SARS-CoV-2 3C-like protease antagonizes interferon-beta production by facilitating the degradation of IRF3.
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ABSTRACT:
SUBMITTER: Zhang W
PROVIDER: S-EPMC8413301 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:UnlabelledPorcine epidemic diarrhea virus (PEDV), a porcine enteropathogenic coronavirus, causes lethal watery diarrhea in piglets and results in large economic losses in many Asian and European countries. A large-scale outbreak of porcine epidemic diarrhea occurred in China in 2010, and the virus emerged in the United States in 2013 and spread rapidly, posing significant economic and public health concerns. Previous studies have shown that PEDV infection inhibits the synthesis of type I interferon (IFN), and viral papain-like protease 2 has been identified as an IFN antagonist. In this study, we found that the PEDV-encoded nucleocapsid (N) protein also inhibits Sendai virus-induced IFN-β production, IFN-stimulated gene expression, and activation of the transcription factors IFN regulatory factor 3 (IRF3) and NF-κB. We also found that N protein significantly impedes the activation of the IFN-β promoter stimulated by TBK1 or its upstream molecules (RIG-I, MDA5, IPS-1, and TRAF3) but does not counteract its activation by IRF3. A detailed analysis revealed that the PEDV N protein targets TBK1 by direct interaction and that this binding sequesters the association between TBK1 and IRF3, which in turn inhibits both IRF3 activation and type I IFN production. Together, our findings demonstrate a new mechanism evolved by PEDV to circumvent the host's antiviral immunity.ImportancePEDV has received increasing attention since the emergence of a PEDV variant in China and the United States. Here, we identify nucleocapsid (N) protein as a novel PEDV-encoded interferon (IFN) antagonist and demonstrate that N protein antagonizes IFN production by sequestering the interaction between IRF3 and TBK1, a critical step in type I IFN signaling. This adds another layer of complexity to the immune evasion strategies evolved by this economically important viral pathogen. An understanding of its immune evasion mechanism may direct us to novel therapeutic targets and more effective vaccines against PEDV infection.
| S-EPMC4136253 | biostudies-literature
Project description:Virus infection leads to the activation of transcription factor IRF3 and subsequent production of type I inteferons, which induce the transcription of various antiviral genes called interferon stimulated genes (ISGs) to eliminate viral infection. IRF3 activation requires phosphorylation, dimerization and nuclear translocation. However, the mechanisms for the termination of IRF3 activation in nucleus are elusive. Here we report the identification of TRIM26 to negatively regulate IFN-β production and antiviral response by targeting nuclear IRF3. TRIM26 bound to IRF3 and promoted its K48-linked polyubiquitination and degradation in nucleus. TRIM26 degraded WT IRF3 and the constitutive active mutant IRF3 5D, but not the phosphorylation deficient mutant IRF3 5A. Furthermore, IRF3 mutant in the Nuclear Localization Signal (NLS), which could not move into nucleus, was not degraded by TRIM26. Importantly, virus infection promoted TRIM26 nuclear translocation, which was required for IRF3 degradation. As a consequence, TRIM26 attenuated IFN-β promoter activation and IFN-β production downstream of TLR3/4, RLR and DNA sensing pathways. TRIM26 transgenic mice showed much less IRF3 activation and IFN-β production, while increased virus replication. Our findings delineate a novel mechanism for the termination of IRF3 activation in nucleus through TRIM26-mediated IRF3 ubiquitination and degradation.
| S-EPMC4357427 | biostudies-literature
Project description:Porcine epidemic diarrhea virus (PEDV) causes a porcine disease associated with swine epidemic diarrhea. The type I interferon (IFN-I or IFN α/β) is a key mediator of innate antiviral response during virus infection. Different antagonistic strategies have been identified and determined as to how PEDV infection inhibits the host's IFN responses to escape the host innate immune pathway, but the pathogenic mechanisms of PEDV infection are not fully elucidated. Our preliminary results revealed that endogenous TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3), the key components in the IFN signaling pathway were downregulated in PEDV infected IPEC-J2 cells by iTRAQ analysis. In this study, we screened nsp15 as the most important viral encoded protein involved in TBK1 and IRF3 reduction. Endoribonuclease (EndoU) activity has been well determined for coronavirus nsp15. Three residues (H226, H241, and K282) of PEDV nsp15 were identified as critical amino acids for PEDV EndoU but not D265, which was not well correlated with published results of other coronaviruses, such as severe acute respiratory syndrome virus (SARS-CoV). Moreover, PEDV nsp15 can directly degrade the RNA levels of TBK1 and IRF3 dependent on its EndoU activity to suppress IFN production and constrain the induction of IFN stimulated genes (ISGs), by which PEDV antagonizes the host innate response to facilitate its replication. Collectively, these results have confirmed that PEDV nsp15 was capable of subverting the IFN response by the RNA degradation of TBK1 and IRF3.
