Interplay among RNA polymerases II, IV and V in RNA-directed DNA methylation at a low copy transgene locus in Arabidopsis thaliana
Interplay among RNA polymerases II, IV and V in RNA-directed DNA methylation at a low copy transgene locus in Arabidopsis thaliana
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DOI：
10.1007/s11103-013-0041-4
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发表时间：
2013-05-01
影响因子：
5.1
通讯作者：
Matzke, Marjori
中科院分区：
文献类型：
--
作者：
You, Wanhui;Lorkovic, Zdravko J.;Matzke, Marjori
RNA 指导的 DNA 甲基化 (RdDM) 是一种表观遗传过程，通过小干扰 RNA (siRNA) 引导同源 DNA 序列的胞嘧啶甲基化。 RdDM 需要两种专门的 RNA 聚合酶：Pol IV 转录 siRNA 前体，而 Pol V 生成支架 RNA，与 siRNA 相互作用并吸引甲基化机制。最近的证据还表明 RNA 聚合酶 II (Pol II) 参与将 Pol IV 和 Pol V 招募到低拷贝、基因间基因座。我们之前证明，拟南芥转基因位点上的 Pol V 介导的甲基化通过 Pol IV/RNA 依赖的 RNA 聚合酶 2 (RDR2) 依赖的 24 nt 二级 siRNA 扩散到最初目标区域的下游。在这里，我们表明，这些二级 siRNA 不仅可以在未连接的靶位点诱导顺式甲基化，还可以诱导反式甲基化，前提是该序列被 Pol II 转录以产生非编码 RNA。 Pol II 转录物似乎对于未连接靶位点的 siRNA 扩增很重要，因为它的存在不仅与甲基化相关，还与 24-nt siRNA 水平升高相关。缺乏重叠 Pol II 转录物并在反式作用 24-nt siRNA 存在下保持未甲基化的潜在靶位点在 21-24-nt 发夹衍生 siRNA 存在下仍能获得甲基化，这表明非转录靶序列的 RdDM 需要多种尺寸类别的 siRNA。我们的研究结果表明，低拷贝位点的 RdDM 并不总是需要 Pol II 转录本，但它们可能通过促进 DNA 靶位点上 Pol IV 依赖性 siRNA 的扩增来强化 RdDM。
RNA-directed DNA methylation (RdDM) is an epigenetic process whereby small interfering RNAs (siRNAs) guide cytosine methylation of homologous DNA sequences. RdDM requires two specialized RNA polymerases: Pol IV transcribes the siRNA precursor whereas Pol V generates scaffold RNAs that interact with siRNAs and attract the methylation machinery. Recent evidence also suggests the involvement of RNA polymerase II (Pol II) in recruiting Pol IV and Pol V to low copy, intergenic loci. We demonstrated previously that Pol V-mediated methylation at a transgene locus in Arabidopsis spreads downstream of the originally targeted region by means of Pol IV/RNA-DEPENDENT RNA POLYMERASE2 (RDR2)-dependent 24-nt secondary siRNAs. Here we show that these secondary siRNAs can not only induce methylation in cis but also in trans at an unlinked target site, provided this sequence is transcribed by Pol II to produce a non-coding RNA. The Pol II transcript appears to be important for amplification of siRNAs at the unlinked target site because its presence correlates not only with methylation but also with elevated levels of 24-nt siRNAs. Potential target sites that lack an overlapping Pol II transcript and remain unmethylated in the presence of trans-acting 24-nt siRNAs can nevertheless acquire methylation in the presence of 21-24-nt hairpin-derived siRNAs, suggesting that RdDM of non-transcribed target sequences requires multiple size classes of siRNA. Our findings demonstrate that Pol II transcripts are not always needed for RdDM at low copy loci but they may intensify RdDM by facilitating amplification of Pol IV-dependent siRNAs at the DNA target site.