IFITM1 and IFITM2 inhibit the replication of senecavirus A by positive feedback with RIG-I signaling pathway.

The role of host factors in the replication of emerging senecavirus A (SVA) which induced porcine idiopathic vesicular disease (PIVD) distributed worldwide remains obscure. Here, interferon-induced transmembrane (IFITM) protein 1 and 2 inhibit SVA replication by positive feedback with RIG-I signaling pathway was reported. The expression levels of IFITM1 and IFITM2 increased significantly in SVA infected 3D4/21 cells. Infection experiments of cells with over and interference expression of IFITM1 and IFITM2 showed that these two proteins inhibit SVA replication by regulating the expression of interferon beta (IFN-ß), IFN-stimulated gene 15 (ISG-15), interleukin 6 (IL-6), IL-8, tumor necrosis factor alpha (TNF-α), IFN regulatory factor-3 (IRF3), and IRF7. Further results showed that antiviral responses of IFITM1 and IFITM2 were achieved by activating retinoic acid-inducible gene I (RIG-I) signaling pathway which in turn enhanced the expression of IFITM1 and IFITM2. It is noteworthy that conserved domains of these two proteins also paly the similar role. These findings provide new data on the role of host factors in infection and replication of SVA and help to develop new agents against the virus.

Novel peripheral blood mononuclear cell mRNA signature for IFN-beta therapy responsiveness prediction in multiple sclerosis.

Interferon-beta (IFN-ß) is one of the classical drugs for immunomodulatory therapy in relapsing-remitting multiple sclerosis (RRMS) patients, but the drug responsiveness of different patients varies. Currently, there is no valid model to predict IFN-ß responsiveness. This research attempted to develop an IFN-ß responsiveness prediction model based on mRNA expression in RRMS patient peripheral blood mononuclear cells. Peripheral blood mononuclear cell mRNA expression datasets including 50 RRMS patients receiving IFN-ß treatment were obtained from GEO. Among the datasets, 24 cases from GSE24427 were included in a training set, and 18 and 9 cases from GSE19285 and GSE33464, respectively, were adopted as two independent test sets. In the training set, blood samples were collected immediately before first, second, month 1, 12, and 24 IFN-ß injection, and the mRNA expression data at four time points, namely, two days, one month, one year and two years after the onset of IFN-ß treatment, were compared with pre-treatment data to identify IFN-stimulated genes (ISGs). The ISGs at the one-month time point were used to construct the drug responsiveness prediction model. Next, the drug responsiveness model was verified in the two independent test sets to examine the performance of the model in predicting drug responsiveness. Finally, we used CIBERSORTx to estimate the content of cell subtypes in samples and evaluated whether differences in the proportions of cell subtypes were related to differences in IFN-ß responsiveness. Among the four time points, one month was the time point when the training set GSE24427 and test set GSE33464 had the highest number of ISGs. Functional analysis showed that these one-month ISGs were enriched in biological functions such as the innate immune response, type-I interferon signalling pathway, and other IFN-ß-associated functions. Based on these ISGs, we obtained a four-factor prediction model for IFN-ß responsiveness including MX1, MX2, XAF1, and LAMP3. In addition, the model demonstrated favourable predictive performance within the training set and two external test sets. A higher proportion of activated NK cells and lower naive CD4/total CD4 ratio might indicate better drug responsiveness. This research developed a polygene-based biomarker model that could predict RRMS patient IFN-ß responsiveness in the early treatment period. This model could probably help doctors screen out patients who would not benefit from IFN-ß treatment early and determine whether a current treatment plan should be continued.

African swine fever virus early protein pI73R suppresses the type-I IFN promoter activities.

African swine fever virus is known to suppress type-I interferon (IFN) responses. The main objective of this study was to screen early-expressed viral genes for their ability to suppress IFN production. Out of 16 early genes examined, I73R exhibited robust suppression of cGAS-STING-induced IFN-ß promoter activities, impeding the function of both IRF3 and NF-κB transcription factors. As a result, I73R obstructed IRF3 nuclear translocation following the treatment of cells with poly(dA:dT), a strong inducer of the cGAS-STING signaling pathway. Although the I73R protein exhibits structural homology with the Zα domain binding to the left-handed helical form of DNA known as Z-DNA, its ability to suppress cGAS-STING induction of IFN-ß was independent of Z-DNA binding activity. Instead, the α3 and ß1 domains of I73R played a significant role in suppressing cGAS-STING induction of IFN-ß. These findings offer insights into the protein's functions and support its role as a virulence factor.

