Wielpütz Mark O, Lee Il-Ha, Dinudom Anuwat, Boulkroun Sheerazed, Farman Nicolette, Cook David I, Korbmacher Christoph, Rauh Robert
Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstrasse 6, Erlangen 91054, Germany.
J Biol Chem. 2007 Sep 21;282(38):28264-73. doi: 10.1074/jbc.M702168200. Epub 2007 Jul 25.
Regulation of the epithelial sodium channel (ENaC) is highly complex and may involve several aldosterone-induced regulatory proteins. The N-Myc downstream-regulated gene 2 (NDRG2) has been identified as an early aldosterone-induced gene. Therefore, we hypothesized that NDRG2 may affect ENaC function. To test this hypothesis we measured the amiloride-sensitive (2 microm) whole cell current (DeltaI(ami)) in Xenopus laevis oocytes expressing ENaC alone or co-expressing ENaC and NDRG2. Co-expression of NDRG2 significantly increased DeltaI(ami) in some, but not, all batches of oocytes tested. An inhibitory effect of NDRG2 was never observed. Using a chemiluminescence assay we demonstrated that the NDRG2-induced increase in ENaC currents was accompanied by a similar increase in channel surface expression. The stimulatory effect of NDRG2 was preserved in oocytes maintained in a low sodium bath solution to prevent sodium feedback inhibition. These findings suggest that the stimulatory effect of NDRG2 is independent of sodium feedback regulation. Furthermore, the stimulatory effect of NDRG2 on ENaC was at least in part additive to that of Sgk1. A short isoform of NDRG2 also stimulated DeltaI(ami). Overexpression of NDRG2 and ENaC in Fisher rat thyroid cells confirmed the stimulatory effect of NDRG2 on ENaC-mediated short-circuit current (I(SC-ami)). In addition, small interference RNA against NDRG2 largely reduced I(SC-ami) in Fisher rat thyroid cells. Our results indicate that NDRG2 is a likely candidate to contribute to aldosterone-mediated ENaC regulation.
上皮钠通道(ENaC)的调节高度复杂,可能涉及多种醛固酮诱导的调节蛋白。N-Myc下游调节基因2(NDRG2)已被确定为一种早期醛固酮诱导基因。因此,我们推测NDRG2可能影响ENaC功能。为了验证这一假设,我们测量了单独表达ENaC或共表达ENaC和NDRG2的非洲爪蟾卵母细胞中的阿米洛利敏感(2微摩尔)全细胞电流(ΔI(ami))。在部分(而非全部)测试批次的卵母细胞中,NDRG2的共表达显著增加了ΔI(ami)。从未观察到NDRG2的抑制作用。使用化学发光测定法,我们证明NDRG2诱导的ENaC电流增加伴随着通道表面表达的类似增加。在维持于低钠浴溶液中的卵母细胞中,NDRG2的刺激作用得以保留,以防止钠反馈抑制。这些发现表明,NDRG2的刺激作用独立于钠反馈调节。此外,NDRG2对ENaC的刺激作用至少部分与Sgk1的作用相加。NDRG2的短异构体也刺激了ΔI(ami)。在Fisher大鼠甲状腺细胞中过表达NDRG2和ENaC证实了NDRG2对ENaC介导的短路电流(I(SC-ami))的刺激作用。此外,针对NDRG2的小干扰RNA在很大程度上降低了Fisher大鼠甲状腺细胞中的I(SC-ami)。我们的结果表明,NDRG2可能是参与醛固酮介导的ENaC调节的一个候选因素。