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Involvement of MEM1 in DNA demethylation in Arabidopsis

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MEM1 participates in ROS1-mediated DNA demethylation pathway, and acts functionally as ROS3 to counteract the effects of RdDM pathway. mem1 mutation leads to large numbers of hyper-DMRs in Arabidopsis genome.

Abstract

In higher plants, DNA methylation performs important functions in silencing transcribed genes and transposable elements (TEs). Active DNA demethylation mediated by REPRESSOR OF SILENCING 1 (ROS1) is able to antagonize the action of DNA methylation caused by RNA-directed DNA methylation (RdDM) pathway, which plays critical roles in keeping DNA methylation at a proper level. In this study, a new mutant named mem1 (for methylation elevated mutant 1) was isolated from a genetic screen of T-DNA insertional mutant population for lines with elevated DNA methylation at a particular locus through Chop-PCR method. MEM1 possesses a Zf-C3HC domain, and is localized in nucleus as well as highly expressed in cotyledons. Whole-genome bisulfite sequencing data showed that knockout mutation of MEM1 leads to 4519 CG, 1793 CHG and 12739 CHH hyper-DMRs (for differentially methylated regions). Further analysis indicated that there are 2751, 2216 and 2042 overlapped CG hyper-DMRs between mem1-1and three mutants, i.e. ros1-4, rdd and ros3-2, respectively; 797, 2514, and 6766 overlapped CHH hyper-DMRs were observed between mem1-1 and three such mutants, respectively; mem1 nrpd1-3 and mem1 rdm1 double mutants showed nearly complete or partial loss of hypermethylation at 4 tested loci, suggesting that MEM1 performs similar functions as DNA glycosylase/lyases in counteracting excessive DNA methylation, and MEM1 plays important roles as REPRESSOR OF SILENCING 3 (ROS3) in erasing CHH methylation caused by the RdDM pathway. Together, these data demonstrate the involvement of MEM1 in ROS1-mediated DNA demethylation pathway and functional connections between MEM1 and ROS3.

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Acknowledgements

This study was funded by the National Natural Science Foundation of China (31771427), the National Natural Science Foundation of China (31471214), the Fundamental Research Funds for the Central Universities (KJSY201704), “333” project of Jiangsu province (BRA2017430), and A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD). Data analysis was supported by the high-performance computing platform of Bioinformatics Center, Nanjing Agricultural University.

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YL and HL designed the experiments; YL, LY, QW and FT performed the experiments; SZ provided technical assistance; JD, YL, YW and WK analyzed the data; YL, SQ and HL wrote the manuscript.

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Correspondence to Honggui La.

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Accession number: The whole-genome bisulfite sequencing data included in this article can be found in SRA database with accession number SRR10489181 (Data Deposition Information—https://trace.ncbi.nlm.nih.gov/Traces/sra/?run=SRR10489181).

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Lu, Y., Dai, J., Yang, L. et al. Involvement of MEM1 in DNA demethylation in Arabidopsis. Plant Mol Biol 102, 307–322 (2020). https://doi.org/10.1007/s11103-019-00949-0

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