Identification of pigeon mitochondrial antiviral signaling protein (MAVS) and its role in antiviral innate immunity.

Pigeons can be infected with various RNA viruses, and their innate immune system responds to viral infection to establish an antiviral response. Mitochondrial antiviral signaling protein (MAVS), an important adaptor protein in signal transduction, plays a pivotal role in amplifying the innate immune response. In this study, we successfully cloned pigeon MAVS (piMAVS) and performed a bioinformatics analysis. The results showed that the caspase recruitment domain (CARD) and transmembrane (TM) domain are highly conserved in poultry and mammals but poorly conserved in other species. Furthermore, we observed that MAVS expression is upregulated both in pigeons and pigeon embryonic fibroblasts (PEFs) upon RNA virus infection. Overexpression of MAVS resulted in increased levels of ß-interferon (IFN-ß), IFN-stimulated genes (ISGs), and interleukin (ILs) mRNA and inhibited Newcastle disease virus (NDV) replication. We also found that piMAVS and human MAVS (huMAVS) induced stronger expression of IFN-ß and ISGs when compared to chicken MAVS (chMAVS), and this phenomenon was also reflected in the degree of inhibition of NDV replication. Our findings demonstrate that piMAVS plays an important role in repressing viral replication by regulating the activation of the IFN signal pathway in pigeons. This study not only sheds light on the function of piMAVS in innate immunity but also contributes to a more comprehensive understanding of the innate immunity system in poultry. Our data also provide unique insights into the differences in innate immunity between poultry and mammal.

NDV inhibited IFN-ß secretion through impeding CHCHD10-mediated mitochondrial fusion to promote viral proliferation.

Newcastle disease virus (NDV) is an RNA virus that can promote its own replication through the inhibition of cellular mitochondrial fusion. The proteins involved in mitochondrial fusion, namely mitofusin 1 (Mfn1) and optic atrophy 1 (OPA1) are associated with interferon-beta (IFN-ß) secretion during NDV infection. However, the precise mechanism by which NDV modulates the Mfn1-mediated or OPA1-mediated fusion of mitochondria, thereby impacting IFN-ß, remains elusive. This study revealed that the downregulation of the mitochondrial protein known as coiled-coil-helix-coiled-coil-helix domain containing 10 (CHCHD10) exerts a negative regulatory effect on OPA1 and Mfn1 in human lung adenocarcinoma (A549) cells during the late stage of NDV infection. This reduction in CHCHD10 expression impeded cellular mitochondrial fusion, subsequently leading to a decline in the activation of interferon regulatory factor 3 (IRF3) and nuclear factor kappa B (NF-κB), ultimately resulting in diminished secretion of IFN-ß. In contrast, the overexpression of CHCHD10 alleviated infection-induced detrimental effect in mitochondrial fusion, thereby impeding viral proliferation. In summary, NDV enhances its replication by inhibiting the CHCHD10 protein, which impedes mitochondrial fusion and suppresses IFN-ß production through the activation of IRF3 and NF-κB.

Nonself RNA rewires IFN-ß signaling: A mathematical model of the innate immune response.

Type I interferons (IFNs) are key coordinators of the innate immune response to viral infection, which, through activation of the transcriptional regulators STAT1 and STAT2 (STAT1/2) in bystander cells, induce the expression of IFN-stimulated genes (ISGs). Here, we showed that in cells transfected with poly(I:C), an analog of viral RNA, the transcriptional activity of STAT1/2 was terminated because of depletion of the interferon-ß (IFN-ß) receptor, IFNAR. Activation of RNase L and PKR, products of two ISGs, not only hindered the replenishment of IFNAR but also suppressed negative regulators of IRF3 and NF-κB, consequently promoting IFNB transcription. We incorporated these findings into a mathematical model of innate immunity. By coupling signaling through the IRF3-NF-κB and STAT1/2 pathways with the activities of RNase L and PKR, the model explains how poly(I:C) switches the transcriptional program from being STAT1/2 induced to being IRF3 and NF-κB induced, which converts IFN-ß-responding cells to IFN-ß-secreting cells.

