engleski

FIRST CHARACTERIZATION OF ZEBRAFISH (Danio rerio) UPTAKE TRANSPORTERS (Solute Carrier families SLC21 and SLC22)

Membrane transporters of families SLC21 (Organic anion polypeptides ; Oatp) and SLC22 (Organic cation/anion/zwitterion transporters) are involved in uptake of various endo- and xenobiotics and are important part of ADME concept. However, their role has been largely neglected in the field of aquatic toxicology. The main goal of our study was molecular characterization of toxicologically relevant uptake transporters in zebrafish (Danio rerio). Through extensive searches of available genome databases and phylogenetic analysis, we identified and annotated 12 and 17 members of SLC21 and SLC22 family, respectively. Tissue expression profiling using qPCR revealed high expression of specific uptake transporter in detoxification tissues: liver (Oatp1d1, Oatp2b1, Oct1/2a), kidney (Oatp1f2, Oatp2b1, /2a, Oct1Orctl3/4a, Orctl3/4c, Oat2c, Oat1/3), intestine (Oatp2b1, Oat2d) and gills (Oatp2b1). Based on these insights we conclude that toxicologically most important uptake transporters in zebrafish are Oatp1d1, Oatp2b1, Oatp1f2 (SLC21 family) and Oat2c, Oat2d, Oat1/3 and Oct1/2a (SLC22 family). Using the transiently transfected HEK293 cells and radioactively labeled model substrate [H3]-estrone-3-sulfate (E3S), we conducted detailed functional characterization of Oatp1d1. We found that Oatp1d1 is a high affinity E3S transporter. Based on this finding the inhibition essay was performed for 50 compounds that included model substrates/inhibitors and a series of priority environmental contaminants. Among other results, we have found that most potent competitive substrates/inhibitors are: bromosulfophthalein (Ki=0.44 μM), estradiol-17ß-glucuronide (Ki=1.91 μM), gemfibrozil (Ki=0.53 μM) and cyclosporine A (Ki=2.99 μM). Furthermore, some of the tested organophosphate pesticides can strongly interfere with transport function of Oatp1d1. Functional characterization of toxicologically important SLC22 members in zebrafish is current focus of our research.

uptake transporters; zebrafish; function

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