Ablation of the calpain-targeted site in cardiac myosin binding protein-C is cardioprotective during ischemia-reperfusion injury

David Y Barefield; James W McNamara; Thomas L Lynch; Diederik W D Kuster; Suresh Govindan; Lauren Haar; Yang Wang; Erik N Taylor; John N Lorenz; Michelle L Nieman; Guangshuo Zhu; Pradeep K Luther; Andras Varró; Dobromir Dobrev; Xun Ai; Paul M L Janssen; David A Kass; Walter Keith Jones; Richard J Gilbert; Sakthivel Sadayappan

doi:10.1016/j.yjmcc.2019.03.006

Ablation of the calpain-targeted site in cardiac myosin binding protein-C is cardioprotective during ischemia-reperfusion injury

J Mol Cell Cardiol. 2019 Apr:129:236-246. doi: 10.1016/j.yjmcc.2019.03.006. Epub 2019 Mar 9.

Authors

David Y Barefield¹, James W McNamara², Thomas L Lynch³, Diederik W D Kuster⁴, Suresh Govindan⁵, Lauren Haar⁶, Yang Wang⁶, Erik N Taylor⁷, John N Lorenz⁸, Michelle L Nieman⁸, Guangshuo Zhu⁹, Pradeep K Luther¹⁰, Andras Varró¹¹, Dobromir Dobrev¹², Xun Ai¹³, Paul M L Janssen¹⁴, David A Kass⁹, Walter Keith Jones⁶, Richard J Gilbert¹⁵, Sakthivel Sadayappan¹⁶

Affiliations

¹ Department of Cell and Molecular Physiology, Loyola University Chicago, Maywood, IL, USA; Center for Genetic Medicine, Northwestern University, Chicago, IL, USA. Electronic address: david.barefield@northwestern.edu.
² Heart, Lung and Vascular Institute, Division of Cardiovascular Health and Disease, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH, USA.
³ Department of Cell and Molecular Physiology, Loyola University Chicago, Maywood, IL, USA; Department of Molecular Pharmacology and Therapeutics, Loyola University Chicago, Maywood, IL, USA.
⁴ Department of Cell and Molecular Physiology, Loyola University Chicago, Maywood, IL, USA; Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, the Netherlands.
⁵ Department of Cell and Molecular Physiology, Loyola University Chicago, Maywood, IL, USA.
⁶ Department of Molecular Pharmacology and Therapeutics, Loyola University Chicago, Maywood, IL, USA.
⁷ Department of Physiology and Biophysics, Boston University, Boston, MA, USA.
⁸ Department of Pharmacology and Systems Physiology, University of Cincinnati College of Medicine, Cincinnati, OH, USA.
⁹ Division of Cardiology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
¹⁰ Molecular Medicine Section, National Heart and Lung Institute, Imperial College London, London, UK.
¹¹ Department of Pharmacology and Pharmacotherapy, Faculty of Medicine, University of Szeged, Szeged, Hungary.
¹² Institute of Pharmacology, West German Heart and Vascular Center, University Duisburg-Essen, Essen, Germany.
¹³ Department of Physiology and Biophysics, Rush University, Chicago, IL, USA.
¹⁴ Department of Physiology and Cell Biology, The Ohio State University College of Medicine, Columbus, OH, USA.
¹⁵ Research Service, Providence VA Medical Center and Brown University, Providence, RI, USA.
¹⁶ Department of Cell and Molecular Physiology, Loyola University Chicago, Maywood, IL, USA; Heart, Lung and Vascular Institute, Division of Cardiovascular Health and Disease, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH, USA. Electronic address: sadayasl@ucmail.uc.edu.

Abstract

Cardiac myosin binding protein-C (cMyBP-C) phosphorylation is essential for normal heart function and protects the heart from ischemia-reperfusion (I/R) injury. It is known that protein kinase-A (PKA)-mediated phosphorylation of cMyBP-C prevents I/R-dependent proteolysis, whereas dephosphorylation of cMyBP-C at PKA sites correlates with its degradation. While sites on cMyBP-C associated with phosphorylation and proteolysis co-localize, the mechanisms that link cMyBP-C phosphorylation and proteolysis during cardioprotection are not well understood. Therefore, we aimed to determine if abrogation of cMyBP-C proteolysis in association with calpain, a calcium-activated protease, confers cardioprotection during I/R injury. Calpain is activated in both human ischemic heart samples and ischemic mouse myocardium where cMyBP-C is dephosphorylated and undergoes proteolysis. Moreover, cMyBP-C is a substrate for calpain proteolysis and cleaved by calpain at residues 272-TSLAGAGRR-280, a domain termed as the calpain-target site (CTS). Cardiac-specific transgenic (Tg) mice in which the CTS motif was ablated were bred into a cMyBP-C null background. These Tg mice were conclusively shown to possess a normal basal structure and function by analysis of histology, electron microscopy, immunofluorescence microscopy, Q-space MRI of tissue architecture, echocardiography, and hemodynamics. However, the genetic ablation of the CTS motif conferred resistance to calpain-mediated proteolysis of cMyBP-C. Following I/R injury, the loss of the CTS reduced infarct size compared to non-transgenic controls. Collectively, these findings demonstrate the physiological significance of calpain-targeted cMyBP-C proteolysis and provide a rationale for studying inhibition of calpain-mediated proteolysis of cMyBP-C as a therapeutic target for cardioprotection.

Keywords: Calpain; Cardioprotection; Ischemia-reperfusion injury; MYBPC3; cMyBP-C.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Calpain / metabolism*
Cardiotonic Agents / metabolism*
Carrier Proteins / metabolism*
Female
Heart Function Tests
Humans
Male
Mice, Transgenic
Middle Aged
Myocardial Infarction / metabolism
Myocardial Reperfusion Injury / metabolism*
Myocardial Reperfusion Injury / physiopathology
Myocardium / metabolism*
Phosphorylation
Proteolysis

Substances

Cardiotonic Agents
Carrier Proteins
myosin-binding protein C
Calpain

Abstract

Publication types

MeSH terms

Substances

Grants and funding