Tethering of synaptic vesicles (SVs) to the active zone determines synaptic strength, although the molecular basis governing SV tethering is elusive. Here, we discover that small unilamellar vesicles (SUVs) and SVs from rat brains coat on the surface of condensed liquid droplets formed by active zone proteins RIM, RIM-BP, and ELKS via phase separation. Remarkably, SUV-coated RIM/RIM-BP condensates are encapsulated by synapsin/SUV condensates, forming two distinct SUV pools reminiscent of the reserve and tethered SV pools that exist in presynaptic boutons. The SUV-coated RIM/RIM-BP condensates can further cluster Ca²⁺ channels anchored on membranes. Thus, we reconstitute a presynaptic bouton-like structure mimicking the SV-tethered active zone with its one side attached to the presynaptic membrane and the other side connected to the synapsin-clustered SV condensates. The distinct interaction modes between membraneless protein condensates and membrane-based organelles revealed here have general implications in cellular processes, including vesicular formation and trafficking, organelle biogenesis, and autophagy.

尽管控制SV系链的分子基础难以捉摸，但将突触小泡（SVs）拴系到活动区决定了突触强度。在这里，我们发现来自大鼠大脑的小的单层囊泡（SUVs）和SVs通过相分离作用覆盖了由活性区蛋白RIM，RIM-BP和ELKS形成的冷凝液滴表面。值得注意的是，SUV涂层的RIM / RIM-BP冷凝液被突触蛋白/ SUV冷凝液封装，形成两个不同的SUV池，使人联想到突触前按钮中存在的储备和系留SV池。SUV涂层的RIM / RIM-BP冷凝水可进一步使Ca ²⁺聚集通道锚定在膜上。因此，我们重建了一个突触前的bouton-like结构，模仿了SV束缚的活性区，其一侧连接到突触前膜上，另一侧连接到突触素簇上的SV冷凝物。本文揭示的无膜蛋白凝析物和基于膜的细胞器之间的独特相互作用模式在细胞过程中具有普遍意义，包括囊泡形成和运输，细胞器生物发生和自噬。