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Subcellular localization of different regions of porcine Six1 gene and its expression analysis in C2C12 myoblasts

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Abstract

Six1 protein belongs to the Six homeoproteins family, exposing typical domain structure. Although the functions of Six1 have been drawn much attention, the roles of its individual domains are not completely elucidated. Here, we first detected the expression patterns of myogenin, MyoD, Myf5, and Six1 genes using real-time PCR in differentiating C2C12 cells cultured in differentiation medium for 2 or 6 days. The results showed that Six1 gene had the similar expression pattern with myogenin, MyoD, and Myf5 genes, which suggests that it may affect the myogenic differentiation. In order to evaluate the role of distinct domains of Six1 protein in subcellular localization, we constructed a series of truncated vectors tagged with green fluorescent proteins expressing various regions of porcine Six1 protein for subcellular localization analysis. Fluorescence confocal microscopy analysis showed that the different regions of Six1 protein displayed discrete distributions throughout the nucleus and the cytoplasm. The full-length CDS was exclusively localized in the nucleus and the individual HD domain was preferentially distributed to the nucleus both in C2C12 cells and in PK cells. However, the SD domain was diffusely distributed to the cytoplasm and the nucleus, and the localization of SD domain was biased to cytoplasm in C2C12 cells. Taken together, we conclude that the HD domain is important for the nuclear localization of porcine Six1 protein.

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Acknowledgments

This work was supported by National Natural Science Foundation of China (31000996) and the National Project for Breeding of Transgenic Pig (2008ZX08006-002).

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Authors and Affiliations

  1. Department of Animal Genetics, Breeding and Reproduction, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, 210095, China

    Wangjun Wu

  2. Key Laboratory of Swine Genetics and Breeding, Ministry of Agriculture, Key Lab of Agriculture Animal Genetics, Breeding and Reproduction, Ministry of Education, College of Animal Science and Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People’s Republic of China

    Wangjun Wu, Zhuqing Ren, Chao Chen, Yang Liu, Lin Zhang, Zhe Chao, Bo Zuo, Dequan Xu, Minggang Lei & Yuanzhu Xiong

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  1. Wangjun Wu
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  2. Zhuqing Ren
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Correspondence to Yuanzhu Xiong.

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Wangjun Wu and Zhuqing Ren contributed equally to this work.

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Wu, W., Ren, Z., Chen, C. et al. Subcellular localization of different regions of porcine Six1 gene and its expression analysis in C2C12 myoblasts. Mol Biol Rep 39, 9995–10002 (2012). https://doi.org/10.1007/s11033-012-1868-5

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  • DOI: https://doi.org/10.1007/s11033-012-1868-5

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