scholarly journals Infectious bronchitis virus regulates cellular stress granule signaling

2019 ◽  
Author(s):  
Matthew J. Brownsword ◽  
Nicole Doyle ◽  
Michèle Brocard ◽  
Nicolas Locker ◽  
Helena J. Maier
Keyword(s):  
Stress Responses ◽  
Infectious Bronchitis Virus ◽  
Cellular Stress ◽  
Stress Granules ◽  
Stress Granule ◽  
Granule Formation ◽  
Translation Machinery ◽  
Infectious Bronchitis ◽  
Eif2α Phosphorylation ◽  
Cellular Translation

AbstractViruses must hijack cellular translation machinery to efficiently express viral genes. In many cases, this is impeded by cellular stress responses. These stress responses swiftly relocate and repurpose translation machinery, resulting in global inhibition of translation and the aggregation of stalled 48S mRNPs into cytoplasmic foci called stress granules. This results in translational silencing of all mRNAs excluding those beneficial for the cell to resolve the specific stress. For example, expression of antiviral factors is maintained during viral infection. Here we investigated stress granule regulation by Gammacoronavirus infectious bronchitis virus (IBV), which causes the economically important poultry disease, infectious bronchitis. Interestingly, we found that IBV is able to inhibit multiple cellular stress granule signaling pathways whilst at the same time IBV replication also results in induction of seemingly canonical stress granules in a proportion of infected cells. Moreover, IBV infection uncouples translational repression and stress granule formation and both processes are independent of eIF2α phosphorylation. These results provide novel insights into how IBV modulates cellular translation and antiviral stress signaling.

scholarly journals Infectious Bronchitis Virus Regulates Cellular Stress Granule Signaling

Viruses ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 536
Author(s):  
Matthew J. Brownsword ◽  
Nicole Doyle ◽  
Michèle Brocard ◽  
Nicolas Locker ◽  
Helena J. Maier
Keyword(s):  
Stress Responses ◽  
Infectious Bronchitis Virus ◽  
Cellular Stress ◽  
Stress Granules ◽  
Stress Granule ◽  
Granule Formation ◽  
Infectious Bronchitis ◽  
Eif2α Phosphorylation ◽  
Cellular Translation ◽  
Cellular Stress Responses

Viruses must hijack cellular translation machinery to express viral genes. In many cases, this is impeded by cellular stress responses. These stress responses result in the global inhibition of translation and the storage of stalled mRNAs, into RNA-protein aggregates called stress granules. This results in the translational silencing of the majority of mRNAs excluding those beneficial for the cell to resolve the specific stress. For example, the expression of antiviral factors is maintained during viral infection. Here we investigated stress granule regulation by Gammacoronavirus infectious bronchitis virus (IBV), which causes the economically important poultry disease, infectious bronchitis. Interestingly, we found that IBV is able to inhibit multiple cellular stress granule signaling pathways, whilst at the same time, IBV replication also results in the induction of seemingly canonical stress granules in a proportion of infected cells. Moreover, IBV infection uncouples translational repression and stress granule formation and both processes are independent of eIF2α phosphorylation. These results provide novel insights into how IBV modulates cellular translation and antiviral stress signaling.

scholarly journals Inhibition of Anti-viral Stress Granule Formation by infectious bronchitis virus endoribonuclease nsp15 Ensures Efficient Virus Replication

2020 ◽  
Author(s):  
Bo Gao ◽  
Xiaoqian Gong ◽  
Shouguo Fang ◽  
Wenlian Weng ◽  
Yingjie Sun ◽  
...  
Keyword(s):  
Virus Replication ◽  
Infectious Bronchitis Virus ◽  
Rna Virus ◽  
Animal Health ◽  
Stress Granules ◽  
Antiviral Response ◽  
Viral Dsrna ◽  
Granule Formation ◽  
Infectious Bronchitis ◽  
Eif2α Phosphorylation

