Transient Receptor Potential Melastatin 1 (TRPM1) Is an Ion-conducting Plasma Membrane Channel Inhibited by Zinc Ions
Transient Receptor Potential Melastatin 1 (TRPM1) Is an Ion-conducting Plasma Membrane Channel Inhibited by Zinc Ions
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DOI：
10.1074/jbc.m110.202945
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发表时间：
2011-04-08
影响因子：
4.8
通讯作者：
Oberwinkler, Johannes
中科院分区：
文献类型：
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作者：
Lambert, Sachar;Drews, Anna;Oberwinkler, Johannes
TRPM1是瞬时受体电位（TRP）蛋白美拉抑素亚群的创始成员，但TRPM1蛋白形成离子通道的机制尚未得到证实。因此，这些蛋白质的生物物理和药理学性质在很大程度上是未知的。在这里，我们表明TRPM1蛋白的异种表达诱导离子电导，可以通过细胞外类固醇应用激活。然而，观察到的电流振幅太小，无法进行可靠的生物物理表征。我们通过以几种独立的方式修改TRPM1通道来克服这一限制，从而增加了与密切相关的TRPM3通道的相似性。结果构建体在过表达后产生了相当大的电流。我们还证明，未经修饰的TRPM1和TRPM3蛋白形成功能性异多聚通道。通过这些方法，我们测量了二价渗透率曲线，发现与仅含有TRPM3孔的通道相比，含有TRPM1孔的通道在生理浓度下受到细胞外锌离子的抑制。将这些发现应用于胰腺β细胞，我们发现TRPM1蛋白在这些细胞的类固醇激活电流中不起主要作用。锌离子对TRPM1的抑制主要是由于7个氨基酸的短链只存在于TRPM1的孔区，而不存在于TRPM3。综上所述，我们的数据表明TRPM1蛋白是真正的离子传导质膜通道。它们独特的生物物理特性允许可靠地识别内源性trpm1介导的电流。
TRPM1 is the founding member of the melastatin subgroup of transient receptor potential (TRP) proteins, but it has not yet been firmly established that TRPM1 proteins form ion channels. Consequently, the biophysical and pharmacological properties of these proteins are largely unknown. Here we show that heterologous expression of TRPM1 proteins induces ionic conductances that can be activated by extracellular steroid application. However the current amplitudes observed were too small to enable a reliable biophysical characterization. We overcame this limitation by modifying TRPM1 channels in several independent ways that increased the similarity to the closely related TRPM3 channels. The resulting constructs produced considerably larger currents after overexpression. We also demonstrate that unmodified TRPM1 and TRPM3 proteins form functional heteromultimeric channels. With these approaches, we measured the divalent permeability profile and found that channels containing the pore of TRPM1 are inhibited by extracellular zinc ions at physiological concentrations, in contrast to channels containing only the pore of TRPM3. Applying these findings to pancreatic beta cells, we found that TRPM1 proteins do not play a major role in steroid-activated currents of these cells. The inhibition of TRPM1 by zinc ions is primarily due to a short stretch of seven amino acids present only in the pore region of TRPM1 but not of TRPM3. Combined, our data demonstrate that TRPM1 proteins are bona fide ion-conducting plasma membrane channels. Their distinct biophysical properties allow a reliable identification of endogenous TRPM1-mediated currents.