OATS/Oats are transmembrane proteins that transport a variety of drugs, environmental toxins and endogenous metabolites into the cell. Zebrafish (Danio rerio) has seven OAT orthologs: Oat1, Oat2a-e and Oat3. In this study we specifically address Oat2 (Slc22a7) family. Conserved synteny analysis showed localization of zebrafish oat2 genes on two chromosomes, 11 and 17. All five zebrafish Oats were localized by live cell imaging in membranes of transiently transfected HEK293-T cells, and Oat2a, b, d, and e were confirmed using western blot analysis. Functional studies using the HEK293T cells overexpressing zebrafish Oats revealed two model fluorescent substrates of three Oats: Lucifer yellow for Oat2a and Oat2d (Km 122, and 49.7μM), and 6-carboxyfluorescein for Oat2b and Oat2d (Km 199.7, and 266.9μM). The initial screening of a series of diverse endo- and xenobiotics showed interaction with a number of compounds, including cGMP and diclofenac (IC50 27.74, and 19.14μM) with Oat2a; estrone-3-sulfate and diclofenac (IC50 30.96, and 12.6μM) with Oat2b; and fumarate and indomethacin (IC50 68.24, and 20.41μM) with Oat2d. This study provides the first comprehensive data set on Oat2 in zebrafish and offers an important basis for more detailed molecular and (eco)toxicological characterizations of these transporters.
Keywords: HEK293 transfection; In vitro characterization; Organic anion transporters 2 (Oat2); Synteny analysis; Transport activity assays; Zebrafish.
Copyright © 2017. Published by Elsevier Ltd.