Institute for Physiology, Center of Integrative Physiology and Molecular Medicine, Saarland University, Homburg, Germany.
Institute for Physiology, Otto-von-Guericke University, Magdeburg, Germany.
Elife. 2020 May 11;9:e55152. doi: 10.7554/eLife.55152.
Vesicle fusion is mediated by assembly of SNARE proteins between opposing membranes. While previous work suggested an active role of SNARE transmembrane domains (TMDs) in promoting membrane merger (Dhara et al., 2016), the underlying mechanism remained elusive. Here, we show that naturally-occurring v-SNARE TMD variants differentially regulate fusion pore dynamics in mouse chromaffin cells, indicating TMD flexibility as a mechanistic determinant that facilitates transmitter release from differentially-sized vesicles. Membrane curvature-promoting phospholipids like lysophosphatidylcholine or oleic acid profoundly alter pore expansion and fully rescue the decelerated fusion kinetics of TMD-rigidifying VAMP2 mutants. Thus, v-SNARE TMDs and phospholipids cooperate in supporting membrane curvature at the fusion pore neck. Oppositely, slowing of pore kinetics by the SNARE-regulator complexin-2 withstands the curvature-driven speeding of fusion, indicating that pore evolution is tightly coupled to progressive SNARE complex formation. Collectively, TMD-mediated support of membrane curvature and SNARE force-generated membrane bending promote fusion pore formation and expansion.
囊泡融合是通过 opposing membranes 之间 SNARE 蛋白的组装来介导的。虽然之前的工作表明 SNARE 跨膜结构域 (TMD) 在促进膜融合中发挥积极作用 (Dhara 等人，2016)，但潜在的机制仍不清楚。在这里，我们表明天然存在的 v-SNARE TMD 变体差异调节小鼠嗜铬细胞中的融合孔动力学，表明 TMD 灵活性作为一种机械决定因素，促进来自不同大小囊泡的递质释放。像溶血磷脂酰胆碱或油酸这样的促进膜曲率的磷脂极大地改变了孔的扩展，并完全挽救了 TMD 僵化 VAMP2 突变体的融合动力学的减速。因此，v-SNARE TMD 和磷脂在支持融合孔颈部的膜曲率方面合作。相反，SNARE 调节剂 complexin-2 减缓孔动力学的速度可以抵抗曲率驱动的融合加速，表明孔的演变与 SNARE 复合物的渐进形成紧密耦合。总的来说，TMD 介导的对膜曲率的支持和 SNARE 力产生的膜弯曲促进了融合孔的形成和扩展。
