The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter

The synthesis of anthocyanin pigments in plants is known to be regulated by multiple mechanisms, including epigenetic regulation; however, the contribution of the RNA-directed DNA methylation (RdDM) pathway is not well understood. Here, we used bisulfite sequencing and Real Time (RT)-quantitative (q...

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Autores principales:	Yifan Xing, Ziyi Xie, Weilei Sun, Yuying Sun, Zhenyun Han, Shiya Zhang, Ji Tian, Jie Zhang, Yuncong Yao
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	RNA directed DNA methylation anthocyanin <i>COP1</i> crabapple Botany QK1-989
Acceso en línea:	https://doaj.org/article/fac8ceb9f82d45aba18cadfe67821dde
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spelling	oai:doaj.org-article:fac8ceb9f82d45aba18cadfe67821dde2021-11-25T18:46:48ZThe RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter10.3390/plants101124662223-7747https://doaj.org/article/fac8ceb9f82d45aba18cadfe67821dde2021-11-01T00:00:00Zhttps://www.mdpi.com/2223-7747/10/11/2466https://doaj.org/toc/2223-7747The synthesis of anthocyanin pigments in plants is known to be regulated by multiple mechanisms, including epigenetic regulation; however, the contribution of the RNA-directed DNA methylation (RdDM) pathway is not well understood. Here, we used bisulfite sequencing and Real Time (RT)-quantitative (q) PCR to analyze the methylation level of the promoter of constitutively photomorphogenic 1 (<i>McCOP1</i>) from <i>Malus</i> cv. spp, a gene involved in regulating anthocyanin biosynthesis. The CHH methylation level of the <i>McCOP1</i> promoter was negatively correlated with <i>McCOP1</i> RNA expression, and inhibiting DNA methylation caused decreased methylation of the <i>McCOP1</i> promoter and asymmetric cytosine CHH methylation. We observed that the <i>McCOP1</i> promoter was a direct target of the RdDM pathway argonaute RISC component 4 (McAGO4) protein, which bound to a <i>McCOP1</i> promoter GGTTCGG site. Bimolecular fluorescence complementation (BIFC) analysis showed that RNA-directed DNA methylation (McRDM1) interacted with McAGO4 and another RdDM protein, domains rearranged methyltransferase 2 (McDRM2), to regulate the CHH methylation of the <i>McCOP1</i> promoter. Detection of CHH methylation and COP1 gene expression in the <i>Arabidopsis thaliana</i><i>atago4</i>, <i>atdrm2</i> and <i>atrdm1</i> mutants showed that <i>RDM1</i> is the effector of the RdDM pathway. This was confirmed by silencing <i>McRDM1</i> in crabapple leaves or apple fruit, which resulted in a decrease in <i>McCOP1</i> CHH methylation and an increase in <i>McCOP1</i> transcript levels, as well as in anthocyanin accumulation. In conclusion, these results show that the RdDM pathway is involved in regulating anthocyanin accumulation through CHH methylation of the <i>McCOP1</i> promoter.Yifan XingZiyi XieWeilei SunYuying SunZhenyun HanShiya ZhangJi TianJie ZhangYuncong YaoMDPI AGarticleRNA directed DNA methylationanthocyanin<i>COP1</i>crabappleBotanyQK1-989ENPlants, Vol 10, Iss 2466, p 2466 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	RNA directed DNA methylation anthocyanin <i>COP1</i> crabapple Botany QK1-989
spellingShingle	RNA directed DNA methylation anthocyanin <i>COP1</i> crabapple Botany QK1-989 Yifan Xing Ziyi Xie Weilei Sun Yuying Sun Zhenyun Han Shiya Zhang Ji Tian Jie Zhang Yuncong Yao The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter
description	The synthesis of anthocyanin pigments in plants is known to be regulated by multiple mechanisms, including epigenetic regulation; however, the contribution of the RNA-directed DNA methylation (RdDM) pathway is not well understood. Here, we used bisulfite sequencing and Real Time (RT)-quantitative (q) PCR to analyze the methylation level of the promoter of constitutively photomorphogenic 1 (<i>McCOP1</i>) from <i>Malus</i> cv. spp, a gene involved in regulating anthocyanin biosynthesis. The CHH methylation level of the <i>McCOP1</i> promoter was negatively correlated with <i>McCOP1</i> RNA expression, and inhibiting DNA methylation caused decreased methylation of the <i>McCOP1</i> promoter and asymmetric cytosine CHH methylation. We observed that the <i>McCOP1</i> promoter was a direct target of the RdDM pathway argonaute RISC component 4 (McAGO4) protein, which bound to a <i>McCOP1</i> promoter GGTTCGG site. Bimolecular fluorescence complementation (BIFC) analysis showed that RNA-directed DNA methylation (McRDM1) interacted with McAGO4 and another RdDM protein, domains rearranged methyltransferase 2 (McDRM2), to regulate the CHH methylation of the <i>McCOP1</i> promoter. Detection of CHH methylation and COP1 gene expression in the <i>Arabidopsis thaliana</i><i>atago4</i>, <i>atdrm2</i> and <i>atrdm1</i> mutants showed that <i>RDM1</i> is the effector of the RdDM pathway. This was confirmed by silencing <i>McRDM1</i> in crabapple leaves or apple fruit, which resulted in a decrease in <i>McCOP1</i> CHH methylation and an increase in <i>McCOP1</i> transcript levels, as well as in anthocyanin accumulation. In conclusion, these results show that the RdDM pathway is involved in regulating anthocyanin accumulation through CHH methylation of the <i>McCOP1</i> promoter.
format	article
author	Yifan Xing Ziyi Xie Weilei Sun Yuying Sun Zhenyun Han Shiya Zhang Ji Tian Jie Zhang Yuncong Yao
author_facet	Yifan Xing Ziyi Xie Weilei Sun Yuying Sun Zhenyun Han Shiya Zhang Ji Tian Jie Zhang Yuncong Yao
author_sort	Yifan Xing
title	The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter
title_short	The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter
title_full	The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter
title_fullStr	The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter
title_full_unstemmed	The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter
title_sort	rna directed dna methylation (rddm) pathway regulates anthocyanin biosynthesis in crabapple (<i>malus</i> cv. spp.) leaves by methylating the <i>mccop1</i> promoter
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/fac8ceb9f82d45aba18cadfe67821dde
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The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (<i>Malus</i> cv. spp.) Leaves by Methylating the <i>McCOP1</i> Promoter

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