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1.
J Immunol ; 208(3): 753-761, 2022 02 01.
Article in English | MEDLINE | ID: covidwho-1614089

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), has seriously threatened global public health. Severe COVID-19 has been reported to be associated with an impaired IFN response. However, the mechanisms of how SARS-CoV-2 antagonizes the host IFN response are poorly understood. In this study, we report that SARS-CoV-2 helicase NSP13 inhibits type I IFN production by directly targeting TANK-binding kinase 1 (TBK1) for degradation. Interestingly, inhibition of autophagy by genetic knockout of Beclin1 or pharmacological inhibition can rescue NSP13-mediated TBK1 degradation in HEK-293T cells. Subsequent studies revealed that NSP13 recruits TBK1 to p62, and the absence of p62 can also inhibit TBK1 degradation in HEK-293T and HeLa cells. Finally, TBK1 and p62 degradation and p62 aggregation were observed during SARS-CoV-2 infection in HeLa-ACE2 and Calu3 cells. Overall, our study shows that NSP13 inhibits type I IFN production by recruiting TBK1 to p62 for autophagic degradation, enabling it to evade the host innate immune response, which provides new insights into the transmission and pathogenesis of SARS-CoV-2 infection.


Subject(s)
Autophagy , COVID-19/immunology , Interferon Type I/biosynthesis , Methyltransferases/physiology , RNA Helicases/physiology , SARS-CoV-2/physiology , Sequestosome-1 Protein/metabolism , Viral Nonstructural Proteins/physiology , Beclin-1/antagonists & inhibitors , Cell Line , Down-Regulation , Humans , Immune Evasion , Immunity, Innate , Immunoprecipitation , Interferon Type I/genetics , Multiprotein Complexes , Protein Aggregates , Protein Interaction Mapping
2.
Front Immunol ; 12: 756262, 2021.
Article in English | MEDLINE | ID: covidwho-1551507

ABSTRACT

A male sex bias has emerged in the COVID-19 pandemic, fitting to the sex-biased pattern in other viral infections. Males are 2.84 times more often admitted to the ICU and mortality is 1.39 times higher as a result of COVID-19. Various factors play a role in this, and novel studies suggest that the gene-dose of Toll-Like Receptor (TLR) 7 could contribute to the sex-skewed severity. TLR7 is one of the crucial pattern recognition receptors for SARS-CoV-2 ssRNA and the gene-dose effect is caused by X chromosome inactivation (XCI) escape. Female immune cells with TLR7 XCI escape have biallelic TLR7 expression and produce more type 1 interferon (IFN) upon TLR7 stimulation. In COVID-19, TLR7 in plasmacytoid dendritic cells is one of the pattern recognition receptors responsible for IFN production and a delayed IFN response has been associated with immunopathogenesis and mortality. Here, we provide a hypothesis that females may be protected to some extend against severe COVID-19, due to the biallelic TLR7 expression, allowing them to mount a stronger and more protective IFN response early after infection. Studies exploring COVID-19 treatment via the TLR7-mediated IFN pathway should consider this sex difference. Various factors such as age, sex hormones and escape modulation remain to be investigated concerning the TLR7 gene-dose effect.


Subject(s)
COVID-19/mortality , Gene Dosage/genetics , Interferon Type I/biosynthesis , Toll-Like Receptor 7/genetics , Toll-Like Receptor 7/metabolism , COVID-19/drug therapy , COVID-19/pathology , Chromosomes, Human, X/genetics , Critical Care/statistics & numerical data , Dendritic Cells/immunology , Female , Humans , Interferon Type I/immunology , Male , RNA, Viral/genetics , Receptors, Pattern Recognition/genetics , Receptors, Pattern Recognition/metabolism , Risk Factors , SARS-CoV-2/immunology , Sex Factors , Signal Transduction/immunology , X Chromosome Inactivation/genetics
3.
Front Immunol ; 12: 750279, 2021.
Article in English | MEDLINE | ID: covidwho-1551505

