Cloning and functional characterization of human sodium-dependent organic anion transporter (SLC10A6).
We have cloned human sodium-dependent organic anion transporter (SOAT) cDNA, which consists of 1502 bp and encodes a 377-amino acid protein. SOAT shows 42% sequence identity to the ileal apical sodium-dependent bile acid transporter ASBT and 33% sequence identity to the hepatic Na(+)/taurocholate-cotransporting polypeptide NTCP. Immunoprecipitation of a SOAT-FLAG-tagged protein revealed a glycosylated form at 46 kDa that decreased to 42 kDa after PNGase F treatment. SOAT exhibits a seven-transmembrane domain topology with an outside-to-inside orientation of the N-terminal and C-terminal ends. SOAT mRNA is most highly expressed in testis. Relatively high SOAT expression was also detected in placenta and pancreas. We established a stable SOAT-HEK293 cell line that showed sodium-dependent transport of dehydroepiandrosterone sulfate, estrone-3-sulfate, and pregnenolone sulfate with apparent K(m) values of 28.7, 12.0, and 11.3 microm, respectively. Although bile acids, such as taurocholic acid, cholic acid, and chenodeoxycholic acid, were not substrates of SOAT, the sulfoconjugated bile acid taurolithocholic acid-3-sulfate was transported by SOAT-HEK293 cells in a sodium-dependent manner and showed competitive inhibition of SOAT transport with an apparent K(i) value of 0.24 mum. Several nonsteroidal organosulfates also strongly inhibited SOAT, including 1-(omega-sulfooxyethyl)pyrene, bromosulfophthalein, 2- and 4-sulfooxymethylpyrene, and alpha-naphthylsulfate. Among these inhibitors, 2- and 4-sulfooxymethylpyrene were competitive inhibitors of SOAT, with apparent K(i) values of 4.3 and 5.5 microm, respectively, and they were also transported by SOAT-HEK293 cells.
Geyer
Joachim
J
Institute of Pharmacology and Toxicology, Justus-Liebig-University of Giessen, Frankfurter Strasse 107, 35392 Giessen, Germany. joachim.m.geyer@vetmed.uni-giessen.de
Döring
Barbara
B
Meerkamp
Kerstin
K
Ugele
Bernhard
B
Bakhiya
Nadiya
N
Fernandes
Carla F
CF
Godoy
José R
JR
Glatt
Hansruedi
H
Petzinger
Ernst
E
D000595:Amino Acid Sequence; D001693:Biological Transport, Active / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D049990:Membrane Transport Modulators / Q000494:pharmacology; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000502:physiology; D027361:Organic Anion Transporters / Q000235:genetics / Q000378:metabolism*; D029363:Organic Anion Transporters, Sodium-Dependent / Q000235:genetics / Q000378:metabolism*; D010179:Pancreas / Q000378:metabolism; D010920:Placenta / Q000378:metabolism; D011247:Pregnancy; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D027981:Symporters / Q000235:genetics; D013737:Testis / Q000378:metabolism
D018076:DNA, Complementary; D049990:Membrane Transport Modulators; D027361:Organic Anion Transporters; D029363:Organic Anion Transporters, Sodium-Dependent; C522730:Slc10a6 protein, human; D027981:Symporters; C071908:sodium-bile acid cotransporter