Bovine herpesvirus-1 gE protein inhibits IFN-ß production to enhance replication by promoting MAVS ubiquitination and interfering with the interaction between IRF3 and CBP/p300.

Bovine herpesvirus-1 (BoHV-1) can infect all breeds of cattle and cause respiratory and genital tract diseases. In the process of viral infection, viruses can use their own proteins to suppress the innate immunity of the host and promote its replication; however, the mechanism by which BoHV-1 evades the innate immune response is not fully understood. In this study, we found that rabbits inoculated with the live gene deletion vaccine BoHV-1-△gI/gE/TK generated higher interferon-ß (IFN-ß) production in the serum, liver, lung and kidney than rabbits inoculated with wt BoHV-1, which led to milder lesions in the lung and kidney. We performed gene deletion and ectopic expression experiments on viral proteins and found that gE was the major protein that inhibited IFN-ß expression. Further studies showed that MAVS and IRF3 were the targets of gE, and the specific mechanism was that gE inhibited IFN-ß production by promoting MAVS ubiquitination and interfering with the interaction between IRF3 and CBP/p300. These results suggest a new way of BoHV-1 inhibition of IFN-ß production to evade the host innate immunity.

The MC160 protein of the molluscum contagiosum virus dampens cGAS/STING-induced interferon-ß activation.

Molluscum contagiosum virus (MCV) is a poxvirus that causes benign, persistent skin lesions. MCV encodes a variety of immune evasion molecules to dampen host immune responses. Two of these proteins are the MC159 and MC160 proteins. Both MC159 and MC160 contain two tandem death effector domains and share homology to the cellular FLIPs, FADD, and procaspase-8. MC159 and MC160 dampen several innate immune responses such as NF-κB activation and mitochondrial antiviral signaling (MAVS)-mediated induction of type 1 interferon (IFN). The type 1 IFN response is also activated by the cytosolic DNA sensors cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING). Both cGAS and STING play a vital role in sensing a poxvirus infection. In this study, we demonstrate that there are nuanced differences between both MC160 and MC159 in terms of how the viral proteins modulate the cGAS/STING and MAVS pathways. Specifically, MC160 expression, but not MC159 expression, dampens cGAS/STING-mediated induction of IFN in HEK 293 T cells. Further, MC160 expression prevented the K63-ubiquitination of both STING and TBK1, a kinase downstream of cGAS/STING. Ectopic expression of the MC160 protein, but not the MC159 protein, resulted in a measurable decrease in the TBK1 protein levels as detected via immunoblotting. Finally, using a panel of MC160 truncation mutants, we report that the MC160 protein requires both DEDs to inhibit cGAS/STING-induced activation of IFN-ß. Our model indicates MC160 likely alters the TBK1 signaling complex to decrease IFN-ß activation at the molecular intersection of the cGAS/STING and MAVS signaling pathways.

Staphylococcus aureus planktonic but not biofilm environment induces an IFN-ß macrophage immune response via the STING/IRF3 pathway.

Chronic implant-related bone infections are a severe complication in orthopaedic surgery. Biofilm formation on the implant impairs the immune response, leading to bacterial persistence. In a previous study, we found that Staphylococcus aureus (SA) induced interferon regulatory factor 3 (IRF3) activation and Ifnb expression only in its planktonic form but not in the biofilm. The aim of this study was to clarify the role of the stimulator of interferon genes (STING) in this process. We treated RAW 264.7 macrophages with conditioned media (CM) generated from planktonic or biofilm cultured SA in combination with agonists or inhibitors of the cyclic GMP-AMP synthase (cGAS)/STING pathway. We further evaluated bacterial gene expression of planktonic and biofilm SA to identify potential mediators. STING inhibition resulted in the loss of IRF3 activation and Ifnb induction in SA planktonic CM, whereas STING activation induced an IRF3 dependent IFN-ß response in SA biofilm CM. The expression levels of virulence-associated genes decreased during biofilm formation, but genes associated with cyclic dinucleotide (CDN) synthesis did not correlate with Ifnb induction. We further observed that cGAS contributed to Ifnb induction by SA planktonic CM, although cGAS activation was not sufficient to induce Ifnb expression in SA biofilm CM. Our data indicate that the different degrees of virulence associated with SA planktonic and biofilm environments result in an altered induction of the IRF3 mediated IFN-ß response via the STING pathway. This finding suggests that the STING/IRF3/IFN-ß axis is a potential candidate as an immunotherapeutic target for implant-related bone infections.

