Abstract
In plants, RNA-directed DNA methylation (RdDM) is a well-known de novo DNA methylation pathway that involves two plant-specific RNA polymerases, Pol IV and Pol V. In this study, we discovered and characterized an RdDM factor, RDM15. Through DNA methylome and genome-wide siRNA analyses, we show that RDM15 is required for RdDM-dependent DNA methylation and siRNA accumulation at a subset of RdDM target loci. We show that RDM15 contributes to Pol V-dependent downstream siRNA accumulation and interacts with NRPE3B, a subunit specific to Pol V. We also show that the C-terminal tudor domain of RDM15 specifically recognizes the histone 3 lysine 4 monomethylation (H3K4me1) mark. Structure analysis of RDM15 in complex with the H3K4me1 peptide showed that the RDM15 tudor domain specifically recognizes the monomethyllysine through an aromatic cage and a specific hydrogen bonding network; this chemical feature-based recognition mechanism differs from all previously reported monomethyllysine recognition mechanisms. RDM15 and H3K4me1 have similar genome-wide distribution patterns at RDM15-dependent RdDM target loci, establishing a link between H3K4me1 and RDM15-mediated RdDM in vivo. In summary, we have identified and characterized a histone H3K4me1-specific binding protein as an RdDM component, and structural analysis of RDM15 revealed a chemical feature-based lower methyllysine recognition mechanism.
Highlights
DNA methylation mainly refers to an addition of a methyl group to the fifth position of cytosine, resulting in 5′methylcytosine (5-mC)
Cytosines in different sequence contexts can be methylated by different methyltransferases: cytosine in all sequence contexts can be de novo methylated by DRM2 (DOMAINS REARRANGED METHYLTRANSFERASE 2) through the RNA-dependent DNA methylation (RdDM) pathway; CG and CHG methylation is maintained by MET1 (METHYLTRANSFERASE 1) and CMT3 (CHROMOMETHYLASE 3), respectively, while CHH methylation is maintained by CMT2 or DRM2, depending on the chromatin context[1,3,4,5,6,7,8]
Through whole-genome bisulfite sequencing and siRNA sequencing of rdm[15] mutants and the wild type (WT), we demonstrate that RDM15 is required for RdDM-dependent DNA methylation and RdDM-dependent siRNA accumulation at a subset of RdDM target regions
Summary
DNA methylation mainly refers to an addition of a methyl group to the fifth position of cytosine, resulting in 5′methylcytosine (5-mC). The second step is siRNA-guided DNA methylation, in which Pol V-transcribed long-noncoding RNA serves as a scaffold RNA, and DRM2 is recruited to catalyze DNA methylation This recruitment of DRM2 involves base-pairing between AGO4 (ARGONAUTE 4)and AGO6-bound siRNAs and the scaffold RNA and the participation of RdDM factors such as KTF1 (KOW DOMAINCONTAINING TRANSCRIPTION FACTOR 1), DRD1, DMS3, and RDM1 (RNA-DIRECTED DNA METHYLATION 1)[10,11]. Different epigenetic marks, such as DNA methylation and histone modifications, can interact to regulate the chromatin state and DNA methylation level in the genome[12]. Our study has identified an RdDM component, RDM15, and has elucidated the molecular mechanism underlying RDM15 function in the RdDM pathway
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