I Interferon Signaling Pathway Research Articles

Metformin activates type I interferon signaling against HCV via activation of adenosine monophosphate-activated protein kinase.

Activation of the type I interferon (IFN) signaling pathway is essential for the eradication of hepatitis C virus (HCV). Metformin can activate adenosine monophosphate-activated protein kinase (AMPK) to reduce insulin resistance. Cross talks between AMPK and IFN signaling remain unclear. To understand the influence of metformin on the type I IFN signaling pathway and HCV infection, the full-length HCV replicon OR6 cells and the infectious HCV clones JFH1 were used to assess the anti-HCV effect of the insulin sensitizers, metformin and pioglitazone. Immunofluorescence staining and the immunoblotting of HCV viral protein demonstrated that metformin, but not pioglitazone, inhibited HCV replication in OR-6 and JFH-1-infected Huh 7.5.1 cells. Immunoblotting data showed that metformin activated the phosphorylation of STAT-1 and STAT-2 in OR-6 and JFH-1 infected Huh 7.5.1 cells. Metformin enhanced the phosphorylation of AMPK, and the metformin-activated IFN signaling was down-regulated by AMPK inhibitor. After treatment of AMPK inhibitor, the level of HCV core protein decreased by metformin can be rescued. In conclusion, metformin activates type I interferon signaling and inhibits the replication of HCV via activation of AMPK.

Rubicon Modulates Antiviral Type I Interferon (IFN) Signaling by Targeting IFN Regulatory Factor 3 Dimerization.

Rubicon is part of a Beclin-1-Vps34-containing autophagy complex. Rubicon induces antimicrobial responses upon Toll-like receptor (TLR) stimulation and functions as a feedback inhibitor to prevent unbalanced proinflammatory responses depending on dectin-1 signaling. However, the role played by Rubicon during antiviral immune responses, particularly the type I interferon (IFN) responses, remains largely unknown. Here, we report that Rubicon acts as a negative regulator for virus-triggered IFN signaling. Knockdown of Rubicon promoted type I interferon signaling and inhibited virus replication, while overexpression of Rubicon had the opposite effect. Rubicon specifically interacts with the interferon regulatory factor (IRF) association domain (IAD) of IRF3, and this interaction leads to inhibition of the dimerization of IRF3, which negatively regulates IFN-mediated antiviral response. Thus, our findings suggest the novel additional role of Rubicon as a negative regulator that inhibits the IFN signaling and cellular antiviral responses, providing a novel cellular mechanism of IRF3 inhibition.IMPORTANCE The type I IFN system is a critical innate immune response that protects organisms against virus infection. However, type I IFN signaling must be tightly regulated to avoid excessive production of IFNs. Hence, negative regulatory mechanisms for type I IFN signaling are important, and to date, several related molecules have been identified. Here, we show that Rubicon is a major negative regulator of type I IFN signaling, and unlike previous reports of cellular molecules that inhibit IRF3 activation via proteasomal degradation or dephosphorylation of IRF3, we show that Rubicon interacts with IRF3 and that ultimately this interaction leads to inhibition of the dimerization of IRF3. Thus, we identified a novel negative regulator of type I IFN signaling pathways and a novel cellular mechanism of IRF3 inhibition. The results of this study will increase our understanding of the role of negative-feedback mechanisms that regulate type I IFN signaling and maintain immune homeostasis.

AXL Promotes Zika Virus Infection in Astrocytes by Antagonizing Type I Interferon Signaling Pathway

