Impact of site-specific phosphorylation of protein kinase A sites Ser23and Ser24of cardiac troponin I in human cardiomyocytes
Open accessAbstract
PKA-mediated phosphorylation of contractile proteins upon β-adrenergic stimulation plays an important role in the regulation of cardiac performance. Phosphorylation of the PKA sites (Ser 23/Ser 24) of cardiac troponin (cTn)I results in a decrease in myofilament Ca 2+sensitivity and an increase in the rate of relaxation. However, the relation between the level of phosphorylation of the sites and the functional effects in the human myocardium is unknown. Therefore, site-directed mutagenesis was used to study the effects of phosphorylation at Ser 23 and Ser 24 of cTnI on myofilament function in human cardiac tissue. Serines were replaced by aspartic acid (D) or alanine (A) to mimic phosphorylation and dephosphorylation, respectively. cTnI-DD mimics both sites phosphorylated, cTnI-AD mimics Ser 23 unphosphorylated and Ser 24 phosphorylated, cTnI-DA mimics Ser 23 phosphorylated and Ser 24 unphosphorylated, and cTnI-AA mimics both sites unphosphorylated. Force development was measured at various Ca 2+concentrations in permeabilized cardiomyocytes in which the endogenous troponin complex was exchanged with these recombinant human troponin complexes. In donor cardiomyocytes, myofilament Ca 2+sensitivity (pCa 50) was significantly lower in cTnI-DD (pCa 50: 5.39 ± 0.01) compared with cTnI-AA (pCa 50: 5.50 ± 0.01), cTnI-AD (pCa 50: 5.48 ± 0.01), and cTnI-DA (pCa 50: 5.51 ± 0.01) at ∼70% cTn exchange. No effects were observed on the rate of tension redevelopment. In cardiomyocytes from idiopathic dilated cardiomyopathic tissue, a linear decline in pCa 50 with cTnI-DD content was observed, saturating at ∼55% bisphosphorylation. Our data suggest that in the human myocardium, phosphorylation of both PKA sites on cTnI is required to reduce myofilament Ca 2+sensitivity, which is maximal at ∼55% bisphosphorylated cTnI. The implications for in vivo cardiac function in health and disease are detailed in the discussion in this article.
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Other studies have shown a much higher extent of human cTnI phosphorylation, with unphosphorylated group as low as ~2-10%.
“other studies which have shown a much higher extent of human cTnI phosphorylation (with un-phosphorylated group accounting for as low as ~2–10%)”
From Small and large animal models in cardiac contraction research: advantages and disadvantages (2014) · cites this paper as Wijnker, et al., 2013
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Small and large animal models in cardiac contraction research: advantages and disadvantages
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- Authors
- Wijnker, P. J. M., Foster, D. B., Tsao, A. L., Frazier, A. H., dos Remedios, C. G., Murphy, A. M., Stienen, G. J. M., & van der Velden, J.
- Affiliations
- Amsterdam Cardiovascular Sciences (NL)
- Johns Hopkins University (US)
- Muscle Research Unit, Institute for Biomedical Research, The University of Sydney, Sydney, New South Wales, Australia; and
- Laboratory for Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands;
- Corresponding author
- Wijnker, Paul J. M.
- ORCID
- Venue
- American Journal of Physiology-Heart and Circulatory Physiology
- Publisher
- American Physiological Society
- Published
- 2013-01-15
- Volume
- 304
- Issue
- 2
- Pages
- H260-H268
- ISSN
- 0363-6135 (ISSN-L)
- 1522-1539
- PMC
- PMC3543672
- OpenAlex
- W2092878566
- Cited by
- 50
- References
- 54
- Citations by year
- 2026: 1 · 2025: 2 · 2024: 1 · 2023: 3 · 2022: 3 · 2021: 1 · 2020: 4 · 2018: 4 · 2017: 5 · 2016: 4 · 2015: 6 · 2014: 8 · 2013: 8
- FWCI
- 4.53
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- American Journal of Physiology-Heart and Circulatory Physiology (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- VU Research Portal (publishedVersion)
- Pure Amsterdam UMC (submittedVersion)
- VU Research Portal (submittedVersion)
- Data Archiving and Networked Services (DANS) (submittedVersion)
- Funders
- National Heart, Lung, and Blood Institute (R01HL063038)
- Topics
- Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Cardiomyopathy and Myosin Studies
- Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Cardiac electrophysiology and arrhythmias
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Phosphodiesterase function and regulation
- Keywords
- cardiac troponin I, protein kinase A, myofilament calcium sensitivity, human cardiomyocytes, dilated cardiomyopathy, beta-adrenergic stimulation, force development, site-directed mutagenesis, aspartic acid substitution
- MeSH terms
- Calcium, Humans, Mutation, Myocardial Contraction, Myofibrils, Phosphorylation, Recombinant Proteins, Serine, Troponin, Mutagenesis, Site-Directed, Cyclic AMP-Dependent Protein Kinases, Myocytes, Cardiac, Muscle Strength
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