Functional expression cloning of the canalicular sulfate transport system of rat hepatocytes.
We have cloned a single cDNA encoding the canalicular sulfate transporter of rat liver using Xenopus laevis oocytes as a functional expression system. The cloned cDNA sulfate anion transporter-1 (sat-1) expresses saturable Na(+)-independent sulfate uptake (Km approximately 0.14 mM) that can be inhibited by 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS, IC50 = 28 microM) and oxalate, but not by succinate or cholate. These properties are very similar to sulfate uptake expressed in oocytes injected with total rat liver mRNA and to the bicarbonate/sulfate exchange system previously characterized in canalicular rat liver plasma membrane vesicles. The cloned sat-1 cDNA has a total length of 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 similar sulfate anion transporters are also present in the kidney, muscle, and brain of rat and in the liver of the mouse. Using antisense oligonucleotides the mRNA-species of the sat-1 analogue in rat kidney has been characterized by hybrid depletion experiments (Markovich, D., Bissig, M., Sorribas, V., Hagenbuch, B., Meier, P. J., and Murer, H. (1994) J. Biol. Chem. 269, 3022-3026).
Bissig
M
M
Department of Medicine, University Hospital, Zurich, Switzerland.
Hagenbuch
B
B
Stieger
B
B
Koller
T
T
Meier
P J
PJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001648:Bile Canaliculi / Q000378:metabolism*; D001693:Biological Transport, Active; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D007668:Kidney / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D026901:Membrane Transport Proteins; D051379:Mice; D008961:Models, Structural; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism; D009928:Organ Specificity; D017433:Protein Structure, Secondary; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D000076988:Sulfate Transporters; D013431:Sulfates / Q000378:metabolism*; D013462:Sulfur Radioisotopes; D014982:Xenopus laevis
D002352:Carrier Proteins; D018076:DNA, Complementary; D026901:Membrane Transport Proteins; D012333:RNA, Messenger; C085609:SLC26A4 protein, human; D000076988:Sulfate Transporters; D013431:Sulfates; D013462:Sulfur Radioisotopes