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Generation of Tg(cyp1a:gfp) Transgenic Zebrafish for Development of a Convenient and Sensitive In Vivo Assay for Aryl Hydrocarbon Receptor Activity

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Abstract

Both dioxins/dioxin-like compounds and polycyclic aromatic hydrocarbons (PAHs) are persistent organic pollutants and cause multiple adverse health effects on human and wildlife. Cyp1a is the most commonly used biomarker induced by these pollutants through activation of the aryl hydrocarbon receptor (AhR) pathway. Here we generated Tg(cyp1a:gfp) transgenic zebrafish for establishing a convenient in vivo assay for analysing these xenobiotic compounds. The Tg(cyp1a:gfp) larvae at 4 day post-fertilization were tested with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and GFP induction was observed mainly in the kidney, liver and gut. Similar GFP expression was also induced strongly by two dioxin-like chemicals, co-planar polychlorinated biphenyl (PCB126) and polychlorinated dibenzo-p-furan (PeCDF) and relatively weakly by two PAHs, 3-methylcholanthrene (3-MC) and benzo[a]pyrene (BAP). The lowest observed effective concentration (LOEC) of TCDD was estimated to be ∼1 pM and the EC50 (effective concentration to induce GFP in 50 % of Tg(cyp1a:gfp) larvae) was ∼10 pM. PCB126 and PeCDF had ∼10× lower potencies in GFP induction than TCDD, while the potencies for 3-MC and BAP were at least 1000× lower. The sensitivity of Tg(cyp1a:gfp) larvae to respond TCDD was also favourable compared to that of ethoxyresorufin-O-deethylase (EROD) assay in both zebrafish larvae and adult livers. As GFP-based assay in transgenic zebrafish can be easily accommodated in multi-well dishes, the Tg(cyp1a:gfp) zebrafish should provide not only a valuable biomonitoring tool for aquatic contaminants but also a potential high-throughput chemical screening platform for identification of new AhR agonists.

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Acknowledgments

This work was supported by the Singapore National Research Foundation under its Environmental and Water Technologies Strategic Research Programme and administered by the Environment and Water Industry Programme Office (EWI) of the PUB, grant number R-154-000-328-272, and National Medical ResearchCouncil of Singapore, grant number R-154-000-547-511.

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    Authors and Affiliations

    1. Department of Biological Sciences, National University of Singapore, S3-Level 6, 14 Science Drive 4, Singapore, 117543, Singapore

      Hongyan Xu, Caixia Li, Yan Li, Grace Hwee Boon Ng, Chunsheng Liu, Xiaoyan Zhang & Zhiyuan Gong

    2. Pearl River Fishery Research Institute, Chinese Academic of Fishery Sciences, Guangzhou, 510380, China

      Hongyan Xu

    3. College of Fisheries, Huazhong Agricultural University, Wuhan, 430070, China

      Chunsheng Liu

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    1. Hongyan Xu
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    Correspondence to Zhiyuan Gong.

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    Hongyan Xu and Caixia Li contributed equally to this work.

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    Xu, H., Li, C., Li, Y. et al. Generation of Tg(cyp1a:gfp) Transgenic Zebrafish for Development of a Convenient and Sensitive In Vivo Assay for Aryl Hydrocarbon Receptor Activity. Mar Biotechnol 17, 831–840 (2015). https://doi.org/10.1007/s10126-015-9669-1

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