AbstractCytoplasmic stress granules (SGs) are generally triggered by stress-induced translation arrest for storing mRNAs. Recently, it has been shown that SGs exert anti-viral functions due to their involvement in protein synthesis shut off and recruitment of innate immune signaling intermediates. The largest RNA virus, coronavirus, mutates frequently and circulates among animals, imposing great threat to public safety and animal health; however, the significance of SGs in coronavirus infections is largely unknown. Infectious bronchitis virus (IBV) is the first identified coronavirus in 1930s and has been prevalent in poultry farm for many years. In this study, we provide evidence that IBV overcomes the host antiviral response by inhibiting SGs formation via the virus-encoded endoribonuclease nsp15. By immunofluorescence analysis, we observed that IBV infection not only did not trigger SGs formation in approximately 80% of the infected cells, but also impaired the formation of SGs triggered by heat shock, sodium arsenite, or NaCl stimuli. We show that the intrinsic endoribonuclease activity of nsp15 is responsible for the inhibition of SGs formation. In fact, nsp15-defective recombinant IBV (rIBV-nsp15-H238A) greatly induced the formation of SGs, along with accumulation of dsRNA and activation of PKR, whereas wild type IBV failed to do so. Consequently, infection with rIBV-nsp15-H238A triggered transcription of IFN-β which in turn greatly affected recombinant virus replication. Further analysis showed that SGs function as antiviral hub, as demonstrated by the attenuated IRF3-IFN response and increased production of IBV in SG-defective cells. Additional evidence includes the aggregation of PRRs and signaling intermediates to the IBV-induced SGs. Collectively, our data demonstrate that the endoribonuclease nsp15 of IBV suppresses the formation of antiviral hub SGs by regulating the accumulation of viral dsRNA and by antagonizing the activation of PKR, eventually ensuring productive virus replication. We speculate that coronaviruses employ similar mechanisms to antagonize the host anti-viral SGs formation for efficient virus replication, as the endoribonuclease function of nsp15 is conserved in all coronaviruses.Author summaryIt has been reported that stress granules (SGs) are part of the host cell antiviral response. Not surprisingly, viruses in turn produce an array of antagonists to counteract such host response. Here, we show that IBV inhibits the formation of SGs through its endoribonuclease nsp15, by reducing the accumulation of viral dsRNA, evading the activation of PKR, and by subsequently inhibiting eIF2α phosphorylation and SGs formation. Nsp15 also inhibits SG formation independent of the eIF2α pathway, probably by targeting host mRNA. Depletion of SG scaffold proteins decreases IRF3-IFN response and increases the production of IBV. All coronaviruses encode a conserved endoribonuclease nsp15, and it will be important to determine whether also other (non-avian) coronaviruses limit the formation of anti-viral SGs in a similar manner.

scholarly journals Puumala and Andes Orthohantaviruses Cause Transient Protein Kinase R-Dependent Formation of Stress Granules

2019 ◽  
Vol 94 (3) ◽  
Author(s):  
Wanda Christ ◽  
Janne Tynell ◽  
Jonas Klingström
Keyword(s):  
Protein Kinase ◽  
Host Cell ◽  
Stress Responses ◽  
Stress Granules ◽  
Cell Stress ◽  
Stress Granule ◽  
Hantavirus Infection ◽  
Stress Signaling ◽  
Protein Kinase R ◽  
Granule Formation

ABSTRACT Virus infection frequently triggers host cell stress signaling resulting in translational arrest; as a consequence, many viruses employ means to modulate the host stress response. Hantaviruses are negative-sense, single-stranded RNA viruses known to inhibit host innate immune responses and apoptosis, but their impact on host cell stress signaling remains largely unknown. In this study, we investigated activation of host cell stress responses during hantavirus infection. We show that hantavirus infection causes transient formation of stress granules (SGs) but does so in only a limited proportion of infected cells. Our data indicate some cell type-specific and hantavirus species-specific variability in SG prevalence and show SG formation to be dependent on the activation of protein kinase R (PKR). Hantavirus infection inhibited PKR-dependent SG formation, which could account for the transient nature and low prevalence of SG formation observed during hantavirus infection. In addition, we report only limited colocalization of hantaviral proteins or RNA with SGs and show evidence indicating hantavirus-mediated inhibition of PKR-like endoplasmic reticulum (ER) kinase (PERK). IMPORTANCE Our work presents the first report on stress granule formation during hantavirus infection. We show that hantavirus infection actively inhibits stress granule formation, thereby escaping the detrimental effects on global translation imposed by host stress signaling. Our results highlight a previously uncharacterized aspect of hantavirus-host interactions with possible implications for how hantaviruses are able to cause persistent infection in natural hosts and for pathogenesis.

scholarly journals Infectious bronchitis virus modulates cellular stress granule signalling

2019 ◽  
Vol 1 (1A) ◽  
Author(s):  
Matthew Brownsword ◽  
Nicole Doyle ◽  
Michele Brocard ◽  
Nicolas Locker ◽  
Helena Maier
Keyword(s):  
Infectious Bronchitis Virus ◽  
Cellular Stress ◽  
Stress Granule ◽  
Infectious Bronchitis

scholarly journals THE TRANSCRIPTIONAL RESPONSE TO OXIDATIVE STRESS IS INDEPENDENT OF STRESS-GRANULE FORMATION

2021 ◽  
Author(s):  
Amanjot Singh ◽  
Arvind Reddy Kandi ◽  
Deepa Jayaprakashappa ◽  
Guillaume Thuery ◽  
Devam J Purohit ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Stress Responses ◽  
Transcriptional Response ◽  
Stress Granules ◽  
Stress Granule ◽  
Cell Recovery ◽  
Granule Formation ◽  
Transcriptional Changes ◽  
Translational Arrest ◽  
Shock Proteins

