Search

Plasma membrane monoamine transporter (PMAT) is a novel polyspecific organic cation transporter that transports organic cations and the purine nucleoside, adenosine. PMAT is expressed in the kidney, but the specific localization and function of this transporter in renal cells are unclear. In this study, we developed a polyclonal antibody toward a 14-amino acid sequence in the last intracellular loop of PMAT and determined the precise cellular localization of PMAT in human and rat kidneys. Surprisingly, we found that the PMAT protein was predominantly expressed in the glomerulus with minimal expression in tubular cells. Within the glomerulus, dual-color immunofluorescence labeling showed that the PMAT protein was specifically localized to the visceral glomerular epithelial cells, i.e., podocytes. There was no significant PMAT immunoreactivity in mesangial or glomerular endothelial cells. We further showed that puromycin aminonucleoside (PAN), a classic podocyte toxin that induces massive proteinuria and severe glomerulopathy, is transported by PMAT. Expression of PMAT in Madin-Darby canine kidney cells significantly increased cell sensitivity to PAN. Decynium 22, a potent PMAT inhibitor, abolished PAN toxicity in PMAT-expressing cells. Together, our data suggest that PMAT is specifically expressed in podocytes and may play an important role in PAN-induced kidney injury. [ABSTRACT FROM AUTHOR]

Human equilibrative nucleoside transporter-3 (hENT3) was recently reported as a pH- dependent, intracellular (lysosomal) transporter capable of transporting anti-human immunodeficiency virus (HIV) dideoxynucleosides (ddNs). Because most anti-HIV ddNs (e.g., zidovudine, AZT) exhibit clinical mitochondrial toxicity, we investigated whether hENT3 facilitates transport of anti-HIV ddNs into the mitochondria. Cellular fractionation and immunofluorescence microscopy studies in several human cell lines identified a substantial presence of hENT3 in the mitochondria, with additional presence at the cell surface of two placental cell lines (JAR, JEG3). Mitochondrial or cell surface hENT3 expression was confirmed in human hepatocytes and placental tissues, respectively. Unlike endogenous hENT3, yellow fluorescent protein (YFP)-tagged hENT3 was partially directed to the lysosomes. Xenopus oocytes expressing NH2-terminal-deleted hENT3 (expressed at the cell surface) showed pH-dependent interaction with several classes of nucleosides (anti-HTV ddNs, gemcitabine, fialuridine, ribavirin) that produce mitochondrial toxicity. Transport studies in hENT3 gene-silenced JAR cells showed significant reduction in mitochondrial transport of nucleosides and nucleoside drugs. Our data suggest that cellular localization of hENT3 is cell type dependent and the native transporter is substantially expressed in mitochondria and/or cell surface. hENT3-mediated mitochondrial transport may play an important role in mediating clinically observed mitochondrial toxicity of nucleoside drugs. In addition, our finding that hENT3 is a mitochondrial transporter is consistent with the recent finding that mutations in the hENT3 gene cause an autosomal recessive disorder in humans called the H syndrome. [ABSTRACT FROM AUTHOR]