Autophagy degrades cytoplasmic cargo by its delivery to lysosomes within double membrane autophagosomes. Synthesis of the phosphoinositide PI(3)P by the autophagic class III phosphatidylinositol-3 kinase complex I (PI3KC3-C1) and conjugation of ATG8/LC3 proteins to phagophore membranes by the ATG12–ATG5-ATG16L1 (E3) complex are two critical steps in autophagosome biogenesis, connected by WIPI2. Here, we present a complete reconstitution of these events. On giant unilamellar vesicles (GUVs), LC3 lipidation is strictly dependent on the recruitment of WIPI2 that in turn depends on PI(3)P. Ectopically targeting E3 to membranes in the absence of WIPI2 is insufficient to support LC3 lipidation, demonstrating that WIPI2 allosterically activates the E3 complex. PI3KC3-C1 and WIPI2 mutually promote the recruitment of each other in a positive feedback loop. When both PI 3-kinase and LC3 lipidation reactions were performed simultaneously, positive feedback between PI3KC3-C1 and WIPI2 led to rapid LC3 lipidation with kinetics similar to that seen in cellular autophagosome formation.

中文翻译：

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PI3K-WIPI2正反馈环变构激活自噬中的LC3脂化

自噬通过将细胞质货物递送至双膜自噬体内的溶酶体来降解细胞质货物。通过自噬 III 类磷脂酰肌醇-3 激酶复合物 I (PI3KC3-C1) 合成磷酸肌醇 PI(3)P 以及通过 ATG12–ATG5-ATG16L1 (E3) 复合物将 ATG8/LC3 蛋白缀合至吞噬细胞膜是两个关键步骤在自噬体生物发生中，通过 WIPI2 连接。在这里，我们对这些事件进行了完整的重构。在巨型单层囊泡 (GUV) 上，LC3 脂化严格依赖于 WIPI2 的募集，而 WIPI2 又依赖于 PI(3)P。在没有 WIPI2 的情况下，将 E3 异位靶向膜不足以支持 LC3 脂化，这表明 WIPI2 以变构方式激活 E3 复合物。PI3KC3-C1 和 WIPI2 在正反馈循环中相互促进彼此的募集。当 PI 3 激酶和 LC3 脂化反应同时进行时，PI3KC3-C1 和 WIPI2 之间的正反馈导致 LC3 快速脂化，其动力学类似于细胞自噬体形成中所见的动力学。