Hrs and STAM function synergistically to bind ubiquitin-modified cargoes in vitro.
Hrs and STAM function synergistically to bind ubiquitin-modified cargoes in vitro.
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DOI：
10.1016/j.bpj.2014.11.004
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发表时间：
2015-01
影响因子：
3.4
通讯作者：
Hirohide Takahashi;J. Mayers;Lei Wang;J. Edwardson;A. Audhya
中科院分区：
文献类型：
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作者：
Hirohide Takahashi;J. Mayers;Lei Wang;J. Edwardson;A. Audhya
整合膜蛋白的周转需要一个专门的运输途径介导的内体分选复合物所需的运输（ESCRT）机制的组成部分。在大多数情况下，进入这一途径需要货物进行泛素修饰，从而促进其隔离的内体膜上的特异性，泛素结合ESCRT亚基。然而，单泛素化货物的初始货物识别的要求仍然定义不清。在这项研究中，我们确定的能力，每个ESCRT复合物，窝藏一个泛素结合域结合重组的整体膜货物（VAMP 2），这已被共价连接到单泛素。我们证明，ESCRT-0，而不是ESCRT-I或ESCRT-II，能够与脂质双层内的单泛素化货物稳定地关联。此外，我们表明，在Hrs和STAM的泛素结合结构域必须是完整的，使货物结合。这些结果表明ESCRT-0的两个亚基一起起作用以结合和隔离货物，用于下游分选到腔内囊泡中。
The turnover of integral membrane proteins requires a specialized transport pathway mediated by components of the endosomal sorting complex required for transport (ESCRT) machinery. In most cases, entry into this pathway requires that cargoes undergo ubiquitin-modification, thereby facilitating their sequestration on endosomal membranes by specific, ubiquitin-binding ESCRT subunits. However, requirements underlying initial cargo recognition of mono-ubiquitinated cargos remain poorly defined. In this study, we determine the capability of each ESCRT complex that harbors a ubiquitin-binding domain to bind a reconstituted integral membrane cargo (VAMP2), which has been covalently linked to mono-ubiquitin. We demonstrate that ESCRT-0, but not ESCRT-I or ESCRT-II, is able to associate stably with the mono-ubiquitinated cargo within a lipid bilayer. Moreover, we show that the ubiquitin-binding domains in both Hrs and STAM must be intact to enable cargo binding. These results indicate that the two subunits of ESCRT-0 function together to bind and sequester cargoes for downstream sorting into intralumenal vesicles.