Methylation of MdMYB1 locus mediated by RdDM pathway regulates anthocyanin biosynthesis in apple
Methylation of MdMYB1 locus mediated by RdDM pathway regulates anthocyanin biosynthesis in apple
复制标题
RdDM通路介导的MdMYB1位点甲基化调控苹果花青素生物合成
DOI：
10.1111/pbi.13337
复制
发表时间：
2020-01-30
影响因子：
13.8
通讯作者：
Chen, Xuesen
中科院分区：
文献类型：
--
作者：
Jiang, Shenghui;Wang, Nan;Chen, Xuesen
据报道，苹果运动中MdMYB1启动子的甲基化是花青素途径的调节因子，但对于甲基化机制如何识别该位点以调节花青素积累知之甚少。在这项研究中，我们分析了三种不同颜色的“富士”苹果，发现MdMYB1的转录水平存在差异，MdMYB1编码花青素生物合成的关键调节因子，控制着果皮中花青素的含量（因此颜色）。MdMYB1启动子MR3区域（- 1246至- 780）的CHH甲基化水平与MdMYB1表达呈负相关。因此，它们是研究苹果运动中DNA甲基化的理想材料。研究发现，负责CHH甲基化的RNA定向DNA甲基化（RdDM）途径蛋白MdAGO4与MdMYB1启动子相互作用。MdAGO4s可以与MdRDM1和MdDRM2s相互作用形成效应复合物，实现CHH甲基化。当MdAGO4s和MdDRM2s在苹果愈伤组织和拟南芥突变体中过表达时，这些蛋白增加了AGO4结合位点的CHH甲基化。在电泳迁移率转移实验中，发现MdAGO4s特异性结合含有ATATCAGA的序列。在35S::AGO4愈伤组织中，敲低MdNRPE1不影响MdAGO4s与MdMYB1启动子c3区域的结合。综上所述，我们的数据表明MdMYB1位点通过MdAGO4s结合MdMYB1启动子来甲基化，从而通过RdDM途径调节花青素的生物合成。
Summary Methylation at the MdMYB1 promoter in apple sports has been reported as a regulator of the anthocyanin pathway, but little is known about how the locus is recognized by the methylation machinery to regulate anthocyanin accumulation. In this study, we analysed three differently coloured ‘Fuji’ apples and found that differences in the transcript levels of MdMYB1, which encodes a key regulator of anthocyanin biosynthesis, control the anthocyanin content (and therefore colour) in fruit skin. The CHH methylation levels in the MR3 region (−1246 to −780) of the MdMYB1 promoter were found to be negatively correlated with MdMYB1 expression. Thus, they were ideal materials to study DNA methylation in apple sports. The protein of RNA‐directed DNA methylation (RdDM) pathway responsible for CHH methylation, MdAGO4, was found to interact with the MdMYB1 promoter. MdAGO4s can interact with MdRDM1 and MdDRM2s to form an effector complex, fulfilling CHH methylation. When MdAGO4s and MdDRM2s were overexpressed in apple calli and Arabidopsis mutants, those proteins increase the CHH methylation of AGO4‐binding sites. In electrophoretic mobility shift assays, MdAGO4s were found to specifically bind to sequence containing ATATCAGA. Knockdown of MdNRPE1 did not affect the binding of MdAGO4s to the c3 region of the MdMYB1 promoter in 35S::AGO4 calli. Taken together, our data show that the MdMYB1 locus is methylated through binding of MdAGO4s to the MdMYB1 promoter to regulate anthocyanin biosynthesis by the RdDM pathway.