SADS-CoV nsp1 inhibits the IFN-ß production by preventing TBK1 phosphorylation and inducing CBP degradation.

Swine acute diarrhea syndrome (SADS) is first reported in January 2017 in Southern China. It subsequently causes widespread outbreaks in multiple pig farms, leading to economic losses. Therefore, it is an urgent to understand the molecular mechanisms underlying the pathogenesis and immune evasion of Swine acute diarrhea syndrome coronavirus (SADS-CoV). Our research discovered that SADS-CoV inhibited the production of interferon-ß (IFN-ß) during viral infection. The nonstructural protein 1 (nsp1) prevented the phosphorylation of TBK1 by obstructing the interaction between TBK1 and Ub protein. Moreover, nsp1 induced the degradation of CREB-binding protein (CBP) through the proteasome-dependent pathway, thereby disrupting the IFN-ß enhancer and inhibiting IFN transcription. Finally, we identified nsp1-Phe39 as the critical amino acid that downregulated IFN production. In conclusion, our findings described two mechanisms in nsp1 that inhibited IFN production and provided new insights into the evasion strategy adopted by SADS-CoV to evade host antiviral immunity.

Host Derivation of Sindbis Virus Influences Mammalian Type I Interferon Response to Infection.

Arboviruses are defined by their ability to replicate in both mosquito vectors and mammalian hosts. There is good evidence that arboviruses "prime" their progeny for infection of the next host, such as via differential glycosylation of their outer glycoproteins or packaging of host ribosomal subunits. We and others have previously shown that mosquito-derived viruses more efficiently infect mammalian cells than mammalian-derived viruses. These observations are consistent with arboviruses acquiring host-specific adaptations, and we hypothesized that a virus derived from either the mammalian host or mosquito vector elicits different responses when infecting the mammalian host. Here, we perform an RNA-sequencing analysis of the transcriptional response of Human Embryonic Kidney 293 (HEK-293) cells to infection with either mosquito (Aedes albopictus, C7/10)- or mammalian (Baby Hamster Kidney, BHK-21)-derived Sindbis virus (SINV). We show that the C7/10-derived virus infection leads to a more robust transcriptional response in HEK-293s compared to infection with the BHK-derived virus. Surprisingly, despite more efficient infection, we found an increase in interferon-ß (IFN-ß) and interferon-stimulated gene (ISG) transcripts in response to the C7/10-derived virus infection versus the BHK-derived virus infection. However, translation of interferon-stimulated genes was lower in HEK-293s infected with the C7/10-derived virus, starkly contrasting with the transcriptional response. This inhibition of ISG translation is reflective of a more rapid overall shut-off of host cell translation following infection with the C7/10-derived virus. Finally, we show that the C7/10-derived virus infection of HEK-293 cells leads to elevated levels of phosphorylated eukaryotic translation elongation factor-2 (eEF2), identifying a potential mechanism leading to the more rapid shut-off of host translation. We postulate that the rapid shut-off of host translation in mammalian cells infected with the mosquito-derived virus acts to counter the IFN-ß-stimulated transcriptional response.

Japanese encephalitis virus NS4B inhibits interferon beta production by targeting TLR3 and TRIF.

Japanese encephalitis virus (JEV) is a flavivirus transmitted by mosquitoes, causing epidemics of encephalitis in humans and reproductive disorders in pigs. This virus is predominantly distributed in Asian countries and causes tens of thousands of infections in humans annually. Interferon (IFN) is an essential component of host defense against viral infection. Multiple studies have indicated that multifunctional nonstructural proteins of flaviviruses suppress the host IFN response via various strategies to facilitate viral replication. The flaviviruses encoded nonstructural protein 4B (NS4B) is a multifunctional hydrophobic nonstructural protein widely involved in viral replication, pathogenesis and host immune evasion. In this study, we demonstrated that NS4B of JEV suppressed the induction of IFN-ß production, mainly through targeting the TLR3 and TRIF (a TIR domain-containing linker that induces IFN-ß) proteins in the TLR3 pathway. In a dual-luciferase reporter assay, JEV NS4B significantly inhibited the activation of IFN-ß promoter induced by TLR3 and simultaneously treated with poly (I:C). Moreover, NS4B also inhibited the activation of IFN-ß promoter triggered by interferon regulatory factor 3 (IRF3)/5D or its upstream molecules in TLR3 signaling pathway. Furthermore, NS4B inhibited the phosphorylation of IRF3 under the stimulation of TLR3 and TRIF molecules. Mechanistically, JEV NS4B interacts with TLR3 and TRIF and confirmed by co-localization and co-immunoprecipitation assay, thereby inhibiting the activation of downstream sensors in the TLR3-mediated pathway. Overall, our results provide a novel mechanism by which JEV NS4B interferes with the host's antiviral response through targeting TLR3 receptor signaling pathway.

