DNA 5-methylcytosine (5-meC) is an important epigenetic mark for transcriptional gene silencing in many eukaryotes. In Arabidopsis, 5-meC DNA glycosylase/lyases actively remove 5-meC to counter-act transcriptional gene silencing in a locus-specific manner, and have been suggested to maintain the expression of transposons. However, it is unclear whether plant DNA demethylases can promote the transposition of transposons. Here we report the functional characterization of the DNA glycosylase/lyase DNG701 in rice. DNG701 encodes a large (1,812 amino acid residues) DNA glycosylase domain protein. Recombinant DNG701 protein showed 5-meC DNA glycosylase and lyase activities in vitro. Knockout or knockdown of DNG701 in rice plants led to DNA hypermethylation and reduced expression of the retrotransposon Tos17. Tos17 showed less transposition in calli derived from dng701 knockout mutant seeds compared with that in wild-type calli. Overexpression of DNG701 in both rice calli and transgenic plants substantially reduced DNA methylation levels of Tos17 and enhanced its expression. The overexpression also led to more frequent transposition of Tos17 in calli. Our results demonstrate that rice DNG701 is a 5-meC DNA glycosylase/lyase responsible for the demethylation of Tos17 and this DNA demethylase plays a critical role in promoting Tos17 transposition in rice calli.
Bibliographical noteKAUST Repository Item: Exported on 2020-10-01
Acknowledgements: We thank Drs. Ko Shimamoto and Michael Goodin for providing the pANDA vector and pGDG vector, respectively; Dr. Steven E. Jacobsen for providing the genomic bisulfite sequencing protocol; Dr. Jiming Jiang for providing the centromeric repeat probe; and the Rice Genome Resource Center in Japan for providing the cDNA clones and Tos17 insertion lines. This research is supported by National Institutes of Health Grant R01GM070795 (to J.-K.Z.), and US Department of Agriculture-Cooperative State Research, Education, and Extension Service Grant 2004-05425 and National Science Foundation-Plant Genome Research Program Grants 0321437 and 0701745 (to G.-L.W.).
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