IFNAR signaling of neuroectodermal cells is essential for the survival of C57BL/6 mice infected with Theiler's murine encephalomyelitis virus.

BACKGROUND: Theiler's murine encephalomyelitis virus (TMEV) is a single-stranded RNA virus that causes encephalitis followed by chronic demyelination in SJL mice and spontaneous seizures in C57BL/6 mice. Since earlier studies indicated a critical role of type I interferon (IFN-I) signaling in the control of viral replication in the central nervous system (CNS), mouse strain-specific differences in pathways induced by the IFN-I receptor (IFNAR) might determine the outcome of TMEV infection. METHODS: Data of RNA-seq analysis and immunohistochemistry were used to compare the gene and protein expression of IFN-I signaling pathway members between mock- and TMEV-infected SJL and C57BL/6 mice at 4, 7 and 14 days post-infection (dpi). To address the impact of IFNAR signaling in selected brain-resident cell types, conditional knockout mice with an IFNAR deficiency in cells of the neuroectodermal lineage (NesCre±IFNARfl/fl), neurons (Syn1Cre±IFNARfl/fl), astrocytes (GFAPCre±IFNARfl/fl), and microglia (Sall1CreER±IFNARfl/fl) on a C57BL/6 background were tested. PCR and an immunoassay were used to quantify TMEV RNA and cytokine and chemokine expression in their brain at 4 dpi. RESULTS: RNA-seq analysis revealed upregulation of most ISGs in SJL and C57BL/6 mice, but Ifi202b mRNA transcripts were only increased in SJL and Trim12a only in C57BL/6 mice. Immunohistochemistry showed minor differences in ISG expression (ISG15, OAS, PKR) between both mouse strains. While all immunocompetent Cre-negative control mice and the majority of mice with IFNAR deficiency in neurons or microglia survived until 14 dpi, lack of IFNAR expression in all cells (IFNAR-/-), neuroectodermal cells, or astrocytes induced lethal disease in most of the analyzed mice, which was associated with unrestricted viral replication. NesCre±IFNARfl/fl mice showed more Ifnb1, Tnfa, Il6, Il10, Il12b and Ifng mRNA transcripts than Cre-/-IFNARfl/fl mice. IFNAR-/- mice also demonstrated increased IFN-α, IFN-ß, IL1-ß, IL-6, and CXCL-1 protein levels, which highly correlated with viral load. CONCLUSIONS: Ifi202b and Trim12a expression levels likely contribute to mouse strain-specific susceptibility to TMEV-induced CNS lesions. Restriction of viral replication is strongly dependent on IFNAR signaling of neuroectodermal cells, which also controls the expression of key pro- and anti-inflammatory cytokines during viral brain infection.

PARP1 Enhances Influenza A Virus Propagation by Facilitating Degradation of Host Type I Interferon Receptor.

Influenza A virus (IAV) utilizes multiple strategies to confront or evade host type I interferon (IFN)-mediated antiviral responses in order to enhance its own propagation within the host. One such strategy is to induce the degradation of type I IFN receptor 1 (IFNAR1) by utilizing viral hemagglutinin (HA). However, the molecular mechanism behind this process is poorly understood. Here, we report that a cellular protein, poly(ADP-ribose) polymerase 1 (PARP1), plays a critical role in mediating IAV HA-induced degradation of IFNAR1. We identified PARP1 as an interacting partner for IAV HA through mass spectrometry analysis. This interaction was confirmed by coimmunoprecipitation analyses. Furthermore, confocal fluorescence microscopy showed altered localization of endogenous PARP1 upon transient IAV HA expression or during IAV infection. Knockdown or inhibition of PARP1 rescued IFNAR1 levels upon IAV infection or HA expression, exemplifying the importance of PARP1 for IAV-induced reduction of IFNAR1. Notably, PARP1 was crucial for the robust replication of IAV, which was associated with regulation of the type I IFN receptor signaling pathway. These results indicate that PARP1 promotes IAV replication by controlling viral HA-induced degradation of host type I IFN receptor. Altogether, these findings provide novel insight into interactions between influenza virus and the host innate immune response and reveal a new function for PARP1 during influenza virus infection.IMPORTANCE Influenza A virus (IAV) infections cause seasonal and pandemic influenza outbreaks, which pose a devastating global health concern. Despite the availability of antivirals against influenza, new IAV strains continue to persist by overcoming the therapeutics. Therefore, much emphasis in the field is placed on identifying new therapeutic targets that can more effectively control influenza. IAV utilizes several tactics to evade host innate immunity, which include the evasion of antiviral type I interferon (IFN) responses. Degradation of type I IFN receptor (IFNAR) is one known method of subversion, but the molecular mechanism for IFNAR downregulation during IAV infection remains unclear. Here, we have found that a host protein, poly(ADP-ribose) polymerase 1 (PARP1), facilitates IFNAR degradation and accelerates IAV replication. The findings reveal a novel cellular target for the potential development of antivirals against influenza, as well as expand our base of knowledge regarding interactions between influenza and the host innate immunity.

