Qi Yijun, He Xingyue, Wang Xiu-Jie, Kohany Oleksiy, Jurka Jerzy, Hannon Gregory J
Cold Spring Harbor Laboratory, Watson School of Biological Sciences and Howard Hughes Medical Institute, 1 Bungtown Road, Cold Spring Harbor, New York 11724, USA.
Nature. 2006 Oct 26;443(7114):1008-12. doi: 10.1038/nature05198. Epub 2006 Sep 24.
DNA methylation has important functions in stable, transcriptional gene silencing, immobilization of transposable elements and genome organization. In Arabidopsis, DNA methylation can be induced by double-stranded RNA through the RNA interference (RNAi) pathway, a response known as RNA-directed DNA methylation. This requires a specialized set of RNAi components, including ARGONAUTE4 (AGO4). Here we show that AGO4 binds to small RNAs including small interfering RNAs (siRNAs) originating from transposable and repetitive elements, and cleaves target RNA transcripts. Single mutations in the Asp-Asp-His catalytic motif of AGO4 do not affect siRNA-binding activity but abolish its catalytic potential. siRNA accumulation and non-CpG DNA methylation at some loci require the catalytic activity of AGO4, whereas others are less dependent on this activity. Our results are consistent with a model in which AGO4 can function at target loci through two distinct and separable mechanisms. First, AGO4 can recruit components that signal DNA methylation in a manner independent of its catalytic activity. Second, AGO4 catalytic activity can be crucial for the generation of secondary siRNAs that reinforce its repressive effects.
DNA甲基化在稳定的转录基因沉默、转座元件固定及基因组组织中具有重要功能。在拟南芥中，双链RNA可通过RNA干扰（RNAi）途径诱导DNA甲基化，这种反应被称为RNA指导的DNA甲基化。这需要一组特殊的RNAi组分，包括AGO4（AGO4蛋白）。我们发现，AGO4可与小RNA结合，这些小RNA包括源自转座元件和重复元件的小干扰RNA（siRNA），并可切割靶RNA转录本。AGO4的天冬氨酸-天冬氨酸-组氨酸催化基序中的单突变不影响siRNA结合活性，但会消除其催化潜能。某些位点的siRNA积累和非CpG DNA甲基化需要AGO4的催化活性，而其他位点则对该活性的依赖性较低。我们的结果与一个模型相符，即AGO4可通过两种不同且可分离的机制在靶位点发挥作用。第一，AGO4可招募以与其催化活性无关的方式发出DNA甲基化信号的组分。第二，AGO4的催化活性对于增强其抑制作用的二级siRNA的产生可能至关重要。
