The sorting nexins (SNXs) are a family of peripheral membrane proteins that direct protein trafficking decisions within the endocytic network. Emerging evidence in yeast and mammalian cells implicates a subgroup of SNXs in selective and non-selective forms of autophagy. Using siRNA and CRISPR-Cas9, we demonstrate that the SNX-BAR protein SNX4 is needed for efficient LC3 (also known as MAP1LC3) lipidation and autophagosome assembly in mammalian cells. SNX-BARs exist as homo- and hetero-dimers, and we show that SNX4 forms functional heterodimers with either SNX7 or SNX30 that associate with tubulovesicular endocytic membranes. Detailed image-based analysis during the early stages of autophagosome assembly reveals that SNX4–SNX7 is an autophagy-specific SNX-BAR heterodimer, required for efficient recruitment and/or retention of core autophagy regulators at the nascent isolation membrane. SNX4 partially colocalises with juxtanuclear ATG9A-positive membranes, with our data linking the autophagy defect upon SNX4 disruption to the mis-trafficking and/or retention of ATG9A in the Golgi region. Taken together, our findings show that the SNX4–SNX7 heterodimer coordinates ATG9A trafficking within the endocytic network to establish productive autophagosome assembly sites, thus extending knowledge of SNXs as positive regulators of autophagy.

分选连接蛋白 (SNX) 是一个外周膜蛋白家族，可指导内吞网络内的蛋白质运输决策。酵母和哺乳动物细胞中的新证据表明 SNX 亚群参与选择性和非选择性自噬形式。使用 siRNA 和 CRISPR-Cas9，我们证明 SNX-BAR 蛋白 SNX4 是哺乳动物细胞中有效 LC3（也称为 MAP1LC3）脂化和自噬体组装所必需的。SNX-BAR 以同源二聚体和异源二聚体的形式存在，我们发现 SNX4 与 SNX7 或 SNX30 形成功能性异源二聚体，与管泡内吞膜相关。自噬体组装早期阶段基于详细图像的分析表明，SNX4–SNX7 是一种自噬特异性 SNX-BAR 异二聚体，是在新生隔离膜上有效招募和/或保留核心自噬调节因子所必需的。SNX4 与核旁 ATG9A 阳性膜部分共定位，我们的数据将 SNX4 破坏时的自噬缺陷与 ATG9A 在高尔基体区域的错误运输和/或保留联系起来。综上所述，我们的研究结果表明，SNX4-SNX7 异二聚体协调内吞网络内的 ATG9A 运输，以建立高效的自噬体组装位点，从而扩展了 SNX 作为自噬正调节因子的知识。