| S-EPMC7354440 | biostudies-literature
Project description:Japanese encephalitis virus (JEV) is a mosquito-borne virus and the major cause of viral encephalitis in Asia. NS1', a 52-amino acid C-terminal extension of NS1, is generated with a -1 programmed ribosomal frameshift and is only present in members of the Japanese encephalitis serogroup of flaviviruses. Previous studies demonstrated that NS1' plays a vital role in virulence, but the mechanism is unclear. In this study, an NS1' defected (rG66A) virus was generated. We found that rG66A virus was less virulent than its parent virus (pSA14) in wild-type mice. However, similar mortality caused by the two viruses was observed in an IFNAR knockout mouse model. Moreover, we found that rG66A virus induced a greater type I interferon (IFN) response than that by pSA14, and JEV NS1' significantly inhibited the production of IFN-? and IFN-stimulated genes. Taken together, our results reveal that NS1' plays a vital role in blocking type I IFN production to help JEV evade antiviral immunity and benefit viral replication.
| S-EPMC6335221 | biostudies-literature
Project description:Interferon regulatory factor 3 (IRF3), one member of the IRF family, plays a central role in induction of type I interferon (IFN) and regulation of apoptosis. Controlled activity of IRF3 is essential for its functions. During reverse transcription (RT)-PCR to clone the full-length open reading frame (ORF) of IRF3, we cloned a full-length ORF encoding an isoform of IRF3, termed as IRF3-CL, and has a unique carboxyl-terminus of 125 amino acids. IRF3-CL is ubiquitously expressed in distinct cell lines. Overexpression of IRF3-CL inhibits Sendai virus (SeV)-triggered induction of IFN-? and SeV-induced and inhibitor of NF-?B kinase-? (IKK?)-mediated nuclear translocation of IRF3. When IKK? is overexpressed, IRF3-CL is associated with IRF3. These results suggest that IRF3-CL, the alternative splicing isoform of IRF-3, may function as a negative regulator of IRF3.
| S-EPMC4002990 | biostudies-literature
Project description:3C protease (3Cpro), a chymotrypsin-like cysteine protease encoded by the foot-and-mouth disease virus (FMDV), plays an essential role in processing the FMDV P1 polyprotein into individual viral capsid proteins in FMDV replication. Previously, it has been shown that 3Cpro is involved in the blockage of the host type-I interferon (IFN) responses by FMDV. However, the underlying mechanisms are poorly understood. Here, we demonstrated that the protease activity of 3Cpro contributed to the degradation of RIG-I and MDA5, key cytosolic sensors of the type-I IFN signaling cascade in proteasome, lysosome and caspase-independent manner. And also, we examined the degradation ability on RIG-I and MDA5 of wild-type FMDV 3Cpro and FMDV 3Cpro C142T mutant which is known to significantly alter the enzymatic activity of 3Cpro. The results showed that the FMDV 3Cpro C142T mutant dramatically reduce the degradation of RIG-I and MDA5 due to weakened protease activity. Thus, the protease activity of FMDV 3Cpro governs its RIG-I and MDA5 degradation ability and subsequent negative regulation of the type-I IFN signaling. Importantly, FMD viruses harboring 3Cpro C142T mutant showed the moderate attenuation of FMDV in a pig model. In conclusion, our results indicate that a novel mechanism evolved by FMDV 3Cpro to counteract host type-I IFN responses and a rational approach to virus attenuation that could be utilized for future vaccine development.
| S-EPMC8639872 | biostudies-literature
Project description:SARS-CoV-2 is the pathogenic agent of COVID-19, which has evolved into a global pandemic. Compared with some other respiratory RNA viruses, SARS-CoV-2 is a poor inducer of type I interferon (IFN). Here, we report that SARS-CoV-2 nsp12, the viral RNA-dependent RNA polymerase (RdRp), suppresses host antiviral responses. SARS-CoV-2 nsp12 attenuated Sendai virus (SeV)- or poly(I:C)-induced IFN-β promoter activation in a dose-dependent manner. It also inhibited IFN promoter activation triggered by RIG-I, MDA5, MAVS, and IRF3 overexpression. Nsp12 did not impair IRF3 phosphorylation but suppressed the nuclear translocation of IRF3. Mutational analyses suggested that this suppression was not dependent on the polymerase activity of nsp12. Given these findings, our study reveals that SARS-CoV-2 RdRp can antagonize host antiviral innate immunity and thus provides insights into viral pathogenesis.