Abstract Zika virus (ZIKV) is known to be associated with neonate microcephaly and Guillain-Barré syndrome in human adults. Despite the progress on understanding the causal link between ZIKV infection and microcephaly, the life cycle and pathogenesis of ZIKV remains elusive. In particular, recent studies have revealed a controversial role of AXL, a TAM family kinase receptor previously deemed as the entry receptor of ZIKV. In this study, we show that though genetic ablation of AXL protects primary human astrocytes and U-251MG astrocytoma cells from ZIKV infection, AXL knockout (KO) does not block the entry of ZIKV, instead it facilitates the up-regulation of type I interferon (IFN) signaling genes, including interferon regulatory factor 7 (IRF7), a key interferon transcriptional factor, and a number of type I interferon and interferon stimulating genes (ISGs). Knockout of type I IFN receptor a chain (IFNAR1) rescues the vulnerability of AXL KO astrocytes to ZIKV infection. In addition, AXL KO in astrocytes results in the down-regulation of SOCS1 in primary astrocytes and the failure to retain the homeostasis of SOCS1 in astrocyte cell lines, a known inhibitor of type I IFN signaling pathway, thus revealing important roles of AXL in maintaining the homeostasis of SOCS1 and thereby in containing type I interferon signaling pathway. Taken together, our study demonstrates that AXL does not function as the sole, if at all, entry receptor of ZIKV, but instead promotes ZIKV infection in astrocytes by SOCS1-mediated antagonism of type I interferon (IFN) signaling.

A MicroRNA Screen Identifies the Wnt Signaling Pathway as a Regulator of the Interferon Response during Flavivirus Infection.

The impact of mosquito-borne flavivirus infections worldwide is significant, and many critical aspects of these viruses' biology, including virus-host interactions, host cell requirements for replication, and how virus-host interactions impact pathology, remain to be fully understood. The recent reemergence and spread of flaviviruses, including dengue virus (DENV), West Nile virus (WNV), and Zika virus (ZIKV), highlight the importance of performing basic research on this important group of pathogens. MicroRNAs (miRNAs) are small, noncoding RNAs that modulate gene expression posttranscriptionally and have been demonstrated to regulate a broad range of cellular processes. Our research is focused on identifying pro- and antiflaviviral miRNAs as a means of characterizing cellular pathways that support or limit viral replication. We have screened a library of known human miRNA mimics for their effect on the replication of three flaviviruses, DENV, WNV, and Japanese encephalitis virus (JEV), using a high-content immunofluorescence screen. Several families of miRNAs were identified as inhibiting multiple flaviviruses, including the miRNA miR-34, miR-15, and miR-517 families. Members of the miR-34 family, which have been extensively characterized for their ability to repress Wnt/β-catenin signaling, demonstrated strong antiflaviviral effects, and this inhibitory activity extended to other viruses, including ZIKV, alphaviruses, and herpesviruses. Previous research suggested a possible link between the Wnt and type I interferon (IFN) signaling pathways. Therefore, we investigated the role of type I IFN induction in the antiviral effects of the miR-34 family and confirmed that these miRNAs potentiate interferon regulatory factor 3 (IRF3) phosphorylation and translocation to the nucleus, the induction of IFN-responsive genes, and the release of type I IFN from transfected cells. We further demonstrate that the intersection between the Wnt and IFN signaling pathways occurs at the point of glycogen synthase kinase 3β (GSK3β)-TANK-binding kinase 1 (TBK1) binding, inducing TBK1 to phosphorylate IRF3 and initiate downstream IFN signaling. In this way, we have identified a novel cellular signaling network with a critical role in regulating the replication of multiple virus families. These findings highlight the opportunities for using miRNAs as tools to discover and characterize unique cellular factors involved in supporting or limiting virus replication, opening up new avenues for antiviral research.IMPORTANCE MicroRNAs are a class of small regulatory RNAs that modulate cellular processes through the posttranscriptional repression of multiple transcripts. We hypothesized that individual miRNAs may be capable of inhibiting viral replication through their effects on host proteins or pathways. To test this, we performed a high-content screen for miRNAs that inhibit the replication of three medically relevant members of the flavivirus family: West Nile virus, Japanese encephalitis virus, and dengue virus 2. The results of this screen identify multiple miRNAs that inhibit one or more of these viruses. Extensive follow-up on members of the miR-34 family of miRNAs, which are active against all three viruses as well as the closely related Zika virus, demonstrated that miR-34 functions through increasing the infected cell's ability to respond to infection through the interferon-based innate immune pathway. Our results not only add to the knowledge of how viruses interact with cellular pathways but also provide a basis for more extensive data mining by providing a comprehensive list of miRNAs capable of inhibiting flavivirus replication. Finally, the miRNAs themselves or cellular pathways identified as modulating virus infection may prove to be novel candidates for the development of therapeutic interventions.

