1 antagonist drugs and to explore underlying molecular mechanisms. APPROACH AND RESULTS—: We identified 6 rare F2RL3 missense variants in 236 cardiac patients, of which the variant causing a tyrosine 157 to cysteine substitution (Y157C) was predicted computationally to affect most on PAR4 structure. Y157C platelets from 3 cases showed reduced responses to PAR4-activating peptide and to α-thrombin compared with controls, but no reduction in responses to PAR1-activating peptide. Pretreatment with the PAR1 antagonist vorapaxar caused lower residual α-thrombin responses in Y157C platelets than in controls, indicating greater platelet inhibition. HEK293 cells transfected with a PAR4 Y157C expression construct had reduced PAR4 functional responses, unchanged total PAR4 expression but reduced surface expression. PAR4 Y157C was partially retained in the endoplasmic reticulum and displayed an expression pattern consistent with defective N-glycosylation. Mutagenesis of Y322, which is the putative hydrogen bond partner of Y157, also reduced PAR4 surface expression in HEK293 cells. CONCLUSIONS—: Reduced PAR4 responses associated with Y157C result from aberrant anterograde surface receptor trafficking, in part, because of disrupted intramolecular hydrogen bonding. Characterization of PAR4 Y157C establishes that rare F2RL3 variants have the potential to markedly alter platelet PAR4 reactivity particularly after exposure to therapeutic PAR1 antagonists.
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Item type: Article ID code: 56487 Dates: DateEvent1 May 2016Published10 March 2016Published Online22 February 2016AcceptedSubjects: Medicine > Pharmacy and materia medica Department: Faculty of Science > Strathclyde Institute of Pharmacy and Biomedical Sciences Depositing user: Pure Administrator Date deposited: 24 May 2016 09:47 Last modified: 07 Jun 2024 01:22 Related URLs: URI: https://strathprints.strath.ac.uk/id/eprint/56487