In recent years, numerous evidence has been accumulated about the extent of A-to-I editing in human RNAs and the key role ADAR1 plays in the cellular editing machinery. It has been shown that A-to-I editing occurrence and frequency are tissue-specific and essential for some tissue development, such as the liver. To study the effect of ADAR1 function in hepatocytes, we have created Huh7.5 ADAR1 KO cell lines. Upon IFN treatment, the Huh7.5 ADAR1 KO cells show rapid arrest of growth and translation, from which they do not recover. We analyzed translatome changes by using a method based on sequencing of separate polysome profile RNA fractions. We found significant changes in the transcriptome and translatome of the Huh7.5 ADAR1 KO cells. The most prominent changes include negatively affected transcription by RNA polymerase III and the deregulation of snoRNA and Y RNA levels. Furthermore, we observed that ADAR1 KO polysomes are enriched in mRNAs coding for proteins pivotal in a wide range of biological processes such as RNA localization and RNA processing, whereas the unbound fraction is enriched mainly in mRNAs coding for ribosomal proteins and translational factors. This indicates that ADAR1 plays a more relevant role in small RNA metabolism and ribosome biogenesis.

• Klíčová slova
• ADAR1, RNA editing, Y RNA, hepatocyte, miRNA, snoRNA,
• MeSH
• adenosindeaminasa * genetika   metabolismus   MeSH
• buněčné linie MeSH
• editace RNA * MeSH
• genový knockout MeSH
• hepatocyty * metabolismus   MeSH
• lidé MeSH
• messenger RNA genetika   metabolismus   MeSH
• polyribozomy metabolismus   genetika   MeSH
• proteiny vázající RNA * genetika   metabolismus   MeSH
• proteosyntéza MeSH
• transkriptom MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ADAR protein, human MeSH Prohlížeč
• adenosindeaminasa * MeSH
• messenger RNA MeSH
• proteiny vázající RNA * MeSH

MicroRNA (miRNA) and RNA interference (RNAi) pathways rely on small RNAs produced by Dicer endonucleases. Mammalian Dicer primarily supports the essential gene-regulating miRNA pathway, but how it is specifically adapted to miRNA biogenesis is unknown. We show that the adaptation entails a unique structural role of Dicer's DExD/H helicase domain. Although mice tolerate loss of its putative ATPase function, the complete absence of the domain is lethal because it assures high-fidelity miRNA biogenesis. Structures of murine Dicer•-miRNA precursor complexes revealed that the DExD/H domain has a helicase-unrelated structural function. It locks Dicer in a closed state, which facilitates miRNA precursor selection. Transition to a cleavage-competent open state is stimulated by Dicer-binding protein TARBP2. Absence of the DExD/H domain or its mutations unlocks the closed state, reduces substrate selectivity, and activates RNAi. Thus, the DExD/H domain structurally contributes to mammalian miRNA biogenesis and underlies mechanistical partitioning of miRNA and RNAi pathways.

• Klíčová slova
• DExD, Dicer, PKR, RNAi, TARBP2, cryo-EM, dsRBD, dsRNA, helicase, miRNA, mirtron,
• MeSH
• mikro RNA * genetika   metabolismus   MeSH
• myši MeSH
• ribonukleasa III * metabolismus   MeSH
• RNA interference MeSH
• savci metabolismus   MeSH
• transportní proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• komentáře MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mikro RNA * MeSH
• ribonukleasa III * MeSH
• transportní proteiny MeSH

In animals and plants, Dicer enzymes collaborate with double-stranded RNA-binding domain (dsRBD) proteins to convert precursor-microRNAs (pre-miRNAs) into miRNA duplexes. We report six cryo-EM structures of Drosophila Dicer-1 that show how Dicer-1 and its partner Loqs‑PB cooperate (1) before binding pre-miRNA, (2) after binding and in a catalytically competent state, (3) after nicking one arm of the pre-miRNA, and (4) following complete dicing and initial product release. Our reconstructions suggest that pre-miRNA binds a rare, open conformation of the Dicer‑1⋅Loqs‑PB heterodimer. The Dicer-1 dsRBD and three Loqs‑PB dsRBDs form a tight belt around the pre-miRNA, distorting the RNA helix to place the scissile phosphodiester bonds in the RNase III active sites. Pre-miRNA cleavage shifts the dsRBDs and partially closes Dicer-1, which may promote product release. Our data suggest a model for how the Dicer‑1⋅Loqs‑PB complex affects a complete cycle of pre-miRNA recognition, stepwise endonuclease cleavage, and product release.

