Abstract
DNA methylation directed by 24-nucleotide small RNAs involves the small RNA-binding protein ARGONAUTE4 (AGO4), and it was previously shown that AGO4 localizes to nucleolus-adjacent Cajal bodies, sites of snRNP complex maturation. Here we demonstrate that AGO4 also localizes to a second class of nuclear bodies, called AB-bodies, which are found immediately adjacent to condensed 45S ribosomal DNA (rDNA) sequences. AB-bodies also contain other proteins involved in RNA-directed DNA methylation including NRPD1b (a subunit of the RNA Polymerase IV complex, RNA PolIV), NRPD2 (a second subunit of this complex), and the DNA methyltransferase DRM2. These two classes of AGO4 bodies are structurally independent—disruption of one class does not affect the other—suggesting a dynamic regulation of AGO4 within two distinct nuclear compartments in Arabidopsis. Abolishing Cajal body formation in a coilin mutant reduced overall AGO4 protein levels, and coilin dicer-like3 double mutants showed a small decrease in DNA methylation beyond that seen in dicer-like3 single mutants, suggesting that Cajal bodies are required for a fully functioning DNA methylation system in Arabidopsis.
Highlights
RNA-directed DNA methylation (RdDM) is a phenomenon discovered in plants in which small RNAs cause DNA methylation and transcriptional gene silencing at complementary sequences in the genome [1,2,3]
The final step of the RdDM pathway is performed by DOMAINS REARRANGED METHYLTRANSFERASE2 (DRM2), a de novo DNA methyltransferase that is required for the establishment of DNA methylation and for the maintenance of DNA methylation in asymmetric CHH and CHG sequence contexts [12,16]
In the flowering plant Arabidopsis thaliana, ARGONAUTE4 (AGO4) is involved in gene silencing at the transcriptional level in a process called RNA-directed DNA methylation (RdDM), during which small interfering RNAs (siRNAs) cause transcriptional gene repression at complementary sequences
Summary
RNA-directed DNA methylation (RdDM) is a phenomenon discovered in plants in which small RNAs cause DNA methylation and transcriptional gene silencing at complementary sequences in the genome [1,2,3]. The final step of the RdDM pathway is performed by DOMAINS REARRANGED METHYLTRANSFERASE2 (DRM2), a de novo DNA methyltransferase that is required for the establishment of DNA methylation and for the maintenance of DNA methylation in asymmetric CHH (where H1⁄4A, T, or C) and CHG sequence contexts [12,16]. Mutations in these RdDM genes result in decreased transcriptional silencing of certain repetitive loci
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