salicylates and estrone-sulfate

Recently, a novel Slc22 gene family member expressed in murine olfactory mucosa was identified and based on sequence homology proposed to be an organic anion transporter [Oat6 (Slc22a20); J. C. Monte, M. A. Nagle, S. A. Eraly, and S. K. Nigam. Biochem Biophys Res Commun 323: 429-436, 2004]. However, no functional data for Oat6 was reported. In the present study, we demonstrate that murine Oat6 mediates the inhibitable transport of estrone sulfate using both Xenopus oocyte expression assay and Chinese hamster ovary (CHO) cells stably transfected with mOat6 (CHO-mOat6). Uptake was virtually eliminated by probenecid and the anionic herbicide 2,4-dichlorophenoxyacetate. The organic anions ochratoxin A, salicylate, penicillin G, p-aminohippurate, and urate inhibited mOat6-mediated accumulation to varying degrees. Transport of estrone sulfate by mOat6 was demonstrated to be saturable, and K(m) estimates of 109.8 +/- 22.6 microM in oocytes and 44.8 +/- 7.3 microM in CHO-mOat6 cells were obtained. Inhibitory constants for 2,4-dichlorophenoxyacetate (15.7 +/- 2.0 microM), salicylate (49.0 +/- 4.4 microM), probenecid (8.3 +/- 2.5 microM), and penicillin G (1,450 +/- 480 microM) were also determined. Accumulation of estrone sulfate mediated by mOat6 was significantly trans-stimulated by glutarate, indicating that mOat6 functions as an organic anion/dicarboxylate exchanger. These data demonstrate for the first time that the novel murine gene Oat6 (Slc22a20) encodes a functional organic anion transporter and mOat6 is indeed the newest member of the OAT gene family.

Topics: 2,4-Dichlorophenoxyacetic Acid; Animals; Biological Transport; Calcium Channel Blockers; CHO Cells; Cricetinae; Cyclooxygenase Inhibitors; Estrone; GABA Modulators; Herbicides; Mice; Ochratoxins; Organic Anion Transporters; p-Aminohippuric Acid; Penicillin G; Probenecid; Salicylates; Transfection; Uricosuric Agents; Xenopus laevis

An uncharacterized murine cDNA clone was identified and, through sequence, phylogenetic, and functional analysis, determined to encode the newest member of the organic anion transporter family, organic anion transporter 5 (Oat5; Slc22a19). The Oat5 cDNA clone contained an insert 1,964 bp in length with a predicted open reading frame (from bp 84 to bp 1,739) coding for a peptide 551 amino acids long. Slc22a19 was localized to mouse chromosome 19 near the genes encoding Oat1 (Slc22a6) and Oat3 (Slc22a8). Northern blot analysis revealed Oat5 is highly expressed in the kidney of adult mice and rats. No sexual dimorphism in renal or hepatic expression of Oat5 was observed. Unlike Oat1-3, Oat5 expression was not detected in the choroid plexus of either mice or rats. Murine Oat5-expressing Xenopus laevis oocytes supported increased accumulation of the mycotoxin ochratoxin A, compared with water-injected control oocytes. This uptake was significantly inhibited by probenecid and the organic anions 2,4-dichlorophenoxyacetic acid, salicylate, and estrone sulfate but not by para-aminohippurate or urate. Transport of ochratoxin A by murine Oat5 was saturable, with an estimated K(m) of 2.0 +/- 0.45 microM. Oat5-mediated transport was neither cis-inhibited nor trans-stimulated by the dicarboxylate glutarate. Uptake was also completely unaffected by short-circuiting of the membrane potential. Thus the motive forces behind Oat5 function, which provide insight into its membrane localization, need to be further resolved. These data demonstrate for the first time that this newly identified gene encodes a protein that functions as an organic anion transporter.

Topics: 2,4-Dichlorophenoxyacetic Acid; Amino Acid Sequence; Animals; Dose-Response Relationship, Drug; Electrochemistry; Estrone; Female; Glutarates; Kidney; Mice; Molecular Sequence Data; Mycotoxins; Ochratoxins; Oocytes; Organic Anion Transporters; Potassium; Rats; RNA, Messenger; Salicylates; Tissue Distribution; Xenopus laevis