Molecular cloning of canalicular multispecific organic anion transporter defective in EHBR
| Autor: | Hiroshi Suzuki, Kazuhiko Kume, Kousei Ito, Takao Shimizu, Yuichi Sugiyama, Tomoko Hirohashi |
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| Rok vydání: | 1997 |
| Předmět: |
DNA
Complementary Transcription Genetic Physiology Sequence analysis Anion Transport Proteins Molecular Sequence Data Biology Molecular cloning Polymerase Chain Reaction Rats Mutant Strains Rats Sprague-Dawley Physiology (medical) Complementary DNA Animals Tissue Distribution Amino Acid Sequence Cloning Molecular Peptide sequence Gene Library Hyperbilirubinemia chemistry.chemical_classification Gene Rearrangement Hepatology Base Sequence cDNA library Gastroenterology Gene rearrangement Blotting Northern Molecular biology Amino acid Rats Open reading frame Biochemistry chemistry Liver Carrier Proteins |
| Zdroj: | The American journal of physiology. 272(1 Pt 1) |
| ISSN: | 0002-9513 |
| Popis: | Several organic anions are excreted into the bile via a canalicular multispecific organic anion transporter (cMOAT), which is hereditarily defective in mutant rats, such as the Eisai hyperbilirubinemic rat (EHBR) and TR- rat. In the present study, we cloned cMOAT from the Sprague-Dawley rat liver cDNA library based on the homology with human multidrug resistance-associated protein (hMRP). cMOAT was encoded by 4,623-base pair (bp) cDNA with a homology of 53.0 and 46.3% with hMRP at the cDNA and deduced amino acid level, respectively. The deduced amino acid sequence was the same as that cloned in Wistar rats (C. C. Paulusma, P. J. Bosma, G. J. Zaman, C. T. Bakker, M. Otter, G. L. Sceffer, P. Borst, and R. P. Oude Elferink. Science Wash. DC 271: 1126, 1996) except for four amino acid substitutions. By screening the library, three kinds of cDNA species for cMOAT with the same open reading frame and different 3'-untranslated region lengths (0.2, 1.5, and 3.5 kbp) were isolated. The Northern blot analysis of poly(A)+ RNA from the liver revealed that the expression of plural bands (approximately 5, 6, and 8 kb) was defective in EHBR, and this may be due to the presence of these cDNA species. Expression of cMOAT was observed almost exclusively in the liver and to a lesser extent in the duodenum, kidney, and jejunum. Reverse transcription-polymerase chain reaction (RT-PCR) and subsequent sequence analysis of EHBR liver, kidney, duodenum, and jejunum revealed that 1-bp replacement from G to A at nucleotide 2564 resulted in the introduction of the premature stop codon in all tissues examined. This mutation was different from that observed in TR (C. C. Paulusma, P. J. Bosma, G. J. Zaman, C. T. Bakker, M. Otter, G. L. Sceffer, P. Borst, and R. P. Oude Elferink. Science Wash. DC 271: 1126, 1996). Because EHBR and TR- are allelic mutants and both strains exhibit an autosomal recessive inheritance in the biliary excretion of organic anions it was concluded that the impaired expression of this particular protein is related to the pathogenesis of hyperbilirubinemia in the mutant animals. |
| Databáze: | OpenAIRE |
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