Gao Tianyu, Li Wang, Shao Shuai, Zhang Zhengyao, Li Na, Zhang Hangyu, Liu Bo
Cancer Hospital of Dalian University of Technology, Shenyang, China.
Faculty of Medicine, Liaoning Key Lab of Integrated Circuit and Biomedical Electronic System, Dalian University of Technology, Dalian, China.
Commun Biol. 2025 Sep 25;8(1):1362. doi: 10.1038/s42003-025-08747-9.
"The last mile" of neuronal vesicles, from being tethered by the active zone filaments to docking at the presynaptic membrane, remains unclear, which limits the deep understanding of synaptic transmission and related physiological changes. Here, we develop two molecular biosensors (BKTS and RKTS) based on fluorescence resonance energy transfer technology according to the structure of RIM-BP2. By detecting the spatial distance between the two ends of the RIM-BP2 and the presynaptic membrane separately, the spatial posture changes in RIM-BP2 are reflected to explore how vesicles are transported to the presynaptic membrane for fusion. In the process of vesicle release, RIM-BP2 in primary cortical neurons and SH-SY5Y cells rotates like a "crane" with amino terminal deviating from the presynaptic membrane while the carboxyl terminal becomes closer. Furthermore, disturbing the microfilament or enhancing cell membrane fluidity inhibits the rotation of RIM-BP2. Through mutating RIM-BP2, we find that actin filaments provide mechanical stress through RIM-BP2 amino terminal, thereby regulating vesicle transport and release. Our work identifies a purely mechanical pathway of vesicle transport, in which microfilaments power the RIM-BP2 to drag vesicles to the presynaptic membrane as a "crane" for further release.
神经元囊泡从被活性区细丝拴系到停靠在突触前膜的“最后一英里”仍不清楚，这限制了对突触传递及相关生理变化的深入理解。在此，我们根据RIM-BP2的结构，利用荧光共振能量转移技术开发了两种分子生物传感器（BKTS和RKTS）。通过分别检测RIM-BP2两端与突触前膜之间的空间距离，反映RIM-BP2的空间姿态变化，以探究囊泡如何被运输到突触前膜进行融合。在囊泡释放过程中，原代皮层神经元和SH-SY5Y细胞中的RIM-BP2像“起重机”一样旋转，其氨基末端偏离突触前膜，而羧基末端则变得更近。此外，干扰微丝或增强细胞膜流动性会抑制RIM-BP2的旋转。通过对RIM-BP2进行突变，我们发现肌动蛋白丝通过RIM-BP2氨基末端提供机械应力，从而调节囊泡运输和释放。我们的工作确定了一种纯粹的囊泡运输机械途径，其中微丝驱动RIM-BP2像“起重机”一样将囊泡拖到突触前膜以便进一步释放。
