A SPOPL/Cullin-3 ubiquitin ligase complex regulates endocytic trafficking by targeting EPS15 at endosomes.
A SPOPL/Cullin-3 ubiquitin ligase complex regulates endocytic trafficking by targeting EPS15 at endosomes.
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DOI：
10.7554/elife.13841
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发表时间：
2016-03-23
期刊：
影响因子：
7.7
通讯作者：
Peter M
中科院分区：
文献类型：
--
作者：
Gschweitl M;Ulbricht A;Barnes CA;Enchev RI;Stoffel-Studer I;Meyer-Schaller N;Huotari J;Yamauchi Y;Greber UF;Helenius A;Peter M
基于 Cullin-3 (CUL3) 的泛素连接酶调节哺乳动物细胞内体的成熟和内吞货物向溶酶体的运输。在这里，我们报告这些功能依赖于 SPOPL，一种特定于基质的 CUL3 适配器。我们发现 SPOPL 与内体相关，并且是多泡体 (MVB) 的形成和甲型流感病毒内吞宿主细胞进入所必需的。在 SPOPL 耗尽的细胞中，内体增大并且无法获得腔内囊泡 (ILV)。我们确定了 CUL3-SPOPL 泛素化的关键底物 EPS15，它是一种内吞接头，也与内体上的 ESCRT-0 复合体成员 HRS 和 STAM 相关联。事实上，EPS15 以 SPOPL 依赖性方式泛素化，并在缺乏 SPOPL 的细胞中与 HRS 一起积累。总之，我们的数据表明，CUL3-SPOPL E3 泛素连接酶复合物通过在内体泛素化和降解 EPS15 来调节内吞运输和 MVB 形成，从而影响甲型流感病毒感染以及 EGFR 和其他 EPS15 靶标的降解。 DOI：http://dx.doi.org/10.7554/eLife.13841.001 单个细胞可以通过称为内吞作用的过程将物质（统称为货物）从外部环境移动到细胞内部。然后，细胞有不同的路线将货物包（称为内吞囊泡）运输到细胞内的特定位置。基于蛋白质的分子机器移动货物并控制如何选择货物并将其运送到不同的目的地。例如，含有一种名为 CUL3 的蛋白质的分子机器用化学标签标记系统的其他组件，以调节哺乳动物细胞中货物的路线。然而，目前尚不清楚 CUL3 如何选择性地附着化学标签。格施韦特尔、乌布利希等人。现在发现另一种名为 SPOPL 的蛋白质在人类细胞内吞作用期间为基于 CUL3 的机器提供了选择性。实验表明，SPOPL 附着于内吞囊泡，并且 CUL3 和 SPOPL 共同作用来标记这些囊泡的特定成分，称为 EPS15。该标签改变了 EPS15 与其他蛋白质相互作用的方式。当细胞中不存在 SPOPL 时，EPS15 会异常稳定，并占据内吞货物使用的许多路线。然后，与 EPS15 直接相互作用的货物将在快车道上运送到目的地，而其他货物则积聚在一种分子交通拥堵中。其他蛋白质如 SPOPL 是内吞系统特有的。 SPOPL 与 CUL3 机器中这些相似的蛋白质的交换可能会化学标记一组不同的内吞蛋白质。 Gschweitl、Ulbricht 等人的下一个挑战是确定内吞系统中这些可交换成分的选择性、靶向性和协调性。 DOI：http://dx.doi.org/10.7554/eLife.13841.002
Cullin-3 (CUL3)-based ubiquitin ligases regulate endosome maturation and trafficking of endocytic cargo to lysosomes in mammalian cells. Here, we report that these functions depend on SPOPL, a substrate-specific CUL3 adaptor. We find that SPOPL associates with endosomes and is required for both the formation of multivesicular bodies (MVBs) and the endocytic host cell entry of influenza A virus. In SPOPL-depleted cells, endosomes are enlarged and fail to acquire intraluminal vesicles (ILVs). We identify a critical substrate ubiquitinated by CUL3-SPOPL as EPS15, an endocytic adaptor that also associates with the ESCRT-0 complex members HRS and STAM on endosomes. Indeed, EPS15 is ubiquitinated in a SPOPL-dependent manner, and accumulates with HRS in cells lacking SPOPL. Together, our data indicates that a CUL3-SPOPL E3 ubiquitin ligase complex regulates endocytic trafficking and MVB formation by ubiquitinating and degrading EPS15 at endosomes, thereby influencing influenza A virus infection as well as degradation of EGFR and other EPS15 targets. DOI: http://dx.doi.org/10.7554/eLife.13841.001 Individual cells can move material, collectively referred to as cargo, from the outside environment into the cell interior via a process known as endocytosis. The cell then has different routes to transport the packages of cargo, called endocytic vesicles, to specific locations within the cell. Protein-based molecular machines move the cargo and control how it is selected and targeted to different destinations. For example, a molecular machine that contains a protein called CUL3 labels other components of the system with a chemical tag to regulate the route cargo takes in mammalian cells. However, it was not clear how CUL3 can selectively attach the chemical labels. Gschweitl, Ulbricht et al. have now found that another protein called SPOPL provides selectivity for the CUL3-based machine during endocytosis in human cells. The experiments show that SPOPL attaches to endocytic vesicles, and that CUL3 and SPOPL work together to label a specific component of these vesicles called EPS15. The label changes how EPS15 interacts with other proteins. When SPOPL is not present in a cell, EPS15 is unnaturally stable and occupies many of the routes used by endocytic cargos. The cargo directly interacting with EPS15 is then routed on the fast lane to its destination, while other cargo accumulate in a kind of molecular traffic jam. Other proteins like SPOPL are specific for the endocytic system. Exchange of SPOPL with these similar proteins in the CUL3 machine is likely to chemically label a different set of endocytic proteins. Gschweitl, Ulbricht et al.’s next challenge is to identify the selectivity, targeting and coordination of these exchangeable components in the endocytic system. DOI: http://dx.doi.org/10.7554/eLife.13841.002