Kamsteeg Erik-Jan, Stoffels Monique, Tamma Grazia, Konings Irene B M, Deen Peter M T
Department of Physiology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.
Hum Mutat. 2009 Oct;30(10):1387-96. doi: 10.1002/humu.21068.
Regulation of body water homeostasis occurs by the vasopressin-dependent sorting of aquaporin-2 (AQP2) water channels to and from the apical membrane of renal principal cells. Mutations in AQP2 cause autosomal nephrogenic diabetes insipidus (NDI), a disease that renders the kidney unresponsive to vasopressin, resulting in polyuria and polydipsia. The AQP2 mutant c.772G>A; p.Glu258Lys (AQP2-E258K) causes dominant NDI by oligomerizing with wild-type AQP2 and missorting of this AQP2 complex to multivesicular bodies instead of the apical membrane. The motif causing this missorting of AQP2-E258K was identified here. Functional analyses and plasma membrane expression studies of truncation mutants in oocytes revealed that AQP2-E258K shortened to Leu259 is still intracellular retained. Alanine scanning and glutamic acid to arginine exchanges revealed increased function and plasma membrane expression for AQP2-E258K mutants with the following additional changes: Leu259Ala, Arg252Glu, Arg253Glu, or Arg252Ala-Arg254Ala, or for the AQP2 mutant p.Glu258Ala, indicating that the motif RRRxxxK(258)L confers AQP2-E258K retention. Fusion of this motif to aquaporin-1 also resulted in missorting of that water channel, indicating that this retention motif is transferable. In conclusion, our data reveal that the RRRxxxKL motif and repulsion between K258 and the arginine-triplet within this motif are the primary cause of missorting of AQP2-E258K in NDI.
机体水稳态的调节是通过血管加压素依赖的水通道蛋白2（AQP2）水通道在肾主细胞顶端膜的往返分选来实现的。AQP2突变会导致常染色体显性遗传性肾性尿崩症（NDI），这种疾病会使肾脏对血管加压素无反应，从而导致多尿和烦渴。AQP2突变体c.772G>A；p.Glu258Lys（AQP2-E258K）通过与野生型AQP2寡聚化，并将该AQP2复合物错误分选至多囊泡体而非顶端膜，从而导致显性NDI。本文确定了导致AQP2-E258K发生这种错误分选的基序。对卵母细胞中截短突变体的功能分析和质膜表达研究表明，截短至Leu259的AQP2-E258K仍保留在细胞内。丙氨酸扫描以及谷氨酸到精氨酸的交换显示，具有以下额外变化的AQP2-E258K突变体的功能和质膜表达有所增加：Leu259Ala、Arg252Glu、Arg253Glu或Arg252Ala-Arg254Ala，或者对于AQP2突变体p.Glu258Ala，这表明基序RRRxxxK(258)L导致AQP2-E258K滞留。将该基序与水通道蛋白-1融合也会导致该水通道的错误分选，这表明这种滞留基序是可转移的。总之，我们的数据表明，RRRxxxKL基序以及该基序内K258与精氨酸三联体之间的排斥作用是NDI中AQP2-E258K错误分选的主要原因。
