Structure and Degradation of Circular RNAs Regulate PKR Activation in Innate Immunity.

Circular RNAs (circRNAs) produced from back-splicing of exons of pre-mRNAs are widely expressed, but current understanding of their functions is limited. These RNAs are stable in general and are thought to have unique structural conformations distinct from their linear RNA cognates. Here, we show that endogenous circRNAs tend to form 16-26 bp imperfect RNA duplexes and act as inhibitors of double-stranded RNA (dsRNA)-activated protein kinase (PKR) related to innate immunity. Upon poly(I:C) stimulation or viral infection, circRNAs are globally degraded by RNase L, a process required for PKR activation in early cellular innate immune responses. Augmented PKR phosphorylation and circRNA reduction are found in peripheral blood mononuclear cells (PBMCs) derived from patients with autoimmune disease systemic lupus erythematosus (SLE). Importantly, overexpression of the dsRNA-containing circRNA in PBMCs or T cells derived from SLE can alleviate the aberrant PKR activation cascade, thus providing a connection between circRNAs and SLE.

RNA helicase SKIV2L limits antiviral defense and autoinflammation elicited by the OAS-RNase L pathway.

The OAS-RNase L pathway is one of the oldest innate RNA sensing pathways that leads to interferon (IFN) signaling and cell death. OAS recognizes viral RNA and then activates RNase L, which subsequently cleaves both cellular and viral RNA, creating "processed RNA" as an endogenous ligand that further triggers RIG-I-like receptor signaling. However, the IFN response and antiviral activity of the OAS-RNase L pathway are weak compared to other RNA-sensing pathways. Here, we discover that the SKIV2L RNA exosome limits the antiviral capacity of the OAS-RNase L pathway. SKIV2L-deficient cells exhibit remarkably increased interferon responses to RNase L-processed RNA, resulting in heightened antiviral activity. The helicase activity of SKIV2L is indispensable for this function, acting downstream of RNase L. SKIV2L depletion increases the antiviral capacity of OAS-RNase L against RNA virus infection. Furthermore, SKIV2L loss exacerbates autoinflammation caused by human OAS1 gain-of-function mutations. Taken together, our results identify SKIV2L as a critical barrier to OAS-RNase L-mediated antiviral immunity that could be therapeutically targeted to enhance the activity of a basic antiviral pathway.

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

Viral and endogenous double-stranded RNA (dsRNA) is a potent trigger for programmed RNA degradation by the 2-5A/RNase L complex in cells of all mammals. This 2-5A-mediated decay (2-5AMD) is a conserved stress response switching global protein synthesis from homeostasis to production of interferons (IFNs). To understand this mechanism, we examined 2-5AMD in human cells and found that it triggers polysome collapse characteristic of inhibited translation initiation. We determined that translation initiation complexes and ribosomes purified from translation-arrested cells remain functional. However, spike-in RNA sequencing (RNA-seq) revealed cell-wide decay of basal mRNAs accompanied by rapid accumulation of mRNAs encoding innate immune proteins. Our data attribute this 2-5AMD evasion to better stability of defense mRNAs and positive feedback in the IFN response amplified by RNase L-resistant molecules. We conclude that 2-5AMD and transcription act in concert to refill mammalian cells with defense mRNAs, thereby "prioritizing" the synthesis of innate immune proteins.

RNase L Reprograms Translation by Widespread mRNA Turnover Escaped by Antiviral mRNAs.

In 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 allowing translation of mRNAs resistant to RNase L degradation, including numerous antiviral mRNAs such as interferon (IFN)-ß. 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 RNase L reprograms translation in response to dsRNA.

Recurrent viral capture of cellular phosphodiesterases that antagonize OAS-RNase L.

