Molecular cloning and characterization of the murine bile salt export pump
Molecular cloning and characterization of the murine bile salt export pump
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DOI：
10.1016/s0378-1119(99)00460-6
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发表时间：
2000-01-04
期刊：
影响因子：
3.5
通讯作者：
Ward, KL
中科院分区：
文献类型：
--
作者：
Green, RM;Hoda, F;Ward, KL
肝胆汁盐分泌和胆汁形成是哺乳动物肝脏的基本功能，肝细胞分泌胆汁盐的限速步骤是小管分泌。最近，大鼠P-糖蛋白/胆盐输出泵（spgp/BSEP）被证明编码大鼠ATP依赖性小管胆盐输出蛋白，人类BSEP突变被确定为PFIC 2的原因。由于小鼠模型对于肝细胞转运和代谢的研究至关重要，因此小鼠基因的克隆和表征是必不可少的。在这项研究中，我们已经克隆了一个全长，功能的cDNA的mBsep。推导的氨基酸序列编码一个1321个氨基酸的蛋白质，与大鼠的相似性为94%，与人的相似性为89%。使用针对mbsep蛋白的羧基末端肽的抗体的Western免疫印迹揭示了160 kDa的蛋白质，其高度富集于小鼠小管膜中。将mBSEP转染到Sf-9昆虫细胞或哺乳动物Balb-3 T3细胞中，赋予胆盐牛磺胆酸盐的功能性转运。mBsep mRNA在小鼠肝脏中表达，但在其他组织中不表达。肝脏mBsep水平出现高度调节，在胆汁淤积的LPS和雌激素模型中均显著降低。这些数据是重要的，为进一步研究小鼠肝细胞转运生理和代谢。(C)2000 Elsevier Science B. V.保留所有权利。
Hepatic bile salt secretion and bile formation are essential functions of the mammalian liver, and the rate-limiting step of hepatocellular secretion of bile salts is canalicular secretion. Recently, the rat sister-of-p-glycoprotein/bile salt export pump (spgp/BSEP) was demonstrated to encode for the rat ATP-dependent canalicular bile salt export protein, and mutations of human BSEP were identified as the cause of PFIC 2. Since mouse models are vital for studies in hepatocellular transport and metabolism, cloning and characterization of the murine gene are essential. In this study, we have cloned a full-length, functional cDNA for the mBsep. The deduced amino acid sequence encodes for a 1321-amino-acid protein and is 94% similar to rat and 89% similar to human bsep. Western immunoblotting using an antibody directed against a carboxy-terminal peptide of mbsep protein reveals a 160 kDa protein, which is highly enriched in mouse canalicular membranes. Transfection of mBSEP into Sf-9 insect cells or mammalian Balb-3T3 cells confers functional transport of the bile salt taurocholate. The mBsep mRNA is expressed in murine liver, but not in other tissues. Hepatic mBsep levels appear highly regulated, bring markedly diminished in both LPS and estrogen models of cholestasis. These data are important for further murine studies of hepatocellular transport physiology and metabolism. (C) 2000 Elsevier Science B.V. All rights reserved.