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Profiling of skeletal muscle Ankrd2 protein in human cardiac tissue and neonatal rat cardiomyocytes

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Abstract

Muscle-specific mechanosensors Ankrd2/Arpp (ankyrin repeat protein 2) and Ankrd1/CARP (cardiac ankyrin repeat protein) have an important role in transcriptional regulation, myofibrillar assembly, cardiogenesis and myogenesis. In skeletal muscle myofibrils, Ankrd2 has a structural role as a component of a titin associated stretch-sensing complex, while in the nucleus it exerts regulatory function as transcriptional co-factor. It is also involved in myogenic differentiation and coordination of myoblast proliferation. Although expressed in the heart, the role of Ankrd2 in the cardiac muscle is completely unknown. Recently, we have shown that hypertrophic and dilated cardiomyopathy pathways are altered upon Ankrd2 silencing suggesting the importance of this protein in cardiac tissue. Here we provide the underlying basis for the functional investigation of Ankrd2 in the heart. We confirmed reduced Ankrd2 expression levels in human heart in comparison with Ankrd1 using RNAseq and Western blot. For the first time we demonstrated that, apart from the sarcomere and nucleus, both proteins are localized to the intercalated disks of human cardiomyocytes. We further tested the expression and localization of endogenous Ankrd2 in rat neonatal cardiomyocytes, a well-established model for studying cardiac-specific proteins. Ankrd2 was found to be expressed in both the cytoplasm and nucleus, independently from maturation status of cardiomyocytes. In contrast to Ankrd1, it is not responsive to the cardiotoxic drug Doxorubicin, suggesting that different mechanisms govern their expression in cardiac cells.

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Acknowledgments

The excellent technical assistance of B Anicic is gratefully acknowledged. We would like to thank M Schwirtlich for connexin 43 and β-catenin antibodies. This study was supported by grants from the Ministry of Education and Science of Serbia (Project No. 173008 to DR), Collaborative Research Programme, ICGEB, Italy (grant CRP/YUG-05-01 to SK), Fondazione Cariparo, Italy (Progetto Eccellenza 2010 CHROMUS to GV and GF) and the Italian Ministry of Research (PRIN 20108XYHJS to GV).

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical standard

All procedures performed in this study involving human participants were in accordance with the ethical standards of the Faculty of Medicine, University of Belgrade, 1964 Helsinki declaration and the Law on Health Care of the Republic of Serbia. For this type of study, formal consent is not required. All applicable international and national guidelines for the care and use of animals were followed.

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Authors and Affiliations

  1. Institute of Molecular Genetics and Genetic Engineering, University of Belgrade, Vojvode Stepe 444a, PO Box 23, 11010, Belgrade, Serbia

    Jovana Jasnic-Savovic, Aleksandra Nestorovic, Dragica Radojkovic & Snezana Kojic

  2. Institute of Forensic Medicine, School of Medicine, University of Belgrade, Deligradska 31a, 11000, Belgrade, Serbia

    Slobodan Savic

  3. Institute for Medical Research, Military Medical Academy, Crnotravska 17, 11000, Belgrade, Serbia

    Sinisa Karasek

  4. CRIBI, Via Ugo Bassi 58, 35121, Padua, Italy

    Nicola Vitulo, Giorgio Valle & Georgine Faulkner

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  1. Jovana Jasnic-Savovic
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  2. Aleksandra Nestorovic
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  3. Slobodan Savic
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  4. Sinisa Karasek
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  5. Nicola Vitulo
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  6. Giorgio Valle
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  7. Georgine Faulkner
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  8. Dragica Radojkovic
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  9. Snezana Kojic
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Correspondence to Snezana Kojic.

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Jasnic-Savovic, J., Nestorovic, A., Savic, S. et al. Profiling of skeletal muscle Ankrd2 protein in human cardiac tissue and neonatal rat cardiomyocytes. Histochem Cell Biol 143, 583–597 (2015). https://doi.org/10.1007/s00418-015-1307-5

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