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Eosinophil cationic protein enhances stabilization of β-catenin during cardiomyocyte differentiation in P19CL6 embryonal carcinoma cells

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Abstract

Prior to gastrulation, the Wnt signaling pathway through stabilized β-catenin enhances the differentiation of mouse ES cell into cardiomyocytes. We have recently shown that cardiomyocyte differentiation is enhanced by eosinophil cationic protein (ECP) through accelerated expression of marker genes of early cardiac differentiation. Furthermore, ECP enhanced the expression of Wnt3a in P19CL6 cells which were stimulated to differentiate into cardiomyocytes by DMSO. Following these findings, we evaluated in this study the potential of ECP to activate the Wnt/β-catenin signaling pathway during cardiomyocyte differentiation. Analysis by real time qPCR revealed that ECP increased the expression of Frizzled genes such as Frizzled-1, -2, -4 and -10 in P19CL6 cells in the presence of DMSO. The increased expression of those Wnt receptors was found to inhibit the phosphorylation of β-catenin resulting in the stabilization and translocation of β-catenin into the nucleus of P19CL6 cells during the early stages of cardiomyocyte differentiation. When assessed for β-catenin/TCF transcriptional activity with a TCF-luciferase (TOP/FOP) assay, ECP enhanced luciferase activity in P19CL6 cells during 48 h after transfection with TOP/FOP flash reporter in a stoichiometric manner. Collectively, this suggests that ECP can activate a canonical Wnt/β-catenin signaling pathway by enhancing the stabilization of β-catenin during cardiomyocyte differentiation.

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Acknowledgments

This research was supported in part by the Ministry of Education, Culture, Sports, Science and Technology Grant-in-Aid for Scientific Research, by Japan Science and Technology Agency (JST), and by Chugoku Industrial Innovation Center.

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Authors and Affiliations

  1. Laboratory of Nano-Biotechnology, Department of Chemistry and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Okayama, 700-8530, Japan

    Guoliang Jin, Akifumi Mizutani, Takayuki Fukuda, Takayuki Otani, Ting Yan, Marta Prieto Vila, Hiroshi Murakami, Takayuki Kudoh, Tomonari Kasai & Masaharu Seno

  2. Department of Neurology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, 221002, Jiangsu, China

    Guoliang Jin

  3. Department of Molecular Biology and Biochemistry, Graduate School of Medicine, Dentistry and Pharmacology, Okayama University, Okayama, 700-8558, Japan

    Satoshi Hirohata

  4. Laboratory of Cancer Prevention, National Cancer Institute, Frederick National Laboratory for Cancer Research, Frederick, MD, 21702, USA

    David S. Salomon

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  1. Guoliang Jin
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  2. Akifumi Mizutani
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  3. Takayuki Fukuda
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  4. Takayuki Otani
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Correspondence to Masaharu Seno.

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Jin, G., Mizutani, A., Fukuda, T. et al. Eosinophil cationic protein enhances stabilization of β-catenin during cardiomyocyte differentiation in P19CL6 embryonal carcinoma cells. Mol Biol Rep 40, 3165–3171 (2013). https://doi.org/10.1007/s11033-012-2390-5

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  • DOI: https://doi.org/10.1007/s11033-012-2390-5

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