title: Functional characterization of rainbow trout (Oncorhynchus mykiss) Abcg2a (Bcrp) transporter.
abstract: ABCG2 (BCRP - breast cancer resistance protein) belongs to the ATP-binding cassette (ABC) superfamily. It plays an important role in the disposition and elimination of xeno- and endobiotics and/or their metabolites in mammals. Likewise, the protective role of ABC transporters, including Abcg2, has been reported for aquatic organisms. In our previous study we have cloned the full gene sequence of rainbow trout (Oncorhynchus mykiss) Abcg2a and showed its high expression in liver and primary hepatocytes. Based on those insights, the main goal of this study was to perform a detailed functional characterization of trout Abcg2a using insect ovary cells (Spodoptera frugiperda, Sf9) as a heterologous expression system. Membrane vesicles preparations from Sf9 cells were used for the ATPase assay determinations and basic biochemical properties of fish Abcg2a versus human ABCG2 have been compared. A series of 39 physiologically and/or environmentally relevant substances was then tested on interaction with trout Abcg2a and human ABCG2. Correlation analysis reveals highly similar pattern of activation and inhibition. Significant activation of trout Abcg2a ATPase was observed for prazosin, doxorubicine, sildenafil, furosemid, propranolol, fenofibrate and pheophorbide. Pesticides showed either a weak activation (malathione) or strong (endosulfan) to weak (chlorpyrifos, fenoxycarb, DDE) inhibition of trout Abcg2a ATPase while the highest activation was obtained for benzo(a)pyrene, curcumine and testosterone. In conclusion, data from this study offer the first characterization of fish Abcg2a, reveal potent interactors among physiologically or environmentally relevant substances and point to similarities regarding strengths and interactor preferences between human ABCG2 and fish Abcg2a.
authors.affiliation: Laboratory for Molecular Ecotoxicology, Division for Marine and Environmental Research, Ruđer Bošković Institute, Zagreb, Croatia.
authors.forename: Roko
authors.initials: R
authors.lastname: Zaja
authors.affiliation: Laboratory for Molecular Ecotoxicology, Division for Marine and Environmental Research, Ruđer Bošković Institute, Zagreb, Croatia.
authors.forename: Marta
authors.initials: M
authors.lastname: Popović
authors.affiliation: Laboratory for Molecular Ecotoxicology, Division for Marine and Environmental Research, Ruđer Bošković Institute, Zagreb, Croatia.
authors.forename: Jovica
authors.initials: J
authors.lastname: Lončar
authors.affiliation: Laboratory for Molecular Ecotoxicology, Division for Marine and Environmental Research, Ruđer Bošković Institute, Zagreb, Croatia. Electronic address: smital@irb.hr.
authors.forename: Tvrtko
authors.initials: T
authors.lastname: Smital
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D029941:Fish Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D006868:Hydrolysis; D009363:Neoplasm Proteins / Q000378:metabolism; D017686:Oncorhynchus mykiss / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D061987:Sf9 Cells; D013045:Species Specificity; D018411:Spodoptera; D013997:Time Factors; D014162:Transfection; D014874:Water Pollutants, Chemical / Q000633:toxicity
chemical_list: C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D029941:Fish Proteins; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D014874:Water Pollutants, Chemical; D000255:Adenosine Triphosphate
keywords: ABCG2/BCRP; ATPase determinations; Breast cancer resistance protein; Insect ovary cells (Spodoptera frugiperda, Sf9); Rainbow trout; Substrate/interactor preferences