Abstract 20476: Cardiac Myosin Heavy Chain Isoforms Are Acetylated at Lysine Residues, Resulting in Enhanced Enzymatic and Contractile Activities of the Myosin Motor.
Background: Reversible lysine-acetylation of proteins is regulated by histone acetyl transferases and deacetylases (HDACs). Previous studies from this laboratory have shown that a class-II HDAC, HDAC4 is associated with cardiac sarcomeres, and that HDAC-inhibitors enhance the contractile activity of myofilaments. Since, HDAC4 has little or no deacetylase activity of its own, this study was undertaken to examine the presence of other HDACs on cardiac sarcomeres.
Methods and Results: We prepared skinned papillary muscle fibers of the mouse heart and subjected them to western analysis. Results showed that a class-I HDAC, HDAC3 was localized to cardiac sarcomeres. By immuno-histochemical and electron microscopic analyses we found that HDAC3 was localized to the A-band of cardiac sarcomeres. We therefore examined the reversible acetylation of the A-band protein, myosin heavy chains (MHCs). MHC isoforms were prepared from control and PTU-treated mice, and examined for acetylation by western analysis with use of an anti-acetyl-lysine antibody. Both α- and β-MHC isoforms were found to be acetylated. By in vitro acetylation assay we found that S1 fragment of MHC was acetylated and HDAC3 was capable of deacetylating it. We then performed bio-mechanical studies to understand the role of acetylation on the motor activity of MHCs. Results showed that acetylation increased the actin-activated ATPase activity of α- and β-MHC isoforms by 1.6 and 2.0 fold, respectively. By in vitro motility assay we found that acetylation increased the actin-sliding velocity of α-MHC by 20% and of β-MHC by 36%, compared to their respective non-acetylated counterparts. To test whether MHC acetylation was sensitive to cardiac stress, we examined MHC acetylation during development of isoproterenol-induced hypertrophy of the heart. MHC acetylation increased progressively with the duration of stress-stimuli, reaching maximum at 4th day of agonist-treatment, suggesting that this posttranslational modification could be an early response of the myofilaments to increase contractile performance of the heart.
Conclusions: These studies provide first evidence for the localization of HDAC3 on sarcomeres and uncover a novel mechanism regulating the motor activity of cardiac MHC isoforms.
- © 2010 by American Heart Association, Inc.