scholarly journals Y-RNA and tRNA Cleavage by RNase L Mediates Terminal dsRNA Response

2016 ◽  
Author(s):  
Jesse Donovan ◽  
Sneha Rath ◽  
David Kolet-Mandrikov ◽  
Alexei Korennykh
Keyword(s):  
Small Rnas ◽  
Mammalian Cells ◽  
Innate Immune ◽  
Rna Cleavage ◽  
Rnase L ◽  
Rna Seq ◽  
Danger Signal ◽  
Double Stranded Rna ◽  
Transfer Rnas ◽  
Non Coding Rna

AbstractDouble-stranded RNA (dsRNA) is a danger signal that triggers endonucleolytic degradation of RNA inside infected and stressed mammalian cells. This mechanism inhibits growth and ultimately removes problematic cells via apoptosis. To elucidate the molecular functions of this program and understand the connection between RNA cleavage and programmed cell death, we visualized dsRNA-induced degradation of human small RNAs using RtcB ligase-assisted RNA sequencing (RtcB RNA-seq). RtcB RNA-seq revealed strong cleavage of select transfer RNAs (tRNAs) and autoantigenic Y-RNAs, and identified the innate immune receptor RNase L as the responsible endoribonuclease. RNase L cleaves the non-coding RNA (ncRNA) targets site-specifically, releasing abundant ncRNA fragments, and downregulating full-length tRNAs and Y-RNAs. The depletion of a single Y-RNA, RNY1, appears particularly important and the loss of this Y-RNA is sufficient to initiate apoptosis. Site-specific cleavage of small ncRNA by RNase L thus emerges as an important terminal step in dsRNA surveillance.

scholarly journals Real-time 2-5A kinetics suggest that interferons β and λ evade global arrest of translation by RNase L

2019 ◽  
Vol 116 (6) ◽  
pp. 2103-2111 ◽  
Author(s):  
Alisha Chitrakar ◽  
Sneha Rath ◽  
Jesse Donovan ◽  
Kaitlin Demarest ◽  
Yize Li ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Innate Immune System ◽  
Innate Immune ◽  
Host Protein ◽  
Rna Cleavage ◽  
Rnase L ◽  
Foreign Material ◽  
Defense Proteins ◽  
Double Stranded Rna ◽  
Host Protein Synthesis

Cells of all mammals recognize double-stranded RNA (dsRNA) as a foreign material. In response, they release interferons (IFNs) and activate a ubiquitously expressed pseudokinase/endoribonuclease RNase L. RNase L executes regulated RNA decay and halts global translation. Here, we developed a biosensor for 2′,5′-oligoadenylate (2-5A), the natural activator of RNase L. Using this biosensor, we found that 2-5A was acutely synthesized by cells in response to dsRNA sensing, which immediately triggered cellular RNA cleavage by RNase L and arrested host protein synthesis. However, translation-arrested cells still transcribed IFN-stimulated genes and secreted IFNs of types I and III (IFN-β and IFN-λ). Our data suggest that IFNs escape from the action of RNase L on translation. We propose that the 2-5A/RNase L pathway serves to rapidly and accurately suppress basal protein synthesis, preserving privileged production of defense proteins of the innate immune system.

scholarly journals Human RNase L tunes gene expression by selectively destabilizing the microRNA-regulated transcriptome

2015 ◽  
Vol 112 (52) ◽  
pp. 15916-15921 ◽  
Author(s):  
Sneha Rath ◽  
Jesse Donovan ◽  
Gena Whitney ◽  
Alisha Chitrakar ◽  
Wei Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Innate Immune System ◽  
Mammalian Cells ◽  
Innate Immune ◽  
Rnase L ◽  
Double Stranded Rna ◽  
Rna Targets ◽  
Human Rnase ◽  
Cell Growth And Differentiation ◽  
Cell Adhesion And Proliferation

