The N domain of Argonaute drives duplex unwinding during RISC assembly
The N domain of Argonaute drives duplex unwinding during RISC assembly
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DOI：
10.1038/nsmb.2232
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发表时间：
2012-02-01
影响因子：
16.8
通讯作者：
Tomari, Yukihide
中科院分区：
文献类型：
--
作者：
Kwak, Pieter Bas;Tomari, Yukihide
小RNA，如microRNA和小干扰RNA，通过Argonaute（Ago）蛋白作为RNA诱导沉默复合物（RISC）的一部分发挥作用。为了制造RISC，Ago蛋白结合并随后解开小RNA双链体，最后留下一条稳定掺入的链。在这里，我们确定了人AGO 2的N结构域作为RISC组装过程中双链体解旋的起始子。我们发现，一个功能性的N结构域是严格需要的小RNA双链体解旋，但不是预先双链体加载或随后的目标切割。我们假设RISC组装是三部分的，包括（i）RISC加载，由此Ago经历构象打开并加载小RNA双链体，形成pre-RISC;（ii）楔入，由此双链体的末端通过N结构域的主动楔入而被撬开，为解旋做准备;和（iii）解旋，由此通过依赖于切割器或不依赖于切割器的解旋除去过客链，形成成熟RISC。
Small RNAs, such as microRNAs and small interfering RNAs, act through Argonaute (Ago) proteins as a part of RNA-induced silencing complexes (RISC). To make RISCs, Ago proteins bind and subsequently unwind small RNA duplexes, finally leaving one strand stably incorporated. Here we identified the N domain of human AGO2 as the initiator of duplex unwinding during RISC assembly. We discovered that a functional N domain is strictly required for small RNA duplex unwinding but not for precedent duplex loading or subsequent target cleavage. We postulate that RISC assembly is tripartite, comprising (i) RISC loading, whereby Ago undergoes conformational opening and loads a small RNA duplex, forming pre-RISC; (ii) wedging, whereby the end of the duplex is pried open through active wedging by the N domain, in preparation for unwinding; and (iii) unwinding, whereby the passenger strand is removed through slicer-dependent or slicer-independent unwinding, forming mature RISC.