Sarkar Susovan, Liu Hao-Yang, Yuan Feng, Malady Brandon T, Wang Liping, Lafer Eileen M, Perez Jessica, Huibregtse Jon M, Stachowiak Jeanne C
Department of Biomedical Engineering, The University of Texas at Austin, Austin, TX, USA.
Department of Biochemistry and Structural Biology, The University of Texas Health, Science Center at San Antonio, San Antonio, TX, USA.
Nat Commun. 2025 Aug 25;16(1):7929. doi: 10.1038/s41467-025-63238-z.
Clathrin-mediated endocytosis internalizes proteins and lipids from the cell surface. A flexible condensate of initiator proteins catalyzes assembly of clathrin-coated vesicles in diverse organisms. Here we reveal that an endocytic adaptor protein, Epsin1, conditionally stabilizes this network, creating a cargo-dependent endocytic checkpoint. Epsin1 recruits ubiquitylated cargo to endocytic sites. Using in vitro assays, we demonstrate that Epsin1 destabilizes condensation of initiator proteins in the absence of ubiquitin. However, when polyubiquitin is present, Epsin1 binds to both ubiquitin and initiator proteins, stabilizing condensation. Similarly, in mammalian cells, endocytosis is disrupted by removal of either ubiquitin or Epsin1. When both components are removed simultaneously, endocytic defects are largely rescued, although the ability to preferentially internalize ubiquitylated cargo is lost. These results suggest that Epsin1 tunes protein condensation to internalize ubiquitylated cargo. More broadly, these findings illustrate how a balance of attractive and repulsive molecular interactions can exert dynamic control over cellular events.
网格蛋白介导的内吞作用可将细胞表面的蛋白质和脂质内化。起始蛋白的柔性凝聚物在多种生物体中催化网格蛋白包被囊泡的组装。在这里，我们揭示了一种内吞衔接蛋白Epsin1可条件性地稳定该网络，从而形成一个依赖货物的内吞检查点。Epsin1将泛素化的货物招募到内吞位点。通过体外实验，我们证明在没有泛素的情况下，Epsin1会破坏起始蛋白的凝聚。然而，当存在多聚泛素时，Epsin1会同时结合泛素和起始蛋白，从而稳定凝聚。同样，在哺乳动物细胞中，去除泛素或Epsin1都会破坏内吞作用。当同时去除这两种成分时，尽管优先内化泛素化货物的能力丧失，但内吞缺陷在很大程度上得到挽救。这些结果表明，Epsin1调节蛋白质凝聚以内化泛素化货物。更广泛地说，这些发现说明了吸引和排斥分子相互作用的平衡如何对细胞事件施加动态控制。
