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Molecular cloning and functional analysis of the zebrafish luteinizing hormone beta subunit (LH<beta>) promoter

Fish Physiology and Biochemistry volume 36, pages 1253–1262 (2010)Cite this article

Abstract

The luteinizing hormone (LH) plays important roles in vertebrate reproduction. In the present study, we cloned and characterized the zebrafish (Danio rerio) LH<beta> subunit gene structure and promoter region. Analysis of 3.0 kb (LH3.4K~5′UTR) of the LH<beta> subunit proximal promoter region displayed maximal promoter activity in a tilapia ovary cell line (TO2 cells) after treatment with gonadotropin-releasing hormone (GnRH). Transient expression experiments with a 5′-deletion revealed at least 10 regulatory regions in the zebrafish LH<beta> subunit gene. Compared to the molecular mechanisms of other vertebrates, GnRH treatment led to the activation of zebrafish LH<beta> subunit gene transcription in ovary cells. We demonstrated that LH<beta> subunit gene transcription increased with 6 h of treatment with GnRH but was repressed by protein kinase C, mitogen-activated protein kinase, and calcium in the TO2 cell line. To study promoter-specific expression, we constructed an LH<beta> subunit (LH3.4k~5′UTR) promoter region-driven green fluorescent protein (GFP), and the results indicated that LH<beta> promoter-driven GFP transcripts appeared in the pituitary gland. For the gene knockdown study, we targeted knockdown of the LH<beta> subunit gene by two antisense morpholino oligonucleotides that resulted in serious abnormalities and death during zebrafish embryogenesis. These results suggest that the LH plays important roles in reproduction and general embryonic development in zebrafish.

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Acknowledgments

This work was supported by grants in 2007 and 2008 from the National Science and Technology Program for Agricultural Biotechnology for the Platform Technology Development for the Biosafety of Aquatic Genetically Modified Organisms: Technology Development for the Sterility of Genetically Modified Fish to Dr. Ching-Ming Kuo and Dr. Jyh-Yih Chen.

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  1. Marine Research Station, Institute of Cellular and Organismic Biology, Academia Sinica, 23-10 Dahuen Rd., Jiaushi, Ilan, 262, Taiwan

    Jyh-Yih Chen & Ming-Jyun Chiou

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  1. Jyh-Yih Chen
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  2. Ming-Jyun Chiou
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Correspondence to Jyh-Yih Chen.

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Chen, JY., Chiou, MJ. Molecular cloning and functional analysis of the zebrafish luteinizing hormone beta subunit (LH<beta>) promoter. Fish Physiol Biochem 36, 1253–1262 (2010). https://doi.org/10.1007/s10695-010-9405-8

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  • DOI: https://doi.org/10.1007/s10695-010-9405-8

Keywords

  • Luteinizing hormone subunit gene
  • Promoter analysis
  • Zebrafish
  • Morpholino