Phosphodiesterases (PDEs) encoded by viruses are putatively acquired by horizontal transfer of cellular PDE ancestor genes. Viral PDEs inhibit the OAS-RNase L antiviral pathway, a key effector component of the innate immune response. Although the function of these proteins is well-characterized, the origins of these gene acquisitions are less clear. Phylogenetic analysis revealed at least five independent PDE acquisition events by ancestral viruses. We found evidence that PDE-encoding genes were horizontally transferred between coronaviruses belonging to different genera. Three clades of viruses within Nidovirales: merbecoviruses (MERS-CoV), embecoviruses (HCoV-OC43), and toroviruses encode independently acquired PDEs, and a clade of rodent alphacoronaviruses acquired an embecovirus PDE via recent horizontal transfer. Among rotaviruses, the PDE of rotavirus A was acquired independently from rotavirus B and G PDEs, which share a common ancestor. Conserved motif analysis suggests a link between all viral PDEs and a similar ancestor among the mammalian AKAP7 proteins despite low levels of sequence conservation. Additionally, we used ancestral sequence reconstruction and structural modeling to reveal that sequence and structural divergence are not well-correlated among these proteins. Specifically, merbecovirus PDEs are as structurally divergent from the ancestral protein and the solved structure of human AKAP7 PDE as they are from each other. In contrast, comparisons of rotavirus B and G PDEs reveal virtually unchanged structures despite evidence for loss of function in one, suggesting impactful changes that lie outside conserved catalytic sites. These findings highlight the complex and volatile evolutionary history of viral PDEs and provide a framework to facilitate future studies.

RNA is required for the integrity of multiple nuclear and cytoplasmic membrane-less RNP granules.

Numerous membrane-less organelles, composed of a combination of RNA and proteins, are observed in the nucleus and cytoplasm of eukaryotic cells. These RNP granules include stress granules (SGs), processing bodies (PBs), Cajal bodies, and nuclear speckles. An unresolved question is how frequently RNA molecules are required for the integrity of RNP granules in either the nucleus or cytosol. To address this issue, we degraded intracellular RNA in either the cytosol or the nucleus by the activation of RNase L and examined the impact of RNA loss on several RNP granules. We find the majority of RNP granules, including SGs, Cajal bodies, nuclear speckles, and the nucleolus, are altered by the degradation of their RNA components. In contrast, PBs and super-enhancer complexes were largely not affected by RNA degradation in their respective compartments. RNA degradation overall led to the apparent dissolution of some membrane-less organelles, whereas others reorganized into structures with altered morphology. These findings highlight a critical and widespread role of RNA in the organization of several RNP granules.

Origin of the OAS-RNase L innate immune pathway before the rise of jawed vertebrates via molecular tinkering.

Discriminating self from nonself is fundamental to immunity. Yet, it remains largely elusive how the mechanisms of self and nonself discrimination originated. Sensing double-stranded RNA as nonself, the 2',5'-oligoadenylate synthetase (OAS)-ribonuclease L (RNase L) pathway represents a crucial component of innate immunity. Here, we combine phylogenomic and functional analyses to show that the functional OAS-RNase L pathway likely originated through tinkering with preexisting proteins before the rise of jawed vertebrates during or before the Silurian period (444 to 419 Mya). Multiple concerted losses of OAS and RNase L occurred during the evolution of jawed vertebrates, further supporting the ancient coupling between OAS and RNase L. Moreover, both OAS and RNase L genes evolved under episodic positive selection across jawed vertebrates, suggesting a long-running evolutionary arms race between the OAS-RNase L pathway and microbes. Our findings illuminate how an innate immune pathway originated via molecular tinkering.

Activation of protein kinase receptor (PKR) plays a pro-viral role in mammarenavirus-infected cells.

