Cao Xiaofeng, Aufsatz Werner, Zilberman Daniel, Mette M Florian, Huang Michael S, Matzke Marjori, Jacobsen Steven E
Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, 917 Datun Road, Beijing 100101, China.
Curr Biol. 2003 Dec 16;13(24):2212-7. doi: 10.1016/j.cub.2003.11.052.
RNA interference is a conserved process in which double-stranded RNA is processed into 21-25 nucleotide siRNAs that trigger posttranscriptional gene silencing. In addition, plants display a phenomenon termed RNA-directed DNA methylation (RdDM) in which DNA with sequence identity to silenced RNA is de novo methylated at its cytosine residues. This methylation is not only at canonical CpG sites but also at cytosines in CpNpG and asymmetric sequence contexts. In this report, we study the role of the DRM and CMT3 DNA methyltransferase genes in the initiation and maintenance of RdDM. Neither drm nor cmt3 mutants affected the maintenance of preestablished RNA-directed CpG methylation. However, drm mutants showed a nearly complete loss of asymmetric methylation and a partial loss of CpNpG methylation. The remaining asymmetric and CpNpG methylation was dependent on the activity of CMT3, showing that DRM and CMT3 act redundantly to maintain non-CpG methylation. These DNA methyltransferases appear to act downstream of siRNAs, since drm1 drm2 cmt3 triple mutants show a lack of non-CpG methylation but elevated levels of siRNAs. Finally, we demonstrate that DRM activity is required for the initial establishment of RdDM in all sequence contexts including CpG, CpNpG, and asymmetric sites.
RNA干扰是一个保守的过程，在这个过程中，双链RNA被加工成21 - 25个核苷酸的小干扰RNA（siRNA），这些siRNA会引发转录后基因沉默。此外，植物表现出一种称为RNA指导的DNA甲基化（RdDM）的现象，在这种现象中，与沉默RNA具有序列同一性的DNA在其胞嘧啶残基处发生从头甲基化。这种甲基化不仅发生在典型的CpG位点，也发生在CpNpG和不对称序列背景下的胞嘧啶处。在本报告中，我们研究了DRM和CMT3 DNA甲基转移酶基因在RdDM起始和维持中的作用。drm和cmt3突变体均不影响预先建立的RNA指导的CpG甲基化的维持。然而，drm突变体显示不对称甲基化几乎完全丧失，CpNpG甲基化部分丧失。剩余的不对称和CpNpG甲基化依赖于CMT3的活性，这表明DRM和CMT3在维持非CpG甲基化方面起冗余作用。这些DNA甲基转移酶似乎在siRNA下游起作用，因为drm1drm2cmt3三突变体显示缺乏非CpG甲基化，但siRNA水平升高。最后，我们证明DRM活性是在包括CpG、CpNpG和不对称位点在内的所有序列背景下RdDM初始建立所必需的。
