The N-terminal extension of Arabidopsis ARGONAUTE 1 is essential for microRNA activities [ncRNA_seq]
Ontology highlight
ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE212364 | GEO | 2023/02/21
REPOSITORIES: GEO
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Project description:We profiled the small RNA of AGO1 NTE truncation mutant lines, and the small RNA associated with AGO1 NTE truncation mutant to study the roles of the NTE region in Arabidopsis AGO1 involved RNA-induced silencing complex (RISC) formation.
2023-02-21 | GSE224480 | GEO
Project description:We profiled gene expression of AGO1 NTE truncation mutant lines to study the roles of the NTE region in Arabidopsis AGO1 involved RNA-induced silencing complex (RISC) formation.
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Project description:We profiled gene expression of AGO1 NTE truncation mutant lines to study the roles of the NTE region in Arabidopsis AGO1 involved RNA-induced silencing complex (RISC) formation.
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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.
2024-05-22 | PXD049971 | Pride
Project description:Leishmania donovani, an intracellular protozoan parasite, is the causative agent of visceral leishmaniasis or kala-azar, the most severe form of leishmaniasis in humans. To date, our understanding of the molecular mechanisms associated with the pathogenicity of Leishmania infection is still limited. RNA interference—collectively RNA silencing pathways—participates in the regulation of various biological processes in most eukaryotic cells. Complexes of Argonaute proteins with small RNAs are core components of the RNA interference system and play a key role in silencing gene expression. It is becoming increasingly clear that several intracellular pathogens target host cell RNA interference pathways to promote their survival. In this study, we investigated the potential role of host macrophage Argonautes in Leishmania pathogenesis. Western blot analysis showed that protein abundance of infected macrophage Argonaute 1 (Ago1) was selectively and significantly higher than that of non-infected control at 24 h post-infection, suggesting that Ago1 plays a role in pathogenicity. In fact, siRNA-mediated downregulation of Ago1 enhanced Leishmania clearance from infected host cells, linking macrophage Ago1 to Leishmania virulence. To investigate the mechanisms of host Ago1 in Leishmania pathogenesis, a stable isotope labeling by amino acids in cell culture (SILAC)-based whole proteome approach was employed, which showed that expression of several previously reported Leishmania pathogenesis-related proteins were dependent on the level of macrophage Ago1. Moreover, the proteomic-based detailed biochemical analysis showed that Leishmania modulated host RNA-induced silencing complex (RISC) composition during infection, strongly suggesting macrophage RISC targeting. Strikingly, Leishmania proteins were detected as part of host RISC in infected cells. Together, our results demonstrate that Leishmania targets host RNA interference machinery to promote its survival inside the host macrophage.
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Project description:Small RNAs regulate the genetic networks through a ribonucleoprotein complex called the RNA induced silencing complexes (RISC), which in mammals contains at its center one of four Argonaute proteins (Ago1-4). A key regulatory event in the RNAi and miRNA pathways is Ago loading, where double stranded small RNA duplexes are incorporated into RISC (pre-RISC) and then become single stranded (mature-RISC), a process that is not well understood. The Agos contain an evolutionary conserved PAZ (Piwi/Argonaute/Zwille) domain whose primary function is to bind the 3’-end of small RNAs. We created multiple Paz domain disrupted Ago mutant proteins and studied their biochemical properties and biological functionality in cells. We found that the Paz domain is dispensable for Ago loading of slicing-competent RISC. In contrast, in the absence of slicer activity or slicer substrate duplex RNAs, Paz-disrupted Agos bound duplex siRNAs but were unable to unwind/eject the passenger strand and form functional RISC complexes. We have discovered that the highly conserved Paz domain plays an important role in RISC activation, providing new mechanistic insights into how miRNAs regulate genes, as well as new insights for future design of miRNA and RNAi-based therapeutics.