ABSTRACTCells respond to stress with translational arrest, robust transcriptional changes, and transcription-independent formation of mRNP assemblies termed stress granules (SGs). Despite considerable interest in the role of SGs in oxidative, unfolded-protein, and viral stress responses, whether and how SGs contribute to stress-induced transcription has not been rigorously examined. To address this issue, we characterized transcriptional changes in Drosophila S2 cells induced by acute oxidative-stress and assessed how these were altered under conditions that disrupted SG assembly. Sodium-arsenite stress for 3 hours predominantly resulted in the induction or upregulation of stress-responsive mRNAs whose levels peaked during cell recovery after stress cessation. The stress-transcriptome is enriched in mRNAs coding for protein chaperones, including HSP70 and low molecular-weight heat shock proteins, glutathione transferases, and several non-coding RNAs. Oxidative stress also induced prominent cytoplasmic stress granules that disassembled 3-hours after stress cessation. As expected, RNAi-mediated knockdown of the conserved G3BP1/ Rasputin protein inhibited stress-granule assembly. However, this disruption had no significant effect on the stress-induced transcriptional response or stress-induced translational arrest. Thus, SG assembly and stress-induced effects on gene expression appear to be driven by distinctive signaling processes. We suggest that while SG assembly represents a fast, transient mechanism, the transcriptional response enables a slower, longer-lasting mechanism for adaptation to and recovery from cell stress.

scholarly journals Stress Granule Formation Attenuates RACK1-Mediated Apoptotic Cell Death Induced by Morusin

2020 ◽  
Vol 21 (15) ◽  
pp. 5360
Author(s):  
Ye-Jin Park ◽  
Dong Wook Choi ◽  
Sang Woo Cho ◽  
Jaeseok Han ◽  
Siyoung Yang ◽  
...  
Keyword(s):  
Cell Death ◽  
Apoptotic Cell ◽  
Stress Granules ◽  
Protective Role ◽  
Stress Granule ◽  
Protein Kinase R ◽  
Granule Formation ◽  
Protective Function ◽  
Eif2α Phosphorylation

Stress granules are membraneless organelles composed of numerous components including ribonucleoproteins. The stress granules are characterized by a dynamic complex assembly in response to various environmental stressors, which has been implicated in the coordinated regulation of diverse biological pathways, to exert a protective role against stress-induced cell death. Here, we show that stress granule formation is induced by morusin, a novel phytochemical displaying antitumor capacity through barely known mechanisms. Morusin-mediated induction of stress granules requires activation of protein kinase R (PKR) and subsequent eIF2α phosphorylation. Notably, genetic inactivation of stress granule formation mediated by G3BP1 knockout sensitized cancer cells to morusin treatment. This protective function against morusin-mediated cell death can be attributed at least in part to the sequestration of receptors for activated C kinase-1 (RACK1) within the stress granules, which reduces caspase-3 activation. Collectively, our study provides biochemical evidence for the role of stress granules in suppressing the antitumor capacity of morusin, proposing that morusin treatment, together with pharmacological inhibition of stress granules, could be an efficient strategy for targeting cancer.

scholarly journals The Leader Protein of Theiler's Virus Prevents the Activation of PKR

2019 ◽  
Vol 93 (19) ◽  
Author(s):  
Fabian Borghese ◽  
Frédéric Sorgeloos ◽  
Teresa Cesaro ◽  
Thomas Michiels
Keyword(s):  
Stress Granule ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Wild Type Virus ◽  
Granule Formation ◽  
Double Stranded Rna ◽  
Eif2α Phosphorylation ◽  
Leader Protein ◽  
Infected Cells ◽  
L Proteins

