Vasopressin increases S261 phosphorylation in AQP2-P262L, a mutant in recessive nephrogenic diabetes insipidus.
Vasopressin increases S261 phosphorylation in AQP2-P262L, a mutant in recessive nephrogenic diabetes insipidus.
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DOI：
10.1093/ndt/gfs292
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发表时间：
2012-12
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影响因子：
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通讯作者：
C. Trimpert;Dennis T M van den Berg;R. Fenton;E. Klussmann;P. Deen
中科院分区：
文献类型：
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作者：
C. Trimpert;Dennis T M van den Berg;R. Fenton;E. Klussmann;P. Deen
水通道蛋白-2（AQP 2）基因突变导致肾源性尿崩症（NDI），这是一种肾脏疾病，其特征是由于缺乏对加压素的抗利尿反应而导致的多尿。虽然大多数AQP 2突变体在隐性NDI是错误折叠，并保留在内质网，AQP 2-P262 L在NDI受损，其加压素依赖性转运从囊泡到质膜。方法加压素诱导的AQP 2易位与AQP 2 S256、S264和T269磷酸化和S261去磷酸化一致。由于P262位于S261附近，我们测试了改变的磷酸化是否是NDI中AQP-P262 L错配的基础。结果在极化细胞中，AQP 2-P262 L呈29/30 kDa双带，而wt-AQP 2仅呈29 kDa双带。磷酸酶处理显示30 kDa AQP 2-P262 L条带是由于磷酸化改变所致。新开发的磷酸特异性抗体的使用表明，毛喉素不仅增加pS256和pT 269，而且与wt-AQP 2相反，还增加AQP 2-P262 L中的pS261。然而，其中S261磷酸化被阻止（S261 A）的AQP 2-P262 L蛋白的表达仍然错误分选为囊泡/基底外侧膜，尽管缺乏30 kDa条带。总之，我们的数据表明，加压素诱导而不是减少AQP 2-P262 L中S261的磷酸化，但仍有待确定的是，改变的磷酸化是否会导致其在NDI中的错误分选。
BACKGROUND Mutations in the aquaporin-2 (AQP2) gene cause nephrogenic diabetes insipidus (NDI), a renal disorder characterized by polyuria due to a lacking antidiuretic response to vasopressin. While most AQP2 mutants in recessive NDI are misfolded and retained in the endoplasmic reticulum, AQP2-P262L in NDI was impaired in its vasopressin-dependent translocation from vesicles to the plasma membrane. METHODS Vasopressin-induced translocation of AQP2 coincides with AQP2 phosphorylation at S256, S264 and T269 and dephosphorylation at S261. Since P262 lies adjacent to S261, we tested whether a changed phosphorylation could underlie AQP-P262L missorting in NDI. RESULTS In polarized cells, AQP2-P262L expressed as a double 29/30 kDa band, whereas wt-AQP2 expressed only as a 29 kDa band. Phosphatase treatment revealed that the 30 kDa AQP2-P262L band was due to changed phosphorylation. The use of newly developed phospho-specific antibodies showed that forskolin not only increased pS256 and pT269, but, in contrast to wt-AQP2, also pS261 in AQP2-P262L. The expression of AQP2-P262L proteins in which S261 phosphorylation was prevented (S261A), however, was still missorted to vesicles/basolateral membrane, despite the absence of the 30 kDa band. CONCLUSIONS Together, our data reveal that vasopressin induces instead of reduces the phosphorylation of S261 in AQP2-P262L, but it remains to be established whether the changed phosphorylation causes its missorting in NDI.