Box C/D guide RNAs recognize a maximum of 10 nt of substrates
Box C/D guide RNAs recognize a maximum of 10 nt of substrates
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Box C/D 引导 RNA 最多可识别 10 nt 的底物
DOI：
10.1073/pnas.1604872113
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发表时间：
2016-09-27
影响因子：
11.1
通讯作者：
Ye, Keqiong
中科院分区：
文献类型：
--
作者：
Yang, Zuxiao;Lin, Jinzhong;Ye, Keqiong
Box C/D RNA 指导古细菌和真核生物中 RNA 的位点特异性 2'-O-甲基化。框 C 至 D' 和框 C' 至 D 之间的间隔区包含引导序列，其可与底物 RNA 形成一段碱基对。 C/D RNA 之间的间隔区和引导底物双链体的长度是可变的。在先前确定的 C/D 核糖核蛋白 (RNP) 结构中，12 nt 长的间隔区与底物形成 10 bp。如何容纳其他长度的垫片和引导基板双工仍然未知。在这里，我们分析了间隔基和引导基底双链体的长度如何影响修饰活性，并确定了用不同间隔基和基底组装的 C/D RNP 的三种结构。我们表明，在修饰过程中，引导物只能与底物形成最多 10 bp 的双链体。稍短的双链体是可以耐受的，但较长的双链体必须解开以适合加帽的蛋白质通道以进行修饰。具有 12 个核苷酸的间隔区，靠近框 C/C' 侧的过量未配对序列被环出。我们的结果提供了对 C/D RNA 底物识别机制的深入了解，并反驳了二聚体 C/D RNP 的 RNA 交换模型。
Box C/D RNAs guide site-specific 2'-O-methylation of RNAs in archaea and eukaryotes. The spacer regions between boxes C to D' and boxes C' to D contain the guide sequence that can form a stretch of base pairs with substrate RNAs. The lengths of spacer regions and guide-substrate duplexes are variable among C/D RNAs. In a previously determined structure of C/D ribonucleoprotein (RNP), a 12-nt-long spacer forms 10 bp with the substrate. How spacers and guide-substrate duplexes of other lengths are accommodated remains unknown. Here we analyze how the lengths of spacers and guide-substrate duplexes affect the modification activity and determine three structures of C/D RNPs assembled with different spacers and substrates. We show that the guide can only form a duplex of a maximum of 10 bp with the substrate during modification. Slightly shorter duplexes are tolerated, but longer duplexes must be unwound to fit into a capped protein channel for modification. Spacers with 12 nucleotides, the excessive unpaired sequences near the box C/C' side are looped out. Our results provide insight into the substrate recognition mechanism of C/D RNA and refute the RNA-swapped model for dimeric C/D RNP.