ABSTRACT Leader (L) proteins encoded by cardioviruses are multifunctional proteins that contribute to innate immunity evasion. L proteins of Theiler’s murine encephalomyelitis virus (TMEV), Saffold virus (SAFV), and encephalomyocarditis virus (EMCV) were reported to inhibit stress granule assembly in infected cells. Here, we show that TMEV L can act at two levels in the stress granule formation pathway: on the one hand, it can inhibit sodium arsenite-induced stress granule assembly without preventing eIF2α phosphorylation and, thus, acts downstream of eIF2α; on the other hand, it can inhibit eucaryotic translation initiation factor 2 alpha kinase 2 (PKR) activation and the consequent PKR-mediated eIF2α phosphorylation. Interestingly, coimmunostaining experiments revealed that PKR colocalizes with viral double-stranded RNA (dsRNA) in cells infected with L-mutant viruses but not in cells infected with the wild-type virus. Furthermore, PKR coprecipitated with dsRNA from cells infected with L-mutant viruses significantly more than from cells infected with the wild-type virus. These data strongly suggest that L blocks PKR activation by preventing the interaction between PKR and viral dsRNA. In infected cells, L also rendered PKR refractory to subsequent activation by poly(I·C). However, no interaction was observed between L and either dsRNA or PKR. Taken together, our results suggest that, unlike other viral proteins, L indirectly acts on PKR to negatively regulate its responsiveness to dsRNA. IMPORTANCE The leader (L) protein encoded by cardioviruses is a very short multifunctional protein that contributes to evasion of the host innate immune response. This protein notably prevents the formation of stress granules in infected cells. Using Theiler’s virus as a model, we show that L proteins can act at two levels in the stress response pathway leading to stress granule formation, the most striking one being the inhibition of eucaryotic translation initiation factor 2 alpha kinase 2 (PKR) activation. Interestingly, the leader protein appears to inhibit PKR via a novel mechanism by rendering this kinase unable to detect double-stranded RNA, its typical activator. Unlike other viral proteins, such as influenza virus NS1, the leader protein appears to interact with neither PKR nor double-stranded RNA, suggesting that it acts indirectly to trigger the inhibition of the kinase.

scholarly journals The Stress Granule Protein G3BP1 Recruits Protein Kinase R To Promote Multiple Innate Immune Antiviral Responses

2014 ◽  
Vol 89 (5) ◽  
pp. 2575-2589 ◽  
Author(s):  
Lucas C. Reineke ◽  
Richard E. Lloyd
Keyword(s):  
Protein Kinase ◽  
Antiviral Activity ◽  
Cellular Stress ◽  
Stress Granules ◽  
Innate Immune ◽  
Stress Granule ◽  
Productive Infection ◽  
Antiviral Protein ◽  
Protein Kinase R ◽  
Transcriptional Responses

ABSTRACTStress granules (SGs) are cytoplasmic storage sites containing translationally silenced mRNPs that can be released to resume translation after stress subsides. We previously showed that poliovirus 3C proteinase cleaves the SG-nucleating protein G3BP1, blocking the ability of cells to form SGs late in infection. Many other viruses also target G3BP1 and inhibit SG formation, but the reasons why these functions evolved are unclear. Previously, we also showed a link between G3BP1-induced SGs and protein kinase R (PKR)-mediated translational control, but the mechanism of PKR interplay with SG and the antiviral consequences are unknown. Here, we show that G3BP1 exhibits antiviral activity against several enteroviruses, whereas truncated G3BP1 that cannot form SGs does not. G3BP1-induced SGs are linked to activation of innate immune transcriptional responses through NF-κB and JNK. The G3BP1-induced SGs also recruit PKR and other antiviral proteins. We show that the PXXP domain within G3BP1 is essential for the recruitment of PKR to SGs, for eIF2α phosphorylation driven by PKR, and for nucleating SGs of normal composition. We also show that deletion of the PXXP domain in G3BP1 compromises its antiviral activity. These findings tie PKR activation to its recruitment to SGs by G3BP1 and indicate that G3BP1 promotes innate immune responses at both the transcriptional and translational levels and integrates cellular stress responses and innate immunity.IMPORTANCEStress granules appear during virus infection, and their importance is not well understood. Previously, it was assumed that they were nonfunctional artifacts associated with cellular stress. PKR is a well-known antiviral protein; however, its regulation in cells is not well understood. Our work links cellular stress granules with activation of PKR and other innate immune pathways through the activity of G3BP1, a critical stress granule component. The ability of stress granules and G3BP1 to activate PKR and other innate immune transcriptional responses indicates that G3BP1 is an antiviral protein. This work helps to refine a longstanding paradigm indicating stress granules are inert structures and explains why G3BP1 is subverted by many viruses to promote a productive infection.

scholarly journals Retinoic Acid Inducible Gene I and Protein Kinase R, but Not Stress Granules, Mediate the Proinflammatory Response to Yellow Fever Virus

