Molecular Screen Identifies Cardiac Myosin-Binding Protein-C as a Protein Kinase G-Ialpha Substrate Journal Article

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Authors: Thoonen, R.; Giovanni, S.; Govindan, S.; Lee, D. I.; Wang, G. R.; Calamaras, T. D.; Takimoto, E.; Kass, D. A.; Sadayappan, S; Blanton, R. M.
Article Title: Molecular Screen Identifies Cardiac Myosin-Binding Protein-C as a Protein Kinase G-Ialpha Substrate
Abstract: BACKGROUND: Pharmacological activation of cGMP-dependent protein kinase G I (PKGI) has emerged as a therapeutic strategy for humans with heart failure. However, PKG-activating drugs have been limited by hypotension arising from PKG-induced vasodilation. PKGIalpha antiremodeling substrates specific to the myocardium might provide targets to circumvent this limitation, but currently remain poorly understood. METHODS AND RESULTS: We performed a screen for myocardial proteins interacting with the PKGIalpha leucine zipper (LZ)-binding domain to identify myocardial-specific PKGI antiremodeling substrates. Our screen identified cardiac myosin-binding protein-C (cMyBP-C), a cardiac myocyte-specific protein, which has been demonstrated to inhibit cardiac remodeling in the phosphorylated state, and when mutated leads to hypertrophic cardiomyopathy in humans. GST pulldowns and precipitations with cGMP-conjugated beads confirmed the PKGIalpha-cMyBP-C interaction in myocardial lysates. In vitro studies demonstrated that purified PKGIalpha phosphorylates the cMyBP-C M-domain at Ser-273, Ser-282, and Ser-302. cGMP induced cMyBP-C phosphorylation at these residues in COS cells transfected with PKGIalpha, but not in cells transfected with LZ mutant PKGIalpha, containing mutations to disrupt LZ substrate binding. In mice subjected to left ventricular pressure overload, PKGI activation with sildenafil increased cMyBP-C phosphorylation at Ser-273 compared with untreated mice. cGMP also induced cMyBP-C phosphorylation in isolated cardiac myocytes. CONCLUSIONS: Taken together, these data support that PKGIalpha and cMyBP-C interact in the heart and that cMyBP-C is an anti remodeling PKGIalpha kinase substrate. This study provides the first identification of a myocardial-specific PKGIalpha LZ-dependent antiremodeling substrate and supports further exploration of PKGIalpha myocardial LZ substrates as potential therapeutic targets for heart failure.
Journal Title: Circulation.Heart failure
Volume: 8
Issue: 6
ISSN: 1941-3297; 1941-3289
Publisher: American Heart Association, Inc  
Journal Place: United States
Date Published: 2015
Start Page: 1115
End Page: 1122
Language: eng
DOI/URL:

10.1161/CIRCHEARTFAILURE.115.002308
Notes: LR: 20151120; CI: (c) 2015; GR: K02 HL114749/HL/NHLBI NIH HHS/United States; GR: R01 HL105826/HL/NHLBI NIH HHS/United States; JID: 101479941; NIHMS731603; OID: NLM: NIHMS731603 [Available on 11/01/16]; OID: NLM: PMC4651836 [Available on 11/01/16]; OTO: NOTNLM; PMCR: 2016/11/01 00:00; 2015/05/05 [received]; 2015/10/08 [accepted]; 2015/10/18 [aheadofprint]; ppublish