Abstract 122 Cardiology: Signaling Mechanisms and Cardiovascular Function Platform, Sunday, 5/2

Hypertensive cardiac hypertrophy is a widespread form of heart disease mediated by angiotensin II (Ang II). Increased protein synthesis (protein translation) is the cardinal feature of cardiac hypertrophy and is regulated by various molecules that interact with the translational machinery of the ribosome. Elongation factor 2 (eEF-2) is an essential translation control protein whose activation by dephosphorylation is required for the synthesis of forming peptide chains. However, the mechanism by which eEF-2 is activated to produce cardiac hypertrophy is unknown. To explore this fundamental question, we determined the role of protein phosphatase 2A (PP2A), a serine/threonine phosphatase, in the activation of eEF-2 in isolated cardiac myocytes stimulated to hypertrophy with Ang II. Activation of eEF-2 was measured by Western blotting using phospho-specific and total eEF-2 antisera. Ventricular cardiac myocytes were isolated from 1-3 day old Sprague-Dawley rat hearts by enzymatic dispersion and cultured in defined, serum free media for 24 hours. Ang II treatment (10-10 -10-7 M) of cardiac myocytes for 30 minutes produced a concentration dependent activation of eEF-2 by Western analysis. Ang II (10-7 M) also produced a time dependent complete activation of eEF-2 by 30 minutes. To determine if PP2A is responsible for the activation of eEF-2 with Ang II stimulation, cardiac myocytes were treated with the specific PP2A inhibitor, okadaic acid (1-9 nM) prior to Ang II stimulation. PP2A inhibition with okadaic acid at 3 nM completely prevented Ang II activation of eEF-2. To demonstrate whether Ang II signals PP2A to activate eEF-2 via the mitogen activated protein kinase (MAPK) signaling pathway, myocytes were pre-treated with the MAPK (MEK1) inhibitor PD98059 (1-20 nM). PD98059 completely blocked Ang II stimulated activation of eEF-2 in a concentration dependent manner. Therefore, the final common pathway of Ang II stimulation of myocyte protein synthesis is by activation of eEF-2 via dephosphorylation by PP2A through the MAPK signaling pathway.