SPT5L also implicated in target locus interactions ( Li et al. 2018), with protein–protein interactions between AGO4 and the Pol V largest subunit (NRPE1) and/or the Pol V-associated protein Loci by base-pairing to transcripts generated by nuclear RNA polymerase V (Pol V) ( Wierzbicki et al. The siRNAs then guide AGO4 to RdDM target 2022), one strand of which ultimately becomes stably associated with ARGONAUTE 4 (AGO4). The resulting dicing products consist of a 24-nt siRNA strand paired with either a 23-nt strand (24/23 duplexes) or a 24-nt DICER-LIKE 3 (DC元) then cuts the dsRNAs into siRNAs, guided by sequence and structural features at the ends of precursorĭuplexes ( Loffer et al. These enzymes physically associate and carry out coupled reactions that transcribe target locus DNA sequences into double-stranded 2009 Haag and Pikaard 2011) and RNA-dependent RNA polymerase 2 (RDR2) ( Zhou and Law 2015 Fukudome et al. RdDM siRNA precursors are synthesized by DNA-dependent nuclear RNA polymerase IV (Pol IV) ( Ream et al. RNA-directed DNA methylation (RdDM) is the major transcriptional gene silencing pathway in plants ( Matzke and Mosher 2014 Zhou and Law 2015 Wendte and Pikaard 2017 Zhang et al. Of the targeted RNA at a phosphodiester bond ( Joshua-Tor and Hannon 2011). An ancestral and fundamental function shared by many Argonaute proteins is small RNA-programmed endonucleolytic slicing Transcription ( Martienssen and Moazed 2015 Iwakawa and Tomari 2022). With complementary target RNAs to interfere with RNA translation or to bring about chromatin modifications that repress gene In eukaryotes, RNA silencing plays a significant role in gene regulation, transposable element repression, and genome stability.Ī common theme among RNA silencing pathways is that small RNAs stably associate with Argonaute family proteins and then interact RNAs are distinct modes by which AGO4 slicing enhances RNA-directed DNA methylation.
We propose that siRNA passenger strand elimination and AGO4 tethering to sliced target In vivo, RdDM target locus RNAs that copurify with AGO4 also display a sequence signature of slicing.Ĭomparing plants expressing slicing-competent versus slicing-defective AGO4 shows that slicing elevates cytosine methylation Slicing recapitulated with recombinantĪGO4 and synthetic RNAs reveals that siRNAs of 21–24 nt, with any 5′-terminal nucleotide, can guide slicing, with sliced RNAs Sliced into 11- and 12-nt fragments, with 12-nt fragments remaining associated with AGO4. We show that 23-nt RNAs function as passenger strands during 24-nt siRNA incorporation into AGO4. RNA-directed DNA methylation in plants is guided by 24-nt siRNAs generated in parallel with 23-nt RNAs of unknown function.
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