Mammalian target of rapamycin complex 2 signaling pathway regulates transient receptor potential cation channel 6 in podocytes.
Mammalian target of rapamycin complex 2 signaling pathway regulates transient receptor potential cation channel 6 in podocytes.
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DOI：
10.1371/journal.pone.0112972
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Ding J
中科院分区：
文献类型：
--
作者：
Ding F;Zhang X;Li X;Zhang Y;Li B;Ding J
瞬时受体电位阳离子通道 6 (TRPC6) 是一种非选择性阳离子通道，TRPC6 的异常表达和功能获得参与遗传性和非遗传性肾脏疾病的发病机制。尽管这些疾病背后的分子机制仍知之甚少，但最近的研究表明许多信号通路参与调节 TRPC6。我们的目的是检查哺乳动物雷帕霉素靶点 (mTOR) 复合物（mTOR 复合物 1 [mTORC1] 或 mTOR 复合物 2 [mTORC2]）信号通路对足细胞中 TRPC6 的影响，足细胞是高度终末分化的肾上皮细胞，对于维持肾小球滤过屏障至关重要。我们应用 mTOR 的药理学抑制剂和针对 mTOR 成分的特异性 siRNA 来探索哪条 mTOR 信号通路参与足细胞中 TRPC6 的调节。将足细胞暴露于雷帕霉素（一种 mTORC1 抑制剂）和 ku0063794（一种 mTORC1 和 mTORC2 双重抑制剂）。此外，还采用了特异性 siRNA 介导的 mTORC1 组分 raptor 和 mTORC2 组分 rictor 敲低。分别通过实时定量PCR和Western blot检测TRPC6 mRNA和蛋白表达水平。此外，还进行荧光钙成像来评估足细胞中 TRPC6 的功能。雷帕霉素对 TRPC6 mRNA 或蛋白质表达水平或足细胞中 TRPC6 依赖性钙流入没有影响。然而，ku0063794 下调了 TRPC6 mRNA 和蛋白质水平，并抑制了足细胞中 TRPC6 依赖性钙流入。此外，raptor 的敲低并不影响 TRPC6 的表达或功能，而 rictor 的敲低则抑制了足细胞中 TRPC6 蛋白的表达和 TRPC6 依赖性钙内流。这些发现表明 mTORC2 信号通路调节足细胞中的 TRPC6，但 mTORC1 信号通路似乎对 TRPC6 没有影响。
Transient receptor potential cation channel 6 (TRPC6) is a nonselective cation channel, and abnormal expression and gain of function of TRPC6 are involved in the pathogenesis of hereditary and nonhereditary forms of renal disease. Although the molecular mechanisms underlying these diseases remain poorly understood, recent investigations revealed that many signaling pathways are involved in regulating TRPC6. We aimed to examine the effect of the mammalian target of rapamycin (mTOR) complex (mTOR complex 1 [mTORC1] or mTOR complex 2 [mTORC2]) signaling pathways on TRPC6 in podocytes, which are highly terminally differentiated renal epithelial cells that are critically required for the maintenance of the glomerular filtration barrier. We applied both pharmacological inhibitors of mTOR and specific siRNAs against mTOR components to explore which mTOR signaling pathway is involved in the regulation of TRPC6 in podocytes. The podocytes were exposed to rapamycin, an inhibitor of mTORC1, and ku0063794, a dual inhibitor of mTORC1 and mTORC2. In addition, specific siRNA-mediated knockdown of the mTORC1 component raptor and the mTORC2 component rictor was employed. The TRPC6 mRNA and protein expression levels were examined via real-time quantitative PCR and Western blot, respectively. Additionally, fluorescence calcium imaging was performed to evaluate the function of TRPC6 in podocytes. Rapamycin displayed no effect on the TRPC6 mRNA or protein expression levels or TRPC6-dependent calcium influx in podocytes. However, ku0063794 down-regulated the TRPC6 mRNA and protein levels and suppressed TRPC6-dependent calcium influx in podocytes. Furthermore, knockdown of raptor did not affect TRPC6 expression or function, whereas rictor knockdown suppressed TRPC6 protein expression and TRPC6-dependent calcium influx in podocytes. These findings indicate that the mTORC2 signaling pathway regulates TRPC6 in podocytes but that the mTORC1 signaling pathway does not appear to exert an effect on TRPC6.
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影响因子：
3.7
作者：
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通讯作者：
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DOI：
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