ABSTRACT

SARS-CoV-2 coronavirus infection induces heterogeneous symptoms, ranging from asymptomatic to lethal forms. Severe forms usually occur in the elderly and/or individuals with comorbidities. Children generally remain asymptomatic to primary infection, suggesting that they may have an effective local innate immune response. IFN-I and -III have non-redundant protective roles against SARS-CoV-2, although sometimes damaging the host. The expression and role of anti-viral peptides during SARS-CoV-2 infection have thus far been little studied. We aimed to identify the innate immune molecules present at the SARS-CoV-2 entry point. We analyzed the mRNA levels of type I (IFN-α and -ß) and type III (IFN-λ1-3) interferons and selected antiviral peptides (i.e., ß-defensins 1-3, α-defensins [HNP1-3, HD5] pentraxin-3, surfactant protein D, the cathelicidin LL-37 and interleukin-26) in nasopharyngeal swabs from 226 individuals of various ages, either infected with SARS-CoV-2 (symptomatic or asymptomatic) or negative for the virus. We observed that infection induced selective upregulation of IFN-λ1 expression in pediatric subjects (≤15 years), whereas IFN-α, IFN-ß, IFN-λ2/λ3, and ß-defensin 1-3 expression was unaffected. Conversely, infection triggered upregulation of IFN-α, IFN-ß, IFN-λ2/λ3, and ß-defensin 1-3 mRNA expression in adults (15-65 years) and the elderly (≥ 65 years), but without modulation of IFN-λ1. The expression of these innate molecules was not associated with gender or symptoms. Expression of the interferon-stimulated genes IFITM1 and IFITM3 was upregulated in SARS-CoV-2-positive subjects and reached similar levels in the three age groups. Finally, age-related differences in nasopharyngeal innate immunity were also observed in SARS-CoV-2-negative subjects. This study shows that the expression patterns of IFN-I/-III and certain anti-viral molecules in the nasopharyngeal mucosa of SARS-CoV-2-infected subjects differ with age and suggests that susceptibility to SARS-CoV-2 may be related to intrinsic differences in the nature of mucosal anti-viral innate immunity.


Subject(s)
/analysis , Interferon Type I/biosynthesis , Interferon-gamma/biosynthesis , Nasal Mucosa/immunology , SARS-CoV-2/immunology , beta-Defensins/biosynthesis , Adolescent , Adult , Age Factors , Aged , COVID-19/immunology , Cells, Cultured , Female , Humans , Immunity, Innate/immunology , Interferon Type I/immunology , Interferon-gamma/immunology , Interferons/biosynthesis , Interferons/immunology , Interleukins/biosynthesis , Interleukins/immunology , Male , Middle Aged , Nasopharynx/immunology , Young Adult , beta-Defensins/immunology
4.
PLoS Comput Biol ; 17(11): e1009587, 2021 11.
Article in English | MEDLINE | ID: covidwho-1533401

ABSTRACT

Patients with coronavirus disease 2019 (COVID-19) often exhibit diverse disease progressions associated with various infectious ability, symptoms, and clinical treatments. To systematically and thoroughly understand the heterogeneous progression of COVID-19, we developed a multi-scale computational model to quantitatively understand the heterogeneous progression of COVID-19 patients infected with severe acute respiratory syndrome (SARS)-like coronavirus (SARS-CoV-2). The model consists of intracellular viral dynamics, multicellular infection process, and immune responses, and was formulated using a combination of differential equations and stochastic modeling. By integrating multi-source clinical data with model analysis, we quantified individual heterogeneity using two indexes, i.e., the ratio of infected cells and incubation period. Specifically, our simulations revealed that increasing the host antiviral state or virus induced type I interferon (IFN) production rate can prolong the incubation period and postpone the transition from asymptomatic to symptomatic outcomes. We further identified the threshold dynamics of T cell exhaustion in the transition between mild-moderate and severe symptoms, and that patients with severe symptoms exhibited a lack of naïve T cells at a late stage. In addition, we quantified the efficacy of treating COVID-19 patients and investigated the effects of various therapeutic strategies. Simulations results suggested that single antiviral therapy is sufficient for moderate patients, while combination therapies and prevention of T cell exhaustion are needed for severe patients. These results highlight the critical roles of IFN and T cell responses in regulating the stage transition during COVID-19 progression. Our study reveals a quantitative relationship underpinning the heterogeneity of transition stage during COVID-19 progression and can provide a potential guidance for personalized therapy in COVID-19 patients.