Type I interferon signaling in systemic immune cells from patients with alcoholic cirrhosis and its association with outcome

In immune cells, constitutively and acutely produced type I interferons (IFNs) engage autocrine/paracrine signaling pathways to induce IFN-stimulated genes (ISGs). Enhanced activity of IFN signaling pathways can cause excessive inflammation and tissue damage. We aimed to investigate ISG expression in systemic immune cells from patients with decompensated alcoholic cirrhosis, and its association with outcome. Peripheral blood mononuclear cells (PBMCs) from patients and heathy subjects were stimulated or not with lipopolysaccharide (LPS, an IFN inducer) or increasing concentrations of IFN-β. The expression of 48 ISGs and ten "non-ISG" inflammatory cytokines were analyzed using RT-qPCR. We developed an 8-ISG signature (IFN score) assessing ISG expression. LPS-stimulated ISG induction was significantly lower in PBMCs from patients with cirrhosis compared to healthy controls. Non-ISGs, however, showed higher induction. Lower induction of ISGs by LPS was not due to decreased IFN production by cirrhotic PBMCs or neutralization of secreted IFN, but a defective PBMC response to IFN. This defect was at least in part due to decreased constitutive ISG expression. Patients with the higher baseline IFN scores and ISG levels had the higher risk of death. At baseline, "non-ISG" cytokines did not correlate with outcome. PBMCs from patients with decompensated alcoholic cirrhosis exhibit downregulated ISG expression, both constitutively and after an acute stimulus. Our finding that higher baseline PBMC ISG expression was associated with higher risk of death, suggests that constitutive ISG expression in systemic immune cells contributes to the prognosis of alcoholic cirrhosis. Enhanced activity of IFN signaling pathways can cause excessive inflammation and tissue damage. Here we show that peripheral blood mononuclear cells (PBMCs) from patients with alcoholic cirrhosis exhibit a defect in interferon-stimulated genes (ISGs). We found that higher baseline ISG expression in PBMCs was associated with higher risk of death, revealing a probable contribution of ISG expression in immune cells to the outcome of alcoholic cirrhosis.

Identification of Novel Inhibitors of the Type I Interferon Induction Pathway Using Cell-Based High-Throughput Screening

Production of type I interferon (IFN) is an essential component of the innate immune response against invading pathogens. However, its production must be tightly regulated to avoid harmful effects. Compounds that modulate the IFN response are potentially valuable for a variety of applications due to IFN’s beneficial and detrimental roles. We developed and executed a cell-based high-throughput screen (HTS) targeting components that participate in and/or regulate the IRF3 and nuclear factor (NF)–κB branches of the IFN induction pathway. The assay detects activation of the IFN induction pathway via an enhanced green fluorescent protein (eGFP) reporter gene under the control of the IFNβ promoter and was optimized, miniaturized, and demonstrated suitable for HTS as robust Z′ factor scores of >0.6 were consistently achieved. A diversity screening set of 15,667 small molecules was assayed and two novel hit compounds validated that specifically inhibit the IFN induction pathway. We demonstrate that one of these compounds acts at or upstream of IRF3 phosphorylation. A second cell-based assay to detect activation of the IFN signaling (Jak-Stat) pathway via an eGFP reporter gene under the control of an IFN-stimulated response element (ISRE) containing MxA promoter also performed well (robust Z′ factor >0.7) and may therefore be similarly used to identify small molecules that modulate the IFN signaling pathway.

Bioinformatics analysis of gene expression profiles of dermatomyositis.

