Down-Regulation of the Na+-Coupled Phosphate Transporter NaPi-IIa by AMP-Activated Protein Kinase
Down-Regulation of the Na+-Coupled Phosphate Transporter NaPi-IIa by AMP-Activated Protein Kinase
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DOI：
10.1159/000355735
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发表时间：
2013-01-01
影响因子：
2.8
通讯作者：
Lang, Florian
中科院分区：
文献类型：
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作者：
Dermaku-Sopjani, Miribane;Almilaji, Ahmad;Lang, Florian
背景/目的：钠偶联磷酸盐转运蛋白NaPi-IIa是完成肾小管磷酸盐重吸收的主要载体。它是由跨顶细胞膜的电化学Na+梯度驱动的，该梯度通过Na+/K+ ATP酶跨基底外侧细胞膜的Na+挤出来维持。因此，NaPi-IIa的操作需要能量，以避免细胞Na+积累和K+损失，最终降低细胞膜电位，Cl-进入和细胞肿胀。在能量耗尽时，Na+偶联转运过程的早期抑制可以延迟细胞肿胀，从而促进细胞存活。能量消耗由AMP活化蛋白激酶（AMPK）感测，AMPK是一种丝氨酸/苏氨酸激酶，其刺激几种细胞机制，增加能量产生并限制能量利用。本研究探讨AMPK是否影响NAPi-IIa的活性。研究方法：将编码NAPi-IIa的cRNA注射到具有或不具有额外表达野生型AMPK（AMPK（α 1）-HA（+）AMPK（β 1）-Flag+AMPK（γ 1）-HA）、无活性AMPK（α 1 K45 R）（AMPK（α 1 K45 R）+AMPK（β 1）-Flag+AMPK（γ 1）-HA）或组成型活性AMPK（γ R70 Q）（AMPK（α 1）-HA+AMPK（β 1）-Flag+AMPK（γ 1 R70 Q））的爪蟾卵母细胞中。在双电极电压钳实验中从磷酸盐诱导的电流估计NaPi-IIa活性。结果如下：在NaPi-IIa表达，但不是在注水的非洲爪蟾卵母细胞，磷酸盐（1 mM）的细胞外浴溶液中添加产生的电流（I-P），这是显着降低野生型AMPK和AMPK（γ R70 Q），但不是AMPK（α K45 R）的共表达。在表达NaPi-IIa和AMPK的非洲爪蟾卵母细胞中磷酸盐诱导电流被AMPK抑制剂化合物C（20 μ M）显著增加。动力学分析显示AMPK显著降低最大转运速率。结论：AMP激活的蛋白激酶AMPK是NaPi-IIa的强大调节剂，从而调节肾小管磷酸盐转运。版权所有（C）2013 S. Karger AG，巴塞尔
Background/Aims: The Na+-coupled phosphate transporter NaPi-IIa is the main carrier accomplishing renal tubular phosphate reabsorption. It is driven by the electrochemical Na+ gradient across the apical cell membrane, which is maintained by Na+ extrusion across the basolateral cell membrane through the Na+/K+ ATPase. The operation of NaPi-IIa thus requires energy in order to avoid cellular Na+ accumulation and K+ loss with eventual decrease of cell membrane potential, Cl-entry and cell swelling. Upon energy depletion, early inhibition of Na+-coupled transport processes may delay cell swelling and thus foster cell survival. Energy depletion is sensed by the AMP-activated protein kinase (AMPK), a serine/threonine kinase stimulating several cellular mechanisms increasing energy production and limiting energy utilization. The present study explored whether AMPK influences the activity of NAPi-IIa. Methods: cRNA encoding NAPi-IIa was injected into Xenopus oocytes with or without additional expression of wildtype AMPK (AMPK(alpha 1)-HA(+)AMPK(beta 1)-Flag+AMPK(gamma 1)-HA), of inactive AMPK(alpha K45R) (AMPK(alpha 1K45R)+AMPK(beta 1)-Flag+AMPK(gamma 1)-HA) or of constitutively active AMPK(gamma R70Q) (AMPK(alpha 1)-HA+AMPK(beta 1)-Flag+AMPK(gamma 1R70Q)). NaPi-IIa activity was estimated from phosphate-induced current in dual electrode voltage clamp experiments. Results: In NaPi-IIa-expressing, but not in water-injected Xenopus oocytes, the addition of phosphate (1 mM) to the extracellular bath solution generated a current (I-p), which was significantly decreased by coexpression of wild-type AMPK and of AMPK(gamma R70Q) but not of AMPK(alpha K45R). The phosphate-induced current in NaPi-IIa- and AMPK-expressing Xenopus ooocytes was significantly increased by AMPK inhibitor Compound C (20 mu M). Kinetic analysis revealed that AMPK significantly decreased the maximal transport rate. Conclusion: The AMP-activated protein kinase AMPK is a powerful regulator of NaPi-IIa and thus of renal tubular phosphate transport. Copyright (C) 2013 S. Karger AG, Basel