Phosphoregulation of Cardiac Inotropy via Myosin Binding Protein-C During Increased Pacing Frequency or β1-Adrenergic Stimulation.
BACKGROUND
Mammalian hearts exhibit positive inotropic responses to β-adrenergic stimulation as a consequence of protein kinase A-mediated phosphorylation or as a result of increased beat frequency (the Bowditch effect). Several membrane and myofibrillar proteins are phosphorylated under these conditions, but the relative contributions of these to increased contractility are not known. Phosphorylation of cardiac myosin-binding protein-C (cMyBP-C) by protein kinase A accelerates the kinetics of force development in permeabilized heart muscle, but its role in vivo is unknown. Such understanding is important because adrenergic responsiveness of the heart and the Bowditch effect are both depressed in heart failure.


RESULTS
The roles of cMyBP-C phosphorylation were studied using mice in which either WT or nonphosphorylatable forms of cMyBP-C [ser273ala, ser282ala, ser302ala: cMyBP-C(t3SA)] were expressed at similar levels on a cMyBP-C null background. Force and [Ca(2+)]in measurements in isolated papillary muscles showed that the increased force and twitch kinetics because increased pacing or β1-adrenergic stimulation were nearly absent in cMyBP-C(t3SA) myocardium, even though [Ca(2+)]in transients under each condition were similar to WT. Biochemical measurements confirmed that protein kinase A phosphorylated ser273, ser282, and ser302 in WT cMyBP-C. In contrast, CaMKIIδ, which is activated by increased pacing, phosphorylated ser302 principally, ser282 to a lesser degree, and ser273 not at all.


CONCLUSIONS
Phosphorylation of cMyBP-C increases the force and kinetics of twitches in living cardiac muscle. Further, cMyBP-C is a principal mediator of increased contractility observed with β-adrenergic stimulation or increased pacing because of protein kinase A and CaMKIIδ phosphorylations of cMyB-C.
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Carl W
CW
Tong
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Xin
X
Wu
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Yang
Y
Liu
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Paola C
PC
Rosas
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Sakthivel
S
Sadayappan
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Andy
A
Hudmon
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Mariappan
M
Muthuchamy
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Patricia A
PA
Powers
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.).
Héctor H
HH
Valdivia
From the Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison (C.W.T., P.A.P., R.L.M.); Department of Medical Physiology (C.W.T., Y.L., P.C.R., M.M.) and Neuroscience and Experimental Therapeutics (X.W.), Texas A&M University Health Science Center College of Medicine, Temple; Baylor Scott & White Health, Temple, TX (C.W.T.); Department of Physiology, Loyola University Chicago Stritch School of Medicine, IL (S.S.); Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis (A.H.); and Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor (H.H.V.). rlmoss@wisc.edu.
Richard L
RL
Moss
D058665:Adrenergic beta-1 Receptor Agonists / Q000494:pharmacology*; D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism; D002304:Cardiac Pacing, Artificial; D002316:Cardiotonic Agents / Q000494:pharmacology*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation; D005838:Genotype; D007700:Kinetics; D008822:Mice, Transgenic; D053580:Muscle Strength / Q000187:drug effects; D009154:Mutation; D009200:Myocardial Contraction / Q000187:drug effects*; D010210:Papillary Muscles / Q000187:drug effects* / Q000378:metabolism; D010641:Phenotype; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D000099163:Myosin Binding Protein C
D058665:Adrenergic beta-1 Receptor Agonists; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D002316:Cardiotonic Agents; D002352:Carrier Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D000099163:Myosin Binding Protein C; C517673:Camk2d protein, mouse
contractility; myosin-binding protein-C; phosphorylation