Size and positional effects of promoter RNA segments on virus-induced RNA-directed DNA methylation and transcriptional gene silencing
Size and positional effects of promoter RNA segments on virus-induced RNA-directed DNA methylation and transcriptional gene silencing
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DOI：
10.4161/epi.6.6.16214
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发表时间：
2011-06-01
期刊：
影响因子：
3.7
通讯作者：
Kanazawa, Akira
中科院分区：
文献类型：
--
作者：
Otagaki, Shungo;Kawai, Miou;Kanazawa, Akira
基因启动子上的DNA甲基化可以被双链RNA(DsRNAs)通过RNA引导的DNA甲基化(RdDM)途径触发，并诱导转录基因沉默(TGS)。虽然参与RdDM途径的基因已被鉴定，但不同启动子区域的dsRNAs是否对RdDM和/或TGS有不同程度的影响尚不清楚。在这里，我们通过使用含有不同部分启动子的重组黄瓜花叶病毒来靶向植物基因组中的CaMV 35S启动子来解决这个问题。TGS的诱导效率取决于启动RdDM的启动子片段的长度，TGS的诱导下限为81-91个核苷酸。当70个核苷酸片段串联时，TGS被诱导，但没有一个片段单独诱导TGS。TGS的诱导并不简单地依赖于与启动子相对应的小干扰RNA的产生。伴随着RdDM的诱导，检测到DNA甲基化从最初的靶点向邻近区域扩散。在缺乏RNA触发的后代中，TGS的维持依赖于前一代触发RdDM的启动子片段，并与靶区对称位置上的胞嘧啶数量相关。这些结果表明，dsRNA阈值以上的长度和dsRNA靶向区域对称位置上胞嘧啶的频率是允许通过RdDM诱导可遗传TGS的主要因素。
DNA methylation at a gene promoter can be triggered by double-stranded RNAs (dsRNAs) through the RNA-directed DNA methylation (RdDM) pathway and induces transcriptional gene silencing (TGS). Although genes involved in the RdDM pathway have been identified, whether dsRNAs of different promoter regions have different extent of effects on RdDM and/or TGS is unknown. Here, we addressed this question by targeting the CaMV 35S promoter in the plant genome using a recombinant Cucumber mosaic virus that contained various portions of the promoter. The efficiency of the induction of TGS depended on the length of the promoter segment triggering the RdDM; the lower size limit for TGS induction was 81-91 nt. TGS was induced when 70-nt fragments were connected in tandem, none of which solely induced TGS. TGS induction did not simply depend on the production of small interfering RNAs corresponding to the promoter. Along with the induction of RdDM, spreading of DNA methylation from the originally targeted site toward the adjacent regions was detected. The maintenance of TGS in the progeny that lacks an RNA trigger depended on the promoter segments triggering the RdDM in the former generation and was correlated with the number of cytosines at symmetrical sites in the targeted region. These results indicate that both the length of dsRNA above the threshold and the frequency of cytosines at symmetric sites in the region targeted by dsRNA are the major factors that allow induction of heritable TGS via RdDM.