Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.
Department of Plant and Microbial Biology, University of California, Berkeley, CA 94720, USA.
Plant Cell. 2022 Mar 29;34(4):1189-1206. doi: 10.1093/plcell/koab319.
Cytosine methylation is a reversible epigenetic modification of DNA. In plants, removal of cytosine methylation is accomplished by the four members of the DEMETER (DME) family of 5-methylcytosine DNA glycosylases, named DME, DEMETER-LIKE2 (DML2), DML3, and REPRESSOR OF SILENCING1 (ROS1) in Arabidopsis thaliana. Demethylation by DME is critical for seed development, preventing experiments to determine the function of the entire gene family in somatic tissues by mutant analysis. Here, we bypassed the reproductive defects of dme mutants to create somatic quadruple homozygous mutants of the entire DME family. dme; ros1; dml2; and dml3 (drdd) leaves exhibit hypermethylated regions compared with wild-type leaves and rdd triple mutants, indicating functional redundancy among all four demethylases. Targets of demethylation include regions co-targeted by RNA-directed DNA methylation and, surprisingly, CG gene body methylation, indicating dynamic methylation at these less-understood sites. Additionally, many tissue-specific methylation differences are absent in drdd, suggesting a role for active demethylation in generating divergent epigenetic states across wild-type tissues. Furthermore, drdd plants display an early flowering phenotype, which involves 5'-hypermethylation and transcriptional down-regulation of FLOWERING LOCUS C. Active DNA demethylation is therefore required for proper methylation across somatic tissues and defines the epigenetic landscape of intergenic and coding regions.
胞嘧啶甲基化是 DNA 的一种可逆转的表观遗传修饰。在植物中，通过 DEMETER（DME）家族的四个 5-甲基胞嘧啶 DNA 糖苷酶成员完成胞嘧啶甲基化的去除，在拟南芥中分别命名为 DME、DEMETER-LIKE2（DML2）、DML3 和 REPRESSOR OF SILENCING1（ROS1）。DME 的去甲基化对种子发育至关重要，通过突变分析来确定整个基因家族在体细胞组织中的功能的实验受到了阻碍。在这里，我们绕过了 dme 突变体的生殖缺陷，创建了整个 DME 家族的体细胞四重纯合突变体。与野生型叶片和 rdd 三重突变体相比，dme;ros1;dml2;和 dml3（drdd）叶片表现出超甲基化区域，表明所有四个去甲基酶之间存在功能冗余。去甲基化的靶标包括 RNA 指导的 DNA 甲基化和 CG 基因体甲基化共同靶向的区域，这表明这些较少被了解的位点存在动态甲基化。此外，在 drdd 中不存在许多组织特异性的甲基化差异，这表明主动去甲基化在产生野生型组织中不同的表观遗传状态方面发挥了作用。此外，drdd 植物表现出早花表型，涉及 5'-超甲基化和开花位点 C 的转录下调。因此，活性 DNA 去甲基化是体细胞组织中正确甲基化所必需的，并定义了基因间和编码区域的表观遗传景观。
