Chronic Exercise Increases Compliant Titin and Kettin Isoform Content in Cardiac Muscle of Rat and Drosophila Models

Abstract

mutations have been characterized in the context of cardiomyopathy pathogenesis, the precise role of individual proteins in regulating length dependence of force remains unclear. Here, we used previously characterized point mutations of regulatory proteins to probe the thin filament and elucidate the role of tropomyosin in modulating the length dependence of cardiac contractility. Twitch amplitude was measured at short ($2.0mm) and long ($2.3mm) sarcomere lengths (SL) of intact cardiac trabeculae from hearts of a transgenic murine model containing a dilated cardiomyopathy-associated Tpm mutation (D230N; denoted Tpm D230N ). Trabeculae were mounted between a force transducer and length-controlling motor, perfused with oxygenated physiological solution (30 C), and electrically stimulated at 1 Hz. At short SL, twitch force between wild-type (WT) and Tpm D230N trabeculae were not significantly different (2454 versus 1653 mN/mm 2 , respectively). At long SL, WT trabeculae produced significantly higher twitch force compared to Tpm D230N (5156 versus 2753 mN/mm 2 , respectively), demonstrating reduced length-dependent augmentation of contractility in Tpm D230N trabeculae. We hypothesized that this is due to reduced azimuthal displacement of Tpm D230N along the thin filament, limiting effects of cross-bridge-mediated cooperative activation at longer SL. Thus, we incorporated an engineered calcium-sensitizing troponin C mutation (L48Q, denoted TnC L48Q ) to aid thin filament activation by developing a Tpm D230N /TnC L48Q double mutant murine model. Twitch forces in intact trabeculae from Tpm D230N /TnC L48Q mice were increased compared to Tpm D230N and were not significantly different than WT at short and long SL (3054 and 4656 mN/mm 2 , respectively). Our results suggest that tropomyosin plays a unique role in modulating the length dependence of contractility in cardiac muscle. 1561-PosWe study the regulation of cardiac contractility by using SAXS-USAXS at the ID02-beamline of the European Synchrotron (ESRF, Grenoble, France) on intact trabeculae isolated from the rat ventricle to record both the nanometerscale X-ray signals from the contractile proteins along the thin (actin) and thick (myosin) filaments and the changes of the sarcomere length (SL). Previously we demonstrated that in diastole (external [Ca 2þ ] 2.5 mM, 27 C) most of the myosin motors are in the off-state (unavailable for actin binding and ATP hydrolysis), packed into helical tracks with 43-nm periodicity on the surface of the thick filament, and that the fraction of myosin motors leaving the off-state during the twitch depends on the load through a rapid positive feedback based on thick-filament mechano-sensing (Reconditi et al. PNAS 114:3240, 2017). This regulatory mechanism occurs downstream with respect to the Ca 2þ -dependent thin-filament activation which controls cardiac contractility via the intracellular [Ca 2þ ] and the Ca 2þ -sensitivity of the filaments. Here we tested the interdependency of the two regulatory mechanisms by recording the X-r...