Cloning and characterisation of type I interferon receptor 1 in orange-spotted grouper (Epinephelus coioides) for response to nodavirus infection.

Grouper is known as a highly economical teleost species in the Asian aquaculture industry; however, intensive culture activities easily cause disease outbreak, especially viral disease. For the prevention of viral outbreaks, interferon (IFN) is among the major defence systems being studied in different species. Fish type I IFNs are known to possess antiviral properties similar to mammalian type I IFNs. In order to stimulate antiviral function, IFN will bind to its cognate receptor, the type I interferon receptor (IFNAR), composed of heterodimeric receptor subunits known as IFNAR1 and IFNΑR2. The binding of type I interferon to receptors assists in the transduction of signals from the external to internal environments of cells to activate biological responses. In order to study the function of IFN, we first need to understand IFN receptors. In this study, we cloned and identified IFNAR1 in orange-spotted grouper (osgIFNAR1) and noted the up-regulated mRNA expression of the receptor and downstream effectors in the head kidney cells with cytokine treatment. The transcriptional expression of osgIFNAR1, which is characterised using polyinosinic-polycytidylic acid (poly[I:C]) and lipopolysaccharide (LPS) treatments, indicated the involvement of osgIFNAR1 in the immune response of grouper. The subcellular localisation of osgIFNAR1 demonstrated scattering across the grouper cell. Viral infection showed the negative feedback regulation of osgIFNAR1 in grouper larvae. Further loss of function of IFNAR1 showed a decreased expression of the virus. This study reported the identification of osgIFNAR1 and characterisation of receptor sensitivity towards immunostimulants, cytokine response, and viral challenge in the interferon pathway of orange-spotted grouper and possible different role of the receptor in viral production. Together, these results provide a frontline report of the potential function of osgIFNAR1 in the innate immunity of teleost.

Pseudorabies Virus dUTPase UL50 Induces Lysosomal Degradation of Type I Interferon Receptor 1 and Antagonizes the Alpha Interferon Response.

Alphaherpesviruses that establish persistent infections rely partly on their ability to evade host antiviral responses, notably the type I interferon (IFN) response. However, the mechanisms employed by alphaherpesviruses to avoid this response are not well understood. Pseudorabies virus (PRV) is an economically important pathogen and a useful model system for studying alphaherpesvirus biology. To identify PRV proteins that antagonize type I IFN signaling, we performed a screen by using an IFN-stimulated response element reporter in the swine cell line CRL. Unexpectedly, we identified the dUTPase UL50 as a strong inhibitor. We confirmed that UL50 has the ability to inhibit type I IFN signaling by performing ectopic expression of UL50 in cells and deletion of UL50 in PRV. Mechanistically, UL50 impeded type I IFN-induced STAT1 phosphorylation, likely by accelerating lysosomal degradation of IFN receptor 1 (IFNAR1). In addition, this UL50 activity was independent of its dUTPase activity and required amino acids 225 to 253 in the C-terminal region. The UL50 encoded by herpes simplex virus 1 (HSV-1) also possessed similar activity. Moreover, UL50-deleted PRV was more susceptible to IFN than UL50-proficient PRV. Our results suggest that in addition to its dUTPase activity, the UL50 protein of alphaherpesviruses possesses the ability to suppress type I IFN signaling by promoting lysosomal degradation of IFNAR1, thereby contributing to immune evasion. This finding reveals UL50 as a potential antiviral target.IMPORTANCE Alphaherpesviruses can establish lifelong infections and cause many diseases in humans and animals. Pseudorabies virus (PRV) is a swine alphaherpesvirus that threatens pig production. Using PRV as a model, we found that this alphaherpesvirus could utilize its encoded dUTPase UL50 to induce IFNAR1 degradation and inhibit type I IFN signaling in an enzymatic activity-independent manner. Our finding reveals a mechanism employed by an alphaherpesvirus to evade the immune response and indicates that UL50 is an important viral protein in pathogenesis and is a potential target for antiviral drug development.

