Analysis of the vectorial transport of amino acid and drugs in epithelial cells.

我们研究了参与氨基酸和药物在上皮细胞中载体转运的多种转运体及其膜转运的调控机制。确定了其根尖分选所需的BCRP区域。在肾脏和脉络丛(分别为OAT1和OAT3，以及PEPT2和OAT3)，研究了转运体对药物转运的贡献。对MRP3和BCRP的功能进行了表征，并推测它们参与了药物在小肠的转运。摄取(OATP2)和外排(MRP2)转运蛋白在极化细胞系(MDCK II)中共表达，作为有机阴离子肝胆转运的体外模型。在试管中。通过P-gp表达的单层的跨细胞转运可以用来预测PIGP对肠道吸收和脑吸收的影响程度。此外，使用双转染体(Oatp4/mrp2)，体外跨细胞转运可用于预测h-…。更多的有机阴离子在胆管上的清除。建立了Ntcp和Bsep双表达载体作为胆汁酸的肝胆转运模型，新克隆了7个氨基酸转运蛋白(ASC-2、AGT1、TAT1、LAT3、LAT4、CAT5和B^0AT1)，并对其功能进行了研究。克隆了肾脏有机阴离子的腔内转运蛋白OATv1、URAT1和OAT7，并对其功能进行了研究。B0AT1和URAT1的突变分别导致Hartnup障碍和肾脏低尿酸血症。RBAT和4F2hc的跨膜区是其与伴侣分子相互作用所必需的。BAT1 C末端的“VVPP”序列是其膜分离所必需的。以该序列为诱饵，用酵母双杂交技术分离到RACK1。RACK1是一个含有多个PDZ基序的支架蛋白，我们证实它与BAT1的VVPP序列相互作用。URAT1 C末端的PDZ相互作用基序与PDZK1相互作用。PDZ相互作用可能在转运蛋白聚集过程中发挥重要作用。较少

We investigated multiplicity of the transporters involved in vectorial transport of amino acids and drugs in epithelial cells as well as the regulatory mechanism of their membrane trafficking. The region of BCRP necessary for its apical sorting was identified. Contribution of transporters to the drug-transport was investigated in kidney and choroid plexus (OAT1 and OAT3, and PEPT2 andOAT3, respectively). Functional characterization of MRP3 and BCRP was performed, and their involvement in drug-transport in the small intestine has been suggested. Uptake (OATP2) and efflux (MRP2) transporters were co-expressed in a polarized cell line (MDCK II) as an in vitro model of hepatobiliary transport of organic anions. In vitro. transcellular transport across P-gp-expressing monolayers can be used to predict the extent to which intestinal absorption and brain uptake is affected by Pigp. Furthermore, using a double transfectant (Oatp4/Mrp2), in vitro transcellular transport can be used to predict h … More epatobiliary clearance of organic anions. Double-transfectants expressing Ntcp and Bsep was established as model of hepatobiliary transport of bile acids.Seven amino acid transporters (Asc-2, AGT1, TAT1, LAT3, LAT4, CAT5 and B^0AT1) were newly cloned, and their functional characterization was performed. As luminal transporter for organic anions in the kidney, OATv1, URAT1 and OAT7 were cloned, and their functional characterization was performed. Mutations in B0AT1 and URAT1 cause Hartnup disorder and renal hypouricemia, respectively. Transmembrane domain of rBAT and 4F2hc is necessary for its interaction with their partner molecules. The "VVPP" sequence at C-terminus of BAT1 is essential for its membrane sorting. RACK1 was isolated by yeast-two-hybrid using the sequence as bait. RACK1 is a scaffold-protein contains multiple PDZ motifs, and we confirmed that it interacts with "VVPP" sequence of BAT1. The PDZ interacting motif at the C-terminus of URAT1 interacts with PDZK1. PDZ interactions may play an important role in clustering transporters. Less