质子活化氯化物(PAC)通道,在组织中普遍表达,调节细胞内Cl-水平和细胞死亡后酸中毒。然而,参与PAC通道调节的分子机制和信号通路在很大程度上是未知的。在这里,我们确定质膜内小叶的磷脂酰肌醇4,5-双磷酸[PI(4,5)P2]对于PAC通道的质子活化至关重要。通过激活磷脂酰肌醇5-磷酸酶或Gq蛋白偶联的毒蕈碱受体来消耗PI(4,5)P2基本上抑制人PAC电流。在切除的内外补丁中,向细胞质侧施加PI(4,5)P2增加电流。结构模拟表明,假定的PI(4,5)P2结合位点在静息状态下定位于胞质溶胶内,但在激活状态下移向细胞膜的内表面,并与内小叶PI(4,5)P2相互作用。跨膜螺旋2的膜-胞质溶胶界面附近的碱性残基的丙氨酸中和作用显着减弱了PAC电流。总的来说,我们的研究揭示了PAC通道通过内膜PI(4,5)P2的调节机制。
Proton-activated chloride (PAC) channels, ubiquitously expressed in tissues, regulate intracellular Cl- levels and cell death following acidosis. However, molecular mechanisms and signaling pathways involved in PAC channel modulation are largely unknown. Herein, we determine that phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] of the plasma membrane inner leaflet is essential for the proton activation of PAC channels. PI(4,5)P2 depletion by activating phosphatidylinositol 5-phosphatases or Gq protein-coupled muscarinic receptors substantially inhibits human PAC currents. In excised inside-out patches, PI(4,5)P2 application to the cytoplasmic side increases the currents. Structural simulation reveals that the putative PI(4,5)P2-binding site is localized within the cytosol in resting state but shifts to the cell membrane\'s inner surface in an activated state and interacts with inner leaflet PI(4,5)P2. Alanine neutralization of basic residues near the membrane-cytosol interface of the transmembrane helice 2 significantly attenuates PAC currents. Overall, our study uncovers a modulatory mechanism of PAC channel through inner membrane PI(4,5)P2.