Estrous cycle stage-dependent manner of type I interferon-stimulated genes induction in the bovine endometrium.

Interferon tau (IFN-τ) is a ruminant-specific type I IFN secreted by a conceptus before its attachment to the uterus. IFN-τ induces the expression of IFN-stimulated genes (ISGs) via the type I IFN receptor (IFNAR), which is composed of IFNAR1 and IFNAR2 subunits in the endometrium. However, expression patterns of IFNARs during the estrous cycle have not been reported. We hypothesized that the response to a type I IFN changes along with IFNARs and the IFN-regulatory factors (IRFs) driving transcription of IFN signal-related genes and modulating a type I IFN signal during the estrous cycle. We investigated the estrous cycle stage-dependent type I IFN induction of ISGs and expression patterns of IFN signal-related genes in bovine endometrial tissues. Endometrial tissue pieces collected from bovine uteri at each estrous stage (early, mid, and late) were cultured with or without recombinant bovine IFN-α or concentrated pregnant uterine flushing (PUF) on day 18 after confirming the presence of a conceptus. IFN-α and PUF each significantly increased the expression of ISGs in endometrial tissues. The induction levels of the typical ISGs (MX1-a and ISG15) were significantly higher at the mid stage and correlated with high expression of IRFs at the mid stage. The immunostaining of IFNARs showed strong fluorescence intensities in luminal and glandular epithelia at the early and mid stages. Collectively, these results suggest that the endometrium exhibits estrous cycle stage-dependent responsiveness to type I IFN that may be associated with the expression of IFNARs and IRFs for pregnancy recognition.

Hepatitis C virus replication in mouse cells is restricted by IFN-dependent and -independent mechanisms.

BACKGROUND & AIMS: Current treatment strategies for hepatitis C virus (HCV) infection include pegylated interferon (IFN)-alfa and ribavirin. Approximately 50% of patients control HCV infection after treatment, but the broad range of patients' outcomes and responses to treatment, among all genotypes, indicates a role for host factors. Although the IFN system is important in limiting HCV replication, the virus has evolved mechanisms to circumvent the IFN response. However, direct, IFN-independent antiviral processes also might help control HCV replication. We examined the role of IFN-independent responses against HCV replication. METHODS: We analyzed replication of the subgenomic JFH1 replicon in embryonic fibroblasts and primary hepatocytes from mice with disruptions in genes encoding factors in the IFN-dependent and alternative antiviral pathways (signal transducers and activators of transcription 1 [STAT1], protein kinase R, interferon regulatory factors (IRF) IRF-1, IRF-3, IRF-5, IRF-7, mitochondrial antiviral signaling molecule [MAVS], and IFN receptor [IFNAR]). We also assessed the effects of expression of these factors by mouse primary hepatocytes on HCV replication. RESULTS: In addition to IRF-3- and IFN-mediated antiviral responses, IFN-independent, but IRF-1- and IRF-5-dependent mechanisms, restrict HCV replication in mouse embryonic fibroblasts. In primary hepatocytes these IFN-independent require MAVS and IRF-1. CONCLUSIONS: HCV replication is limited by interferon-mediated pathways as well pathways that are independent of type I IFNs. IRF1 and IRF5 control IFN-independent signaling events that lead to antiviral responses. We observed antiviral roles of IRF1 and IRF5 that were IFN-independent and cell-type specific. These mechanisms are important in controlling viruses that interfere with the IFN signaling because cells retain the ability to induce functional but local antiviral states through expression of interferon-stimulated genes.

Interferon-ß modulates microglial polarization to ameliorate delayed tPA-exacerbated brain injury in ischemic stroke.