Dermatomyositis (DM) is a type of autoimmune inflammatory myopathy, which primarily affects the skin and muscle. The underlying mechanisms of DM remain poorly understood. The present study aimed to explore gene expression profile alterations, investigate the underlying mechanisms, and identify novel targets for DM. The GSE48280 dataset, which includes data from fiveDM and fivenormal muscle tissue samples, was obtained from the Gene Expression Omnibus. Firstly, differentially expressed genes (DEGs) were screened by limma package in R. Subsequently, functional and pathway enrichment analyses were performed using ClueGO from Cytoscape. Finally, protein‑protein interaction (PPI) networks were constructed using STRING and Cytoscape, in order to identify hub genes. As a result, 180upregulated and 21downregulated genes were identified in the DM samples. The Gene Ontology enrichment analysis revealed that the typeI interferon (IFN) signaling pathway was the most significantly enriched term within the DEGs. The Kyoto Encyclopedia of Genes and Genomes pathway analysis identified 27significant pathways, the majority of which can be divided into the infectious diseases and immune system categories. Following construction of PPI networks, 24hub genes were selected, all of which were associated with the typeI IFN signaling pathway in DM. The findings of the present study indicated that typeI IFNs may have a central role in the induction of DM. In addition, other DEGs, including chemokine (C‑C motif) ligand5, C‑X‑C motif chemokine10, Toll‑like receptor3, DEXD/H‑Box helicase58, interferon induced with helicaseC domain1, interferon‑stimulated gene 15 and MXdynamin‑like GTPase1, may be potential targets for DM diagnosis and treatment.

Type I Interferon-Mediated Induction of Antiviral Genes and Proteins Fails to Protect Cells from the Cytopathic Effects of Sendai Virus Infection.

Sendai virus (SeV), a murine paramyxovirus, has been used to study the induction of type I interferon (IFN) subtypes in robust quantities. Few studies have measured whether the IFN that SeV induces actually fulfills its intended purpose of interfering with virus-mediated effects in the cells in which it is produced. We determined the effects of IFN on SeV-mediated cytopathic effects (CPE) and the ability of IFN to protect against virus infection. SeV-induced biologically active IFN resulted in Jak/STAT activation and the production of a number of interferon-stimulated genes (ISGs). However, these responses did not inhibit SeV replication or CPE. This observation was not due to SeV effects on canonical IFN signaling. Furthermore, pretreating cells with type I IFN and establishing an antiviral state before infection did not mediate SeV effects. Therefore, the induction of canonical IFN signaling pathways and ISGs does not always confer protection against the IFN-inducing virus. Because type I IFNs are approved to treat various infections, our findings suggest that typical markers of IFN activity may not be indicative of a protective antiviral response and should not be used alone to determine whether an antiviral state against a particular virus is achieved.

Porcine Epidemic Diarrhea Virus Infection Inhibits Interferon Signaling by Targeted Degradation of STAT1.

Porcine epidemic diarrhea virus (PEDV) is a worldwide-distributed alphacoronavirus, but the pathogenesis of PEDV infection is not fully characterized. During virus infection, type I interferon (IFN) is a key mediator of innate antiviral responses. Most coronaviruses develop some strategy for at least partially circumventing the IFN response by limiting the production of IFN and by delaying the activation of the IFN response. However, the molecular mechanisms by which PEDV antagonizes the antiviral effects of interferon have not been fully characterized. Especially, how PEDV impacts IFN signaling components has yet to be elucidated. In this study, we observed that PEDV was relatively resistant to treatment with type I IFN. Western blot analysis showed that STAT1 expression was markedly reduced in PEDV-infected cells and that this reduction was not due to inhibition of STAT1 transcription. STAT1 downregulation was blocked by a proteasome inhibitor but not by an autophagy inhibitor, strongly implicating the ubiquitin-proteasome targeting degradation system. Since PEDV infection-induced STAT1 degradation was evident in cells pretreated with the general tyrosine kinase inhibitor, we conclude that STAT1 degradation is independent of the IFN signaling pathway. Furthermore, we report that PEDV-induced STAT1 degradation inhibits IFN-α signal transduction pathways. Pharmacological inhibition of STAT1 degradation rescued the ability of the host to suppress virus replication. Collectively, these data show that PEDV is capable of subverting the type I interferon response by inducing STAT1 degradation. In this study, we show that PEDV is resistant to the antiviral effect of IFN. The molecular mechanism is the degradation of STAT1 by PEDV infection in a proteasome-dependent manner. This PEDV infection-induced STAT1 degradation contributes to PEDV replication. Our findings reveal a new mechanism evolved by PEDV to circumvent the host antiviral response.