Subject(s)
COVID-19/etiology , SARS-CoV-2 , Antiviral Agents/therapeutic use , COVID-19/immunology , COVID-19/therapy , Computational Biology , Computer Simulation , Disease Progression , Host Microbial Interactions/immunology , Humans , Interferon Type I/biosynthesis , Lymphocyte Activation , Models, Immunological , Models, Statistical , Pandemics/statistics & numerical data , Prognosis , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Severity of Illness Index , T-Lymphocytes/immunology , Treatment Outcome
5.
Viruses ; 13(10)2021 10 14.
Article in English | MEDLINE | ID: covidwho-1470993

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a member of the Coronaviridae family, which is responsible for the COVID-19 pandemic followed by unprecedented global societal and economic disruptive impact. The innate immune system is the body's first line of defense against invading pathogens and is induced by a variety of cellular receptors that sense viral components. However, various strategies are exploited by SARS-CoV-2 to disrupt the antiviral innate immune responses. Innate immune dysfunction is characterized by the weak generation of type I interferons (IFNs) and the hypersecretion of pro-inflammatory cytokines, leading to mortality and organ injury in patients with COVID-19. This review summarizes the existing understanding of the mutual effects between SARS-CoV-2 and the type I IFN (IFN-α/ß) responses, emphasizing the relationship between host innate immune signaling and viral proteases with an insight on tackling potential therapeutic targets.


Subject(s)
COVID-19/immunology , Immune Evasion/immunology , Immunity, Innate/immunology , Interferon Type I/immunology , SARS-CoV-2/immunology , Antiviral Agents/therapeutic use , COVID-19/drug therapy , COVID-19/pathology , Cytokines/metabolism , Drug Combinations , Humans , Interferon Type I/biosynthesis , Lopinavir/therapeutic use , Ribavirin/therapeutic use , Ritonavir/therapeutic use , Signal Transduction/immunology
6.
Viruses ; 13(9)2021 09 15.
Article in English | MEDLINE | ID: covidwho-1411083

ABSTRACT

Type I Interferons (IFN-I) are a family of potent antiviral cytokines that act through the direct restriction of viral replication and by enhancing antiviral immunity. However, these powerful cytokines are a caged lion, as excessive and sustained IFN-I production can drive immunopathology during infection, and aberrant IFN-I production is a feature of several types of autoimmunity. As specialized producers of IFN-I plasmacytoid (p), dendritic cells (DCs) can secrete superb quantities and a wide breadth of IFN-I isoforms immediately after infection or stimulation, and are the focus of this review. Notably, a few days after viral infection pDCs tune down their capacity for IFN-I production, producing less cytokines in response to both the ongoing infection and unrelated secondary stimulations. This process, hereby referred to as "pDC exhaustion", favors viral persistence and associates with reduced innate responses and increased susceptibility to secondary opportunistic infections. On the other hand, pDC exhaustion may be a compromise to avoid IFN-I driven immunopathology. In this review we reflect on the mechanisms that initially induce IFN-I and subsequently silence their production by pDCs during a viral infection. While these processes have been long studied across numerous viral infection models, the 2019 coronavirus disease (COVID-19) pandemic has brought their discussion back to the fore, and so we also discuss emerging results related to pDC-IFN-I production in the context of COVID-19.


Subject(s)
COVID-19/metabolism , COVID-19/virology , Host-Pathogen Interactions , Interferon Type I/biosynthesis , SARS-CoV-2/physiology , Biomarkers , COVID-19/immunology , Cytokines/metabolism , Dendritic Cells/immunology , Dendritic Cells/metabolism , Host-Pathogen Interactions/immunology , Humans , Immunity, Innate/immunology , Immunomodulation , Toll-Like Receptors/metabolism
7.
Int J Mol Sci ; 22(9)2021 Apr 28.
Article in English | MEDLINE | ID: covidwho-1359279