Double-stranded RNA (dsRNA) activates the innate immune system of mammalian cells and triggers intracellular RNA decay by the pseudokinase and endoribonuclease RNase L. RNase L protects from pathogens and regulates cell growth and differentiation by destabilizing largely unknown mammalian RNA targets. We developed an approach for transcriptome-wide profiling of RNase L activity in human cells and identified hundreds of direct RNA targets and nontargets. We show that this RNase L-dependent decay selectively affects transcripts regulated by microRNA (miR)-17/miR-29/miR-200 and other miRs that function as suppressors of mammalian cell adhesion and proliferation. RNase L mimics the effects of these miRs and acts as a suppressor of proliferation and adhesion in mammalian cells. Our data suggest that RNase L-dependent decay serves to establish an antiproliferative state via destabilization of the miR-regulated transcriptome.

scholarly journals Curcumin, a Natural Antioxidant, Acts as a Noncompetitive Inhibitor of Human RNase L in Presence of Its Cofactor 2-5AIn Vitro

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Ankush Gupta ◽  
Pramod C. Rath
Keyword(s):  
Innate Immune System ◽  
Mammalian Cells ◽  
Viral Infections ◽  
Innate Immune ◽  
Therapeutic Interventions ◽  
Active Principle ◽  
Rnase L ◽  
Double Stranded Rna ◽  
Human Rnase ◽  
Noncompetitive Inhibitor

Ribonuclease L (RNase L) is an antiviral endoribonuclease of the innate immune system, which is induced and activated by viral infections, interferons, and double stranded RNA (dsRNA) in mammalian cells. Although, RNase L is generally protective against viral infections, abnormal RNase L expression and activity have been associated with a number of diseases. Here, we show that curcumin, a natural plant-derived anti-inflammatory active principle, inhibits RNase L activity; hence, it may be exploited for therapeutic interventions in case of pathological situations associated with excess activation of RNase L.

scholarly journals Concerted 2-5A-Mediated mRNA Decay and Transcription Reprogram Protein Synthesis in dsRNA Response

2018 ◽  
Author(s):  
Sneha Rath ◽  
Eliza Prangley ◽  
Jesse Donovan ◽  
Kaitlin Demarest ◽  
Yigal Meir ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Translation Initiation ◽  
Mrna Stability ◽  
Rna Degradation ◽  
Innate Immune ◽  
Messenger Rnas ◽  
Rnase L ◽  
Rna Seq ◽  
Defense Proteins ◽  
Double Stranded Rna

RNA degradation by RNase L during 2-5A-mediated decay (2-5AMD) is a conserved mammalian stress response to viral and endogenous double-stranded RNA (dsRNA). 2-5AMD onsets rapidly and facilitates a switch of protein synthesis from homeostasis to production of interferons (IFNs). To understand the mechanism of this protein synthesis reprogramming, we examined 2-5AMD in human cells. 2-5AMD triggers polysome collapse characteristic of a translation initiation defect, but translation initiation complexes and ribosomes purified from the translation-arrested cells remain functional. Using spike-in RNA-seq we found that basal messenger RNAs (mRNAs) rapidly decay, while mRNAs encoding IFNs and IFN-stimulated genes evade 2-5AMD and accumulate. The IFN evasion results from the combined effect of better mRNA stability and positive feedback amplification in the IFN response. Therefore, 2-5AMD and transcription act in concert to revamp the cellular mRNA composition. The resulting preferential accumulation of innate immune mRNAs establishes 'prioritized' synthesis of defense proteins.

scholarly journals E. coli OxyS non-coding RNA does not trigger RNAi in C. elegans

2014 ◽  
Author(s):  
Alper Akay ◽  
Peter Sarkies ◽  
Eric Alexander Miska
Keyword(s):  
Small Rnas ◽  
Biological Function ◽  
Rapid Development ◽  
Rnai Pathway ◽  
Regulate Gene Expression ◽  
Double Stranded Rna ◽  
E Coli ◽  
C Elegans ◽  
Non Coding Rna ◽  
Rnai Response

The discovery of RNA interference (RNAi) in C. elegans has had a major impact on scientific research, led to the rapid development of RNAi tools and has inspired RNA-based therapeutics. Astonishingly, nematodes, planaria and many insects take up double-stranded RNA (dsRNA) from their environment to elicit RNAi; the biological function of this mechanism is unclear. Recently, the E. coli OxyS non-coding RNA was shown to regulate gene expression in C. elegans when E. coli is offered as food. This was surprising given that C. elegans is unlikely to encounter E. coli in nature. To directly test the hypothesis that the E. coli OxyS non-coding RNA triggers the C. elegans RNAi pathway, we sequenced small RNAs from C. elegans after feeding with bacteria. We clearly demonstrate that the OxyS non-coding RNA does not trigger an RNAi response in C. elegans. We conclude that the biology of environmental RNAi remains to be discovered.