Role of glucose-6-phosphate dehydrogenase in hepatitis B virus replication and its possible mechanism of action

To investigate the role of glucose-6-phosphate dehydrogenase (G6PD) in hepatitis B virus (HBV) replication and its possible mechanism of action. Tissue microarray, quantitative real-time PCR, and Western blot were performed to analyze the differences in G6PD expression levels in the HBV-positive and HBV-negative liver tissues, HepG2.2.15 cells, and HepG2 cells. The siRNA transfection technique was used to knock down G6PD gene in HepG2.2.15 cells for 48 hours. Chemiluminescence was used for HBsAg and HBeAg quantification in supernatant, and quantitative real-time PCR was used to measure HBV DNA, type I interferon (IFN), and downstream IFN-stimulated genes. The t-test was used for comparison between groups. G6PD expression was significantly upregulated in the HBV-positive liver tissues and cells compared with HBV-negative liver tissues and cells, and the stain intensity and immunohistochemical scores were 89.69±54.92 and 31.90±18.62, respectively (P < 0.05). After G6PD expression in HepG2.2.15 cells was interfered by siRNA, the quantitative levels of HBV DNA, HBsAg, and HBeAg in supernatant were reduced significantly, and the mRNA expression levels of IFNα1, IFNβ1, and five downstream IFN-stimulated genes (OAS1, ISG15, OAS3, EIF2α, and PKR) increased significantly (all P < 0.05). G6PD plays a vital role in HBV replication, and its mechanism of action in regulating HBV replication may be related to type I IFN signaling pathway.

Identification of Cyclin-Dependent Kinase 1 as a Novel Regulator of Type I Interferon Signaling in Systemic Lupus Erythematosus.

Type I interferon (IFN) signaling is regarded as a central pathogenic pathway in systemic lupus erythematosus (SLE). Specific inhibition of this pathway is a core area for the development of new therapies for SLE. This study was undertaken to clarify the pathogenic mechanism involved and to identify new therapeutic targets, using a high-throughput screening platform to determine novel regulators that contribute to the overactivation of the type I IFN signaling pathway in SLE. A high-throughput IFN-stimulated response element (ISRE)-luciferase assay was used to screen for candidate genes that regulate the IFN signaling pathway. Western blotting was used to confirm the regulatory function of CDK1. SYBR Green quantitative reverse transcriptase-polymerase chain reaction was used to detect the expression of individual IFN-stimulated genes (ISGs). The differential expression of CDK1 and ISGs in SLE patients and healthy controls was analyzed using RNA sequencing data and a microarray. The high-throughput ISRE-luciferase assay revealed that CDK1 enhanced type I IFN signaling. Consistent with this finding, CDK1 promoted the type I IFN-induced phosphorylation of STAT-1 and the up-regulated expression of ISGs. CDK1 expression was elevated in peripheral blood mononuclear cells (PBMCs) and kidney biopsy specimens from SLE patients and correlated positively with their IFN scores. A CDK1 inhibitor reduced the expression of ISGs in PBMCs from SLE patients and in renal cells from mice with lupus. Our findings indicate that CDK1 is a positive regulator of the IFN signaling pathway. The overexpression of CDK1 might contribute to the abnormally amplified type I IFN signaling in SLE, and the inhibition of CDK1 could be used to down-regulate type I IFN signaling in SLE.

Cause and consequences of the activated type I interferon system in SLE.

Patients with systemic lupus erythematosus (SLE) have an increased expression of type I interferon (IFN)-regulated genes (an IFN signature), which is caused by an ongoing production of type I IFNs by plasmacytoid dendritic cells (pDCs). The reasons behind the continuous IFN production in SLE are the presence of self-derived IFN inducers and a lack of negative feed-back signals that downregulate the IFN response. In addition, several cells in the immune system promote the IFN production by pDCs and gene variants in the type I IFN signaling pathway contribute to the IFN signature. The type I IFNs act as an immune adjuvant and stimulate T cells, B cells, and monocytes, which all play an important role in the loss of tolerance and persistent autoimmune reaction in SLE. Consequently, new treatments aiming to inhibit the activated type I IFN system in SLE are now being developed and investigated in clinical trials.