ABSTRACT

Deeply understanding the virus-host interaction is a prerequisite for developing effective anti-viral strategies. Traditionally, the transporter associated with antigen processing type 1 (TAP1) is critical for antigen presentation to regulate adaptive immunity. However, its role in controlling viral infections through modulating innate immune signaling is not yet fully understood. In the present study, we reported that TAP1, as a product of interferon-stimulated genes (ISGs), had broadly antiviral activity against various viruses such as herpes simplex virus 1 (HSV-1), adenoviruses (AdV), vesicular stomatitis virus (VSV), dengue virus (DENV), Zika virus (ZIKV), and influenza virus (PR8) etc. This antiviral activity by TAP1 was further confirmed by series of loss-of-function and gain-of-function experiments. Our further investigation revealed that TAP1 significantly promoted the interferon (IFN)-ß production through activating the TANK binding kinase-1 (TBK1) and the interferon regulatory factor 3 (IRF3) signaling transduction. Our work highlighted the broadly anti-viral function of TAP1 by modulating innate immunity, which is independent of its well-known function of antigen presentation. This study will provide insights into developing novel vaccination and immunotherapy strategies against emerging infectious diseases.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 2/immunology , Antiviral Agents/immunology , Host Microbial Interactions/immunology , Interferon Type I/biosynthesis , ATP Binding Cassette Transporter, Subfamily B, Member 2/antagonists & inhibitors , ATP Binding Cassette Transporter, Subfamily B, Member 2/deficiency , ATP Binding Cassette Transporter, Subfamily B, Member 2/genetics , Animals , Gene Knockout Techniques , HEK293 Cells , Humans , Immunity, Innate , Interferon Regulatory Factor-3/immunology , Mice , Models, Immunological , RAW 264.7 Cells , Toll-Like Receptors/agonists , Virus Diseases/immunology
8.
Emerg Microbes Infect ; 10(1): 1626-1637, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1348038

ABSTRACT

Coronaviruses (CoVs) can infect a variety of hosts, including humans, livestock and companion animals, and pose a serious threat to human health and the economy. The current COVID-19 pandemic, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has killed millions of people. Unfortunately, effective treatments for CoVs infection are still lacking, suggesting the importance of coronavirus vaccines. Our previous work showed that CoV nonstuctural protein 14 (nsp14) functions as (guanine-N7)-methyltransferase (N7-MTase), which is involved in RNA cap formation. Moreover, we found that N7-MTase is well conserved among different CoVs and is a universal target for developing antivirals against CoVs. Here, we show that N7-MTase of CoVs can be an ideal target for designing live attenuated vaccines. Using murine hepatitis virus strain A59 (MHV-A59), a representative and well-studied model of coronaviruses, we constructed N7-MTase-deficient recombinant MHV D330A and Y414A. These two mutants are highly attenuated in mice and exhibit similar replication efficiency to the wild-type (WT) virus in the cell culture. Furthermore, a single dose immunization of D330A or Y414A can induce long-term humoral immune responses and robust CD4+ and CD8+ T cell responses, which can provide full protection against the challenge of a lethal-dose of MHV-A59. Collectively, this study provides an ideal strategy to design live attenuated vaccines for coronavirus by abolishing viral RNA N7-MTase activity. This approach may apply to other RNA viruses that encode their own conservative viral N7-methyltransferase.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , SARS-CoV-2/immunology , Vaccines, Attenuated/immunology , Animals , COVID-19 Vaccines/administration & dosage , Cytokines/biosynthesis , Humans , Immunity, Cellular , Immunity, Humoral , Immunogenicity, Vaccine , Interferon Type I/biosynthesis , Male , Mice , Mutation , Vaccines, Attenuated/administration & dosage , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/immunology
9.
Front Immunol ; 12: 685344, 2021.
Article in English | MEDLINE | ID: covidwho-1295638

ABSTRACT

Vaccination is the best prophylaxis for the prevention of infectious diseases, including coronavirus disease 2019. However, the efficacy of vaccines and onset of adverse reactions vary among individuals. Circulating extracellular vesicles (EVs) regulate the immune responses after vaccination by delivering microRNAs (miRNAs) to myeloid and lymphoid cells. Among these, miR-192 levels in serum EVs increase with aging, in an IL-6-dependent manner, reducing excessive IL-6 expression in aged mice, creating a negative feedback loop. Excessive IL-6 expression reduces vaccination efficacy in aged mice, while EV miR-192 improves efficacy in these aged mice as well, making this miRNA an interesting focus of study. miR-21 levels in serum EVs also increase with aging, and regulates the expression of IL-12 required for Th1 responses; therefore, EV miR-21 is expected to regulate vaccine efficacy. miR-451a, another important miRNA, is abundant in serum EVs and controls the expression of cytokines, such as type I interferon and IL-6. However, levels differ among individuals and correlate with local inflammatory symptoms experienced after a seasonal flu vaccination. These findings suggest the importance of EV miRNAs as a tool to improve vaccine efficacy and also as biomarkers to predict the immune response and adverse reactions after vaccinations.


