Protease-activated receptor 1 (PAR1) signalling-desensitization is counteracted via PAR4 signalling in human platelets
Résumé
Protease-activated receptors 1 and 4 belong to the family of G protein-coupled receptors which induce both Gα12/13 and Gαq signalling. By applying the specific PAR1- and PAR4-activating hexapeptides SFLLRN and AYPGKF, respectively, we found that aggregation of isolated human platelets mediated via PAR1, but not via PAR4, is abolished upon homologous receptor-activation in a concentration- and time-dependent fashion. This effect was not due to receptor internalization, but to a decrease in Ca2+ mobilization, protein kinase C signalling, α granules secretion, as well as a complete lack of dense granules secretion. Interestingly, subthreshold PAR4 activation rapidly abrogated PAR1 signalling-desensitization by differentially reconstituting these affected signalling events and functional responses, sufficient to re-establish aggregation. The lack of ADP release and P2Y12 receptor-induced Gαi signalling accounted for the loss in the aggregation response, as mimicking Gαi/z signalling with 2-MeS-ADP or epinephrine, respectively, could substitute for intermediate PAR4 activation. Finally, we found that the re-sensitization of PAR1 signalling-induced aggregation via PAR4 relied on PKC-mediated release of both ADP from dense granules and fibrinogen from α granules. Our studies further elucidate differences in human platelet PAR signalling regulation and provide evidence for a cross-talk in which PAR4 signalling counteracts mechanisms involved in PAR1 signalling down-regulation.
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