Requirement for the N-terminal coiled-coil domain for expression and function, but not subunit interaction of, the ADPR-activated TRPM2 channel.
Requirement for the N-terminal coiled-coil domain for expression and function, but not subunit interaction of, the ADPR-activated TRPM2 channel.
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DOI：
10.1007/s00232-009-9190-4
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发表时间：
2009-07
影响因子：
2.4
通讯作者：
Jiang, Lin-Hua
中科院分区：
文献类型：
--
作者：
Mei, Zhu-Zhong;Jiang, Lin-Hua
瞬时受体潜能Melastatin 2(TRPM2)蛋白形成多亚基复合体，最有可能的是同源异构体，它们作为细胞内ADP-核糖(ADPR)和氧化应激激活的钙离子渗透的非选择性阳离子通道发挥作用。每个TRPM2通道亚基被预测含有两个螺旋线圈(CC)结构域，一个在N-末端，另一个在C-末端。我们最近的研究表明，C-末端CC结构域在TRPM2通道组装中起着重要但不是排他性的作用。本研究旨在研究N-末端CC结构域的潜在作用。结构域的缺失显著降低了蛋白质的表达，并消除了ADPR诱发的电流，但不改变亚基的相互作用。两个CC结构域的缺失强烈减弱了亚基的相互作用，证实了C-末端CC结构域在亚基相互作用中起着关键作用。将谷氨酰胺替换为七肽重复序列中a和d位置的单个疏水残基来破坏CC的形成，对蛋白质表达、亚基相互作用或ADPR诱发的电流没有影响。Ile658突变为谷氨酰胺，不干扰CC的形成，减少了ADPR诱发的电流，而不影响蛋白表达、亚单位相互作用或膜运输。这些结果共同表明，TRPM2通道的N-末端CC结构域对蛋白质表达和功能是必需的，但不是亚单位相互作用。
Transient receptor potential melastatin 2 (TRPM2) proteins form multiple-subunit complexes, most likely homotetramers, which operate as Ca2+-permeable, nonselective cation channels activated by intracellular ADP-ribose (ADPR) and oxidative stress. Each TRPM2 channel subunit is predicted to contain two coiled-coil (CC) domains, one in the N-terminus and the other in the C-terminus. Our recent study has shown that the C-terminal CC domain plays an important, but not exclusive, role in the TRPM2 channel assembly. This study aimed to examine the potential role of the N-terminal CC domain. Domain deletion dramatically reduced protein expression and abolished ADPR-evoked currents but did not alter the subunit interaction. Deletion of both CC domains strongly attenuated the subunit interaction, confirming that the C-terminal CC domain is critical in the subunit interaction. Glutamine substitutions into individual hydrophobic residues at positions a and d in the heptad repeats to disrupt the CC formation had no effect on protein expression, subunit interaction, or ADPR-evoked currents. Mutation of Ile658 to glutamine, which did not perturb the CC formation, decreased ADPR-evoked currents without affecting protein expression, subunit interaction, or membrane trafficking. These results collectively suggest the requirement for the N-terminal CC domain for protein expression and function, but not subunit interaction, of the TRPM2 channel.
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