Subject(s)
Extracellular Vesicles/metabolism , Interferon Type I/immunology , Interleukin-6/immunology , MicroRNAs/blood , Aging/blood , Aging/immunology , COVID-19/immunology , COVID-19/prevention & control , COVID-19 Vaccines/immunology , Humans , Interferon Type I/biosynthesis , Interleukin-12 Subunit p35/biosynthesis , Interleukin-12 Subunit p35/immunology , Interleukin-6/biosynthesis , MicroRNAs/genetics , SARS-CoV-2/immunology , Th1 Cells/immunology , Vaccination
10.
Sci Immunol ; 6(60)2021 06 22.
Article in English | MEDLINE | ID: covidwho-1280392

ABSTRACT

The side effects of SARS-CoV-2 vaccines are often troubling but may merely reflect transient production of type I interferons, a normal physiological response to contact with invading microorganisms.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/immunology , Interferon Type I/biosynthesis , COVID-19/prevention & control , COVID-19/psychology , COVID-19 Vaccines/adverse effects , Humans
11.
Front Immunol ; 12: 662989, 2021.
Article in English | MEDLINE | ID: covidwho-1256380

ABSTRACT

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the causative pathogen of current COVID-19 pandemic, and insufficient production of type I interferon (IFN-I) is associated with the severe forms of the disease. Membrane (M) protein of SARS-CoV-2 has been reported to suppress host IFN-I production, but the underlying mechanism is not completely understood. In this study, SARS-CoV-2 M protein was confirmed to suppress the expression of IFNß and interferon-stimulated genes induced by RIG-I, MDA5, IKKϵ, and TBK1, and to inhibit IRF3 phosphorylation and dimerization caused by TBK1. SARS-CoV-2 M could interact with MDA5, TRAF3, IKKϵ, and TBK1, and induce TBK1 degradation via K48-linked ubiquitination. The reduced TBK1 further impaired the formation of TRAF3-TANK-TBK1-IKKε complex that leads to inhibition of IFN-I production. Our study revealed a novel mechanism of SARS-CoV-2 M for negative regulation of IFN-I production, which would provide deeper insight into the innate immunosuppression and pathogenicity of SARS-CoV-2.


Subject(s)
Interferon Type I/biosynthesis , SARS-CoV-2/immunology , Ubiquitin/metabolism , Viral Matrix Proteins/immunology , DEAD Box Protein 58/metabolism , HEK293 Cells , Humans , I-kappa B Kinase/metabolism , Interferon Regulatory Factor-3/metabolism , Interferon-Induced Helicase, IFIH1/metabolism , Proteolysis , Receptors, Immunologic/metabolism , Signal Transduction , TNF Receptor-Associated Factor 3/metabolism
12.
J Virol ; 94(16)2020 07 30.
Article in English | MEDLINE | ID: covidwho-1214962

ABSTRACT

The 5' cap methylation of viral RNA plays important roles in RNA stability, efficient translation, and immune evasion. Thus, RNA cap methylation is an attractive target for antiviral discovery and development of new live attenuated vaccines. For coronaviruses, RNA cap structure is first methylated at the guanine-N-7 (G-N-7) position by nonstructural protein 14 (nsp14), which facilitates and precedes the subsequent ribose 2'-O methylation by the nsp16-nsp10 complex. Using porcine epidemic diarrhea virus (PEDV), an Alphacoronavirus, as a model, we showed that G-N-7 methyltransferase (G-N-7 MTase) of PEDV nsp14 methylated RNA substrates in a sequence-unspecific manner. PEDV nsp14 can efficiently methylate RNA substrates with various lengths in both neutral and alkaline pH environments and can methylate cap analogs (GpppA and GpppG) and single-nucleotide GTP but not ATP, CTP, or UTP. Mutations to the S-adenosyl-l-methionine (SAM) binding motif in the nsp14 abolished the G-N-7 MTase activity and were lethal to PEDV. However, recombinant rPEDV-D350A with a single mutation (D350A) in nsp14, which retained 29.0% of G-N-7 MTase activity, was viable. Recombinant rPEDV-D350A formed a significantly smaller plaque and had significant defects in viral protein synthesis and viral replication in Vero CCL-81 cells and intestinal porcine epithelial cells (IPEC-DQ). Notably, rPEDV-D350A induced significantly higher expression of both type I and III interferons in IPEC-DQ cells than the parental rPEDV. Collectively, our results demonstrate that G-N-7 MTase activity of PEDV modulates viral replication, gene expression, and innate immune responses.IMPORTANCE Coronaviruses (CoVs) include a wide range of important human and animal pathogens. Examples of human CoVs include severe acute respiratory syndrome coronavirus (SARS-CoV-1), Middle East respiratory syndrome coronavirus (MERS-CoV), and the most recently emerged SARS-CoV-2. Examples of pig CoVs include porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and swine enteric alphacoronavirus (SeACoV). There are no vaccines or antiviral drugs for most of these viruses. All known CoVs encode a bifunctional nsp14 protein which possesses ExoN and guanine-N-7 methyltransferase (G-N-7 MTase) activities, responsible for replication fidelity and RNA cap G-N-7 methylation, respectively. Here, we biochemically characterized G-N-7 MTase of PEDV nsp14 and found that G-N-7 MTase-deficient PEDV was defective in replication and induced greater responses of type I and III interferons. These findings highlight that CoV G-N-7 MTase may be a novel target for rational design of live attenuated vaccines and antiviral drugs.