Parainfluenza Virus 3 Blocks Antiviral Mediators Downstream of the Interferon Lambda Receptor by Modulating Stat1 Phosphorylation.

Parainfluenza viruses are known to inhibit type I interferon (IFN) production; however, there is a lack of information regarding the type III IFN response during infection. Type III IFNs signal through a unique heterodimeric receptor, IFN-λR1/interleukin-10R2 (IL-10R2), which is primarily expressed by epithelial cells. Parainfluenza virus 3 (PIV-3) infection is highly restricted to the airway epithelium. We therefore sought to examine type III IFN signaling pathways during PIV-3 infection of epithelial cells. We used three strains of PIV-3: human PIV-3 (HPIV-3), bovine PIV-3 (BPIV-3), and dolphin PIV-1 (Tursiops truncatus PIV-1, or TtPIV-1). Here, we show that message levels of IL-29 are significantly increased during PIV-3 infection, yet downstream antiviral signaling molecules are not upregulated to levels similar to those of the positive control. Furthermore, in Vero cells infected with PIV-3, stimulation with recombinant IL-29/-28A/-28B does not cause upregulation of downstream antiviral molecules, suggesting that PIV-3 interferes with the JAK/STAT pathway downstream of the IFN-λR1/IL-10R2 receptor. We used Western blotting to examine the phosphorylation of Stat1 and Stat2 in Vero cells and the bronchial epithelial cell line BEAS-2B. In Vero cells, we observed reduced phosphorylation of the serine 727 (S727) site on Stat1, while in BEAS-2B cells Stat1 phosphorylation was decreased at the tyrosine 701 (Y701) site during PIV-3 infection. PIV-3 therefore interferes with the phosphorylation of Stat1 downstream of the type III IFN receptor. These data provide new evidence regarding strategies employed by parainfluenza viruses to effectively circumvent respiratory epithelial cell-specific antiviral immunity. Parainfluenza virus (PIV) in humans is associated with bronchiolitis and pneumonia and can be especially problematic in infants and the elderly. Also seen in cattle, bovine PIV-3 causes respiratory infections in young calves. In addition, PIV-3 is one of a number of pathogens that contribute to the bovine respiratory disease complex (BRDC). As their name suggests, interferons (IFNs) are produced by cells to interfere with viral replication. Paramyxoviruses have previously been shown to block production and downstream signaling of type I IFNs. For the first time, it is shown here that PIV-3 can induce protective type III IFNs in epithelial cells, the primary site of PIV-3 infection. However, we found that PIV-3 modulates signaling pathways downstream of the type III IFN receptor to block production of several specific molecules that aid in a productive antiviral response. Importantly, this work expands our understanding of how PIV-3 effectively evades host innate immunity.

The Golgi-Associated Plant Pathogenesis-Related Protein GAPR-1 Enhances Type I Interferon Signaling Pathway in Response to Toll-Like Receptor 4.

Lipopolysaccharide (LPS) activates Toll-like receptor 4 (TLR4) through the TIRAP-MyD88 dependent and TRAM-TRIF dependent signaling pathways, respectively. However, the underlying relevance between two signaling pathways remains largely elusive. Here, we investigated the role of the Golgi-Associated plant Pathogenesis-Related protein (GAPR-1) in type I interferon (IFN) signaling pathway in response to TLR4. We found that TIRAP-MyD88 dependent kinase IRAK1 phosphorylated GAPR-1 at Serine 58 site. The phosphorylation of GAPR-1 promoted its interaction with TRAM-TRIF dependent inhibitor TMED7, and impaired TMED7-mediated disruption of the TRAM-TRIF complex to trigger IFN-β and the IL10 secretion. Collectively, our study identified a previously unrecognized role for GAPR-1 to control a unifying TLR4 signaling complex and to regulate type I IFN signaling activation. Understanding the mechanism of GAPR-1 in type I IFN signaling pathway would provide strategies for treatment of infectious diseases.