Many viruses, including mammarenaviruses, have evolved mechanisms to counteract different components of the host cell innate immunity, which is required to facilitate robust virus multiplication. The double-stranded RNA (dsRNA) sensor protein kinase receptor (PKR) pathway plays a critical role in the cell anti-viral response. Whether PKR can restrict the multiplication of the Old World mammarenavirus lymphocytic choriomeningitis virus (LCMV) and the mechanisms by which LCMV may counteract the anti-viral functions of PKR have not yet been investigated. Here we present evidence that LCMV infection results in very limited levels of PKR activation, but LCMV multiplication is enhanced in the absence of PKR. In contrast, infection with a recombinant LCMV with a mutation affecting the 3'-5' exonuclease (ExoN) activity of the viral nucleoprotein resulted in robust PKR activation in the absence of detectable levels of dsRNA, which was associated with severely restricted virus multiplication that was alleviated in the absence of PKR. However, pharmacological inhibition of PKR activation resulted in reduced levels of LCMV multiplication. These findings uncovered a complex role of the PKR pathway in LCMV-infected cells involving both pro- and anti-viral activities.IMPORTANCEAs with many other viruses, the prototypic Old World mammarenavirus LCMV can interfere with the host cell innate immune response to infection, which includes the dsRNA sensor PKR pathway. A detailed understanding of LCMV-PKR interactions can provide novel insights about mammarenavirus-host cell interactions and facilitate the development of effective anti-viral strategies against human pathogenic mammarenaviruses. In the present work, we present evidence that LCMV multiplication is enhanced in PKR-deficient cells, but pharmacological inhibition of PKR activation unexpectedly resulted in severely restricted propagation of LCMV. Likewise, we document a robust PKR activation in LCMV-infected cells in the absence of detectable levels of dsRNA. Our findings have revealed a complex role of the PKR pathway during LCMV infection and uncovered the activation of PKR as a druggable target for the development of anti-viral drugs against human pathogenic mammarenaviruses.

MERS-CoV endoribonuclease and accessory proteins jointly evade host innate immunity during infection of lung and nasal epithelial cells.

Middle East respiratory syndrome coronavirus (MERS-CoV) emerged into humans in 2012, causing highly lethal respiratory disease. The severity of disease may be, in part, because MERS-CoV is adept at antagonizing early innate immune pathways­interferon (IFN) production and signaling, protein kinase R (PKR), and oligoadenylate synthetase/ribonuclease L (OAS/RNase L)­activated in response to viral double-stranded RNA (dsRNA) generated during genome replication. This is in contrast to severe acute respiratory syndrome CoV-2 (SARS-CoV-2), which we recently reported to activate PKR and RNase L and, to some extent, IFN signaling. We previously found that MERS-CoV accessory proteins NS4a (dsRNA binding protein) and NS4b (phosphodiesterase) could weakly suppress these pathways, but ablation of each had minimal effect on virus replication. Here we investigated the antagonist effects of the conserved coronavirus endoribonuclease (EndoU), in combination with NS4a or NS4b. Inactivation of EndoU catalytic activity alone in a recombinant MERS-CoV caused little if any effect on activation of the innate immune pathways during infection. However, infection with recombinant viruses containing combined mutations with inactivation of EndoU and deletion of NS4a or inactivation of the NS4b phosphodiesterase promoted robust activation of dsRNA-induced innate immune pathways. This resulted in at least tenfold attenuation of replication in human lung­derived A549 and primary nasal cells. Furthermore, replication of these recombinant viruses could be rescued to the level of wild-type MERS-CoV by knockout of host immune mediators MAVS, PKR, or RNase L. Thus, EndoU and accessory proteins NS4a and NS4b together suppress dsRNA-induced innate immunity during MERS-CoV infection in order to optimize viral replication.

Role of helical structure and dynamics in oligoadenylate synthetase 1 (OAS1) mismatch tolerance and activation by short dsRNAs.