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Project description:Small RNAs regulate the genetic networks through a ribonucleoprotein complex called the RNA induced silencing complexes (RISC), which in mammals contains at its center one of four Argonaute proteins (Ago1-4). A key regulatory event in the RNAi and miRNA pathways is Ago loading, where double stranded small RNA duplexes are incorporated into RISC (pre-RISC) and then become single stranded (mature-RISC), a process that is not well understood. The Agos contain an evolutionary conserved PAZ (Piwi/Argonaute/Zwille) domain whose primary function is to bind the 3’-end of small RNAs. We created multiple Paz domain disrupted Ago mutant proteins and studied their biochemical properties and biological functionality in cells. We found that the Paz domain is dispensable for Ago loading of slicing-competent RISC. In contrast, in the absence of slicer activity or slicer substrate duplex RNAs, Paz-disrupted Agos bound duplex siRNAs but were unable to unwind/eject the passenger strand and form functional RISC complexes. We have discovered that the highly conserved Paz domain plays an important role in RISC activation, providing new mechanistic insights into how miRNAs regulate genes, as well as new insights for future design of miRNA and RNAi-based therapeutics. Various Argonautes associated small RNA profiles were generated by deep sequencing the Agos-IP samples in HEK293 Cells transfected with corresponding Argonaute.
2012-07-12 | E-GEOD-39146 | biostudies-arrayexpress
Project description:Argonuate proteins are the central components of the RNA-induced silencing complex (RISC) which execute small RNA-mediated gene regulation mainly in the cytoplasm and their nuclear functions in mammalian cells remain largely unknown. Here we show that in human cancer cells Ago1 is pervasively associated with chromosomal loci throughout the genome with the association centered mostly on actively transcribed promoters. Ago1 but not Ago2 physically interacts with RNA polymerase II (RNAP II) in the nucleus. Moreover, genes bound by Ago1 are significantly enriched for several cancer related pathways. Detailed analysis of these genes including PIK3CA, PRKCH, CDC6 and RRM1 verified Ago1-mediated transcriptional gene regulation through Ago1-promoter interactions. We also found a strong correlation between Ago1-bound genes and genes downregulated by Ago1 depletion which, phenotypically, leads to inhibition of proliferation in different cancer cells. Therefore, our findings reveal the first landscape of human Ago1-chromosomal interactions which may drive oncogenic programs in cancer cells Examination of Ago-DNA interactions in human cancer cells
2013-09-27 | E-GEOD-40536 | biostudies-arrayexpress
Project description:Argonuate proteins are the central components of the RNA-induced silencing complex (RISC) which execute small RNA-mediated gene regulation mainly in the cytoplasm and their nuclear functions in mammalian cells remain largely unknown. Here we show that in human cancer cells Ago1 is pervasively associated with chromosomal loci throughout the genome with the association centered mostly on actively transcribed promoters. Ago1 but not Ago2 physically interacts with RNA polymerase II (RNAP II) in the nucleus. Moreover, genes bound by Ago1 are significantly enriched for several cancer related pathways. Detailed analysis of these genes including PIK3CA, PRKCH, CDC6 and RRM1 verified Ago1-mediated transcriptional gene regulation through Ago1-promoter interactions. We also found a strong correlation between Ago1-bound genes and genes downregulated by Ago1 depletion which, phenotypically, leads to inhibition of proliferation in different cancer cells. Therefore, our findings reveal the first landscape of human Ago1-chromosomal interactions which may drive oncogenic programs in cancer cells
2013-09-27 | GSE40536 | GEO
Project description:microRNAs (miRNAs) regulate target gene expression through their ARGONAUTE (AGO) effector protein, mainly AGO1 in Arabidopsis thaliana. In addition to the highly conserved N, PAZ, MID and PIWI domains with known roles in RNA silencing, AGO1 contains a long, unstructured N-terminal extension (NTE) of little-known function. Here, we show that the NTE is indispensable for the functions of Arabidopsis AGO1, as a lack of the NTE leads to seedling lethality. Within the NTE, the region containing amino acids (a.a.) 91 to 189 is essential for rescuing an ago1 null mutant. Through global analyses of small RNAs, AGO1-associated small RNAs, and miRNA target gene expression, we show that the region containing a.a. 91-189 is required for the loading of miRNAs into AGO1. Moreover, we show that reduced nuclear partitioning of AGO1 did not affect its profiles of miRNA and ta-siRNA association. Furthermore, we show that the 1-to-90a.a. and 91-to-189a.a. regions of the NTE redundantly promote the activities of AGO1 in the biogenesis of trans-acting siRNAs. Together, we report novel roles of the NTE of Arabidopsis AGO1.
| S-EPMC9994745 | biostudies-literature