ABSTRACT Cardiac hypertrophy may result from extracellular stimuli activating intracellular signalling pathways, including the mitogen-activated protein kinase (MAPK) pathways. The MAPK pathways include three cascades: ERK1/2, JNK, and p38. ERK1/2 is activated by mitogens whereas stress-related stimuli activate JNK and p38. In cardiac myocytes, activation of ERK1/2 by H 2 O 2 is attenuated by p38. This study was to determine if p38 inhibits activation of ERK1/2 by PMA, a mitogen and potent activator of ERK1/2, in adult cardiac myocytes. Inhibition of p38α/β with SB203580 increased PMA-stimulated ERK1/2 and MEK1/2 phosphorylation. FPLC of myocyte lysates on MonoQ revealed two peaks of PMA-stimulated MBP kinase activity, ERK2 and ERK1. PD 98059, which blocks MEK1/2 activation, attenuated PMA-induced ERK1/2 activation. SB203580 did not increase ERK1/2 activity. Three peaks of PMA-stimulated MEK activity were detected following chromatography on Mono Q. Peaks 1, 2, and 3 eluted at 20-25 mM, 80-100 mM, and 210 mM NaCl, respectively. PD 98059 inhibited each of these activities. Peaks 2 and 3 were increased three-fold by SB203580. In addition, a new peak was detected eluting at 40-70 mM NaCl. Hence, in adult cardiac myocytes, p38α/β attenuates PMA- dependent MEK1/2 and ERK1/2 phosphorylation. p38 MAP kinase attenuates phorbol ester-induced ERK MAP kinase activation in adult cardiac ventricular myocytes KEYWORDS: cardiac myocyte, p38 MAP kinase, ERK1/2MAP kinase, cross talk ABBREVIATIONS ACVMs, adult cardiac ventricular myocytes; DMSO, dimethylsulfoxide; DTT, dithiothreitol; ERK, extracellular signal-regulated kinase; FPLC, fast protein liquid chromatography; GST, glutathione S-transferase; MAPK, mitogen activated protein kinase; MBP, myelin basic protein; PAGE, polyacrylamide gel electrophoresis; PKI, cyclic AMP-dependent protein kinase inhibitory peptide; PMA, phorbol 12-myristate 13-acetate; PMSF, phenylmethylsulfonyl fluoride; TX-100, Triton X-100 INTRODUCTION Cardiac hypertrophy results from hemodynamic overload and/or increases in neurohormonal factors [1, 2]. Endothelin-1, angiotensin II, α 1 -adrenergic agonists and phorbol esters all induce hypertrophy [3-8] and activate the Raf-MEK-ERK cascade. Hypertrophic growth of cardiac myocytes is associated with distinct changes in cell morphology and the pattern of gene expression. Constitutive activation of Ras [9, 10], Raf-1 [11], or MEK1 [12] is sufficient to induce a hypertrophic response. Interference with ERK activation using pharmacological inhibitors of MEK1/2, antisense oligonucleotides to ERK1/2, or expression of catalytically inactive Raf, MEK, ERK1, or the MAPK phosphatase CL100, can inhibit the hypertrophic response (reviewed in [13, 14]). 1 Montreal Heart Institute, 5000 Bélanger St., Montreal, Quebec, H1T 1C8, 2 Departments of Biochemistry and 3 Medicine, Université de Montréal, Montréal, Québec, H3C 3J7, 4 Department of Pharmacology and Therapeutics, McGill University, Montréal, Québec, H3G 1Y6, Canada Benoit Boivin 1,2 and Bruce G. Allen 1,2,3,4, * *Correspondence author bruce.g.allen@umontreal.ca Current Topics in Biochemical Research Vol. 14, 1, 2012