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Nature Communications Feb 2020PIWI-clade Argonaute proteins associate with PIWI-interacting RNAs (piRNAs), and silence transposons in animal gonads. Here, we report the crystal structure of the...
PIWI-clade Argonaute proteins associate with PIWI-interacting RNAs (piRNAs), and silence transposons in animal gonads. Here, we report the crystal structure of the Drosophila PIWI-clade Argonaute Piwi in complex with endogenous piRNAs, at 2.9 Å resolution. A structural comparison of Piwi with other Argonautes highlights the PIWI-specific structural features, such as the overall domain arrangement and metal-dependent piRNA recognition. Our structural and biochemical data reveal that, unlike other Argonautes including silkworm Siwi, Piwi has a non-canonical DVDK tetrad and lacks the RNA-guided RNA cleaving slicer activity. Furthermore, we find that the Piwi mutant with the canonical DEDH catalytic tetrad exhibits the slicer activity and readily dissociates from less complementary RNA targets after the slicer-mediated cleavage, suggesting that the slicer activity could compromise the Piwi-mediated co-transcriptional silencing. We thus propose that Piwi lost the slicer activity during evolution to serve as an RNA-guided RNA-binding platform, thereby ensuring faithful co-transcriptional silencing of transposons.
Topics: Animals; Argonaute Proteins; Bombyx; Cell Line; Crystallography, X-Ray; DNA Transposable Elements; Drosophila; Drosophila Proteins; Drosophila melanogaster; Gene Silencing; Hydrogen Bonding; Models, Molecular; Protein Conformation; Protein Domains; RNA, Small Interfering; RNA, Untranslated; RNA, Guide, CRISPR-Cas Systems
PubMed: 32051406
DOI: 10.1038/s41467-020-14687-1 -
RNA Biology Jan 2022PIWI proteins and their associated PIWI-interacting RNAs (piRNAs) constitute a small RNA-based adaptive immune system that restricts the deleterious activity of mobile... (Review)
Review
PIWI proteins and their associated PIWI-interacting RNAs (piRNAs) constitute a small RNA-based adaptive immune system that restricts the deleterious activity of mobile genetic elements to protect genome integrity. Self/nonself discrimination is at the very core of successful defence and relies on complementary base-pairing in RNA-guided immunity. How the millions of piRNA sequences faithfully discriminate between self and nonself and how they adapt to novel genomic invaders remain key outstanding questions in genome biology. This review aims to introduce principles of piRNA silencing in the context of metazoan small RNA pathways. A distinct feature of piRNAs is their origin from single-stranded instead of double-stranded RNA precursors, and piRNAs require a unique set of processing factors. Novel nucleases, helicases and RNA binding proteins have been identified in piRNA biology, and while we are starting to understand some mechanisms of piRNA biogenesis and function, this diverse and prolific class of small RNAs remains full of surprises.
Topics: Animals; Argonaute Proteins; DNA Helicases; RNA Interference; RNA, Double-Stranded; RNA, Small Interfering; RNA-Binding Proteins
PubMed: 36217279
DOI: 10.1080/15476286.2022.2132359 -
Microbiology Spectrum Jun 2023Prokaryotic Argonaute (pAgo) proteins are guide-dependent nucleases that function in host defense against invaders. Recently, it was shown that TtAgo from Thermus...
