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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Role of PDZK1 in membrane expression of renal brush border ion exchangers
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Role of PDZK1 in membrane expression of renal brush border ion exchangers

机译：PDZK1在肾刷边界离子交换剂膜表达中的作用

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摘要

Na-H exchanger NHE3 and Cl-anion exchanger CFEX (SLC26A6, PAT1) play principal roles in the reabsorption of Na and Cl in the proximal tubule of the mammalian kidney. The mechanisms by which NHE3 and CFEX are localized to and maintained in the brush border of the proximal tubule are largely unknown. To investigate the possible interaction of NHE3 and CFEX with the PDZ-domain-containing scaffolding protein PDZK1, we performed a series of in vitro interaction assays with GST-fusion proteins and native brush border membrane proteins. These studies demonstrated that, not only were NHE3 and CFEX capable of directly interacting with PDZK1, but that this interaction was mediated through their C-terminal PDZ-interaction sites. To determine whether PDZK1 interaction is essential for brush border localization of NHE3 and CFEX in vivo, we examined the expression of NHE3 and CFEX in kidneys of wild-type and PDZK1 -null mutant mice by both Western analysis and immunocytochemistry. These studies indicated that, although brush border expression of NHE3 was unaffected by the loss of PDZK1, the expression of CFEX was markedly reduced. Finally, we assayed CFEX functional activity as Cl-oxalate exchange in brush border membrane vesicles and oxalate-stimulated volume absorption in microperfused proximal tubules. Consistent with the observed decrease in CFEX protein expression, both measures of CFEX functional activity were dramatically reduced in PDZK1 -null animals. In conclusion, the scaffolding protein PDZK1 is essential for the normal expression and function of Cl-anion exchanger CFEX in the proximal tubule of the mammalian kidney.
机译：Na-H交换器NHE3和Cl-阴离子交换器CFEX（SLC26A6，PAT1）在哺乳动物肾脏近端小管对Na和Cl的重吸收中起主要作用。 NHE3和CFEX定位并保持在近端小管的刷状边界中的机制在很大程度上尚不清楚。为了研究NHE3和CFEX与包含PDZ域的支架蛋白PDZK1的可能相互作用，我们使用GST融合蛋白和天然刷状缘膜蛋白进行了一系列体外相互作用测定。这些研究表明，不仅NHE3和CFEX能够直接与PDZK1相互作用，而且这种相互作用是通过其C端PDZ相互作用位点介导的。为了确定PDZK1相互作用对于体内NHE3和CFEX的笔刷边界定位是否必不可少，我们通过Western分析和免疫细胞化学检查了野生型和PDZK1-null突变小鼠肾脏中NHE3和CFEX的表达。这些研究表明，尽管NHE3的刷状边界表达不受PDZK1丢失的影响，但CFEX的表达明显降低。最后，我们测定了CFEX的功能活性，如刷状边界膜囊泡中的Cl-草酸盐交换以及微灌流的近端小管中草酸盐刺激的体积吸收。与观察到的CFEX蛋白表达的减少相一致，在PDZK1缺失的动物中，两种CFEX功能活性的测定均显着降低。总之，支架蛋白PDZK1对于C1阴离子交换剂CFEX在哺乳动物肾脏近端小管中的正常表达和功能至关重要。

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