Myosin-Binding Protein C Corrects an Intrinsic Non-Uniformity in Cardiac Excitation-Contraction Coupling

Abstract

Cardiac contraction is initiated by calcium release from the sarcoplasmic reticulum through channels (ryanodine receptors) that are located near the sarcomere ends. Once released, calcium must diffuse towards the sarcomere center to fully activate the actomyosin contractile system. This physical arrangement should lead to a spatial calcium gradient and thus non-uniform contractile activation. We hypothesize that myosin-binding protein C (MyBP-C), a potent thin filament activator, is localized to the sarcomere center (C-zone) to mitigate the potential deficit in calcium activation. We used EM and super-resolution STORM microscopy to visualize the relative positions of the ryanodine receptors and MyBP-C within the sarcomere. Laser scanning confocal microscopy of calcium transients in isolated cardiac cells showed that calcium concentrations at the center of each sarcomere lagged those near the ends of each sarcomere by as much as 150 nM. The functional impact of this calcium gradient was determined by examining the sliding of native, calcium-sensitive actin-thin filament shards over native mouse cardiac myosin-thick filaments using a TIRFM-based motility assay. The presence of MyBP-C enhanced the fraction of thin filaments moving within the thick filament C-zone (pCa50 6.5 ± 0.04 vs 6.4 ± 0.02; p<0.01). 3D EM reconstructions of native thin filaments suggest this calcium sensitization results from MyBP-C's N-terminal domains shifting tropomyosin on the thin filament. Using an analytic model, we show that MyBP-C residing within the C-zone can counterbalance differences in calcium activation within the sarcomere during the early phase of contraction. Thus, MyBP-C's localization to the C-zone may function to correct an intrinsic defect in cardiac excitation-contraction coupling, and any disturbance of MyBP-C localization or function will contribute to the consequent cardiac pathologies.