MiR-744 enhances type I interferon signaling pathway by targeting PTP1B in primary human renal mesangial cells.

Renal mesangial cells (RMCs) constitute a population of cells in glomerular mesangium. Inflammatory cytokines produced by RMCs play a vital role in renal inflammation. miRNAs are key regulators of inflammatory cytokine expression. The abnormal expression of renal miRNAs and the consequent changes in inflammatory signal transduction are closely associated with renal inflammation. However, our knowledge of the functions of renal miRNAs is still limited. In this study, we investigated the role of miR-744 in type I interferon (IFN) signaling pathway in primary human RMCs. We show that overexpression of miR-744 enhances IFN-induced CCL2, CCL5, CXCL10, and IL6 expression specifically in RMCs. We found that the activation of TYK2, STAT1 and STAT3 was significantly enhanced by miR-744. miR-744 also enhanced the activation of non-classical signal components, such as ERK and p38. We then identified PTP1B, a ubiquitously expressed phosphatase, as the target of miR-744 that is responsible for enhancing type I IFN response. Finally, miR-744 expression was induced by type I IFN in RMCs. Collectively, our data indicate that by targeting PTP1B, miR-744 plays a feed-forward role in regulating type I IFN signaling pathway. These findings give us new insights into the functions of renal miRNAs in regulating important signaling pathways.

Negative feedback contributes to the stochastic expression of the interferon-β gene in virus-triggered type I interferon signaling pathways

Type I interferon (IFN) signaling pathways play an essential role in the defense against early viral infections; however, the diverse and intricate molecular mechanisms of virus-triggered type I IFN responses are still poorly understood. In this study, we analyzed and compared two classes of models i.e., deterministic ordinary differential equations (ODEs) and stochastic models to elucidate the dynamics and stochasticity of type I IFN signaling pathways. Bifurcation analysis based on an ODE model reveals that the system exhibits a bistable switch and a one-way switch at high or low levels when the strengths of the negative and positive feedbacks are tuned. Furthermore, we compared the stochastic simulation results under the Master and Langevin equations. Both of the stochastic equations generate the bistable switch phenomenon, and the distance between two stable states are smaller than normal under the simulation of the Langevin equation. The quantitative computations also show that a moderate ratio between positive and negative feedback strengths is required to ensure a reliable switch between the different IFN concentrations that regulate the immune response. Moreover, we propose a multi-state stochastic model based on the above deterministic model to describe the multi-cellular system coupled with the diffusion of IFNs. The perturbation and inhibition analysis showed that the positive feedback, as well as noises, has little effect on the stochastic expression of IFNs, but the negative feedback of ISG56 on the activation of IRF7 has a great influence on IFN stochastic expression. Together, these results reveal that positive feedback stabilizes IFN gene expression, and negative feedback may be the main contribution to the stochastic expression of the IFN gene in the virus-triggered type I IFN response. These findings will provide new insight into the molecular mechanisms of virus-triggered type I IFN signaling pathways.

Type I Interferon Controls Propagation of Long Interspersed Element-1

Type I interferons (IFN) including IFNα and IFNβ are critical for the cellular defense against viruses. Here we report that increased levels of IFNβ were found in testes from mice deficient in MOV10L1, a germ cell-specific RNA helicase that plays a key role in limiting the propagation of retrotransposons including Long Interspersed Element-1 (LINE-1). Additional experiments revealed that activation of LINE-1 retrotransposons increases the expression of IFNβ and of IFN-stimulated genes. Conversely, pretreatment of cells with IFN suppressed the replication of LINE-1. Furthermore, the efficacy of LINE-1 replication was increased in isogenic cell lines harboring inactivating mutations in diverse elements of the IFN signaling pathway. Knockdown of the IFN receptor chain IFNAR1 also stimulated LINE-1 propagation in vitro. Finally, a greater accumulation of LINE-1 was found in mice that lack IFNAR1 compared with wild type mice. We propose that LINE-1-induced IFN plays an important role in restricting LINE-1 propagation and discuss the putative role of IFN in preserving the genome stability.

