Myosin binding protein C, a phosphorylation-dependent force regulator in muscle that controls the attachment of myosin heads by its interaction with myosin S2

TY - JOUR

T1 - Myosin binding protein C, a phosphorylation-dependent force regulator in muscle that controls the attachment of myosin heads by its interaction with myosin S2

AU - Kunst, G

AU - Kress, K R

AU - Gruen, M

AU - Uttenweiler, D

AU - Gautel, M

AU - Fink, R H

PY - 2000/3/22

Y1 - 2000/3/22

N2 - Myosin binding protein C (MyBP-C) is one of the major sarcomeric proteins involved in the pathophysiology of familial hypertrophic cardiomyopathy (FHC). The cardiac isoform is tris-phosphorylated by cAMP-dependent protein kinase (cAPK) on beta-adrenergic stimulation at a conserved N-terminal domain (MyBP-C motif), suggesting a role in regulating positive inotropy mediated by cAPK. Recent data show that the MyBP-C motif binds to a conserved segment of sarcomeric myosin S2 in a phosphorylation-regulated way. Given that most MyBP-C mutations that cause FHC are predicted to result in N-terminal fragments of the protein, we investigated the specific effects of the MyBP-C motif on contractility and its modulation by cAPK phosphorylation. The diffusion of proteins into skinned fibers allows the investigation of effects of defined molecular regions of MyBP-C, because the endogenous MyBP-C is associated with few myosin heads. Furthermore, the effect of phosphorylation of cardiac MyBP-C can be studied in a defined unphosphorylated background in skeletal muscle fibers only. Triton skinned fibers were tested for maximal isometric force, Ca(2+)/force relation, rigor force, and stiffness in the absence and presence of the recombinant cardiac MyBP-C motif. The presence of unphosphorylated MyBP-C motif resulted in a significant (1) depression of Ca(2+)-activated maximal force with no effect on dynamic stiffness, (2) increase of the Ca(2+) sensitivity of active force (leftward shift of the Ca(2+)/force relation), (3) increase of maximal rigor force, and (4) an acceleration of rigor force and rigor stiffness development. Tris-phosphorylation of the MyBP-C motif by cAPK abolished these effects. This is the first demonstration that the S2 binding domain of MyBP-C is a modulator of contractility. The anchorage of the MyBP-C motif to the myosin filament is not needed for the observed effects, arguing that the mechanism of MyBP-C regulation is at least partly independent of a "tether," in agreement with a modulation of the head-tail mobility. Soluble fragments occurring in FHC, lacking the spatial specificity, might therefore lead to altered contraction regulation without affecting sarcomere structure directly.

AB - Myosin binding protein C (MyBP-C) is one of the major sarcomeric proteins involved in the pathophysiology of familial hypertrophic cardiomyopathy (FHC). The cardiac isoform is tris-phosphorylated by cAMP-dependent protein kinase (cAPK) on beta-adrenergic stimulation at a conserved N-terminal domain (MyBP-C motif), suggesting a role in regulating positive inotropy mediated by cAPK. Recent data show that the MyBP-C motif binds to a conserved segment of sarcomeric myosin S2 in a phosphorylation-regulated way. Given that most MyBP-C mutations that cause FHC are predicted to result in N-terminal fragments of the protein, we investigated the specific effects of the MyBP-C motif on contractility and its modulation by cAPK phosphorylation. The diffusion of proteins into skinned fibers allows the investigation of effects of defined molecular regions of MyBP-C, because the endogenous MyBP-C is associated with few myosin heads. Furthermore, the effect of phosphorylation of cardiac MyBP-C can be studied in a defined unphosphorylated background in skeletal muscle fibers only. Triton skinned fibers were tested for maximal isometric force, Ca(2+)/force relation, rigor force, and stiffness in the absence and presence of the recombinant cardiac MyBP-C motif. The presence of unphosphorylated MyBP-C motif resulted in a significant (1) depression of Ca(2+)-activated maximal force with no effect on dynamic stiffness, (2) increase of the Ca(2+) sensitivity of active force (leftward shift of the Ca(2+)/force relation), (3) increase of maximal rigor force, and (4) an acceleration of rigor force and rigor stiffness development. Tris-phosphorylation of the MyBP-C motif by cAPK abolished these effects. This is the first demonstration that the S2 binding domain of MyBP-C is a modulator of contractility. The anchorage of the MyBP-C motif to the myosin filament is not needed for the observed effects, arguing that the mechanism of MyBP-C regulation is at least partly independent of a "tether," in agreement with a modulation of the head-tail mobility. Soluble fragments occurring in FHC, lacking the spatial specificity, might therefore lead to altered contraction regulation without affecting sarcomere structure directly.

KW - Calcium/physiology

KW - Carrier Proteins/metabolism

KW - Elasticity

KW - Histological Techniques

KW - Isometric Contraction/physiology

KW - Kinetics

KW - Muscle Fibers, Skeletal/physiology

KW - Muscle, Skeletal/metabolism

KW - Myocardial Contraction/physiology

KW - Myosins/physiology

KW - Peptide Fragments/physiology

KW - Phosphorylation

KW - Recombinant Proteins/metabolism

KW - Sarcomeres/metabolism

KW - Solubility

U2 - 10.1161/01.res.86.1.51

DO - 10.1161/01.res.86.1.51

M3 - Article

C2 - 10625305

SN - 0009-7330

VL - 86

SP - 51

EP - 58

JO - Circulation Research

JF - Circulation Research

IS - 1

ER -