ZNF205 positively regulates RLR antiviral signaling by targeting RIG-I.

Retinoic acid-inducible gene I (RIG-I) is a cytosolic viral RNA receptor. Upon viral infection, the protein recognizes and then recruits adapter protein mitochondrial antiviral signaling (MAVS) protein, initiating the production of interferons and proinflammatory cytokines to establish an antiviral state. In the present study, we identify zinc finger protein 205 (ZNF205) which associates with RIG-I and promotes the Sendai virus (SeV)-induced antiviral innate immune response. Overexpression of ZNF205 facilitates interferon-beta (IFN-ß) introduction, whereas ZNF205 deficiency restricts its introduction. Mechanistically, the C-terminal zinc finger domain of ZNF205 interacts with the N-terminal tandem caspase recruitment domains (CARDs) of RIG-I; this interaction markedly promotes K63 ubiquitin-linked polyubiquitination of RIG-I, which is crucial for RIG-I activation. Thus, our results demonstrate that ZNF205 is a positive regulator of the RIG-I-mediated innate antiviral immune signaling pathway.

Nei-like DNA glycosylase 2 selectively antagonizes interferon-ß expression upon respiratory syncytial virus infection.

As part of the antiviral response, cells activate the expressions of type I interferons (IFNs) and proinflammatory mediators to control viral spreading. Viral infections can impact DNA integrity; however, how DNA damage repair coordinates antiviral response remains elusive. Here we report Nei-like DNA glycosylase 2 (NEIL2), a transcription-coupled DNA repair protein, actively recognizes the oxidative DNA substrates induced by respiratory syncytial virus (RSV) infection to set the threshold of IFN-ß expression. Our results show that NEIL2 antagonizes nuclear factor κB (NF-κB) acting on the IFN-ß promoter early after infection, thus limiting gene expression amplified by type I IFNs. Mice lacking Neil2 are far more susceptible to RSV-induced illness with an exuberant expression of proinflammatory genes and tissue damage, and the administration of NEIL2 protein into the airway corrected these defects. These results suggest a safeguarding function of NEIL2 in controlling IFN-ß levels against RSV infection. Due to the short- and long-term side effects of type I IFNs applied in antiviral therapy, NEIL2 may provide an alternative not only for ensuring genome fidelity but also for controlling immune responses.

Optineurin deficiency impairs autophagy to cause interferon beta overproduction and increased survival of mice following viral infection.

BACKGROUND: Optineurin (OPTN) is associated with several human diseases, including amyotrophic lateral sclerosis (ALS), and is involved in various cellular processes, including autophagy. Optineurin regulates the expression of interferon beta (IFNß), which plays a central role in the innate immune response to viral infection. However, the role of optineurin in response to viral infection has not been fully clarified. It is known that optineurin-deficient cells produce more IFNß than wild-type cells following viral infection. In this study, we investigate the reasons for, and effects of, IFNß overproduction during optineurin deficiency both in vitro and in vivo. METHODS: To investigate the mechanism of IFNß overproduction, viral nucleic acids in infected cells were quantified by RT-qPCR and the autophagic activity of optineurin-deficient cells was determined to understand the basis for the intracellular accumulation of viral nucleic acids. Moreover, viral infection experiments using optineurin-disrupted (Optn-KO) animals were performed with several viruses. RESULTS: IFNß overproduction following viral infection was observed not only in several types of optineurin-deficient cell lines but also in Optn-KO mice and human ALS patient cells carrying mutations in OPTN. IFNß overproduction in Optn-KO cells was revealed to be caused by excessive accumulation of viral nucleic acids, which was a consequence of reduced autophagic activity caused by the loss of optineurin. Additionally, IFNß overproduction in Optn-KO mice suppressed viral proliferation, resulting in increased mouse survival following viral challenge. CONCLUSION: Our findings indicate that the combination of optineurin deficiency and viral infection leads to IFNß overproduction in vitro and in vivo. The effects of optineurin deficiency are elicited by viral infection, therefore, viral infection may be implicated in the development of optineurin-related diseases.