Subject(s)
Exoribonucleases/metabolism , Interferon Type I/biosynthesis , Interferons/biosynthesis , Porcine epidemic diarrhea virus/physiology , RNA Caps/metabolism , Viral Nonstructural Proteins/metabolism , Animals , Binding Sites , Cell Line , Chlorocebus aethiops , Exoribonucleases/genetics , Gene Expression , Guanine/metabolism , Immunity, Innate , Methylation , Mutation , Porcine epidemic diarrhea virus/enzymology , Porcine epidemic diarrhea virus/genetics , Porcine epidemic diarrhea virus/pathogenicity , RNA, Viral/metabolism , S-Adenosylmethionine/metabolism , Swine , Vero Cells , Viral Nonstructural Proteins/genetics , Virus Replication
13.
Int J Biol Sci ; 17(6): 1547-1554, 2021.
Article in English | MEDLINE | ID: covidwho-1206441

ABSTRACT

Suppression of type I interferon (IFN) response is one pathological outcome of the infection of highly pathogenic human coronaviruses. To effect this, severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2 encode multiple IFN antagonists. In this study, we reported on the IFN antagonism of SARS-CoV-2 main protease NSP5. NSP5 proteins of both SARS-CoV and SARS-CoV-2 counteracted Sendai virus-induced IFN production. NSP5 variants G15S and K90R commonly seen in circulating strains of SARS-CoV-2 retained the IFN-antagonizing property. The suppressive effect of NSP5 on IFN-ß gene transcription induced by RIG-I, MAVS, TBK1 and IKKϵ suggested that NSP5 likely acts at a step downstream of IRF3 phosphorylation in the cytoplasm. NSP5 did not influence steady-state expression or phosphorylation of IRF3, suggesting that IRF3, regardless of its phosphorylation state, might not be the substrate of NSP5 protease. However, nuclear translocation of phosphorylated IRF3 was severely compromised in NSP5-expressing cells. Taken together, our work revealed a new mechanism by which NSP5 proteins encoded by SARS-CoV and SARS-CoV-2 antagonize IFN production by retaining phosphorylated IRF3 in the cytoplasm. Our findings have implications in rational design and development of antiviral agents against SARS-CoV-2.


Subject(s)
Cell Nucleus/metabolism , Coronavirus 3C Proteases/metabolism , Interferon Regulatory Factor-3/metabolism , Interferon Type I/biosynthesis , SARS-CoV-2/enzymology , Animals , COVID-19/virology , Chlorocebus aethiops , Humans , Phosphorylation , Protein Transport , Vero Cells
14.
Signal Transduct Target Ther ; 5(1): 299, 2020 12 28.
Article in English | MEDLINE | ID: covidwho-997814