Prokaryotic Argonaute (pAgo) proteins are guide-dependent nucleases that function in host defense against invaders. Recently, it was shown that TtAgo from Thermus thermophilus also participates in the completion of DNA replication by decatenating chromosomal DNA. Here, we show that two pAgos from cyanobacteria Synechococcus elongatus (SeAgo) and Limnothrix rosea (LrAgo) are active in heterologous Escherichia coli and aid cell division in the presence of the gyrase inhibitor ciprofloxacin, depending on the host double-strand break repair machinery. Both pAgos are preferentially loaded with small guide DNAs (smDNAs) derived from the sites of replication termination. Ciprofloxacin increases the amounts of smDNAs from the termination region and from the sites of genomic DNA cleavage by gyrase, suggesting that smDNA biogenesis depends on DNA replication and is stimulated by gyrase inhibition. Ciprofloxacin enhances asymmetry in the distribution of smDNAs around Chi sites, indicating that it induces double-strand breaks that serve as a source of smDNA during their processing by RecBCD. While active in E. coli, SeAgo does not protect its native host from ciprofloxacin. These results suggest that pAgo nucleases may help to complete replication of chromosomal DNA by promoting chromosome decatenation or participating in the processing of gyrase cleavage sites, and may switch their functional activities depending on the host species. Prokaryotic Argonautes (pAgos) are programmable nucleases with incompletely understood functions . In contrast to eukaryotic Argonautes, most studied pAgos recognize DNA targets. Recent studies suggested that pAgos can protect bacteria from invader DNA and counteract phage infection and may also have other functions including possible roles in DNA replication, repair, and gene regulation. Here, we have demonstrated that two cyanobacterial pAgos, SeAgo and LrAgo, can assist DNA replication and facilitate cell division in the presence of topoisomerase inhibitors in Escherichia coli. They are specifically loaded with small guide DNAs from the region of replication termination and protect the cells from the action of the gyrase inhibitor ciprofloxacin, suggesting that they help to complete DNA replication and/or repair gyrase-induced breaks. The results show that pAgo proteins may serve as a backup to topoisomerases under conditions unfavorable for DNA replication and may modulate the resistance of host bacterial strains to antibiotics.
Topics: Bacterial Proteins; Escherichia coli; Argonaute Proteins; Topoisomerase Inhibitors; Bacteria; Ciprofloxacin; DNA; Cell Division
PubMed: 37102866
DOI: 10.1128/spectrum.04146-22 -
Epigenomics 2015Epigenetic mechanisms work in an orchestrated fashion to control gene expression in both homeostasis and diseases. Among small noncoding RNAs, piRNAs seem to meet the... (Review)
Review
Epigenetic mechanisms work in an orchestrated fashion to control gene expression in both homeostasis and diseases. Among small noncoding RNAs, piRNAs seem to meet the necessary requirements to be included in this epigenetic network due to their role in both transcriptional and post-transcriptional regulation. piRNAs and PIWI proteins might play important roles in cancer occurrence, prognosis and treatment as reported previously. Nevertheless, the potential clinical relevance of these molecules has yet been elucidated. A brief overview of piRNA biogenesis and their potential roles as part of an epigenetic network that is possibly involved in cancer is provided. Moreover, potential strategies based on the use of piRNAs and PIWI proteins as diagnostic and prognostic biomarkers as well as for cancer therapeutics are discussed.
Topics: Argonaute Proteins; Biomarkers; Epigenesis, Genetic; Epigenomics; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Molecular Targeted Therapy; Neoplasms; Organ Specificity; Prognosis; RNA Interference; RNA, Small Interfering; RNA, Untranslated; Transcriptome
PubMed: 25929784
DOI: 10.2217/epi.15.37 -
Trends in Biochemical Sciences Jan 2023MicroRNAs (miRNAs) post-transcriptionally repress gene expression by guiding Argonaute (AGO) proteins to target mRNAs. While much is known about the regulation of miRNA... (Review)
Review
MicroRNAs (miRNAs) post-transcriptionally repress gene expression by guiding Argonaute (AGO) proteins to target mRNAs. While much is known about the regulation of miRNA biogenesis, miRNA degradation pathways are comparatively poorly understood. Although miRNAs generally exhibit slow turnover, they can be rapidly degraded through regulated mechanisms that act in a context- or sequence-specific manner. Recent work has revealed a particularly important role for specialized target interactions in controlling rates of miRNA degradation. Engagement of these targets is associated with the addition and removal of nucleotides from the 3' ends of miRNAs, a process known as tailing and trimming. Here we review these mechanisms of miRNA modification and turnover, highlighting the contexts in which they impact miRNA stability and discussing important questions that remain unanswered.
