Renal secretory transport of organic anions (OA) and organic cations (OC) controls the excretion of most foreign chemicals and/or their metabolites. We have previously characterized the mechanisms and energetics of both processes. Our current focus is the biochemistry of these transport proteins; their development, expression, and control; and their impact on the toxicity of xenobiotics. We have cloned one of the OC transport proteins (OCT2) and for the first time in any species, one of the OA transporters (ROAT1), i.e., the basolateral a-ketoglutarate/OA exchanger responsible for the uphill accumulation of OA which drives their renal secretion. Both OCT2 abd ROAT1 have been fully sequenced and the functional prooerties defined using expressed activity in Xenopus oocytes. In addition, stably transfected lines have been prepared in MDCK cells. Our cloned OC transporter (OCT2) has proven to mediate potential driven entry of OC into renal, hepatic, choroid plexus. Its role in relation to other cloned OC transporters and their relative importance in xenobiotic excretion is under evaluation. We have also generated constructs containing the gene for green fluorescent protein (GFP) linked to our transport genes. When transfected into oocytes, cell lines, or isolated teleost and mammalinan tubules, these constructs produce a fluorescent protein which can be followed optically and fortunately, still retain transport actaivity. Initial studies using these constructs have confirmed the basolateral targeting of our OA clone, as excpected based on its physiology. However, the OCT2 clone clearly shows preferential distribution to intracellular organelles, possibly endosomes. This result suggests that the multiple carriers now cloned for OC may well function at different subcellular locations as well as in different tissues or organs. Polyclonal antibodies against C-terminal peptides for this protein confirm this distribution in transfected cells, as well as in the native kidney.
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