Tissue plasminogen activator (tPA) is the only FDA-approved drug for the treatment of ischemic stroke. Delayed tPA administration is associated with increased risks of blood-brain barrier (BBB) disruption and hemorrhagic transformation. Studies have shown that interferon beta (IFNß) or type I IFN receptor (IFNAR1) signaling confers protection against ischemic stroke in preclinical models. In addition, we have previously demonstrated that IFNß can be co-administered with tPA to alleviate delayed tPA-induced adverse effects in ischemic stroke. In this study, we investigated the time limit of IFNß treatment on the extension of tPA therapeutic window and assessed the effect of IFNß on modulating microglia (MG) phenotypes in ischemic stroke with delayed tPA treatment. Mice were subjected to 40 minutes transient middle cerebral artery occlusion (MCAO) followed by delayed tPA treatment in the presence or absence of IFNß at 3h, 4.5h or 6h post-reperfusion. In addition, mice with MG-specific IFNAR1 knockdown were generated to validate the effects of IFNß on modulating MG phenotypes, ameliorating brain injury, and lessening BBB disruption in delayed tPA-treated MCAO mice. Our results showed that IFNß extended tPA therapeutic window to 4.5h post-reperfusion in MCAO mice, and that was accompanied with attenuated brain injury and lessened BBB disruption. Mechanistically, our findings revealed that IFNß modulated MG polarization, leading to the suppression of inflammatory MG and the promotion of anti-inflammatory MG, in delayed tPA-treated MCAO mice. Notably, these effects were abolished in MG-specific IFNAR1 knockdown MCAO mice. Furthermore, the protective effect of IFNß on the amelioration of delayed tPA-exacerbated ischemic brain injury was also abolished in these mice. Finally, we identified that IFNß-mediated modulation of MG phenotypes played a role in maintaining BBB integrity, because the knockdown of IFNAR1 in MG partly reversed the protective effect of IFNß on lessening BBB disruption in delayed tPA-treated MCAO mice. In summary, our study reveals a novel function of IFNß in modulating MG phenotypes, and that may subsequently confer protection against delayed tPA-exacerbated brain injury in ischemic stroke.

Type I Interferon Receptor Subunit 1 Deletion Attenuates Experimental Abdominal Aortic Aneurysm Formation.

OBJECTIVE: Type I interferon receptor signaling contributes to several autoimmune and vascular diseases such as lupus, atherosclerosis and stroke. The purpose of this study was to assess the influence of type I interferon receptor deficiency on the formation and progression of experimental abdominal aortic aneurysms (AAAs). METHODS: AAAs were induced in type I interferon receptor subunit 1 (IFNAR1)-deficient and wild type control male mice via intra-infrarenal aortic infusion of porcine pancreatic elastase. Immunostaining for IFNAR1 was evaluated in experimental and clinical aneurysmal abdominal aortae. The initiation and progression of experimental AAAs were assessed via ultrasound imaging prior to (day 0) and days 3, 7 and 14 following elastase infusion. Aneurysmal histopathology was analyzed at sacrifice. RESULTS: Increased aortic medial and adventitial IFNAR1 expression was present in both clinical AAAs harvested at surgery and experimental AAAs. Following AAA induction, wild type mice experienced progressive, time-dependent infrarenal aortic enlargement. This progression was substantially attenuated in IFNAR1-deficient mice. On histological analyses, medial elastin degradation, smooth muscle cell depletion, leukocyte accumulation and neoangiogenesis were markedly diminished in IFNAR1-deficient mice in comparison to wild type mice. CONCLUSION: IFNAR1 deficiency limited experimental AAA progression in response to intra-aortic elastase infusion. Combined with clinical observations, these results suggest an important role for IFNAR1 activity in AAA pathogenesis.

The Th1/Tfh-like biased responses elicited by the rASP-1 innate adjuvant are dependent on TRIF and Type I IFN receptor pathways.