ABSTRACT

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has quickly spread worldwide and has affected more than 10 million individuals. A typical feature of COVID-19 is the suppression of type I and III interferon (IFN)-mediated antiviral immunity. However, the molecular mechanism by which SARS-CoV-2 evades antiviral immunity remains elusive. Here, we reported that the SARS-CoV-2 membrane (M) protein inhibits the production of type I and III IFNs induced by the cytosolic dsRNA-sensing pathway mediated by RIG-I/MDA-5-MAVS signaling. In addition, the SARS-CoV-2 M protein suppresses type I and III IFN induction stimulated by SeV infection or poly (I:C) transfection. Mechanistically, the SARS-CoV-2 M protein interacts with RIG-I, MAVS, and TBK1, thus preventing the formation of the multiprotein complex containing RIG-I, MAVS, TRAF3, and TBK1 and subsequently impeding the phosphorylation, nuclear translocation, and activation of IRF3. Consequently, ectopic expression of the SARS-CoV-2 M protein facilitates the replication of vesicular stomatitis virus. Taken together, these results indicate that the SARS-CoV-2 M protein antagonizes type I and III IFN production by targeting RIG-I/MDA-5 signaling, which subsequently attenuates antiviral immunity and enhances viral replication. This study provides insight into the interpretation of SARS-CoV-2-induced antiviral immune suppression and illuminates the pathogenic mechanism of COVID-19.


Subject(s)
COVID-19/metabolism , DEAD Box Protein 58/metabolism , Interferon Type I/biosynthesis , Interferon-Induced Helicase, IFIH1/metabolism , Interferons/biosynthesis , SARS-CoV-2/metabolism , Signal Transduction , Viral Matrix Proteins/metabolism , Animals , COVID-19/genetics , Chlorocebus aethiops , DEAD Box Protein 58/genetics , HEK293 Cells , HeLa Cells , Humans , Interferon Type I/genetics , Interferon-Induced Helicase, IFIH1/genetics , Interferons/genetics , Receptors, Immunologic , SARS-CoV-2/genetics , Vero Cells , Viral Matrix Proteins/genetics
15.
EMBO J ; 40(5): e105912, 2021 03 01.
Article in English | MEDLINE | ID: covidwho-962496

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), which may result in acute respiratory distress syndrome (ARDS), multiorgan failure, and death. The alveolar epithelium is a major target of the virus, but representative models to study virus host interactions in more detail are currently lacking. Here, we describe a human 2D air-liquid interface culture system which was characterized by confocal and electron microscopy and single-cell mRNA expression analysis. In this model, alveolar cells, but also basal cells and rare neuroendocrine cells, are grown from 3D self-renewing fetal lung bud tip organoids. These cultures were readily infected by SARS-CoV-2 with mainly surfactant protein C-positive alveolar type II-like cells being targeted. Consequently, significant viral titers were detected and mRNA expression analysis revealed induction of type I/III interferon response program. Treatment of these cultures with a low dose of interferon lambda 1 reduced viral replication. Hence, these cultures represent an experimental model for SARS-CoV-2 infection and can be applied for drug screens.


Subject(s)
Alveolar Epithelial Cells/metabolism , COVID-19/metabolism , Models, Biological , Organoids/metabolism , SARS-CoV-2/physiology , Virus Replication , Alveolar Epithelial Cells/pathology , Alveolar Epithelial Cells/virology , Animals , COVID-19/virology , Chlorocebus aethiops , Gene Expression Regulation , Humans , Interferon Type I/biosynthesis , Interferons/biosynthesis , Organoids/pathology , Organoids/virology , Vero Cells
16.
Pharmacotherapy ; 40(8): 843-856, 2020 08.
Article in English | MEDLINE | ID: covidwho-602791

ABSTRACT

A hyperinflammatory response to severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) infection, reminiscent of cytokine release syndrome, has been implicated in the pathophysiology of acute respiratory distress syndrome and organ damage in patients with coronavirus disease 2019 (COVID-19). Agents that inhibit components of the pro-inflammatory cascade have garnered interest as potential treatment options with hopes that dampening the proinflammatory process may improve clinical outcomes. Baricitinib is a reversible Janus-associated kinase (JAK)-inhibitor that interrupts the signaling of multiple cytokines implicated in COVID-19 immunopathology. It may also have antiviral effects by targeting host factors that viruses rely for cell entry and by suppressing type I interferon driven angiotensin-converting-enzyme-2 upregulation. However, baricitinib's immunosuppressive effects may be detrimental during acute viral infections by delaying viral clearance and increasing vulnerability to secondary opportunistic infections. The lack of reliable biomarkers to monitor patients' immune status as illness evolves complicates deployment of immunosuppressive drugs like baricitinib. Furthermore, baricitinib carries the risk of increased thromboembolic events, which is concerning given the proclivity towards a hypercoagulable state in patients with COVID-19. In this article, we review available data on baricitinib with an emphasis on immunosuppressive and antiviral pharmacology, pharmacokinetics, safety, and current progress in COVID-19 clinical trials.


