The de novo cytosine methyltransferase DRM2 requires intact UBA domains and a catalytically mutated paralog DRM3 during RNA-directed DNA methylation in Arabidopsis thaliana.
Eukaryotic DNA cytosine methylation can be used to transcriptionally silence repetitive sequences, including transposons and retroviruses. This silencing is stable between cell generations as cytosine methylation is maintained epigenetically through DNA replication. The Arabidopsis thaliana Dnmt3 cytosine methyltransferase ortholog DOMAINS rearranged methyltransferase2 (DRM2) is required for establishment of small interfering RNA (siRNA) directed DNA methylation. In mammals PIWI proteins and piRNA act in a convergently evolved RNA-directed DNA methylation system that is required to repress transposon expression in the germ line. De novo methylation may also be independent of RNA interference and small RNAs, as in Neurospora crassa. Here we identify a clade of catalytically mutated DRM2 paralogs in flowering plant genomes, which in A.thaliana we term domains rearranged methyltransferase3 (DRM3). Despite being catalytically mutated, DRM3 is required for normal maintenance of non-CG DNA methylation, establishment of RNA-directed DNA methylation triggered by repeat sequences and accumulation of repeat-associated small RNAs. Although the mammalian catalytically inactive Dnmt3L paralogs act in an analogous manner, phylogenetic analysis indicates that the DRM and Dnmt3 protein families diverged independently in plants and animals. We also show by site-directed mutagenesis that both the DRM2 N-terminal UBA domains and C-terminal methyltransferase domain are required for normal RNA-directed DNA methylation, supporting an essential targeting function for the UBA domains. These results suggest that plant and mammalian RNA-directed DNA methylation systems consist of a combination of ancestral and convergent features.
Department of Molecular, Cell, and Developmental Biology, University of California Los Angeles, Los Angeles, California, United States of America.
Ian R
IR
Henderson
Angelique
A
Deleris
William
W
Wong
Xuehua
X
Zhong
Hang Gyeong
HG
Chin
Gregory A
GA
Horwitz
Krystyna A
KA
Kelly
Sriharsa
S
Pradhan
Steven E
SE
Jacobsen
D000818:Animals; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D055162:Biocatalysis; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation; D015257:DNA-Cytosine Methylases / Q000235:genetics* / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008780:Methyltransferases / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D010802:Phylogeny; D030821:Plants, Genetically Modified; D034622:RNA Interference; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D029681:Arabidopsis Proteins; D007527:Isoenzymes; D018749:RNA, Plant; D015257:DNA-Cytosine Methylases; C556304:DRM3 protein, Arabidopsis; D008780:Methyltransferases; D004248:DNA (Cytosine-5-)-Methyltransferases; C470732:DRM2 protein, Arabidopsis