MiR-122 directly inhibits human papillomavirus E6 gene and enhances interferon signaling through blocking suppressor of cytokine signaling 1 in SiHa cells.

Human Papillomavirus (HPV) 16 infection is considered as one of the significant causes of human cervical cancer. The expression of the viral oncogenes like E6 and E7 play an important role in the development of the cancer. MiR-122 has been reported to exhibit a strong relationship with hepatitis viruses and take part in several tumor development, while the effects of miR-122 on HPV infection and the HPV viral oncogenes expression still remain unexplored. In this study, using RNAhybrid software, the potential binding sites between miR-122 and HPV16 E6 and E7 mRNAs were identified. Over and loss of miR-122 function showed that miR-122 could directly bind with HPV16 E6 mRNA and significantly inhibit its expression in SiHa cells, which was further confirmed by constructing the miR-122-E6-mu to eliminate the miR-122 binding effects with E6. The increase of the expression of type I interferon (IFN) and its classical effective molecules and the phosphorylation of signal transducers and activators of transcription (STAT1) protein indicated that miR-122 might enhance type I interferon in cervical carcinoma cells, which explained the significant reduction of HPV16 E7 and E6*I mRNA expression. This might be due to the binding between miR-122 and suppressor of cytokine signaling 1 (SOCS1) mRNA, which is the suppressor of interferon signaling pathway. Moreover, it was identified that the miR-122 binding position was nt359-nt375 in SOCS1 mRNA. Taken together, this study indicated that HPV16 could be effectively inhibited by miR-122 through both direct binding with E6 mRNA and promoting SOCS1-dependent IFN signaling pathway. Thus, miR-122 may serve as a new therapeutic option for inhibiting HPV infection.

Molecular cloning and functional characterization of porcine DEAD (Asp–Glu–Ala–Asp) box polypeptide 41 (DDX41)

DEAD (Asp–Glu–Ala–Asp) box polypeptide 41 (DDX41), a member of the DEXDc helicase family, was recently identified as an intracellular DNA sensor in mouse myeloid dendritic cells. In this study, porcine DDX41 (poDDX41) was cloned and its role in the type I interferon (IFN) signaling pathway was investigated in porcine kidney (PK-15) cells. Full-length poDDX41 cDNA encodes 622 amino acid residues and contains a DEADc domain and a HELICc domain. poDDX41 mRNA is widely expressed in different tissues, especially the stomach and liver. Overexpression of poDDX41 in PK-15 cells induced IFN-β by activating transcription factors IRF3 and NF-κB. Knockdown of poDDX41 with siRNA significantly reduced IFN-β expression induced by poly(dA:dT), a double-stranded DNA (dsDNA) analogue, or pseudorabies virus, a dsDNA swine virus. Therefore, poDDX41 is involved in the dsDNA- and dsDNA-virus-mediated type I IFN signaling pathway in porcine kidney cells.

Transcription Factor STAT3 and Type I Interferons Are Corepressive Insulators for Differentiation of Follicular Helper and T Helper 1 Cells

Follicular helper T (Tfh) cells are required for the establishment of T-dependent B cell memory and high affinity antibody-secreting cells. We have revealed herein opposing roles for signal transducer and activator of transcription 3 (STAT3) and type I interferon (IFN) signaling in the differentiation of Tfh cells following viral infection. STAT3-deficient CD4(+) Tcells had a profound defect in Tfh cell differentiation, accompanied by decreased germinal center (GC) B cells and antigen-specific antibody production during acute infection with lymphocytic choriomeningitis virus. STAT3-deficient Tfh cells had strikingly increased expression of a number of IFN-inducible genes, in addition to enhanced T-bet synthesis, thus adopting a T helper 1 (Th1) cell-like effector phenotype. Conversely, IFN-αβ receptor blockade restored Tfh and GC B cell phenotypes in mice containing STAT3-deficient CD4(+) Tcells. These data suggest mutually repressive roles for STAT3 andtype I IFN signaling pathways in the differentiation of Tfh cells following viral infection.