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Zebrafish (Danio rerio) Oatp2b1 as a functional ortholog of the human OATP2B1 transporter

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Abstract

OATP2B1 belongs to a highly conserved organic anion transporting polypeptide (OATP) family of transporters, involved in the cellular uptake of both endogenous and exogenous compounds. The reported substrates of human OATP2B1 include steroid conjugates, bile salts, and thyroid hormones, as well as pharmaceuticals. Human OATP2B1 has orthologous genes in other vertebrate species, including zebrafish (Danio rerio), a widely used model organism in biomedical and environmental research. Our previous studies showed that zebrafish Oatp2b1 was phylogenetically closest to mammalian OATP2B1/Oatp2b1 and that it shares a similar tissue expression pattern. In this study, we aimed at discovering whether zebrafish Oatp2b1 could be a functional ortholog of human and rodent OATP2B1. To test this hypothesis, our primary goal was to obtain the first in vitro and in silico insights related to the structure and potential substrate preferences of zebrafish Oatp2b1. We generated cells transiently and stably transfected with zebrafish Oatp2b1 cloned from zebrafish liver, constructed an Oatp2b1 homology model, developed transport activity assays with model fluorescent substrate Lucifer yellow, and finally utilized this assay to analyze the interaction of zebrafish Oatp2b1 with both physiological and xenobiotic substances. Apart from structure similarities, our data revealed the strongest interaction of zebrafish Oatp2b1 with bile acids, steroid sulfates, thyroid hormones, and bilirubin, as well as xenobiotics bromosulfophthalein and sulfasalazine, which indicates its functional orthology with human OATP2B1.

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Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

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Funding

This study was financed by the Croatian Science Foundation (Project No. IP-2019–04-1147) and partially supported under the STIM-REI project, Contract Number: KK.01.1.1.01.0003, a project funded by the European Union through the European Regional Development Fund – Operational Programme Competitiveness and Cohesion 2014–2020 (KK.01.1.1.01). The computational resources and Biovia Discovery Studio Client v18.1 software (Dassault Systèmes, Vélizy-Villacoublay, France), used for homology modeling and molecular docking studies, were provided through Croatian Science Foundation projects (grant numbers HrZZ-IP-2013–11-4307 and HrZZ-IP-2018–01-7683).

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Authors and Affiliations

  1. Laboratory for Molecular Ecotoxicology, Division for Marine and Environmental Research, Ruđer Bošković Institute, Bijenička cesta 54, 10000, Zagreb, Croatia

    Jelena Dragojević, Marta Popović & Tvrtko Smital

  2. Institute for Medical Research and Occupational Health, Ksaverska cesta 2, 10000, Zagreb, Croatia

    Nikola Marakovic

Authors
  1. Jelena Dragojević
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  2. Nikola Marakovic
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  3. Marta Popović
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  4. Tvrtko Smital
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Contributions

JD designed and performed majority of cloning, transfection, and interaction experiments and wrote the manuscript; NM performed homology modeling and molecular docking studies; MP did initial cloning of zebrafish Oatp2b1; TS supervised the project, study conception, and design and wrote the manuscript with input from all authors. All of the authors reviewed the results and approved the final version of the manuscript.

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Correspondence to Tvrtko Smital.

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Dragojević, J., Marakovic, N., Popović, M. et al. Zebrafish (Danio rerio) Oatp2b1 as a functional ortholog of the human OATP2B1 transporter. Fish Physiol Biochem 47, 1837–1849 (2021). https://doi.org/10.1007/s10695-021-01015-7

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  • DOI: https://doi.org/10.1007/s10695-021-01015-7

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