Homers regulate calcium entry and aggregation in human platelets: a role for Homers in the association between STIM1 and Orai1

Abstract

Homer is a family of cytoplasmic adaptor proteins that play different roles in cell function, including the regulation of G-protein-coupled receptors. These proteins contain an Ena (Enabled)/VASP (vasodilator-stimulated phosphoprotein) homology 1 domain that binds to the PPXXF sequence motif, which is present in different Ca2+-handling proteins such as IP3 (inositol 1,4,5-trisphosphate) receptors and TRPC (transient receptor potential canonical) channels. In the present study we show evidence for a role of Homer proteins in the STIM1 (stromal interaction molecule 1)–Orai1 association, as well as in the TRPC1–IP3RII (type II IP3 receptor) interaction, which might be of relevance in platelet function. Treatment of human platelets with thapsigargin or thrombin results in a Ca2+-independent association of Homer1 with TRPC1 and IP3RII. In addition, thapsigargin and thrombin enhanced the association of Homer1 with STIM1 and Orai1 in a Ca2+-dependent manner. Interference with Homer function by introduction of the synthetic PPKKFR peptide into cells, which emulates the proline-rich sequences of the PPXXF motif, reduced STIM1–Orai1 and TRPC1– IP3RII associations, as compared with the introduction of the inactive PPKKRR peptide. The PPKKFR peptide attenuates thrombin-evoked Ca2+ entry and the maintenance of thapsigargin-induced store-operated Ca2+ entry. Finally, the PPKKFR peptide attenuated thrombin-induced platelet aggregation. The findings of the present study support an important role for Homer proteins in thrombin-stimulated platelet function, which is likely to be mediated by the support of agonist-induced Ca2+ entry.