Subject(s)
Azetidines/pharmacology , Azetidines/therapeutic use , Coronavirus Infections/complications , Inflammation/drug therapy , Inflammation/etiology , Janus Kinases/antagonists & inhibitors , Pneumonia, Viral/complications , Sulfonamides/pharmacology , Sulfonamides/therapeutic use , Angiotensin-Converting Enzyme 2 , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , Area Under Curve , Azetidines/administration & dosage , Azetidines/adverse effects , Betacoronavirus , COVID-19 , Clinical Trials as Topic , Cytokines/metabolism , Drug Interactions , Humans , Interferon Type I/biosynthesis , Metabolic Clearance Rate , Pandemics , Peptidyl-Dipeptidase A/biosynthesis , Purines , Pyrazoles , SARS-CoV-2 , Signal Transduction/drug effects , Sulfonamides/administration & dosage , Sulfonamides/adverse effects
17.
J Virol ; 94(13)2020 06 16.
Article in English | MEDLINE | ID: covidwho-60496

ABSTRACT

Type I and type III interferons (IFNs) are the frontline of antiviral defense mechanisms that trigger hundreds of downstream antiviral genes. In this study, we observed that MERS-CoV nucleocapsid (N) protein suppresses type I and type III IFN gene expression. The N protein suppresses Sendai virus-induced IFN-ß and IFN-λ1 by reducing their promoter activity and mRNA levels, as well as downstream IFN-stimulated genes (ISGs). Retinoic acid-inducible gene I (RIG-I) is known to recognize viral RNA and induce IFN expression through tripartite motif-containing protein 25 (TRIM25)-mediated ubiquitination of RIG-I caspase activation and recruitment domains (CARDs). We discovered that MERS-CoV N protein suppresses RIG-I-CARD-induced, but not MDA5-CARD-induced, IFN-ß and IFN-λ1 promoter activity. By interacting with TRIM25, N protein impedes RIG-I ubiquitination and activation and inhibits the phosphorylation of transcription factors IFN-regulatory factor 3 (IRF3) and NF-κB that are known to be important for IFN gene activation. By employing a recombinant Sindbis virus-EGFP replication system, we showed that viral N protein downregulated the production of not only IFN mRNA but also bioactive IFN proteins. Taken together, MERS-CoV N protein functions as an IFN antagonist. It suppresses RIG-I-induced type I and type III IFN production by interfering with TRIM25-mediated RIG-I ubiquitination. Our study sheds light on the pathogenic mechanism of how MERS-CoV causes disease.IMPORTANCE MERS-CoV causes death of about 35% of patients. Published studies showed that some coronaviruses are capable of suppressing interferon (IFN) expression in the early phase of infection and MERS-CoV proteins can modulate host immune response. In this study, we demonstrated that MERS-CoV nucleocapsid (N) protein suppresses the production of both type I and type III IFNs via sequestering TRIM25, an E3 ubiquitin ligase that is essential for activating the RIG-I signaling pathway. Ectopic expression of TRIM25 rescues the suppressive effect of the N protein. In addition, the C-terminal domain of the viral N protein plays a pivotal role in the suppression of IFN-ß promoter activity. Our findings reveal how MERS-CoV evades innate immunity and provide insights into the interplay between host immune response and viral pathogenicity.


Subject(s)
Coronavirus Infections/metabolism , Coronavirus Infections/virology , DEAD Box Protein 58/metabolism , Interferon Type I/biosynthesis , Interferons/biosynthesis , Middle East Respiratory Syndrome Coronavirus/physiology , Nucleocapsid Proteins/metabolism , Signal Transduction , CARD Signaling Adaptor Proteins/metabolism , Cell Line , Coronavirus Infections/genetics , Gene Expression Regulation , Host-Pathogen Interactions/genetics , Humans , Interferon Regulatory Factor-3/metabolism , Interferon Type I/genetics , Interferons/genetics , Promoter Regions, Genetic , Protein Binding , Receptors, Immunologic , Transcription Factors , Tripartite Motif Proteins , Ubiquitin-Protein Ligases
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