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An Investigation of the Molecular Mechanism of Double cMyBP-C Mutation in a Patient with End-Stage Hypertrophic Cardiomyopathy

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Abstract

Mutations in the gene coding for cardiac myosin binding protein-C (cMyBP-C), a multi-domain (C0-C10) protein, are a major causative factor for inherited hypertrophic cardiomyopathy. Patients carrying mutations in this gene have an extremely heterogeneous clinical course, with some progressing to end-stage heart failure. The cause of this variability is unknown. We here describe molecular modeling of a double mutation in domains C1 (E258K) and C2 (E441K) in a patient with severe HCM phenotype. The three-dimensional structure for the C1-motif-C2 complex was constructed with double and single mutations being introduced. Molecular dynamic simulations were performed for 10 ns under physiological conditions. The results showed that both E258K and E441K in isolation can predominantly affect the native domain as well as the nearby motif via conformational changes and result in an additive effect when they coexist. These changes involve important regions of the motif such as phosphorylation and potential actin-binding sites. Moreover, the charge reversal mutations altered the surface electrostatic properties of the complex. In addition, we studied protein expression, which showed that the mutant proteins were expressed and we can suppose that the severe phenotype was not due to haploinsufficiency. However, additional studies on human gene expression will need to confirm this hypothesis. The double mutation affecting the regulatory N-terminal of cMyBP-C have the potential of synergistically interfering with the binding to neighbouring domains and other sarcomeric proteins. These effects may account for the severe phenotype and clinical course observed in the complex cMyBP-C genotypes.

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Acknowledgments

We would like to thank Dr. Othmane Bouhali, Director of the Research Computing and Mr. Faisal Chaudhry, Senior Lead Systems Engineer, Texas A&M University in Qatar for providing supercomputing facility. We also extend our thanks to Dr. Brian P. Mitchelson, QCRC and Mr. Mark Radford for their suggestions. No human or animal studies were carried out by the authors of this article.

Funding

This work was supported by Qatar Foundation through Qatar Cardiovascular Research Center, Doha, Qatar, and it was also supported by Magdi Yacoub Research Network, London, UK. IO and FG are supported by the Italian Ministry of Health (RF 2010 – 2313451 “Hypertrophic cardiomyopathy: new insights from deep sequencing and psychosocial evaluation”) and NET-2011-02347173 (Mechanisms and treatment of coronary microvascular dysfunction in patients with genetic or secondary left ventricular hypertrophy). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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None declared.

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    Authors and Affiliations

    1. Qatar Cardiovascular Research Centre, Qatar Foundation, Doha, Qatar

      Poornima Gajendrarao, Navaneethakrishnan Krishnamoorthy, Senthil Selvaraj & Magdi Yacoub

    2. Referral Centre for Myocardial Diseases, Careggi University Hospital, Florence, Italy

      Francesca Girolami, Franco Cecchi & Iacopo Olivotto

    3. Genetic Unit, Careggi University Hospital, Florence, Italy

      Francesca Girolami

    4. Heart Science Centre, National Heart and Lung Institute, Imperial College London, Harefield, UB9 6JH, UK

      Poornima Gajendrarao, Navaneethakrishnan Krishnamoorthy & Magdi Yacoub

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    1. Poornima Gajendrarao
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    Correspondence to Magdi Yacoub.

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    Associate Editor Daniel P. Judge oversaw the review of this article

    Poornima Gajendrarao and Navaneethakrishnan Krishnamoorthy contributed equally to this work.

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    Supplemental Figure 1

    The effect of MYBPC3 variants on protein stability in H9C2 cells. H9C2 cells were either transfected with wild type (WT) or MYBPC3 variants or untransfected (control). After 36h of transfection, cells were lysed and immunoblotted with MYBPC3 antibody (DOCX 263 kb)

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    Gajendrarao, P., Krishnamoorthy, N., Selvaraj, S. et al. An Investigation of the Molecular Mechanism of Double cMyBP-C Mutation in a Patient with End-Stage Hypertrophic Cardiomyopathy. J. of Cardiovasc. Trans. Res. 8, 232–243 (2015). https://doi.org/10.1007/s12265-015-9624-6

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