HYPERTROPHIC AGONISTS STIMULATE THE ACTIVITIES OF THE PROTEIN-KINASES C-RAF AND A-RAF IN CULTURED VENTRICULAR MYOCYTES

We detected expression of two Raf isoforms, c-Raf and A Raf, in neonatal rat heart, Both isoforms phosphorylated, activated, and formed complexes with mitogen-activated protein kinase kinase 1 in vitro. However, these isoforms were differentially activated by hypertrophic stimuli such as peptide growth factors, endothelin-l (ET1), or 12-O-tetradecanoylphorbol-13-acetate (TPA) that activate the mitogen-activated protein kinase cascade, Exposure of cultured ventricular myocytes to acidic fibroblast growth factor activated c-Raf but not A-Raf, In contrast, TPA produced a sustained activation of A-Raf and only transiently activated c-Raf. ET1 transiently activated both isoforms. TPA and ET1 were the most potent activators of c-Raf and A-Raf. Both utilized protein kinase C dependent pathways, but stimulation by ET1 was also partially sensitive to pertussis toxin pretreatment. c-Raf was inhibited by activation of cAMP-dependent protein kinase although A-Raf was less affected. Fetal calf serum, phenylephrine, and carbachol were less potent activators of c-Raf and A-Raf. These results demonstrate that A-Raf and c-Raf are differentially regulated and that A-Raf may be an important mediator of mitogen activated protein kinase cas cade activation when cAMP is elevated.