The Cytomegalovirus M35 Protein Directly Binds to the Interferon-ß Enhancer and Modulates Transcription of Ifnb1 and Other IRF3-Driven Genes.

Induction of type I interferon (IFN) gene expression is among the first lines of cellular defense a virus encounters during primary infection. We previously identified the tegument protein M35 of murine cytomegalovirus (MCMV) as an essential antagonist of this antiviral system, showing that M35 interferes with type I IFN induction downstream of pattern-recognition receptor (PRR) activation. Here, we report structural and mechanistic details of M35's function. Determination of M35's crystal structure combined with reverse genetics revealed that homodimerization is a key feature for M35's immunomodulatory activity. In electrophoretic mobility shift assays (EMSAs), purified M35 protein specifically bound to the regulatory DNA element that governs transcription of the first type I IFN gene induced in nonimmune cells, Ifnb1. DNA-binding sites of M35 overlapped with the recognition elements of interferon regulatory factor 3 (IRF3), a key transcription factor activated by PRR signaling. Chromatin immunoprecipitation (ChIP) showed reduced binding of IRF3 to the host Ifnb1 promoter in the presence of M35. We furthermore defined the IRF3-dependent and the type I IFN signaling-responsive genes in murine fibroblasts by RNA sequencing of metabolically labeled transcripts (SLAM-seq) and assessed M35's global effect on gene expression. Stable expression of M35 broadly influenced the transcriptome in untreated cells and specifically downregulated basal expression of IRF3-dependent genes. During MCMV infection, M35 impaired expression of IRF3-responsive genes aside of Ifnb1. Our results suggest that M35-DNA binding directly antagonizes gene induction mediated by IRF3 and impairs the antiviral response more broadly than formerly recognized. IMPORTANCE Replication of the ubiquitous human cytomegalovirus (HCMV) in healthy individuals mostly goes unnoticed but can impair fetal development or cause life-threatening symptoms in immunosuppressed or -deficient patients. Like other herpesviruses, CMV extensively manipulates its hosts and establishes lifelong latent infections. Murine CMV (MCMV) presents an important model system as it allows the study of CMV infection in the host organism. We previously showed that during entry into host cells, MCMV virions release the evolutionary conserved protein M35 protein to immediately dampen the antiviral type I interferon (IFN) response induced by pathogen detection. Here, we show that M35 dimers bind to regulatory DNA elements and interfere with recruitment of interferon regulatory factor 3 (IRF3), a key cellular factor for antiviral gene expression. Thereby, M35 interferes with expression of type I IFNs and other IRF3-dependent genes, reflecting the importance for herpesviruses to avoid IRF3-mediated gene induction.

SARS-CoV-2 omicron sub-lineages differentially modulate interferon response in human lung epithelial cells.

Although most of the attention was focused on the characterization of changes in the Spike protein among variants of SARS-CoV-2 virus, mutations outside the Spike region are likely to contribute to virus pathogenesis, virus adaptation and escape to the immune system. Phylogenetic analysis of SARS-CoV-2 Omicron strains reveals that several virus sub-lineages could be distinguished, from BA.1 up to BA.5. Regarding BA.1, BA.2 and BA.5, several mutations concern viral proteins with antagonistic activity to the innate immune system, such as NSP1 (S135R), which is involved in mRNAs translation, exhibiting a general shutdown in cellular protein synthesis. Additionally, mutations and/or deletions in the ORF6 protein (D61L) and in the nucleoprotein N (P13L, D31-33ERS, P151S, R203K, G204R and S413R) have been reported, although the impact of such mutations on protein function has not been further studied. The aim of this study was to better investigate the innate immunity modulation by different Omicron sub-lineages, in the attempt to identify viral proteins that may affect virus fitness and pathogenicity. Our data demonstrated that, in agreement with a reduced Omicron replication in Calu-3 human lung epithelial cells compared to the Wuhan-1 strain, a lower secretion of interferon beta (IFN-ß) from cells was observed in all sub-lineages, except for BA.2. This evidence might be correlated with the presence of a mutation within the ORF6 protein (D61L), which is strikingly associated to the antagonistic function of the viral protein, since additional mutations in viral proteins acting as interferon antagonist were not detected or did not show significant influence. Indeed, the recombinant mutated ORF6 protein failed to inhibit IFN-ß production in vitro. Furthermore, we found an induction of IFN-ß transcription in BA.1 infected cells, that was not correlated with the cytokine release at 72 h post-infection, suggesting that post-transcriptional events can be involved in controlling the innate immunity.

