Clathrin-mediated endocytosis (CME) is the main route of internalization from the plasma membrane. It is known that the heterotetrameric AP2 clathrin adaptor must open to simultaneously engage membrane and endocytic cargo, yet it is unclear how transmembrane cargos are captured to catalyze CME. Using cryogenic-electron microscopy, we discover a new way in which mouse AP2 can reorganize to expose membrane- and cargo-binding pockets, which is not observed in clathrin-coated structures. Instead, it is stimulated by endocytic pioneer proteins called muniscins, which do not enter vesicles. Muniscin-engaged AP2 is primed to rearrange into the vesicle-competent conformation on binding the tyrosine cargo internalization motif (YxxΦ). We propose adaptor priming as a checkpoint to ensure cargo internalization.

网格蛋白介导的内吞作用（CME）是质膜内化的主要途径。众所周知，异四聚体 AP2 网格蛋白接头必须打开才能同时接合膜和内吞货物，但尚不清楚跨膜货物如何被捕获以催化 CME。使用低温电子显微镜，我们发现了小鼠 AP2 可以重组以暴露膜和货物结合袋的新方法，这在网格蛋白包被的结构中没有观察到。相反，它受到称为粘菌素的内吞先驱蛋白的刺激，这种蛋白不会进入囊泡。粘菌素结合的 AP2 在结合酪氨酸货物内化基序 (YxxΦ) 时会重新排列成囊泡活性构象。我们建议将适配器启动作为检查点，以确保货物内部化。