The Nociceptor Ion Channel TRPA1 Is Potentiated and Inactivated by Permeating Calcium Ions
The Nociceptor Ion Channel TRPA1 Is Potentiated and Inactivated by Permeating Calcium Ions
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DOI：
10.1074/jbc.m803568200
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发表时间：
2008-11-21
影响因子：
4.8
通讯作者：
Liman, Emily R.
中科院分区：
文献类型：
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作者：
Wang, Yuanyuan Y.;Chang, Rui B.;Liman, Emily R.
瞬时受体电位A1（TRPA 1）通道是环境刺激物和刺激性化学物质（如肉桂醛和芥子油）的分子靶点。细胞外Ca 2+是TRPA 1活性的关键调节因子，既增强又随后使其失活。在这份报告中，我们提供的证据表明，细胞外Ca 2+对这些过程的影响是间接的，可以完全归因于通过TRPA 1和随后的细胞内钙离子的升高进入。具体而言，我们发现，在孔突变体的TRPA 1，D918 A，其中Ca 2+渗透性大大降低，细胞外Ca 2+既不产生增强作用，也不失活。通过减少细胞内Ca 2+缓冲，这使得细胞内Ca 2+水平在通过D918 A通道进入时升高，这两个过程都得以恢复。将Ca 2+应用于切除的斑块的胞质面足以产生TRPA 1通道的增强和失活。此外，在全细胞记录中，通过1-（4，5-二甲氧基-2-硝基苯基）-EDTA-的UV释放而升高的细胞内Ca 2+增强TRPA 1电流。此外，我们的数据表明，增强和失活是独立的过程。TRPA 1电流可被Mg ~（2+）、Ba ~（2+）和Ca ~（2+）失活，但仅被Ba ~（2+）和Ca ~（2+）增强。饱和激活肉桂醛或芥子油闭塞增强，但不干扰失活。最后，这两个过程都没有受到一个假定的细胞内Ca 2+结合EF-手基序突变的影响。总之，我们进一步阐明了使用D918 A孔突变体的TRPA 1的增强和失活机制，这是研究通过TRPA 1的Ca 2+内流对伤害性信号传导的贡献的重要工具。
The transient receptor potential A1 (TRPA1) channel is the molecular target for environmental irritants and pungent chemicals, such as cinnamaldehyde and mustard oil. Extracellular Ca2+ is a key regulator of TRPA1 activity, both potentiating and subsequently inactivating it. In this report, we provide evidence that the effect of extracellular Ca2+ on these processes is indirect and can be entirely attributed to entry through TRPA1 and subsequent elevation of intracellular calcium. Specifically, we found that in a pore mutant of TRPA1, D918A, in which Ca2+ permeability was greatly reduced, extracellular Ca2+ produced neither potentiation nor inactivation. Both processes were restored by reducing intracellular Ca2+ buffering, which allowed intracellular Ca2+ levels to become elevated upon entry through D918A channels. Application of Ca2+ to the cytosolic face of excised patches was sufficient to produce both potentiation and inactivation of TRPA1 channels. Moreover, in whole cell recordings, elevation of intracellular Ca2+ by UV uncaging of 1-(4,5-dimethoxy-2-nitrophenyl)-EDTA-potentiated TRPA1 currents. In addition, our data show that potentiation and inactivation are independent processes. TRPA1 currents could be inactivated by Mg2+, Ba2+, and Ca2+ but potentiated only by Ba2+ and Ca2+. Saturating activation by cinnamaldehyde or mustard oil occluded potentiation but did not interfere with inactivation. Last, neither process was affected by mutation of a putative intracellular Ca2+-binding EF-hand motif. In conclusion, we have further clarified the mechanisms of potentiation and inactivation of TRPA1 using the D918A pore mutant, an important tool for investigating the contribution of Ca2+ influx through TRPA1 to nociceptive signaling.