COPI-coated vesicles mediate trafficking within the Golgi apparatus and from the Golgi to the endoplasmic reticulum. The structures of membrane protein coats, including COPI, have been extensively studied with in vitro reconstitution systems using purified components. In a previous paper (Dodonova et al., 2017), we determined a complete structural model of the in vitro reconstituted COPI coat. Here, we applied cryo-focused ion beam milling, cryo-electron tomography and subtomogram averaging to determine the native structure of the COPI coat within vitrified Chlamydomonas reinhardtii cells. The native algal structure resembles the in vitro mammalian structure, but additionally reveals cargo bound beneath β'-COP. We find that all coat components disassemble simultaneously and relatively rapidly after budding. Structural analysis in situ, maintaining Golgi topology, shows that vesicles change their size, membrane thickness, and cargo content as they progress from cis to trans, but the structure of the coat machinery remains constant.

COPI涂层的囊泡介导高尔基体内部以及从高尔基体到内质网的运输。膜蛋白涂层的结构（包括COPI）已通过使用纯化成分的体外重构系统进行了广泛研究。在之前的一篇论文中（Dodonova et al。，2017），我们确定了体外重建的COPI涂层的完整结构模型。在这里，我们应用了低温聚焦离子束铣削，低温电子断层扫描和断层图平均法来确定玻璃化莱茵衣藻细胞内COPI涂层的天然结构。天然藻类结构类似于体外哺乳动物的结构，但另外揭示了绑定在β'-COP下的货物。我们发现，所有涂层成分在出芽后会同时且相对快速地分解。原位结构分析（保持高尔基体拓扑结构）显示，囊泡从顺式转变为反式时会改变其大小，膜厚度和货物含量，但包衣机械的结构保持不变。