White K Ian, Khan Yousuf A, Qiu Kangqiang, Balaji Ashwin, Couoh-Cardel Sergio, Esquivies Luis, Pfuetzner Richard A, Diao Jiajie, Brunger Axel T
Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA, USA.
Department of Neurology and Neurological Sciences, Stanford University, Stanford, CA, USA.
Nat Commun. 2025 Sep 24;16(1):8371. doi: 10.1038/s41467-025-62764-0.
Synaptic vesicles containing neurotransmitters fuse with the plasma membrane upon the arrival of an action potential at the active zone. Multiple proteins organize trans-SNARE complex assembly and priming, leading to fusion. One target membrane SNARE, syntaxin, forms nanodomains at the active zone, and another, SNAP-25, enters non-fusogenic complexes with it. Here, we reveal mechanistic details of AAA+ protein NSF (N-ethylmaleimide sensitive factor) and SNAP (soluble NSF attachment protein) action before fusion. We show that syntaxin clusters are conserved, that NSF colocalizes with them, and characterize SNARE populations that may exist within or near them using cryo-EM. Supercomplexes of NSF, α-SNAP, and either a syntaxin tetramer or one of two binary complexes of syntaxin-SNAP-25 reveal atomic details of SNARE processing and show how sequential ATP hydrolysis drives disassembly. These results suggest a functional role for syntaxin clusters as reservoirs and a corresponding role for NSF in syntaxin liberation and SNARE protein quality control preceding fusion.
当动作电位到达活性区时，含有神经递质的突触小泡与质膜融合。多种蛋白质参与跨SNARE复合体的组装和引发，从而导致融合。一种靶膜SNARE蛋白—— syntaxin（突触融合蛋白）在活性区形成纳米结构域，另一种SNARE蛋白——SNAP-25（突触小体相关蛋白25）则与之形成非融合复合体。在此，我们揭示了AAA+蛋白NSF（N-乙基马来酰亚胺敏感因子）和SNAP（可溶性NSF附着蛋白）在融合前作用的机制细节。我们发现syntaxin簇是保守的，NSF与它们共定位，并使用冷冻电镜对可能存在于它们内部或附近的SNARE群体进行了表征。NSF、α-SNAP以及syntaxin四聚体或syntaxin-SNAP-25两种二元复合体之一的超复合体揭示了SNARE加工的原子细节，并展示了连续的ATP水解如何驱动解聚。这些结果表明syntaxin簇作为储存库的功能作用，以及NSF在融合前syntaxin释放和SNARE蛋白质量控制中的相应作用。
