Topography and motion of acid-sensing ion channel intracellular domains.
Topography and motion of acid-sensing ion channel intracellular domains.
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DOI：
10.7554/elife.68955
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发表时间：
2021-07-22
期刊：
影响因子：
7.7
通讯作者：
Maclean DM
中科院分区：
文献类型：
--
作者：
Couch T;Berger KD;Kneisley DL;McCullock TW;Kammermeier P;Maclean DM
酸感应离子通道（asic）是由细胞外ph降低激活的三聚体阳离子选择性通道。ASIC1的细胞内N和C末端尾部影响缺血性细胞死亡中的通道门控、运输和信号传导。尽管ASIC1的细胞外和跨膜片段有几个x射线和低温电镜结构，但这些重要的细胞内尾巴仍未得到解决。在这里，我们描述了通过荧光共振能量转移（FRET）确定的鸡ASIC1细胞内结构域的粗略地形，使用荧光寿命成像或膜片钳荧光法测量。我们发现C端尾部向胞质中伸出约35 Å，并且来自相同亚基的N和C尾部比相邻亚基更接近。使用ph不敏感的荧光蛋白，我们没有检测到细胞外酸化时N和C尾部之间的任何相对运动，但确实观察到膜近端段向质膜的轴向运动。综上所述，我们的研究提供了ASIC细胞内结构域的粗略地形图，同时为细胞外酸化诱导的细胞内构象变化提供了方向性和背景。
Acid-sensing ion channels (ASICs) are trimeric cation-selective channels activated by decreases in extracellular pH. The intracellular N and C terminal tails of ASIC1 influence channel gating, trafficking, and signaling in ischemic cell death. Despite several X-ray and cryo-EM structures of the extracellular and transmembrane segments of ASIC1, these important intracellular tails remain unresolved. Here, we describe the coarse topography of the chicken ASIC1 intracellular domains determined by fluorescence resonance energy transfer (FRET), measured using either fluorescent lifetime imaging or patch clamp fluorometry. We find the C terminal tail projects into the cytosol by approximately 35 Å and that the N and C tails from the same subunits are closer than adjacent subunits. Using pH-insensitive fluorescent proteins, we fail to detect any relative movement between the N and C tails upon extracellular acidification but do observe axial motions of the membrane proximal segments toward the plasma membrane. Taken together, our study furnishes a coarse topographic map of the ASIC intracellular domains while providing directionality and context to intracellular conformational changes induced by extracellular acidification.