Ebolavirus Species-Specific Interferon Antagonism Mediated by VP24.

Members of the Ebolavirus genus demonstrate a marked differences in pathogenicity in humans with Ebola (EBOV) being the most pathogenic, Bundibugyo (BDBV) less pathogenic, and Reston (RESTV) is not known to cause a disease in humans. The VP24 protein encoded by members of the Ebolavirus genus blocks type I interferon (IFN-I) signaling through interaction with host karyopherin alpha nuclear transporters, potentially contributing to virulence. Previously, we demonstrated that BDBV VP24 (bVP24) binds with lower affinities to karyopherin alpha proteins relative to EBOV VP24 (eVP24), and this correlated with a reduced inhibition in IFN-I signaling. We hypothesized that modification of eVP24-karyopherin alpha interface to make it similar to bVP24 would attenuate the ability to antagonize IFN-I response. We generated a panel of recombinant EBOVs containing single or combinations of point mutations in the eVP24-karyopherin alpha interface. Most of the viruses appeared to be attenuated in both IFN-I-competent 769-P and IFN-I-deficient Vero-E6 cells in the presence of IFNs. However, the R140A mutant grew at reduced levels even in the absence of IFNs in both cell lines, as well as in U3A STAT1 knockout cells. Both the R140A mutation and its combination with the N135A mutation greatly reduced the amounts of viral genomic RNA and mRNA suggesting that these mutations attenuate the virus in an IFN-I-independent attenuation. Additionally, we found that unlike eVP24, bVP24 does not inhibit interferon lambda 1 (IFN-λ1), interferon beta (IFN-ß), and ISG15, which potentially explains the lower pathogenicity of BDBV relative to EBOV. Thus, the VP24 residues binding karyopherin alpha attenuates the virus by IFN-I-dependent and independent mechanisms.

ß-Glucan Induces Training Immunity to Promote Antiviral Activity by Activating TBK1.

Many studies have shown that ß-glucan induces a trained immune phenotype in innate immune cells to defend against bacterial and fungal infections. The specific mechanism involves cellular metabolism and epigenetic reprogramming. However, it is unclear whether ß-glucan plays a role in antiviral infection. Therefore, this study investigated the role of trained immunity induced by Candida albicans and ß-glucan in antiviral innate immunity. It showed that C. albicans and ß-glucan promoted the expression of interferon-ß (IFN-ß) and interleukin-6 (IL-6) in mouse macrophages triggered by viral infection. In addition, ß-glucan pretreatment attenuated the pathological damage induced by the virus in mouse lungs and promoted the expression of IFN-ß. Mechanistically, ß-glucan could promote the phosphorylation and ubiquitination of TANK Binding Kinase 1 (TBK1), a key protein of the innate immune pathway. These results suggest that ß-glucan can promote innate antiviral immunity, and this bioactive material may be a potential therapeutic target for antiviral treatment.

Zika virus replication on endothelial cells and invasion into the central nervous system by inhibiting interferon ß translation.

The blood-brain barrier (BBB) is one of the tightest physical barriers to prevent pathogens from invading the central nervous system (CNS). However, the mechanism by which Zika virus (ZIKV) crossing the BBB remains unresolved. We found ZIKV induced high morbidity and mortality in newborn mice, accompanied by inflammatory injury on CNS. ZIKV was found to replicate primarily in the cortex and hippocampus in neonatal mouse brains. An in vitro model revealed that ZIKV had no impact on hBMECs permeability but led to endothelial activation, as shown by the enhancement of adhesion molecules expression and F-actin redistribution. ZIKV replication in hBMECs might be associated with the suppression of IFN-ß translation via inhibiting RPS6 phosphorylation. On the other hand, ZIKV infection induced IFN-stimulated genes (ISGs), activated the mitogen-activated protein kinase (MAPK) signaling pathway, and promoted chemokine secretion. This study provides an understanding of virus replication and transmigration across the BBB during ZIKV infection.