• Klíčová slova
• Dcr-1, Dicer, Dicer-partner proteins, Loqs-PB, Loquacious, RNase III, cryo-EM, dsRBD, isomiR, miRNA, microRNA,
• MeSH
• Drosophila genetika   MeSH
• mikro RNA * genetika   metabolismus   MeSH
• proteiny Drosophily * genetika   metabolismus   MeSH
• proteiny vázající RNA metabolismus   MeSH
• ribonukleasa III genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, N.I.H., Intramural MeSH
• Názvy látek
• mikro RNA * MeSH
• proteiny Drosophily * MeSH
• proteiny vázající RNA MeSH
• ribonukleasa III MeSH

ADAR RNA editing enzymes are high-affinity dsRNA-binding proteins that deaminate adenosines to inosines in pre-mRNA hairpins and also exert editing-independent effects. We generated a Drosophila AdarE374A mutant strain encoding a catalytically inactive Adar with CRISPR/Cas9. We demonstrate that Adar adenosine deamination activity is necessary for normal locomotion and prevents age-dependent neurodegeneration. The catalytically inactive protein, when expressed at a higher than physiological level, can rescue neurodegeneration in Adar mutants, suggesting also editing-independent effects. Furthermore, loss of Adar RNA editing activity leads to innate immune induction, indicating that Drosophila Adar, despite being the homolog of mammalian ADAR2, also has functions similar to mammalian ADAR1. The innate immune induction in fly Adar mutants is suppressed by silencing of Dicer-2, which has a RNA helicase domain similar to MDA5 that senses unedited dsRNAs in mammalian Adar1 mutants. Our work demonstrates that the single Adar enzyme in Drosophila unexpectedly has dual functions.

• MeSH
• adenosindeaminasa chemie   genetika   MeSH
• adenosinmonofosfát metabolismus   MeSH
• bodová mutace genetika   MeSH
• degenerace nervu patologie   MeSH
• Drosophila melanogaster genetika   imunologie   MeSH
• editace RNA genetika   MeSH
• katalýza MeSH
• lokomoce MeSH
• messenger RNA genetika   metabolismus   MeSH
• mozek metabolismus   MeSH
• přirozená imunita genetika   MeSH
• proteinové domény MeSH
• proteiny Drosophily chemie   genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• ribonukleasa III metabolismus   MeSH
• RNA-helikasy metabolismus   MeSH
• stárnutí patologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• Adar protein, Drosophila MeSH Prohlížeč
• adenosindeaminasa MeSH
• adenosinmonofosfát MeSH
• DCR-2 protein, Drosophila MeSH Prohlížeč
• messenger RNA MeSH
• proteiny Drosophily MeSH
• ribonukleasa III MeSH
• RNA-helikasy MeSH

ADAR RNA editing enzymes (adenosine deaminases acting on RNA) that convert adenosine bases to inosines were first identified biochemically 30 years ago. Since then, studies on ADARs in genetic model organisms, and evolutionary comparisons between them, continue to reveal a surprising range of pleiotropic biological effects of ADARs. This review focuses on Drosophila melanogaster, which has a single Adar gene encoding a homolog of vertebrate ADAR2 that site-specifically edits hundreds of transcripts to change individual codons in ion channel subunits and membrane and cytoskeletal proteins. Drosophila ADAR is involved in the control of neuronal excitability and neurodegeneration and, intriguingly, in the control of neuronal plasticity and sleep. Drosophila ADAR also interacts strongly with RNA interference, a key antiviral defense mechanism in invertebrates. Recent crystal structures of human ADAR2 deaminase domain-RNA complexes help to interpret available information on Drosophila ADAR isoforms and on the evolution of ADARs from tRNA deaminase ADAT proteins. ADAR RNA editing is a paradigm for the now rapidly expanding range of RNA modifications in mRNAs and ncRNAs. Even with recent progress, much remains to be understood about these groundbreaking ADAR RNA modification systems.

