The m 6 A pathway protects the transcriptome integrity by restricting RNA chimera formation in plants

International audienceGlobal, segmental, and gene duplication-related processes are driving genome size and complexity in plants. Despite their evolutionary potentials, those processes can also have adverse effects on genome regulation, thus implying the existence of specialized corrective mechanisms. Here, we report that an N6-methyladenosine (m 6 A)-assisted polyadenylation (m-ASP) pathway ensures tran-scriptome integrity in Arabidopsis thaliana. Efficient m-ASP pathway activity requires the m 6 A methyltransferase-associated factor FIP37 and CPSF30L, an m 6 A reader corresponding to an YT512-B Homology Domain-containing protein (YTHDC)-type domain containing isoform of the 30-kD subunit of cleavage and polyadenylation specificity factor. Targets of the m-ASP pathway are enriched in recently rearranged gene pairs, displayed an atypical chromatin signature, and showed transcriptional readthrough and mRNA chimera formation in FIP37-and CPSF30L-deficient plants. Furthermore, we showed that the m-ASP pathway can also restrict the formation of chimeric gene/transposable-element transcript, suggesting a possible implication of this pathway in the control of transposable elements at specific locus. Taken together, our results point to selective recognition of 39-UTR m 6 A as a safeguard mechanism ensuring transcriptome integrity at rearranged genomic loci in plants

A Plant-Specific Transcription Factor IIB-Related Protein, pBRP2, Is Involved in Endosperm Growth Control

General transcription factor IIB (TFIIB) and TFIIB-related factor (BRF), are conserved RNA polymerase II/III (RNAPII/III) selectivity factors that are involved in polymerase recruitment and transcription initiation in eukaryotes. Recent findings have shown that plants have evolved a third type of B-factor, plant-specific TFIIB-related protein 1 (pBRP1), which seems to be involved in RNAPI transcription. Here, we extend the repertoire of B-factors in plants by reporting the characterization of a novel TFIIB-related protein, plant-specific TFIIB-related protein 2 (pBRP2), which is found to date only in the Brassicacea family. Unlike other B-factors that are ubiquitously expressed, PBRP2 expression is restricted to reproductive organs and seeds as shown by RT-PCR, immunofluorescence labelling and GUS staining experiments. Interestingly, pbrp2 loss-of-function specifically affects the development of the syncytial endosperm, with both parental contributions required for wild-type development. pBRP2, is the first B-factor to exhibit cell-specific expression and regulation in eukaryotes, and might play a role in enforcing bi-parental reproduction in angiosperms

RNA-directed DNA methylation requires an AGO4-interacting member of the SPT5 elongation factor family

Recent studies have identified a conserved WG/GW-containing motif, known as the Argonaute (AGO) hook, which is involved in the recruitment of AGOs to distinct components of the eukaryotic RNA silencing pathways. By using this motif as a model to detect new components in plant RNA silencing pathways, we identified SPT5-like, a plant-specific AGO4-interacting member of the nuclear SPT5 (Suppressor of Ty insertion 5) RNA polymerase (RNAP) elongation factor family that is characterized by the presence of a carboxy-terminal extension with more than 40 WG/GW motifs. Knockout SPT5-like mutants show a decrease in the accumulation of several 24-nt RNAs and hypomethylation at different loci revealing an implication in RNA-directed DNA methylation (RdDM). Here, we propose that SPT5-like emerged in plants as a facultative RNAP elongation factor. Its plant-specific origin and role in RdDM might reflect functional interactions with plant-specific RNA Pols required for RdDM

Reiterated WG/GW motifs form functionally and evolutionarily conserved ARGONAUTE-binding platforms in RNAi-related components

Two forms of RNA Polymerase IV (PolIVa/PolIVb) have been implicated in RNA-directed DNA methylation (RdDM) in Arabidopsis. Prevailing models imply a distinct function for PolIVb by association of Argonaute4 (AGO4) with the C-terminal domain (CTD) of its largest subunit NRPD1b. Here we show that the extended CTD of NRPD1b-type proteins exhibits conserved Argonaute-binding capacity through a WG/GW-rich region that functionally distinguishes Pol IVb from Pol IVa, and that is essential for RdDM. Site-specific mutagenesis and domain-swapping experiments between AtNRPD1b and the human protein GW182 demonstrated that reiterated WG/GW motifs form evolutionarily and functionally conserved Argonaute-binding platforms in RNA interference (RNAi)-related components

Evidence for ARGONAUTE4–DNA interactions in RNA-directed DNA methylation in plants

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