title: Cardiac Myosin-binding Protein C and Troponin-I Phosphorylation Independently Modulate Myofilament Length-dependent Activation.
abstract: β-Adrenergic stimulation in heart leads to increased contractility and lusitropy via activation of protein kinase A (PKA). In the cardiac sarcomere, both cardiac myosin binding protein C (cMyBP-C) and troponin-I (cTnI) are prominent myofilament targets of PKA. Treatment of permeabilized myocardium with PKA induces enhanced myofilament length-dependent activation (LDA), the cellular basis of the Frank-Starling cardiac regulatory mechanism. It is not known, however, which of these targets mediates the altered LDA and to what extent. Here, we employed two genetic mouse models in which the three PKA sites in cMyBP-C were replaced with either phospho-mimic (DDD) or phospho-null (AAA) residues. AAA- or DDD-permeabilized myocytes (n = 12-17) were exchanged (~93%) for recombinant cTnI in which the two PKA sites were mutated to either phospho-mimic (DD) or phospho-null (AA) residues. Force-[Ca(2+)] relationships were determined at two sarcomere lengths (SL = 1.9 μm and SL = 2.3 μm). Data were fit to a modified Hill equation for each individual cell preparation at each SL. LDA was indexed as ΔEC50, the difference in [Ca(2+)] required to achieve 50% force activation at the two SLs. We found that PKA-mediated phosphorylation of cMyBP-C and cTnI each independently contribute to enhance myofilament length-dependent activation properties of the cardiac sarcomere, with relative contributions of ~67 and ~33% for cMyBP-C for cTnI, respectively. We conclude that β-adrenergic stimulation enhances the Frank-Starling regulatory mechanism predominantly via cMyBP-C PKA-mediated phosphorylation. We speculate that this molecular mechanism enhances cross-bridge formation at long SL while accelerating cross-bridge detachment and relaxation at short SLs.
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153.
authors.forename: Mohit
authors.initials: M
authors.lastname: Kumar
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153.
authors.forename: Suresh
authors.initials: S
authors.lastname: Govindan
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153.
authors.forename: Mengjie
authors.initials: M
authors.lastname: Zhang
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153.
authors.forename: Ramzi J
authors.initials: RJ
authors.lastname: Khairallah
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153.
authors.forename: Jody L
authors.initials: JL
authors.lastname: Martin
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153.
authors.forename: Sakthivel
authors.initials: S
authors.lastname: Sadayappan
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Ilinois 60153 pdetombe@luc.edu.
authors.forename: Pieter P
authors.initials: PP
authors.lastname: de Tombe
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D005260:Female; D006352:Heart Ventricles / Q000473:pathology; D007537:Isometric Contraction; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D032342:Muscle Cells / Q000166:cytology / Q000378:metabolism; D009200:Myocardial Contraction; D009206:Myocardium / Q000378:metabolism; D009210:Myofibrils / Q000378:metabolism*; D010766:Phosphorylation; D011943:Receptors, Adrenergic, beta / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012518:Sarcomeres / Q000378:metabolism; D015398:Signal Transduction; D013314:Stress, Mechanical; D019210:Troponin I / Q000378:metabolism*; D000099163:Myosin Binding Protein C
chemical_list: D002118:Calcium; D002352:Carrier Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D011943:Receptors, Adrenergic, beta; D011994:Recombinant Proteins; D019210:Troponin I; D000099163:Myosin Binding Protein C
keywords: Frank-Starling Mechanism; contractile protein; heart; mouse; protein kinase A (PKA); signaling; β-adrenergic stimulation