Topics: MicroRNAs; Argonaute Proteins; RNA Stability; Nucleotides; RNA, Messenger
PubMed: 35811249
DOI: 10.1016/j.tibs.2022.06.005 -
Database : the Journal of Biological... Sep 2022Argonaute (Ago) proteins are widely expressed in almost all organisms. Eukaryotic Ago (eAgo) proteins bind small RNA guides forming RNA-induced silencing complex that...
UNLABELLED
Argonaute (Ago) proteins are widely expressed in almost all organisms. Eukaryotic Ago (eAgo) proteins bind small RNA guides forming RNA-induced silencing complex that silence gene expression, and prokaryotic Ago (pAgo) proteins defend against invading nucleic acids via binding small RNAs or DNAs. pAgo proteins have shown great potential as a candidate 'scissors' for gene editing. Protein domains are fundamental units of protein structure, function and evolution; however, the domains of Ago proteins are not well annotated/curated currently. Therefore, full functional domain annotation of Ago proteins is urgently needed for researchers to understand the function and mechanism of Ago proteins. Herein, we constructed the first comprehensive domain annotation database of Ago proteins (AGODB). The database curates detailed information of 1902 Ago proteins, including 1095 eAgos and 807 pAgos. Especially for long pAgo proteins, all six domains are annotated and curated. Gene Ontology (GO) enrichment analysis revealed that Ago genes in different species were enriched in the following GO terms: biological processes (BPs), molecular function and cellular compartment. GO enrichment analysis results were integrated into AGODB, which provided insights into the BP that Ago genes may participate in. AGODB also allows users to search the database with a variety of options and download the search results. We believe that the AGODB will be a useful resource for understanding the function and domain components of Ago proteins. This database is expected to cater to the needs of scientific community dedicated to the research of Ago proteins.
DATABASE URL
http://i.uestc.edu.cn/agodb/.
Topics: Argonaute Proteins; DNA; Eukaryota
PubMed: 36068786
DOI: 10.1093/database/baac078 -
Human Gene Therapy Jun 2016Genome engineering has gone mainstream because of breakthroughs in defining and harnessing naturally occurring, customizable DNA recognition cursors (protein or... (Review)
Review
Genome engineering has gone mainstream because of breakthroughs in defining and harnessing naturally occurring, customizable DNA recognition cursors (protein or RNA-guided). At present, most gene editing relies on these cursors to direct custom DNA endonucleases to a specific genomic sequence to induce a double-strand break. New tools for genome engineering are continuously being explored, and another advance in DNA targeting has recently been described. Argonaute isolated from Natronobacterium gregoryi (NgAgo) is an ssDNA-based cursor that thus far has no known limitations in sequence recognition, shows promise for high specificity, and for many applications may represent a potentially more accessible genome-editing system over prior tools as it requires only a single, 24-base, 5' phosphorylated ssDNA for DNA targeting. Genome engineering is in a remarkable moment of unprecedented growth with exponential reduction in costs reminiscent of Moore's law in electronics. Many questions remain with regard to Argonaute utility in specific systems, but there is no doubt that genome engineering is expanding into new and exciting areas from synthetic biology to gene therapy.
Topics: Animals; Argonaute Proteins; DNA, Single-Stranded; Gene Editing; Gene Targeting; Humans
PubMed: 27230540
DOI: 10.1089/hum.2016.071 -
Current Opinion in Plant Biology Oct 2015ARGONAUTES (AGOs) are the effector proteins functioning in eukaryotic RNA silencing pathways. AGOs associate with small RNAs and are programmed to target complementary... (Review)
Review
ARGONAUTES (AGOs) are the effector proteins functioning in eukaryotic RNA silencing pathways. AGOs associate with small RNAs and are programmed to target complementary RNA or DNA. Plant viruses induce a potent and specific antiviral RNA silencing host response in which AGOs play a central role. Antiviral AGOs associate with virus-derived small RNAs to repress complementary viral RNAs or DNAs, or with endogenous small RNAs to regulate host gene expression and promote antiviral defense. Here, we review recent progress towards understanding the roles of plant AGOs in antiviral defense. We also discuss the strategies that viruses have evolved to modulate, attenuate or suppress AGO antiviral functions.
