Vlachová Viktorie, Teisinger Jan, Susánková Klára, Lyfenko Alla, Ettrich Rüdiger, Vyklický Ladislav
Institute of Physiology, Academy of Sciences, 142 20 Prague 4, Czech Republic.
J Neurosci. 2003 Feb 15;23(4):1340-50. doi: 10.1523/JNEUROSCI.23-04-01340.2003.
The vanilloid receptor [transient receptor potential (TRP)V1, also known as VR1] is a member of the TRP channel family. These receptors share a significant sequence homology, a similar predicted structure with six transmembrane-spanning domains (S1-S6), a pore-forming region between S5 and S6, and the cytoplasmically oriented C- and N-terminal regions. Although structural/functional studies have identified some of the key amino acids influencing the gating of the TRPV1 ion channel, the possible contributions of terminal regions to vanilloid receptor function remain elusive. In the present study, C-terminal truncations of rat TRPV1 have been constructed to characterize the contribution of the cytoplasmic C-terminal region to TRPV1 function and to delineate the minimum amount of C tail necessary to form a functional channel. The truncation of 31 residues was sufficient to induce changes in functional properties of TRPV1 channel. More pronounced effects of C-terminal truncation were seen in mutants lacking the final 72 aa. These changes were characterized by a decline of capsaicin-, pH-, and heat-sensitivity; progressive reduction of the activation thermal threshold (from 41.5 to 28.6 degrees C); and slowing of the activation rate of heat-evoked membrane currents (Q10 from 25.6 to 4.7). The voltage-induced currents of the truncated mutants exhibited a slower onset, markedly reduced outward rectification, and significantly smaller peak tail current amplitudes. Truncation of the entire TRPV1 C-terminal domain (155 residues) resulted in a nonfunctional channel. These results indicate that the cytoplasmic COOH-terminal domain strongly influences the TRPV1 channel activity, and that the distal half of this structural domain confers specific thermal sensitivity.
香草酸受体[瞬时受体电位（TRP）V1，也称为VR1]是TRP通道家族的成员。这些受体具有显著的序列同源性、类似的预测结构，包括六个跨膜结构域（S1-S6）、S5和S6之间的孔形成区域以及面向细胞质的C端和N端区域。尽管结构/功能研究已经确定了一些影响TRPV1离子通道门控的关键氨基酸，但末端区域对香草酸受体功能的可能贡献仍不清楚。在本研究中，构建了大鼠TRPV1的C端截短体，以表征细胞质C端区域对TRPV1功能的贡献，并确定形成功能性通道所需的最小C尾长度。截短31个残基足以诱导TRPV1通道功能特性的变化。在缺失最后72个氨基酸的突变体中，C端截短的影响更为明显。这些变化的特征是辣椒素、pH和热敏感性下降；激活热阈值逐渐降低（从41.5摄氏度降至28.6摄氏度）；热诱发膜电流的激活速率减慢（Q10从25.6降至4.7）。截短突变体的电压诱导电流起始较慢，外向整流明显降低，峰值尾电流幅度显著减小。截短整个TRPV1 C端结构域（155个残基）导致通道无功能。这些结果表明，细胞质COOH端结构域强烈影响TRPV1通道活性，并且该结构域的远端一半赋予特定的热敏感性。
