In hypertrophy of cultured rat cardiac myocytes, alpha 1-adrenergic agonists activate protein kinase C (PKC) and up-regulate beta-myosin heavy chain (MHC). The 3300-base pair (bp) rat beta-MHC promoter is stimulated by both an alpha 1-agonist and a constitutively activated mutant of beta-PKC (Kariya, K., Karns, L. R., Simpson, P. C. (1991) J. Biol. Chem. 266, 10023-10026). Here, we report the convergence of alpha 1-adrenergic and beta-PKC signaling on the same element of the beta-MHC promoter. A 20-bp sequence in the beta-MHC promoter (-215/-196) was required for induction by both alpha 1-adrenergic stimulation and beta-PKC and conferred induction on a heterologous promoter. This sequence bound myocyte nuclear factor(s) through a 9-bp "enhancer core" (5'-TGTGGTATG-3'). A 3-bp mutation within the enhancer core which abolished factor binding also abolished inducibility of a 215-bp beta-MHC promoter. These results support the idea that beta-PKC is in the pathway for alpha 1-adrenergic regulation of beta-MHC transcription during cardiac myocyte hypertrophy. The enhancer core is the first PKC response element mapped by transfection of an activated PKC mutant, rather than by treatment with phorbol esters.