| S-EPMC7907794 | biostudies-literature
Project description:Human norovirus (HuNoV) is the leading cause of epidemic acute gastroenteritis worldwide. Type I interferons (IFN)-α/β are highly potent cytokines that are initially identified for their essential roles in antiviral defense. It was reported that HuNoV infection did not induce IFN-β expression but was controlled in the presence of IFN-β in human intestinal enteroids and a gnotobiotic pig model, suggesting that HuNoV has likely developed evasion countermeasures. In this study, we found that a cDNA clone of GII.4 HuNoV, the predominantly circulating genotype worldwide, inhibits the production of IFN-β and identified the viral NTPase as a key component responsible for such inhibition. HuNoV NTPase not only inhibits the activity of IFN-β promoter but also the mRNA and protein production of IFN-β. Additional studies indicate that NTPase inhibits the phosphorylation and nuclear translocation of interferon-regulatory factor-3 (IRF-3), leading to the suppression of IFN-β promoter activation. Mechanistically, NTPase interacts with IkB kinase ε (IKKε), an important factor for IRF-3 phosphorylation, and such interaction blocks the association of IKKε with unanchored K48-linked polyubiquitin chains, resulting in the inhibition of IKKε phosphorylation. Further studies demonstrated that the 1-179 aa domain of NTPase which interacts with IKKε is critical for the suppression of IFN-β production. Our findings highlight the role of HuNoV NTPase in the inhibition of IFN-β production, providing insights into a novel mechanism underlying how HuNoV evades the host innate immunity.
| S-EPMC8319745 | biostudies-literature
Project description:Equine arteritis virus (EAV) and porcine reproductive and respiratory syndrome virus (PRRSV) represent two members of the family Arteriviridae and pose major threats for the horse- and swine-breeding industries worldwide. A previous study suggested that PRRSV nsp4, a 3C-like protease, antagonizes interferon beta (IFN-β) production by cleaving the NF-κB essential modulator (NEMO) at a single site, glutamate 349 (E349). Here, we demonstrated that EAV nsp4 also inhibited virus-induced IFN-β production by targeting NEMO for proteolytic cleavage and that the scission occurred at four sites: E166, E171, glutamine 205 (Q205), and E349. Additionally, we found that, besides the previously reported cleavage site E349 in NEMO, scission by PRRSV nsp4 took place at two additional sites, E166 and E171. These results imply that while cleaving NEMO is a common strategy utilized by EAV and PRRSV nsp4 to antagonize IFN induction, EAV nsp4 adopts a more complex substrate recognition mechanism to target NEMO. By analyzing the abilities of the eight different NEMO fragments resulting from EAV or PRRSV nsp4 scission to induce IFN-β production, we serendipitously found that a NEMO fragment (residues 1 to 349) could activate IFN-β transcription more robustly than full-length NEMO, whereas all other NEMO cleavage products were abrogated for the IFN-β-inducing capacity. Thus, NEMO cleavage at E349 alone may not be sufficient to completely inactivate the IFN response via this signaling adaptor. Altogether, our findings suggest that EAV and PRRSV nsp4 cleave NEMO at multiple sites and that this strategy is critical for disarming the innate immune response for viral survival.IMPORTANCE The arterivirus nsp4-encoded 3C-like protease (3CLpro) plays an important role in virus replication and immune evasion, making it an attractive target for antiviral therapeutics. Previous work suggested that PRRSV nsp4 suppresses type I IFN production by cleaving NEMO at a single site. In contrast, the present study demonstrates that both EAV and PRRSV nsp4 cleave NEMO at multiple sites and that this strategy is essential for disruption of type I IFN production. Moreover, we reveal that EAV nsp4 also cleaves NEMO at glutamine 205 (Q205), which is not targeted by PRRSV nsp4. Notably, targeting a glutamine in NEMO for cleavage has been observed only with picornavirus 3C proteases (3Cpro) and coronavirus 3CLpro In aggregate, our work expands knowledge of the innate immune evasion mechanisms associated with NEMO cleavage by arterivirus nsp4 and describes a novel substrate recognition characteristic of EAV nsp4.
| S-EPMC6613749 | biostudies-literature
Project description:Coronaviruses (CoVs), such as human coronavirus NL63 (HCoV-NL63), severe acute respiratory syndrome CoV (SARS-CoV), murine hepatitis virus (MHV), porcine epidemic diarrhea virus (PEDV), and Middle East Respiratory Syndrome Coronavirus (MERS-CoV), encode papain-like (PL) proteases that inhibit Sendai virus- (SeV-) induced interferon (IFN-?) production. Recently, the crystal structure of transmissible gastroenteritis virus (TGEV) PL1 has been solved, which was similar to that of SARS-CoV PL2pro, which may antagonize host innate immunity. However, very little is known about whether TGEV PL1 can antagonize host innate immune response. Here, we presented evidence that TGEV PL1 encoded by the replicase gene could suppress the IFN-? expression and inhibit the nuclear translocation of interferon regulatory factor 3 (IRF3). The ability to antagonize IFN-? production was dependent on the intact catalytic activity of PL1. Furthermore, TGEV PL1 exerted deubiquitinase (DUB) activity which strongly inhibited the retinoic acid-induced gene I- (RIG-1-) and stimulator of interferon gene- (STING-) dependent IFN expression. Our data collectively suggest that TGEV PL1 can inhibit the IFN-? expression and interfere with RIG-1- and STING-mediated signaling through a viral DUB activity. Our study has yielded strong evidence for the TGEV PL1 mechanisms that counteract the host innate immunity.
| S-EPMC5672592 | biostudies-literature