H2O2 alters rat cardiac sarcomere function and protein phosphorylation through redox signaling

Avner BS, Hinken AC, Yuan C, Solaro RJ. H2O2 alters rat cardiac sarcomere function and protein phosphorylation through redox signaling. Am J Physiol Heart Circ Physiol 299: H723-H730, 2010. First published June 18, 2010; doi:10.1152/ajpheart.00050.2010.-ROS, such as H2O2, are a component of pathological conditions in many organ systems and have been reported to be elevated in cardiac pathophysiology. The experiments presented here test the hypothesis that H2O2 induces alterations in cardiac myofilament function by the posttranslational modification of sarcomeric proteins indirectly through PKC signaling. In vitro assessment of actomyosin Mg2+-ATPase activity of myofibrillar fractions showed blunted relative ATP consumption in the relaxed state (pCa 8.0) in response to treatment with 0.5 mM H2O2 before myofilament isolation. The effect was attributable to downstream "redox signaling," inasmuch as the direct application of H2O2 to isolated myofibrils did not alter Mg2+-ATPase activity. Ca2+-ATPase activity, which was used as a measure of myofibrillar myosin function, was unaffected by H2O2. Functional experiments using rat cardiac trabeculae treated with 0.5 or 5 mM H2O2 followed by detergent extraction of membranes demonstrated increased Ca2+ sensitivity of force production, a faster rate of force redevelopment, and (for 5 mM) decreased maximum tension. Biochemical analysis of myocardial samples treated with 0.5 mM H2O2 demonstrated increased phosphorylation of two sarcomeric proteins: cardiac troponin I and myosin-binding protein-C. These changes were eliminated by a general PKC inhibitor. However, H2O2 and the general PKC activator PMA induced different phosphorylation patterns in cardiomyocytes in which PKC-delta was elevated by viral infection. These data provide evidence that PKC-dependent redox signaling affects the function of cardiac myofilaments and indicate modification of specific proteins through this signaling mechanism.