Deen P M, Croes H, van Aubel R A, Ginsel L A, van Os C H
Department of Cell Physiology, University of Nijmegen, The Netherlands.
J Clin Invest. 1995 May;95(5):2291-6. doi: 10.1172/JCI117920.
Congenital nephrogenic diabetes insipidus is a recessive hereditary disorder characterized by the inability of the kidney to concentrate urine in response to vasopressin. Recently, we reported mutations in the gene encoding the water channel of the collecting duct, aquaporin-2 (AQP-2) causing an autosomal recessive form of nephrogenic diabetes insipidus (NDI). Expression of these mutant AQP-2 proteins (Gly64Arg, Arg187Cys, Ser216Pro) in Xenopus oocytes revealed nonfunctional water channels. Here we report further studies into the inability of these missense AQP-2 proteins to facilitate water transport in Xenopus oocytes. cRNAs encoding the missense AQPs were translated with equal efficiency as cRNAs encoding wild-type AQP-2 and were equally stable. Arg187Cys AQP2 was more stable and Gly6-4Arg and Ser216Pro AQP2 were less stable when compared to wild-type AQP2 protein. On immunoblots, oocytes expressing missense AQP-2 showed, besides the wild-type 29 kDa band, an endoplasmic reticulum-retarded form of AQP-2 of approximately 32 kD. Immunoblots and immunocytochemistry demonstrated only intense labeling of the plasma membranes of oocytes expressing wild-type AQP-2. Therefore, we conclude that in Xenopus oocytes the inability of Gly64-Arg, Arg187Cys or Ser216Pro substituted AQP-2 proteins to facilitate water transport is caused by an impaired routing to the plasma membrane.
先天性肾性尿崩症是一种隐性遗传性疾病，其特征是肾脏无法对血管加压素作出反应而浓缩尿液。最近，我们报道了编码集合管水通道蛋白水通道蛋白-2（AQP-2）的基因突变，该突变导致了常染色体隐性形式的肾性尿崩症（NDI）。这些突变的AQP-2蛋白（Gly64Arg、Arg187Cys、Ser216Pro）在非洲爪蟾卵母细胞中的表达显示出无功能的水通道。在此，我们报告了对这些错义AQP-2蛋白在非洲爪蟾卵母细胞中无法促进水运输的进一步研究。编码错义AQP的cRNA与编码野生型AQP-2的cRNA翻译效率相同，且同样稳定。与野生型AQP2蛋白相比，Arg187Cys AQP2更稳定，而Gly6-4Arg和Ser216Pro AQP2则较不稳定。在免疫印迹上，表达错义AQP-2的卵母细胞除了显示出野生型的29 kDa条带外，还显示出一条约32 kD的内质网滞留形式的AQP-2。免疫印迹和免疫细胞化学仅显示表达野生型AQP-2的卵母细胞质膜有强烈标记。因此，我们得出结论，在非洲爪蟾卵母细胞中，Gly64-Arg、Arg187Cys或Ser216Pro取代的AQP-2蛋白无法促进水运输是由于其向质膜的转运受损所致。
