Laboratory of Molecular Biology, Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto, Japan.
Cell Calcium. 2012 Feb;51(2):179-85. doi: 10.1016/j.ceca.2011.12.011. Epub 2012 Jan 4.
The transient receptor potential M2 channel (TRPM2) is the Ca(2+)-permeable cation channel controlled by cellular redox status via β-NAD(+) and ADP-ribose (ADPR). TRPM2 activity has been reported to underlie susceptibility to cell death and biological processes such as inflammatory cell migration and insulin secretion. However, little is known about the intracellular mechanisms that regulate oxidative stress-induced cell death via TRPM2. We report here a molecular and functional interaction between the TRPM2 channel and EF-hand motif-containing protein EFHC1, whose mutation causes juvenile myoclonic epilepsy (JME) via mechanisms including neuronal apoptosis. In situ hybridization analysis demonstrates TRPM2 and EFHC1 are coexpressed in hippocampal neurons and ventricle cells, while immunoprecipitation analysis demonstrates physical interaction of the N- and C-terminal cytoplasmic regions of TRPM2 with the EFHC1 protein. Coexpression of EFHC1 significantly potentiates hydrogen peroxide (H(2)O(2))- and ADPR-induced Ca(2+) responses and cationic currents via recombinant TRPM2 in HEK293 cells. Furthermore, EFHC1 enhances TRPM2-conferred susceptibility of HEK293 cells to H(2)O(2)-induced cell death, which is reversed by JME mutations. These results reveal a positive regulatory action of EFHC1 on TRPM2 activity, suggesting that TRPM2 contributes to the expression of JME phenotypes by mediating disruptive effects of JME mutations of EFHC1 on biological processes including cell death.
瞬时受体电位 M2 通道（TRPM2）是一种钙离子通透性阳离子通道，其活性受细胞氧化还原状态调控，该调控通过 β-NAD（+）和 ADP-核糖（ADPR）实现。TRPM2 活性与细胞死亡易感性以及炎症细胞迁移和胰岛素分泌等生物学过程有关。然而，目前对于通过 TRPM2 调节氧化应激诱导的细胞死亡的细胞内机制知之甚少。我们在此报告了 TRPM2 通道与 EF 手模蛋白 EFHC1 之间的分子和功能相互作用，其突变通过包括神经元凋亡在内的机制导致青少年肌阵挛性癫痫（JME）。原位杂交分析表明 TRPM2 和 EFHC1 在海马神经元和脑室细胞中共表达，而免疫沉淀分析表明 TRPM2 的 N 和 C 末端细胞质区域与 EFHC1 蛋白发生物理相互作用。EFHC1 的共表达显著增强了重组 TRPM2 在 HEK293 细胞中对过氧化氢（H2O2）和 ADPR 诱导的 Ca2+反应和阳离子电流的作用。此外，EFHC1 增强了 TRPM2 介导的 HEK293 细胞对 H2O2 诱导的细胞死亡的敏感性，而 JME 突变则逆转了这种敏感性。这些结果揭示了 EFHC1 对 TRPM2 活性的正向调节作用，表明 TRPM2 通过介导 EFHC1 的 JME 突变对包括细胞死亡在内的生物学过程的破坏作用，参与了 JME 表型的表达。
