Nerve growth factor-stimulated B-Raf catalytic activity is refractory to inhibition by cAMP-dependent protein kinase

The cAMP-dependent protein kinase (PKA) exhibits both inhibitory and stimulatory effects upon growth factor signaling mediated by the mitogen-activated protein kinase signaling pathway. PKA has been demonstrated to inhibit Raf-l-mediated cellular proliferation. PKA can both prevent Res-dependent Raf-l activation and directly inhibit Raf-l catalytic activity. In contrast to the inhibitory effect of PKA on Raf-l-dependent processes, PKA potentiates nerve growth factor-stimulated PC12 cell differentiation, a B-Raf mediated process. This potentiation, rather than inhibition, of PC12 cell differentiation is curious in light of the ability of PKA to inhibit Raf-l catalytic activity. The kinase domains of Raf-l and B-Raf are highly conserved, and it has been predicted that B-Raf catalytic activity would also be inhibited by PKA. In this study we examined the ability of PRA to regulate the kinase activity of the B-raf protooncogene, We report that nerve growth factor-stimulated B-Raf activity is not inhibited by PKA. By contrast, an N-terminally truncated, constitutively active form of B-Raf is inhibited by PKA both in vitro and in transfected PC12 cells. These results suggest that the N-terminal regulatory domain interferes with the ability of PRA to modulate B-Raf catalytic activity and provide an explanation for the observed resistance of B-Raf-dependent processes to PKA inhibition.