Exocytosis is a crucial cellular process involved in the release of neural transmitters or signaling hormones, and disposal of waste or toxic materials. The relationship between structural transition and temporal progression of this process is poorly understood, partly due to lack of adequate tools to resolve such dynamic structures at sufficient resolution in 3D. Exocytosis can be hijacked by some viruses, exemplified by the widely used model α-herpesvirus pseudorabies virus (PRV), which relies on exocytosis for trans-synaptic spread across neurons. Here, we have used cryo electron tomography (cryoET) to capture 199 events of PRV exocytosis from cultured hippocampal neurons. We established cumulative frequency analysis to estimate the relative duration of an exocytosis stage based on the frequency of observed viral particles at that stage. This analysis revealed that PRV exocytosis is biphasic, including a fast, "release phase" driven by fusion proteins and fused membranes, and a slow, "recovery phase" driven by flattening of curved membranes. The biphasic property of exocytosis discovered here appears to be conserved for membrane fusion during viral entry, and our approach of cumulative frequency analysis should have general utility for characterizing other membrane fusion events.

中文翻译：

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海马神经元疱疹病毒的双相胞吐作用及其对膜融合的机制意义。

胞吐作用是一个重要的细胞过程，涉及神经递质或信号激素的释放以及废物或有毒物质的处理。人们对这一过程的结构转变和时间进程之间的关系知之甚少，部分原因是缺乏足够的工具来以足够的 3D 分辨率解析此类动态结构。胞吐作用可能被一些病毒劫持，例如广泛使用的α-疱疹病毒模型伪狂犬病病毒（PRV），它依赖胞吐作用在神经元之间进行跨突触传播。在这里，我们使用冷冻电子断层扫描 (cryoET) 从培养的海马神经元中捕获了 199 个 PRV 胞吐作用事件。我们建立了累积频率分析，根据该阶段观察到的病毒颗粒的频率来估计胞吐阶段的相对持续时间。该分析表明，PRV 胞吐作用是双相的，包括由融合蛋白和融合膜驱动的快速“释放阶段”，以及由弯曲膜变平驱动的缓慢“恢复阶段”。这里发现的胞吐作用的双相特性似乎对于病毒进入期间的膜融合是保守的，并且我们的累积频率分析方法对于表征其他膜融合事件应该具有通用性。