Topics: Antiviral Agents; Argonaute Proteins; Gene Expression Regulation, Plant; Plant Diseases; Plant Proteins; Plant Viruses; RNA, Small Interfering; RNA, Viral
PubMed: 26190744
DOI: 10.1016/j.pbi.2015.06.013 -
The Journal of Pharmacology and... Jan 2023Argonautes (AGOs) are a highly conserved family of proteins found in most eukaryotes and involved in mechanisms of gene regulation, both at the transcriptional and... (Review)
Review
Argonautes (AGOs) are a highly conserved family of proteins found in most eukaryotes and involved in mechanisms of gene regulation, both at the transcriptional and post-transcriptional level. Among other functions, AGO proteins associate with microRNAs (miRNAs) to mediate the post-transcriptional repression of protein-coding genes. In this process, AGOs associate with members of the trinucleotide repeat containing 6 protein (TNRC6) family to form the core of the RNA-induced silencing complex (RISC), the effector machinery that mediates miRNA function. However, the description of the exact composition of the RISC has been a challenging task due to the fact the AGO's interactome is dynamically regulated in a cell type- and condition-specific manner. Here, we summarize some of the most significant studies that have identified AGO complexes in mammalian cells, as well as the approaches used to characterize them. Finally, we discuss possible opportunities to exploit what we have learned on the properties of the RISC to develop novel anti-cancer therapies. SIGNIFICANCE STATEMENT: The RNA-induced silencing complex (RISC) is the molecular machinery that mediates miRNA function in mammals. Studies over the past two decades have shed light on important biochemical and functional properties of this complex. However, many aspects of this complex await further elucidation, mostly due to technical limitations that have hindered full characterization. Here, we summarize some of the most significant studies on the mammalian RISC and discuss possible sources of biases in the approaches used to characterize it.
Topics: Animals; Argonaute Proteins; MicroRNAs; RNA-Induced Silencing Complex; Gene Expression Regulation; Mammals
PubMed: 35667689
DOI: 10.1124/jpet.122.001158 -
Cold Spring Harbor Perspectives in... Oct 2011RNAi has existed at least since the divergence of prokaryotes and eukaryotes. This collection of pathways responds to a diversity of "abberant" RNAs and generally... (Review)
Review
RNAi has existed at least since the divergence of prokaryotes and eukaryotes. This collection of pathways responds to a diversity of "abberant" RNAs and generally silences or eliminates genes sharing sequence content with the silencing trigger. In the canonical pathway, double-stranded RNAs are processed into small RNAs, which guide effector complexes to their targets by complementary base pairing. Many alternative routes from silencing trigger to small RNA are continuously being uncovered. Though the triggers of the pathway and the mechanisms of small RNA production are many, all RNAi-related mechanisms share Argonaute proteins as the heart of their effector complexes. These can act as self-contained silencing machines, binding directly to small RNAs, carrying out homology-based target recognition, and in some cases cleaving targets using an endogenous nuclease domain. Here, we discuss the diversity of Argonaute proteins from a structural and functional perspective.
Topics: Argonaute Proteins; Conserved Sequence; Evolution, Molecular; Models, Genetic; Models, Molecular; Phylogeny; Protein Structure, Tertiary; RNA Interference; RNA, Small Interfering; RNA-Induced Silencing Complex
PubMed: 20810548
DOI: 10.1101/cshperspect.a003772