The 2'-5'-oligoadenylate synthetases (OAS) are innate immune sensors of cytosolic double-stranded RNA (dsRNA) that play a critical role in limiting viral infection. How these proteins are able to avoid aberrant activation by cellular RNAs is not fully understood, but adenosine-to-inosine (A-to-I) editing has been proposed to limit accumulation of endogenous RNAs that might otherwise cause stimulation of the OAS/RNase L pathway. Here, we aim to uncover whether and how such sequence modifications can restrict the ability of short, defined dsRNAs to activate the single-domain form of OAS, OAS1. Unexpectedly, we find that all tested inosine-containing dsRNAs have an increased capacity to activate OAS1, whether in a destabilizing (I•U) or standard Watson-Crick-like base pairing (I-C) context. Additional variants with strongly destabilizing A•C mismatches or stabilizing G-C pairs also exhibit increased capacity to activate OAS1, eliminating helical stability as a factor in the relative ability of the dsRNAs to activate OAS1. Using thermal difference spectra and molecular dynamics simulations, we identify both increased helical dynamics and specific local changes in helical structure as important factors in the capacity of short dsRNAs to activate OAS1. These helical features may facilitate more ready adoption of the distorted OAS1-bound conformation or stabilize important structures to predispose the dsRNA for optimal binding and activation of OAS1. These studies thus reveal the molecular basis for the greater capacity of some short dsRNAs to activate OAS1 in a sequence-independent manner.

2-5A-Mediated decay (2-5AMD): from antiviral defense to control of host RNA.

Mammalian cells are exquisitely sensitive to the presence of double-stranded RNA (dsRNA), a molecule that they interpret as a signal of viral presence requiring immediate attention. Upon sensing dsRNA cells activate the innate immune response, which involves transcriptional mechanisms driving inflammation and secretion of interferons (IFNs) and interferon-stimulated genes (ISGs), as well as synthesis of RNA-like signaling molecules comprised of three or more 2'-5'-linked adenylates (2-5As). 2-5As were discovered some forty years ago and described as IFN-induced inhibitors of protein synthesis. The efforts of many laboratories, aimed at elucidating the molecular mechanism and function of these mysterious RNA-like signaling oligonucleotides, revealed that 2-5A is a specific ligand for the kinase-family endonuclease RNase L. RNase L decays single-stranded RNA (ssRNA) from viruses and mRNAs (as well as other RNAs) from hosts in a process we proposed to call 2-5A-mediated decay (2-5AMD). During recent years it has become increasingly recognized that 2-5AMD is more than a blunt tool of viral RNA destruction, but a pathway deeply integrated into sensing and regulation of endogenous RNAs. Here we present an overview of recently emerged roles of 2-5AMD in host RNA regulation.

A closer look at mammalian antiviral condensates.

Several biomolecular condensates assemble in mammalian cells in response to viral infection. The most studied of these are stress granules (SGs), which have been proposed to promote antiviral innate immune signaling pathways, including the RLR-MAVS, the protein kinase R (PKR), and the OAS-RNase L pathways. However, recent studies have demonstrated that SGs either negatively regulate or do not impact antiviral signaling. Instead, the SG-nucleating protein, G3BP1, may function to perturb viral RNA biology by condensing viral RNA into viral-aggregated RNA condensates, thus explaining why viruses often antagonize G3BP1 or hijack its RNA condensing function. However, a recently identified condensate, termed double-stranded RNA-induced foci, promotes the activation of the PKR and OAS-RNase L antiviral pathways. In addition, SG-like condensates known as an RNase L-induced bodies (RLBs) have been observed during many viral infections, including SARS-CoV-2 and several flaviviruses. RLBs may function in promoting decay of cellular and viral RNA, as well as promoting ribosome-associated signaling pathways. Herein, we review these recent advances in the field of antiviral biomolecular condensates, and we provide perspective on the role of canonical SGs and G3BP1 during the antiviral response.

The nuclear matrix protein HNRNPU restricts hepatitis B virus transcription by promoting OAS3-based activation of host innate immunity.

Heterogeneous nuclear protein U (HNRNPU) plays a pivotal role in innate immunity by facilitating chromatin opening to activate immune genes during host defense against viral infection. However, the mechanism by which HNRNPU is involved in Hepatitis B virus (HBV) transcription regulation through mediating antiviral immunity remains unknown. Our study revealed a significant decrease in HNRNPU levels during HBV transcription, which depends on HBx-DDB1-mediated degradation. Overexpression of HNRNPU suppressed HBV transcription, while its knockdown effectively promoted viral transcription, indicating HNRNPU as a novel host restriction factor for HBV transcription. Mechanistically, HNRNPU inhibits HBV transcription by activating innate immunity through primarily the positive regulation of the interferon-stimulating factor 2'-5'-oligoadenylate synthetase 3, which mediates an ribonuclease L-dependent mechanism to enhance innate immune responses. This study offers new insights into the host immune regulation of HBV transcription and proposes potential targets for therapeutic intervention against HBV infection.

