Locus-Specific Requirements of DDR Complexes for Gene-Body Methylation of TAS Genes in Arabidopsis thaliana

Abstract

Posttranscriptional gene silencing via 21-nucleotide (nt) trans-acting small interfering RNAs (tasiRNAs) is a plant-specific RNA silencing mechanism. tasiRNAs are generated by RNA-DEPENDENT RNA POLYMERASE 6, SUPPRESSOR OF GENE SILENCING 3, and DICER-LIKE 4 using TAS transcripts as templates and degrade their target mRNA in a manner similar to micro-RNA. TAS gene-derived small RNAs also trigger cytosine methylation in cis together with known components of RNA-directed DNA methylation (RdDM). RdDM is known to be required for silencing of repetitive elements and transposons via a 24-nt small interfering RNA complementary to the target DNA. Plant-specific DNA-dependent RNA polymerase V (Pol V) is thought to be required for the recruitment of other RdDM components to the target region, together with the DDR complex, which contains DEFECTIVE IN RNA-DIRECTED DNA METHYLATION 1, DEFECTIVE IN MERISTEM SILENCING 3, and RNA-DIRECTED DNA METHYLATION 1. Although genome-wide studies indicate that Pol V and the DDR complex mostly share endogenous targets, the requirement for each component of RdDM at TAS genes remains obscure. Here, we investigated the involvement of components of the DDR complex in RdDM triggered by TAS gene-derived small RNAs. Our results show that methylation of the TAS3a gene was reduced in dms3 and drd1, whereas methylation of the TAS1c gene was decreased in dms3 but not in drd1. These findings indicate that Pol V and components of the DDR complex act separately in a locus-specific manner.