Assembly and analysis of eukaryotic Argonaute-RNA complexes in microRNA-target recognition.
Assembly and analysis of eukaryotic Argonaute-RNA complexes in microRNA-target recognition.
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DOI：
10.1093/nar/gkv990
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发表时间：
2015-11-16
影响因子：
14.9
通讯作者：
Gunsalus KC
中科院分区：
文献类型：
--
作者：
Gan HH;Gunsalus KC
实验研究发现了多种microRNA(MiRNA)靶向双链结构，包括完美的、不完美的和无核的双链。然而，主要依赖于序列和二级结构特征的计算方法并不能很好地预测不完美/无籽双链的非正则结合模式，也没有表征它们的三级结构，因为到目前为止所解决的结构仅限于ArgAertes(AGOS)中的接近完美的直链双链。在这里，我们使用结构建模来研究AGO动力学在组装可存活的真核miRNA诱导沉默复合体(MiRISCs)中的作用。我们表明，在模型人类和线虫AGO结构中，需要低频、全局运动模式的组合来适应RNA双链。活性miRISCs的模型表明，AGO在不同的靶点采用不同的构象，从而产生扭曲的、不完美的miRNA-靶向双链。AGO容纳双链的能力取决于发生结构扭曲的区域：暴露在溶剂中的种子区域和3‘-末端区域中的扭曲比中央双链区域中的扭曲更不可能产生空间冲突。对组装的miRISCs的能量分析表明，目标识别也受到良好的AGO-双链相互作用的驱动。这种对AGO加载和靶标识别机制的结构性洞察可能会为miRNA功能提供更准确的评估。
Experimental studies have uncovered a variety of microRNA (miRNA)–target duplex structures that include perfect, imperfect and seedless duplexes. However, non-canonical binding modes from imperfect/seedless duplexes are not well predicted by computational approaches, which rely primarily on sequence and secondary structural features, nor have their tertiary structures been characterized because solved structures to date are limited to near perfect, straight duplexes in Argonautes (Agos). Here, we use structural modeling to examine the role of Ago dynamics in assembling viable eukaryotic miRNA-induced silencing complexes (miRISCs). We show that combinations of low-frequency, global modes of motion of Ago domains are required to accommodate RNA duplexes in model human and C. elegans Ago structures. Models of viable miRISCs imply that Ago adopts variable conformations at distinct target sites that generate distorted, imperfect miRNA-target duplexes. Ago's ability to accommodate a duplex is dependent on the region where structural distortions occur: distortions in solvent-exposed seed and 3′-end regions are less likely to produce steric clashes than those in the central duplex region. Energetic analyses of assembled miRISCs indicate that target recognition is also driven by favorable Ago-duplex interactions. Such structural insights into Ago loading and target recognition mechanisms may provide a more accurate assessment of miRNA function.