Gα12/13-mediated Up-regulation of TRPC6 Negatively Regulates Endothelin-1-induced Cardiac Myofibroblast Formation and Collagen Synthesis through Nuclear Factor of Activated T Cells Activation

Sustained elevation of [Ca 2+ ] i has been implicated in many cellular events. We previously reported that α subunits of G 12 family G proteins (Gα 12/13 ) participate in sustained Ca 2+ influx required for the activation of nuclear factor of activated T cells (NFAT), a Ca 2+ -responsive transcriptional factor, in rat neonatal cardiac fibroblasts. Here, we demonstrate that Gα 12/13 -mediated up-regulation of canonical transient receptor potential 6 (TRPC6) channels participates in sustained Ca 2+ influx and NFAT activation by endothelin (ET)-1 treatment. Expression of constitutively active Gα 12 or Gα 13 increased the expression of TRPC6 proteins and basal Ca 2+ influx activity. The treatment with ET-1 increased TRPC6 protein levels through Gα 12/13 , reactive oxygen species, and c-Jun N-terminal kinase (JNK)-dependent pathways. NFAT is activated by sustained increase in [Ca 2+ ] i through up-regulated TRPC6. A Gα 12/13 -inhibitory polypeptide derived from the regulator of the G-protein signaling domain of p115-Rho guanine nucleotide exchange factor and a JNK inhibitor, SP600125, suppressed the ET-1-induced increase in expression of marker proteins of myofibroblast formation through a Gα 12/13 -reactive oxygen species-JNK pathway. The ET-1-induced myofibroblast formation was suppressed by overexpression of TRPC6 and CA NFAT, whereas it was enhanced by TRPC6 small interfering RNAs and cyclosporine A. These results suggest two opposite roles of Gα 12/13 in cardiac fibroblasts. First, Gα 12/13 mediate ET-1-induced myofibroblast formation. Second, Gα 12/13 mediate TRPC6 up-regulation and NFAT activation that negatively regulates ET-1-induced myofibroblast formation. Furthermore, TRPC6 mediates hypertrophic responses in cardiac myocytes but suppresses fibrotic responses in cardiac fibroblasts. Thus, TRPC6 mediates opposite responses in cardiac myocytes and fibroblasts.