Accurate maintenance of organelle identity in the secretory pathway relies on retention and retrieval of resident proteins. In the endoplasmic reticulum (ER), secretory proteins are packaged into COPII vesicles that largely exclude ER residents and misfolded proteins by mechanisms that remain unresolved. Here we combined biochemistry and genetics with correlative light and electron microscopy (CLEM) to explore how selectivity is achieved. Our data suggest that vesicle occupancy contributes to ER retention: in the absence of abundant cargo, nonspecific bulk flow increases. We demonstrate that ER leakage is influenced by vesicle size and cargo occupancy: overexpressing an inert cargo protein or reducing vesicle size restores sorting stringency. We propose that cargo recruitment into vesicles creates a crowded lumen that drives selectivity. Retention of ER residents thus derives in part from the biophysical process of cargo enrichment into a constrained spherical membrane-bound carrier.

中文翻译：

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货物拥挤有助于 COPII 囊泡的分选严格性

分泌途径中细胞器特性的准确维持依赖于驻留蛋白的保留和检索。在内质网 (ER) 中，分泌蛋白被包装到 COPII 囊泡中，该囊泡很大程度上排除了 ER 驻留蛋白和错误折叠蛋白，其机制尚未解决。在这里，我们将生物化学和遗传学与相关光学和电子显微镜 (CLEM) 结合起来，探索如何实现选择性。我们的数据表明，囊泡占据有助于 ER 保留：在缺乏丰富货物的情况下，非特异性总体流量增加。我们证明内质网渗漏受到囊泡大小和货物占用的影响：过度表达惰性货物蛋白或减少囊泡大小可以恢复分选严格性。我们建议将货物招募到囊泡中会产生一个拥挤的内腔，从而提高选择性。因此，ER 居民的保留部分源于货物富集到受限球形膜结合载体中的生物物理过程。