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Establishment of the expression system for studying the function of active caspase-3 in zebrafish

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Abstract

Caspase-3, a key molecule in apoptosis, has been extensively studied in cell culture system; however, it has been less well characterized in vivo because certain mediators are required for the proteolytic activation of effector caspases, including caspase-3. In this study, various forms of caspase-3 with the C-terminal GFP tag were inserted into the pCS2+ plasmid, and the expression patterns of caspase-3 proteins were characterized in a zebrafish model system using microinjection of nucleic acids into zebrafish embryos. We have verified that active caspase-3 was generated by its autocatalytic activity under the condition of caspase-2 prodomain (C2P)-caspase-3-GFP overexpression, indicating that the C2P domain is crucial for the activation of caspase-3. We also confirmed that the C2P domain plays an important role in regulating the nuclear localization of the C2P-caspase-3 chimeric protein. We used this expression system to establish an animal model system suitable for the investigation of the functional characteristics of caspase-3 in vivo. Thus, our study provides a useful and specific tool for investigating the molecular mechanisms by which active caspase-3 regulates apoptosis during embryonic development.

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Acknowledgement

We thank Dr. Cheol-Hee Kim for the pCS2+ and pCS2+ GFP plasmids. This work was supported by a Grant (20050401-034-658-135-03-00) from BioGreen 21 Program, Rural Development Administration, Republic of Korea and National R&D Program for Cancer Control, Ministry of Health & Welfare, Republic of Korea (0720300).

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Authors and Affiliations

  1. Department of Biomedical Sciences, The Catholic University of Korea, Seoul, 137-701, Korea

    Ho-Young Kim, Goo-Young Kim, Sang-Soo Kim, Min-Kyung Nam & Hyangshuk Rhim

  2. Research Institute of Molecular Genetics, College of Medicine, The Catholic University of Korea, Seoul, 137-701, Korea

    Ho-Young Kim, Goo-Young Kim, Sang-Soo Kim, Min-Kyung Nam & Hyangshuk Rhim

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  1. Ho-Young Kim
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  2. Goo-Young Kim
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Correspondence to Hyangshuk Rhim.

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Kim, HY., Kim, GY., Kim, SS. et al. Establishment of the expression system for studying the function of active caspase-3 in zebrafish. Mol Biol Rep 36, 405–413 (2009). https://doi.org/10.1007/s11033-007-9194-z

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  • DOI: https://doi.org/10.1007/s11033-007-9194-z

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