Introns encode dsRNAs undetected by RIG-I/MDA5/interferons and sensed via RNase L

2021 ◽  
Vol 118 (46) ◽  
pp. e2102134118
Author(s):  
Alisha Chitrakar ◽  
Kristina Solorio-Kirpichyan ◽  
Eliza Prangley ◽  
Sneha Rath ◽  
Jin Du ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Immune Responses ◽  
Innate Immune ◽  
Rna Decay ◽  
Antiviral Defense ◽  
Rnase L ◽  
Innate Immune Responses ◽  
Double Stranded Rna ◽  
Turn On ◽  
Nuclear Rna

Double-stranded RNA (dsRNA), a hallmark viral material that activates antiviral interferon (IFN) responses, can appear in human cells also in the absence of viruses. We identify phosphorothioate DNAs (PS DNAs) as triggers of such endogenous dsRNA (endo-dsRNA). PS DNAs inhibit decay of nuclear RNAs and induce endo-dsRNA via accumulation of high levels of intronic and intergenic inverted retroelements (IIIR). IIIRs activate endo-dsRNA responses distinct from antiviral defense programs. IIIRs do not turn on transcriptional RIG-I/MDA5/IFN signaling, but they trigger the dsRNA-sensing pathways of OAS3/RNase L and PKR. Thus, nuclear RNA decay and nuclear-cytosolic RNA sorting actively protect from these innate immune responses to self. Our data suggest that the OAS3/RNase L and PKR arms of innate immunity diverge from antiviral IFN responses and monitor nuclear RNA decay by sensing cytosolic escape of IIIRs. OAS3 provides a receptor for IIIRs, whereas RNase L cleaves IIIR-carrying introns and intergenic RNAs.

scholarly journals SARS-CoV-2 induces double-stranded RNA-mediated innate immune responses in respiratory epithelial-derived cells and cardiomyocytes

2021 ◽  
Vol 118 (16) ◽  
pp. e2022643118
Author(s):  
Yize Li ◽  
David M. Renner ◽  
Courtney E. Comar ◽  
Jillian N. Whelan ◽  
Hanako M. Reyes ◽  
...  
Keyword(s):  
Induced Pluripotent Stem Cell ◽  
Cell Types ◽  
Innate Immune ◽  
Alveolar Type ◽  
Rnase L ◽  
Oligoadenylate Synthetase ◽  
Protein Kinase R ◽  
Double Stranded Rna ◽  
Induced Pluripotent ◽  
Host Virus Interactions

Coronaviruses are adept at evading host antiviral pathways induced by viral double-stranded RNA, including interferon (IFN) signaling, oligoadenylate synthetase–ribonuclease L (OAS-RNase L), and protein kinase R (PKR). While dysregulated or inadequate IFN responses have been associated with severe coronavirus infection, the extent to which the recently emerged SARS-CoV-2 activates or antagonizes these pathways is relatively unknown. We found that SARS-CoV-2 infects patient-derived nasal epithelial cells, present at the initial site of infection; induced pluripotent stem cell-derived alveolar type 2 cells (iAT2), the major cell type infected in the lung; and cardiomyocytes (iCM), consistent with cardiovascular consequences of COVID-19 disease. Robust activation of IFN or OAS-RNase L is not observed in these cell types, whereas PKR activation is evident in iAT2 and iCM. In SARS-CoV-2–infected Calu-3 and A549ACE2 lung-derived cell lines, IFN induction remains relatively weak; however, activation of OAS-RNase L and PKR is observed. This is in contrast to Middle East respiratory syndrome (MERS)-CoV, which effectively inhibits IFN signaling and OAS-RNase L and PKR pathways, but is similar to mutant MERS-CoV lacking innate immune antagonists. Remarkably, OAS-RNase L and PKR are activated in MAVS knockout A549ACE2 cells, demonstrating that SARS-CoV-2 can induce these host antiviral pathways despite minimal IFN production. Moreover, increased replication and cytopathic effect in RNASEL knockout A549ACE2 cells implicates OAS-RNase L in restricting SARS-CoV-2. Finally, while SARS-CoV-2 fails to antagonize these host defense pathways, which contrasts with other coronaviruses, the IFN signaling response is generally weak. These host–virus interactions may contribute to the unique pathogenesis of SARS-CoV-2.