A novel role of RNase L in the development of nonalcoholic steatohepatitis.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease and affects about 25% of the population globally. NAFLD has the potential to cause significant liver damage in many patients because it can progress to nonalcoholic steatohepatitis (NASH) and cirrhosis, which substantially increases disease morbidity and mortality. Despite the key role of innate immunity in the disease progression, the underlying molecular and pathogenic mechanisms remain to be elucidated. RNase L is a key enzyme in interferon action against viral infection and displays pleiotropic biological functions such as control of cell proliferation, apoptosis, and autophagy. Recent studies have demonstrated that RNase L is involved in innate immunity. In this study, we revealed that RNase L contributed to the development of NAFLD, which further progressed to NASH in a time-dependent fashion after RNase L wild-type (WT) and knockout mice were fed with a high-fat and high-cholesterol diet. RNase L WT mice showed significantly more severe NASH, evidenced by widespread macro-vesicular steatosis, hepatocyte ballooning degeneration, inflammation, and fibrosis, although physiological and biochemical data indicated that both types of mice developed obesity, hyperglycemia, hypercholesterolemia, dysfunction of the liver, and systemic inflammation at different extents. Further investigation demonstrated that RNase L was responsible for the expression of some key genes in lipid metabolism, inflammation, and fibrosis signaling. Taken together, our results suggest that a novel therapeutic intervention for NAFLD may be developed based on regulating the expression and activity of RNase L.

Serum RNase L levels in patients with chronic hepatitis B virus infection.

BACKGROUND/AIMS: Chronic hepatitis B virus (HBV) infection still poses a major threat to global health. Oligoadenylate synthetase-ribonuclease L (RNase L) antiviral pathway is one of interferon-induced antiviral effectors. The relationship between RNase L and HBV has never been investigated and we aim to examine the serum RNase L levels in patients with different stages of chronic HBV infection. METHODS: The patients were enrolled from 1985 to 2000, who had been HBsAg positive for longer than 6 months, at the National Taiwan University Hospital. In total, 426 patients with chronic HBV infection were included in this study, including 135 inactive carriers, 148 cirrhosis, and 143 hepatocellular carcinoma (HCC) cases. RESULTS: The RNase L levels increase as the disease severity increases. Higher RNase L levels were associated with higher HBV viral load, and the HBV-RNase L relationship was replaced by the disease severity status when adding disease status into the model. Compared with inactive carriers, the risk of liver cirrhosis was 60-fold (odds ratio = 60.8, 95% confidence interval = 3.49-1061) with the highest quintile of RNase L levels, after the adjustment of HBV DNA. The dose-response trend was statistically significant with quintiles and one increment of RNase L level in relation to liver cirrhosis. Similar results were found when HCC was compared with inactive carriers, while there was no association when compared between liver cirrhosis and HCC. CONCLUSIONS: A positive relationship between serum RNase L and HBV viral titers or advanced disease status is uncovered in this study. Further investigation in this area may provide more details of an innate immune response for HBV and opportunity for novel therapeutic strategy.

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

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.

Zika virus employs the host antiviral RNase L protein to support replication factory assembly.