• Klíčová slova
• ADAR, Drosophila melanogaster, RNA editing, RNA modification, dsRNA, epitranscriptome,
• MeSH
• adenosindeaminasa chemie   genetika   metabolismus   MeSH
• Drosophila melanogaster genetika   metabolismus   MeSH
• editace RNA * MeSH
• exprese genu MeSH
• interakční proteinové domény a motivy MeSH
• izoenzymy MeSH
• lidé MeSH
• messenger RNA genetika   MeSH
• molekulární evoluce MeSH
• nervový systém metabolismus   MeSH
• obratlovci MeSH
• proteiny Drosophily genetika   metabolismus   MeSH
• proteiny vázající RNA genetika   metabolismus   MeSH
• RNA interference MeSH
• substrátová specifita MeSH
• vazba proteinů MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• Adar protein, Drosophila MeSH Prohlížeč
• adenosindeaminasa MeSH
• izoenzymy MeSH
• messenger RNA MeSH
• proteiny Drosophily MeSH
• proteiny vázající RNA MeSH

Our knowledge of the variety and abundances of RNA base modifications is rapidly increasing. Modified bases have critical roles in tRNAs, rRNAs, translation, splicing, RNA interference, and other RNA processes, and are now increasingly detected in all types of transcripts. Can new biological principles associated with this diversity of RNA modifications, particularly in mRNAs and long non-coding RNAs, be identified? This review will explore this question by focusing primarily on adenosine to inosine (A-to-I) RNA editing by the adenine deaminase acting on RNA (ADAR) enzymes that have been intensively studied for the past 20 years and have a wide range of effects. Over 100 million adenosine to inosine editing sites have been identified in the human transcriptome, mostly in embedded Alu sequences that form potentially innate immune-stimulating dsRNA hairpins in transcripts. Recent research has demonstrated that inosine in the epitranscriptome and ADAR1 protein establish innate immune tolerance for host dsRNA formed by endogenous sequences. Innate immune sensors that detect viral nucleic acids are among the readers of epitranscriptome RNA modifications, though this does preclude a wide range of other modification effects.

• MeSH
• adenosin genetika   MeSH
• adenosindeaminasa genetika   MeSH
• aminohydrolasy genetika   MeSH
• editace RNA genetika   MeSH
• inosin genetika   MeSH
• lidé MeSH
• messenger RNA genetika   MeSH
• posttranskripční úpravy RNA genetika   MeSH
• přirozená imunita genetika   MeSH
• proteiny vázající RNA genetika   MeSH
• transkriptom genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• ADAR protein, human MeSH Prohlížeč
• adenine deaminase MeSH Prohlížeč
• adenosin MeSH
• adenosindeaminasa MeSH
• aminohydrolasy MeSH
• inosin MeSH
• messenger RNA MeSH
• proteiny vázající RNA MeSH

The ADAR proteins deaminate adenosine to inosine in double-stranded RNA which is one of the most abundant modifications present in mammalian RNA. Inosine can have a profound effect on the RNAs that are edited, not only changing the base-pairing properties, but can also result in recoding, as inosine behaves as if it were guanosine. In mammals there are three ADAR proteins and two ADAR-related proteins (ADAD) expressed. All have a very similar modular structure; however, both their expression and biological function differ significantly. Only two of the ADAR proteins have enzymatic activity. However, both ADAR and ADAD proteins possess the ability to bind double-strand RNA. Mutations in ADARs have been associated with many diseases ranging from cancer, innate immunity to neurological disorders. Here, we will discuss in detail the domain structure of mammalian ADARs, the effects of RNA editing, and the role of ADARs in human diseases.

• Klíčová slova
• ADAR, Alu elements, RNA editing, cancer, deaminase domain, dsRBDs,
• MeSH
• adenosindeaminasa metabolismus   MeSH
• dvouvláknová RNA metabolismus   MeSH
• editace RNA genetika   MeSH
• lidé MeSH
• proteiny vázající RNA metabolismus   MeSH
• savci MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• adenosindeaminasa MeSH
• dvouvláknová RNA MeSH
• proteiny vázající RNA MeSH