scholarly journals Identification of RNase L-Dependent, 3'-End-Modified, Viral Small RNAs in Sindbis Virus-Infected Mammalian Cells

mBio ◽  
2013 ◽  
Vol 4 (6) ◽  
Author(s):  
E. Girardi ◽  
B. Chane-Woon-Ming ◽  
M. Messmer ◽  
P. Kaukinen ◽  
S. Pfeffer
Keyword(s):  
Small Rnas ◽  
Mammalian Cells ◽  
Sindbis Virus ◽  
Rnase L

scholarly journals A comprehensive profile of circulating RNAs in human serum

2017 ◽  
Author(s):  
Sinan Uğur Umu ◽  
Hilde Langseth ◽  
Cecilie Bucher-Jonannessen ◽  
Bastian Fromm ◽  
Andreas Keller ◽  
...  
Keyword(s):  
Human Serum ◽  
Population Based ◽  
Rna Seq ◽  
Serum Samples ◽  
Transfer Rnas ◽  
Rna Molecules ◽  
Non Coding Rna ◽  
Norwegian Population ◽  
Micro Rnas ◽  
Nucleolar Rnas

ABSTRACTNon-coding RNA (ncRNA) molecules have fundamental roles in cells and many are also stable in body fluids as extracellular RNAs. In this study, we used RNA sequencing (RNA-seq) to investigate the profile of small non-coding RNA (sncRNA) in human serum. We analyzed 10 billion lllumina reads from 477 serum samples, included in the Norwegian population-based Janus Serum Bank (JSB). We found that the core serum RNA repertoire includes 258 micro RNAs (miRNA), 441 piwi-interacting RNAs (piRNA), 411 transfer RNAs (tRNA), 24 small nucleolar RNAs (snoRNA), 125 small nuclear RNAs (snRNA) and 123 miscellaneous RNAs (misc-RNA). We also investigated biological and technical variation in expression, and the results suggest that many RNA molecules identified in serum contain signs of biological variation. They are therefore unlikely to be random degradation by-products. In addition, the presence of specific fragments of tRNA, snoRNA, Vault RNA and Y_RNA indicates protection from degradation. Our results suggest that many circulating RNAs in serum can be potential biomarkers.

scholarly journals RNase L reprograms translation by widespread mRNA turnover escaped by antiviral mRNAs

2018 ◽  
Author(s):  
James M Burke ◽  
Stephanie L Moon ◽  
Evan T Lester ◽  
Tyler Matheny ◽  
Roy Parker
Keyword(s):  
Translation Initiation ◽  
Mammalian Cells ◽  
Mrna Degradation ◽  
Stress Granule ◽  
Mrna Turnover ◽  
Rnase L ◽  
Protein Kinase R ◽  
Double Stranded Rna ◽  
Translation Repression ◽  
Eif2α Phosphorylation

SUMMARYIn response to foreign and endogenous double-stranded RNA (dsRNA), protein kinase R (PKR) and ribonuclease L (RNase L) reprogram translation in mammalian cells. PKR inhibits translation initiation through eIF2α phosphorylation, which triggers stress granule (SG) formation and promotes translation of stress responsive mRNAs. The mechanisms of RNase L-driven translation repression, its contribution to SG assembly, and its regulation of dsRNA stress-induced mRNAs are unknown. We demonstrate that RNase L drives translational shut-off in response to dsRNA by promoting widespread turnover of mRNAs. This alters stress granule assembly and reprograms translation by only allowing for the translation of mRNAs resistant to RNase L degradation, including numerous antiviral mRNAs such asIFN-β. Individual cells differentially activate dsRNA responses revealing variation that can affect cellular outcomes. This identifies bulk mRNA degradation and the resistance of antiviral mRNAs as the mechanism by which RNaseL reprograms translation in response to dsRNA.

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