FUNCTIONAL EXPRESSION CLONING OF THE CANALICULAR SULFATE TRANSPORT-SYSTEM OF RAT HEPATOCYTES

We have cloned a single cDNA encoding the canalicular sulfate transpor ter of rat liver using Xenopus laevis oocytes as a functional expressi on system. The cloned cDNA sulfate anion transporter-1 (sat-1) express es saturable Na+-independent sulfate uptake (K(m) approximately 0.14 m M) that can be inhibited by, 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS, IC50 = 28 muM) and oxalate, but not by succinate or ch olate. These properties are very similar to sulfate uptake expressed i n oocytes injected with total rat liver mRNA and to the bicarbonate/su lfate exchange system previously characterized in canalicular rat live r plasma membrane vesicles. The cloned sat-1 cDNA has a total length o f 3726 base pairs (bp) with an open reading frame encompassing 2109 bp , a 5'-untranslated region of 367 bp, and a 3'-untranslated region of 1250 bp. The coding region predicts a protein of 703 amino acids with a calculated molecular mass of 75.4 kDa. Computer-based hydrophobicity analysis suggests the presence of 12 putative transmembrane spanning domains. Furthermore, three potential glycosylation sites are detected (Asn-158, Asn-163, Asn-587). Northern blot analysis indicates that si milar sulfate anion transporters are also present in the kidney, muscl e, and brain of rat and in the liver of the mouse. Using antisense oli gonucleotides the mRNA-species of the sat-1 analogue in rat kidney has been characterized by hybrid depletion experiments (Markovich, D., Bi ssig, M., Sorribas, V., Hagenbuch, B., Meier, P. J., and Murer, H. (19 94) J. Biol. Chem. 269, 3022-3026).