Expression of organic anion transporter Oat3 in rat liver is gender-dependent

In the mammalian liver, elimination of endogenous and exogenous organic anions (metabolic products, various drugs, environmental toxins) is mediated by various organic anion transporters (OATs), transmembrane proteins that belong to solute carrier family Slc22. The rat liver organic anion transporter 3 (rOat3, Slc22a8) has been cloned, characterized as a dicarboxylate exchanger, and defined as “ male predominant” due to much higher expression of its mRNA in males (M) than in females (F). However, cellular localization and the levels of rOat3 protein expression in rat hepatocytes are unknown. This study was designed to examine the expression of rOat3 in the rat liver at the level of: a) mRNA in the tissue using RT-PCR, and b) protein in isolated total cell membranes (TCM) using Western blotting (WB) and in tissue cryosections using immunocytochemistry (IC). For this purpose we used the following experimental groups of rats: a) intact adult M and F, b) castrated M treated with oil (O ; control) or with oil solutions of testosterone (T), progesterone (P) and estradiol (E), and c) adult M treated with O, E and P. The RT-PCR data confirmed the male-predominant expression of rOat3 mRNA in the rat liver. Castration markedly decreased the expression of Oat3 mRNA ; the treatment of castrated rats with T restored it to the level of intact M, while the treatment with E and P had no effect. However, the treatment of adult M with E and P strongly downregulated the expression of rOat3 mRNA as compared to that in O-treated M. By WB in TCM prepared in non-reducing conditions, a polyclonal anti-rOat3 antibody labeled a single peptide-blockable protein band of ~50 kDa. In various experimental conditions the density pattern of this protein resembled the pattern of rOat3 mRNA. In IC studies, the anti-rOat3 antibody strongly stained some intracellular organelles in the M but not F liver. The double staining studies with organelle-specific marker antibodies (plasma membranes, endocytic vesicles, mitochondria, peroxisomes, lysosomes) indicated that rOat3 might be localized in lysosomes. We conclude that rOat3 in the rat liver exhibits strong gender differences (M>F) at both mRNA and protein levels due to testosterone stimulation and estradiol and progesterone inhibition, and may be localized in lysosomes.