Groupe d'étude des protéines membranaires, Université de Montréal, Succ. Centre-Ville, Montreal, Quebec, Canada.
J Membr Biol. 2010 Jul;236(2):225-32. doi: 10.1007/s00232-010-9295-9. Epub 2010 Aug 14.
Myo-inositol (MI; hexahydroxycyclohexane, C(6)H(6)O(12)) is a small neutral molecule used as a compatible osmolyte in the kidney medulla. At high concentrations, MI appears to act as a chemical chaperone and was shown to promote plasma membrane expression of the impaired cystic fibrosis chloride channel (Delta508-CFTR). In the present study, we measured whether MI could increase expression of two human aquaporin 2 (AQP2) mutants which were recently identified as causing nephrogenic diabetes insipidus (NDI). Both proteins (D150E and G196D) were expressed in Xenopus laevis oocytes, but only D150E displayed an increase in oocyte water permeability (P (f)). Adding 5 mM MI to the bathing solution for 24 h produced a 50% increase in the D150E-associated P (f), while it had no effect on noninjected oocytes or on oocytes expressing wt-AQP2 or G196D. Western blots performed on purified plasma membrane preparations confirmed that MI increased the amount of D150E present at the plasma membrane, while G196D was always undetectable. X. laevis oocytes are remarkably impermeable to MI, and the effect of MI on D150E expression does not require the presence of intracellular MI. The effect of external MI was dose-dependent (K (0.5) was 130 microM) and specific with respect to other forms of inositols. Further studies on a second group of AQP2 mutants causing NDI showed that K228E activity was similarly stimulated by MI, while V71M, A70D and S256L were not. It is concluded that physiological concentrations of extracellular MI can stimulate the expression of a specific subgroup of AQP2 mutants.
肌醇（MI；六羟基环己烷，C(6)H(6)O(12))是一种小的中性分子，用作肾髓质中的相容渗透剂。在高浓度下，MI 似乎作为一种化学伴侣，并且被证明可促进受损囊性纤维化氯离子通道（Delta508-CFTR）的质膜表达。在本研究中，我们测量了 MI 是否可以增加最近被鉴定为引起肾性尿崩症（NDI）的两种人水通道蛋白 2（AQP2）突变体的表达。两种蛋白质（D150E 和 G196D）都在非洲爪蟾卵母细胞中表达，但只有 D150E 显示卵母细胞水通透性（P（f））增加。将 5 mM MI 添加到浴液中 24 小时可使 D150E 相关的 P（f）增加 50%，而对未注射的卵母细胞或表达 wt-AQP2 或 G196D 的卵母细胞没有影响。在纯化的质膜制剂上进行的 Western blot 证实 MI 增加了质膜上 D150E 的含量，而 G196D 始终无法检测到。非洲爪蟾卵母细胞对 MI 非常不渗透，并且 MI 对 D150E 表达的作用不需要细胞内 MI 的存在。外部 MI 的作用是剂量依赖性的（K（0.5）为 130 microM），并且对其他形式的肌醇具有特异性。对引起 NDI 的另一组 AQP2 突变体的进一步研究表明，MI 同样刺激 K228E 的活性，而 V71M、A70D 和 S256L 则没有。结论是，细胞外 MI 的生理浓度可以刺激特定亚组 AQP2 突变体的表达。
