VHL-box and SOCS-box domains determine binding specificity for Cul2-Rbx1 and Cul5-Rbx2 modules of ubiquitin ligases
VHL-box and SOCS-box domains determine binding specificity for Cul2-Rbx1 and Cul5-Rbx2 modules of ubiquitin ligases
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DOI：
10.1101/gad.1252404
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发表时间：
2004-12-15
影响因子：
10.5
通讯作者：
Nakayama, KI
中科院分区：
文献类型：
--
作者：
Kamura, T;Maenaka, K;Nakayama, KI
ECS（Elongin B/C-Cul 2/Cul 5-SOCS-box protein）复合物是共享Cullin-Rbx模块的泛素连接酶家族的成员。SOCS盒蛋白将底物募集到ECS复合物中，并通过延伸蛋白B/C与Cullin-Rbx连接。VHL被认为是SOCS盒蛋白，但缺乏SOCS盒的C-末端序列（BC盒的下游）。我们现在表明，VHL特异性地与哺乳动物细胞中的内源性Cul 2-Rbx 1相互作用，而SOCS盒蛋白与Cul 5-Rbx 2相关。我们还鉴定了LRR-1和FEM 1B作为与VHL（VHL盒，包括BC盒和下游残基）共享同源区域并与Cul 2-Rbx 1相关的蛋白。因此，ECS复合物可以分为两种不同的蛋白质组装体，即，含有具有与Cul 2-Rbx 1相互作用的VHL盒（由BC盒和下游Cul 2盒组成）的亚基的那些，以及含有具有与Cul 5-Rbx 2相互作用的SOCS盒（BC盒和下游Cul 5盒）的亚基的那些。结构域交换分析表明，VHL盒和SOCS盒蛋白与Cullin-Rbx模块的相互作用的特异性分别由Cul 2和Cul 5盒决定。最后，RNAi介导的Cul 2-Rbx 1的敲除抑制了VHL介导的HIF-2 α的降解，而Cul 5-Rbx 2的敲除没有影响它。这些数据表明Cul 2-Rbx 1和Cul 5-Rbx 2模块的功能是不同的。
The ECS (Elongin B/C-Cul2/Cul5-SOCS-box protein) complex is a member of a family of ubiquitin ligases that share a Cullin-Rbx module. SOCS-box proteins recruit substrates to the ECS complex and are linked to Cullin-Rbx via Elongin B/C. VHL has been implicated as a SOCS-box protein, but lacks a C-terminal sequence (downstream of the BC box) of the SOCS box. We now show that VHL specifically interacts with endogenous Cul2-Rbx1 in mammalian cells, whereas SOCS-box proteins associate with Cul5-Rbx2. We also identify LRR-1 and FEM1B as proteins that share a region of homology with VHL (the VHL box, including the BC box and downstream residues) and associate with Cul2-Rbx1. ECS complexes can thus be classified into two distinct protein assemblies, that is, those that contain a subunit with a VHL box (composed of the BC box and a downstream Cul2 box) that interacts with Cul2-Rbx1, and those that contain a subunit with a SOCS box (BC box and downstream Cul5 box) that interacts with Cul5-Rbx2. Domain-swapping analyses showed that the specificity of interaction of VHL-box and SOCS-box proteins with Cullin-Rbx modules is determined by the Cul2 and Cul5 boxes, respectively. Finally, RNAi-mediated knockdown of the Cul2-Rbx1 inhibited the VHL-mediated degradation of HIF-2alpha, whereas knockdown of Cul5-Rbx2 did not affect it. These data suggest that the functions of the Cul2-Rbx1 and Cul5-Rbx2 modules are distinct.