ARGONAUTE 6 bridges transposable element m RNA ‐derived si RNA s to the establishment of DNA methylation

Abstract

Transposable elements (TEs) generate mutations and chromosomal instability when active. To repress TE activity, eukaryotic cells evolved mechanisms to both degrade TE mRNAs into small interfering RNAs (siRNAs) and modify TE chromatin to epigenetically inhibit transcription. Since the populations of small RNAs that participate in TE post-transcriptional regulation differ from those that establish RNA-directed DNA methylation (RdDM), the mechanism through which transcriptionally active TEs transition from post-transcriptional RNAi regulation to chromatin level control has remained unclear. We have identified the molecular mechanism of a plant pathway that functions to direct DNA methylation to transcriptionally active TEs. We demonstrated that 21-22 nucleotide (nt) siRNA degradation products from the RNAi of TE mRNAs are directly incorporated into the ARGONAUTE 6 (AGO6) protein and direct AGO6 to TE chromatin to guide its function in RdDM. We find that this pathway functions in reproductive precursor cells to primarily target long centromeric high-copy transcriptionally active TEs for RdDM prior to gametogenesis. This study provides a direct mechanism that bridges the gap between the posttranscriptional regulation of TEs and the establishment of TE epigenetic silencing.

RNAi in Plants: An Argonaute-Centered View

“…It was also shown that AGO4 and AGO6 differ in their subnuclear colocalization and in interactions with Pol II and Pol IV, and it was proposed that they may act sequentially to mediate DNA methylation . AGO6 also binds RDR6-dependent 21-to 22-nucleotide siRNAs generated from transcriptionally active TEs and is thought to trigger the initiation of de novo TE DNA methylation (Nuthikattu et al, 2013;McCue et al, 2015), which may provide a link between PTGS and TGS of TEs.…”

The First Rule of Plant Transposable Element Silencing: Location, Location, Location

“…Although it is well established that 24-nucleotide siRNAs can target RdDM, recently other types of siRNAs have also been implicated in directing RdDM. Diced 21-to 22-nucleotide TE siRNAs incorporate into AGO6 to target RdDM to active TEs (McCue et al, 2015), and when the Dicer proteins are absent, Dicer-independent longer RNAs can incorporate into AGO4 and then be subsequently trimmed to the 21-to 24-nucleotide size (Ye et al, 2016). Targeting of RdDM acts through an RNA scaffold at the target locus produced by the plant-specific Pol V, which transcribes methylated regions ( Figure 2A; Johnson et al, 2014).…”

“…Several articles have recently described two mutually exclusive classes of TEs in Arabidopsis: one where CHH methylation is maintained by CMT2 and the other where CHH methylation is constantly targeted via RdDM ( Figure 5) (Stroud et al, 2014;Li et al, 2015;McCue et al, 2015). These two distinct classes represent (1) TEs present in large constitutive heterochromatic blocks found in the pericentromere, knobs, and TE islands regulated via CMT2 and (2) TEs near genes regulated by RdDM and the DNA methyltransferase DRM2 (see above).…”

“…How the H3K9me2 mark is differentially interpreted and leads to either CMT2-based CHH methylation or DRM2/RdDM-based CHH methylation is currently unknown and represents a key branch point in TE silencing ( Figure 5). This distinction is not primarily due to TE type or location, as a silenced TE regulated by CMT2 can switch to RdDM regulation upon transcriptional activation (McCue et al, 2015). In addition, although they are not targeted via RdDM, pericentromeres, knobs, and TE islands are thought to be transcribed by RNA Pol IV because they still produce 24-nucleotide siRNAs (Law et al, 2013;Li et al, 2015).…”

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

“…AGO6 mediates RNA-directed DNA methylation Havecker et al, 2010;Eun et al, 2011;McCue et al, 2015). AGO6 participates in RNA-directed DNA methylation of transcriptionally active transposable elements through incorporation of fragments of PTGS-targeted transcripts of transposable elements onto AGO6 (McCue et al, 2015).…”