Nerve growth factor stimulation of p42/p44 mitogen-activated protein kinase in PC12 cells: Role of G(i/o), G protein-coupled receptor kinase 2, beta-arrestin I, and endocytic processing

In this study, we have shown that nerve growth factor (NGF)dependent activa tion of the p42/p44 mitogen-activated protein kinase (p42/p44 MAPK) pathway in PC12 cells can be partially blocked by pertussis toxin (which inactivat es the G proteins G,,). This suggests that the Trk A receptor may use a G p rotein-coupled receptor pathway to signal to p42/p44 MAPK. This was support ed by data showing that the NGF-dependent activation of p42/p44 MAPK is pot entiated in cells transfected with G protein-coupled receptor kinase 2 (GRK 2) or beta -arrestin I. Moreover, GRK2 is constitutively bound with the Trk A receptor, whereas NGF stimulates the pertussis toxin-sensitive binding o f beta -arrestin I to the TrkA receptor-GRK2 complex. Both GRK2 and beta -a rrestin I are involved in clathrin-mediated endocytic signaling to p42/p44 MAPK. Indeed, inhibitors of clathrin-mediated endocytosis (e.g., monodansyl cadaverine, concanavalin A, and hyperosmolar sucrose) reduced the NGF-depen dent activation of p42/p44 MAPK. Finally, we have found that the G protein- coupled receptor-dependent component regulating p42/p44 MAPK is required fo r NGF-induced differentiation of PC12 cells. Thus, NGF-dependent inhibition of DNA synthesis was partially blocked by PD098059 (inhibitor of MAPK kina se-1 activation) and pertussis toxin. Our findings are the first to show th at the Trk A receptor uses a classic G protein-coupled receptor-signaling p athway to promote differentiation of PC12 cells.