Project description:Conventional RNA interference (RNAi) pathways suppress eukaryotic gene expression at the post-transcriptional or transcriptional level. At the core of RNAi are small RNAs (sRNAs) and effector Argonaute proteins. Arabidopsis AGO1 is known to bind microRNAs (miRNAs) and post-transcriptionally repress target genes via mRNA cleavage and/or translational inhibition in the cytoplasm. Here, we report that AGO1 binds to the chromatin of transcriptionally active genes and promotes their transcription. We show that sRNAs and SWI/SNF are required for AGO1 binding to chromatin and its function in promoting gene transcription. Moreover, we show that various stimuli including plant hormones and stresses specifically trigger AGO1 binding to the stimulus-responsive genes. Finally, we show that AGO1 facilitates the induction of genes in the jasmonate (JA) signaling pathways and the activation of JA responses. Our findings reveal an unsuspected role for AGO1 in facilitating gene transcription in response to plant hormones and stresses.
Project description:In Arabidopsis thaliana, ARGONAUTE1 (AGO1) plays a central role[AQ1] in microRNA (miRNA) and small interfering RNA (siRNA)- mediated silencing and is a key component in antiviral responses. The polerovirus F-box P0 protein triggers AGO1 degradation as a viral counterdefense. Here, we identified a motif in AGO1 that is required for its interaction with the S phase kinase-associated protein1-cullin 1-F-box protein (SCF) P0 (SCFP0) complex and subsequent degradation. The AGO1 P0 degron is conserved and confers P0-mediated degradation to other AGO[AQ2] proteins. Interestingly, the degron motif is localized in the DUF1785 domain of AGO1, in which a single point mutation (ago1-57, obtained by forward genetic screening) compromises recognition by SCFP0. Recapitulating formation of the RNA-induced silencing complex in a cell-free system revealed that this mutation impairs RNA unwinding, leading to stalled forms of AGO1 still bound to double-stranded RNAs. In vivo, the DUF1785 is required for unwinding perfectly matched siRNA duplexes, but is mostly dispensable for unwinding imperfectly matched miRNA duplexes. Consequently, its mutation nearly abolishes phased siRNA production and sense transgene posttranscriptional gene silencing. Overall, our work sheds new light on the mode of AGO1 recognition by P0 and the in vivo function of DUF1785 in RNA silencing.
Project description:Plant ARGONAUTE (AGO) proteins play pivotal roles in gene expression regulation through small (s)RNA-guided mechanisms. Among the ten AGO proteins in Arabidopsis thaliana, AGO1 stands out as the main effector of post-transcriptional gene silencing. Intriguingly, a specific region of AGO1, its N-terminal extension (NTE), has gained prominence in recent studies linked to diverse regulatory functions, including subcellular localization, sRNA loading, and interactions with regulatory factors. In the realm of post-translational modifications (PTMs), little is known about arginine methylation in Arabidopsis AGOs. This study reveals a novel and intricate landscape of AGO1 methylation. Here, we have elucidated that NTEAGO1 undergoes symmetric arginine dimethylation on specific residues, and interacts with the methyltransferase PRMT5, which catalyzes its methylation. Notably, we observed that the lack of symmetric dimethylarginine has no discernible impact on AGO1's subcellular localization or miRNA loading capabilities. However, the absence of PRMT5 significantly alters the loading of a subgroup of sRNAs into AGO1 and reshapes the NTEAGO1 interactome. Importantly, our research extends beyond AGO1, illustrating that symmetric arginine dimethylation of NTEs is a common process across Arabidopsis AGOs, taking place in AGO1, AGO2, AGO3, and AGO5, deepening our understanding of PTMs in the intricate landscape of RNA-associated gene regulation.
Project description:Plant microRNAs (miRNAs) have been implicated in plant immunity. These mainly focusing Arabidopsis thaliana threatened by (hemi-)biotrophic pathogens such as the bacterial pathogen Pseudomonas syringae. Here, we show that the Arabidopsis miRNA pathway is important for defense responses against the necrotrophic fungus Alternaria brassicicola. The miRNA pathway mutant ago1 exhibits an exaggerated response when treated with A. brassicicola, proposing that AGO1 is positive regulator. We found a subset of Arabidopsis miRNAs that quickly change their expression and their abundance in AGO1 complexes in plants exposed to A. brassicicola. The miRNAs responding to pathogen treatment are mainly targeting genes encoding metabolic enzymes, proteins involved protein degradation or transposons. In case of miR163, A. brassicicola infection results in increased levels of miRNA precursors and preferential accumulation of an unspliced form of pri-miR163, suggesting that A. brassicicola infection changes the transcriptional and post-regulation of pri-miRNAs. miR163 acts as a negative regulator of plant defense because mir163 mutants are more resistant when treated with A. brassicicola. Taken together, our results reveal the existence of positively and negatively acting Arabidopsis miRNA modulating the defense responses against A. brassicicola and highlight the importance of host miRNAs in the interaction between plants and necrotrophic pathogens.
Project description:We report flg22 regulate the accumulation of AGO1-bound small RNA in arabidopsis. We find that a number of miRNAs are up- or down-regulated by flg22, a well-studied PAMP.
Project description:We profiled total sRNA and root cell layer specific AGO1-bound sRNA in ktn1 mutant to study the role of KTN1 in miRNA non-cell autonomous activiy in Arabidopsis.
Project description:AGO3 predominantly bound 24-nt sRNAs with 5â terminal adenine. The spectrum of AGO3-associated sRNAs was different from those bound to AGO2. By contrast, approximately 30% of AGO3-bound 24-nt sRNAs overlapped with those bound to AGO4 and over 60% of AGO3-associated 24-nt sRNA enriched loci were identical to those of AGO4. In addition, expression of AGO3 driven by AGO4 native promoter partially complemented AGO4 function and rescued DNA methylation defect in ago4-1 background. Examination of DNA methylation using bisulfite conversion of unmethylated cytosines in three genetic backgrounds of Arabidopsis thaliana.
Project description:We report flg22 regulate the accumulation of AGO1-bound small RNA in arabidopsis. We find that a number of miRNAs are up- or down-regulated by flg22, a well-studied PAMP. Examination of AGO1-bound small RNAs with or without flg22 treatment.