Infection with the flavivirus Zika virus (ZIKV) can result in tissue tropism, disease outcome, and route of transmission distinct from those of other flaviviruses; therefore, we aimed to identify host machinery that exclusively promotes the ZIKV replication cycle, which can inform on differences at the organismal level. We previously reported that deletion of the host antiviral ribonuclease L (RNase L) protein decreases ZIKV production. Canonical RNase L catalytic activity typically restricts viral infection, including that of the flavivirus dengue virus (DENV), suggesting an unconventional, proviral RNase L function during ZIKV infection. In this study, we reveal that an inactive form of RNase L supports assembly of ZIKV replication factories (RFs) to enhance infectious virus production. Compared with the densely concentrated ZIKV RFs generated with RNase L present, deletion of RNase L induced broader subcellular distribution of ZIKV replication intermediate double-stranded RNA (dsRNA) and NS3 protease, two constituents of ZIKV RFs. An inactive form of RNase L was sufficient to contain ZIKV genome and dsRNA within a smaller RF area, which subsequently increased infectious ZIKV release from the cell. Inactive RNase L can interact with cytoskeleton, and flaviviruses remodel cytoskeleton to construct RFs. Thus, we used the microtubule-stabilization drug paclitaxel to demonstrate that ZIKV repurposes RNase L to facilitate the cytoskeleton rearrangements required for proper generation of RFs. During infection with flaviviruses DENV or West Nile Kunjin virus, inactive RNase L did not improve virus production, suggesting that a proviral RNase L role is not a general feature of all flavivirus infections.

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

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.

Unraveling the Role of RNase L Knockout in Alleviating Immune Response Activation in Mice Bone Marrow after Irradiation.

Ionizing radiation (IR) induces severe hematopoietic injury by causing DNA and RNA damage as well as activating the immune responses, necessitating the development of effective therapeutic strategies. Ribonuclease L (RNase L) as an innate immune response pathway is triggered by exogenous and endogenous abnormal dsRNA under viral infection and dyshomeostasis, thereby activating the immune responses. Thus, we investigated the effect of RNase L on irradiation-induced bone marrow damage using RNase L knockout (RNase L-/-) mice. Phenotypic analysis revealed that RNase L knockout mitigates irradiation-induced injury in the bone marrow. Further investigation into the mechanism of RNase L by RNA-seq, qRT-PCR, and CBA analysis demonstrated that RNase L deficiency counteracts the upregulation of genes related to immune responses induced by irradiation, including cytokines and interferon-stimulated genes. Moreover, RNase L deficiency inhibits the increased levels of immunoglobulins in serum induced by irradiation. These findings indicate that RNase L plays a role in the immune response induced by irradiation in the bone marrow. This study further enhances our understanding of the biological functions of RNase L in the immune response induced by irradiation and offers a novel approach for managing irradiation-induced bone marrow injury through the regulation of RNase L activation.

SARS-CoV-2 infection triggers widespread host mRNA decay leading to an mRNA export block.

The transcriptional induction of interferon (IFN) genes is a key feature of the mammalian antiviral response that limits viral replication and dissemination. A hallmark of severe COVID-19 disease caused by SARS-CoV-2 is the low presence of IFN proteins in patient serum despite elevated levels of IFN-encoding mRNAs, indicative of post-transcriptional inhibition of IFN protein production. Here, we performed single-molecule RNA visualization to examine the expression and localization of host mRNAs during SARS-CoV-2 infection. Our data show that the biogenesis of type I and type III IFN mRNAs is inhibited at multiple steps during SARS-CoV-2 infection. First, translocation of the interferon regulatory factor 3 (IRF3) transcription factor to the nucleus is limited in response to SARS-CoV-2, indicating that SARS-CoV-2 inhibits RLR-MAVS signaling and thus weakens transcriptional induction of IFN genes. Second, we observed that IFN mRNAs primarily localize to the site of transcription in most SARS-CoV-2 infected cells, suggesting that SARS-CoV-2 either inhibits the release of IFN mRNAs from their sites of transcription and/or triggers decay of IFN mRNAs in the nucleus upon exiting the site of transcription. Lastly, nuclear-cytoplasmic transport of IFN mRNAs is inhibited during SARS-CoV-2 infection, which we propose is a consequence of widespread degradation of host cytoplasmic basal mRNAs in the early stages of SARS-CoV-2 replication by the SARS-CoV-2 Nsp1 protein, as well as the host antiviral endoribonuclease, RNase L. Importantly, IFN mRNAs can escape SARS-CoV-2-mediated degradation if they reach the cytoplasm, making rescue of mRNA export a viable means for promoting the immune response to SARS-CoV-2.