2020 ◽  
Vol 94 (22) ◽  
Author(s):  
Guillaume Beauclair ◽  
Felix Streicher ◽  
Maxime Chazal ◽  
Daniela Bruni ◽  
Sarah Lesage ◽  
...  
Keyword(s):  
Yellow Fever ◽  
Yellow Fever Virus ◽  
Rna Virus ◽  
Stress Granules ◽  
Cytokine Response ◽  
Fever Virus ◽  
Stress Granule ◽  
Proinflammatory Response ◽  
Granule Formation ◽  
Proinflammatory Mediators

ABSTRACT Yellow fever virus (YFV) is an RNA virus primarily targeting the liver. Severe YF cases are responsible for hemorrhagic fever, plausibly precipitated by excessive proinflammatory cytokine response. Pathogen recognition receptors (PRRs), such as the cytoplasmic retinoic acid inducible gene I (RIG-I)-like receptors (RLRs), and the viral RNA sensor protein kinase R (PKR), are known to initiate a proinflammatory response upon recognition of viral genomes. Here, we sought to reveal the main determinants responsible for the acute cytokine expression occurring in human hepatocytes following YFV infection. Using a RIG-I-defective human hepatoma cell line, we found that RIG-I largely contributes to cytokine secretion upon YFV infection. In infected RIG-I-proficient hepatoma cells, RIG-I was localized in stress granules. These granules are large aggregates of stalled translation preinitiation complexes known to concentrate RLRs and PKR and are so far recognized as hubs orchestrating RNA virus sensing. Stable knockdown of PKR in hepatoma cells revealed that PKR contributes to both stress granule formation and cytokine induction upon YFV infection. However, stress granule disruption did not affect the cytokine response to YFV infection, as assessed by small interfering RNA (siRNA)-knockdown-mediated inhibition of stress granule assembly. Finally, no viral RNA was detected in stress granules using a fluorescence in situ hybridization approach coupled with immunofluorescence. Our findings suggest that both RIG-I and PKR mediate proinflammatory cytokine induction in YFV-infected hepatocytes, in a stress granule-independent manner. Therefore, by showing the uncoupling of the cytokine response from the stress granule formation, our model challenges the current view in which stress granules are required for the mounting of the acute antiviral response. IMPORTANCE Yellow fever is a mosquito-borne acute hemorrhagic disease caused by yellow fever virus (YFV). The mechanisms responsible for its pathogenesis remain largely unknown, although increased inflammation has been linked to worsened outcome. YFV targets the liver, where it primarily infects hepatocytes. We found that two RNA-sensing proteins, RIG-I and PKR, participate in the induction of proinflammatory mediators in human hepatocytes infected with YFV. We show that YFV infection promotes the formation of cytoplasmic structures, termed stress granules, in a PKR- but not RIG-I-dependent manner. While stress granules were previously postulated to be essential platforms for immune activation, we found that they are not required for the production of proinflammatory mediators upon YFV infection. Collectively, our work uncovered molecular events triggered by the replication of YFV, which could prove instrumental in clarifying the pathogenesis of the disease, with possible repercussions for disease management.

scholarly journals Stable Formation of Compositionally Unique Stress Granules in Virus-Infected Cells

2009 ◽  
Vol 84 (7) ◽  
pp. 3654-3665 ◽  
Author(s):  
Joanna Piotrowska ◽  
Spencer J. Hansen ◽  
Nogi Park ◽  
Katarzyna Jamka ◽  
Peter Sarnow ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Heat Shock ◽  
Viral Infections ◽  
De Novo ◽  
Stress Granules ◽  
Stress Granule ◽  
Initiation Factor ◽  
Granule Formation ◽  
Infected Cells ◽  
Positive Strand Rna

ABSTRACT Stress granules are sites of mRNA storage formed in response to a variety of stresses, including viral infections. Here, the mechanisms and consequences of stress granule formation during poliovirus infection were examined. The results indicate that stress granules containing T-cell-restricted intracellular antigen 1 (TIA-1) and mRNA are stably constituted in infected cells despite lacking intact RasGAP SH3-domain binding protein 1 (G3BP) and eukaryotic initiation factor 4G. Fluorescent in situ hybridization revealed that stress granules in infected cells do not contain significant amounts of viral positive-strand RNA. Infection does not prevent stress granule formation in response to heat shock, indicating that poliovirus does not block de novo stress granule formation. A mutant TIA-1 protein that prevents stress granule formation during oxidative stress also prevents formation in infected cells. However, stress granule formation during infection is more dependent upon ongoing transcription than is formation during oxidative stress or heat shock. Furthermore, Sam68 is recruited to stress granules in infected cells but not to stress granules formed in response to oxidative stress or heat shock. These results demonstrate that stress granule formation in poliovirus-infected cells utilizes a transcription-dependent pathway that results in the appearance of stable, compositionally unique stress granules.

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