Ov-ASP-1 (rASP-1), a parasite-derived protein secreted by the helminth Onchocerca volvulus, is an adjuvant which enhances the potency of the influenza trivalent vaccine (IIV3), even when used with 40-fold less IIV3. This study is aimed to provide a deeper insight into the molecular networks that underline the adjuvanticity of rASP-1. Here we show that rASP-1 stimulates mouse CD11c+ bone marrow-derived dendritic (BMDCs) to secrete elevated levels of IL-12p40, TNF-α, IP-10 and IFN-ß in a TRIF-dependent but MyD88-independent manner. rASP-1-activated BMDCs promoted the differentiation of naïve CD4+ T cells into Th1 cells (IFN-γ+) that was TRIF- and type I interferon receptor (IFNAR)-dependent, and into Tfh-like cells (IL21+) and Tfh1 (IFN-γ+ IL21+) that were TRIF-, MyD88- and IFNAR-dependent. rASP-1-activated BMDCs promoted the differentiation of naïve CD4+ T cells into Th17 (IL-17+) cells only when the MyD88 pathway was inhibited. Importantly, rASP-1-activated human blood cDCs expressed upregulated genes that are associated with DC maturation, type I IFN and type II IFN signaling, as well as TLR4-TRIF dependent signaling. These activated cDCs promoted the differentiation of naïve human CD4+ T cells into Th1, Tfh-like and Th17 cells. Our data thus confirms that the rASP-1 is a potent innate adjuvant that polarizes the adaptive T cell responses to Th1/Tfh1 in both mouse and human DCs. Notably, the rASP-1-adjuvanted IIV3 vaccine elicited protection of mice from a lethal H1N1 infection that is also dependent on the TLR4-TRIF axis and IFNAR signaling pathway, as well as on its ability to induce anti-IIV3 antibody production.

The Role of Type I Interferon Signaling in Regulating Cytokine Production and Cell Survival in Bone Marrow-Derived Macrophages.

During viral infections, cells produce type I interferons (IFNs), which are detected by the IFNα/ß receptor (IFNAR). To survive in hosts, viruses have strategies to downregulate IFN-mediated signaling. We hypothesized that macrophages, which are among the first myeloid cells to respond to viral infections, would produce a different cytokine profile if responding to ligation of pattern recognition receptors (PRRs) while IFNAR-mediated signaling was compromised. Specifically, IFNAR-mediated regulation of interleukin (IL)-1α, IL-6, IL-12, and tumor necrosis factor-α was studied in cultured murine bone marrow-derived macrophages. Since viruses like vesicular stomatitis virus can ligate PRRs such as Toll-like receptor (TLR)4 and 7, macrophages were stimulated with the TLR4 and TLR7 agonists lipopolysaccharide (LPS) or imiquimod, respectively, with or without antibody-mediated IFNAR-blockade. Cytokines and viability were assessed for 3 days poststimulation. Blocking IFNARs acutely exacerbated cytokine production by macrophages and aided their survival when they were treated with LPS. In contrast, cytokine concentrations were unaffected or slightly reduced by IFNAR blockade, but macrophages died at a greater rate when imiquimod was the stimulus. This demonstrated a differential role of IFNAR signaling in regulating PRR-induced cytokines. This suggests potential mechanisms whereby macrophages responding to viruses that inhibit type I IFN responses might contribute to excessive inflammation in some cases and inappropriately low-magnitude responses in others. This also provides a well-defined cell-based model for further dissecting the role of type I IFN signaling in macrophages responding to viral and other infections.

Disruption of type I interferon signaling causes sexually dimorphic dysregulation of anti-viral cytokines.

Type I interferons (IFNs) play a crucial role in the establishment of an antiviral state via signaling through their cognate type I IFN receptor (IFNAR). In this study, a replication-competent but highly attenuated strain of VSV (rVSVΔm51) carrying a deletion at position 51 of the matrix protein to remove suppression of anti-viral type I IFN responses was used to explore the effect of disrupted IFNAR signaling on inflammatory cytokine responses in mice. The kinetic responses of interleukin-6, tumor necrosis factor-α and interleukin-12 were evaluated in virus-infected male and female mice with or without concomitant antibody-mediated IFNAR-blockade. Unlike controls, both male and female IFNAR-blocked mice showed signs of sickness by 24-hours post-infection. Female IFNAR-blocked mice experienced greater morbidity as demonstrated by a significant decrease in body temperature. This was not the case for males. In addition, females with IFNAR-blockade mounted prolonged and exaggerated systemic inflammatory cytokine responses to rVSVΔm51. This was in stark contrast to controls with intact IFNAR signaling and males with IFNAR-blockade; they were able to down-regulate virus-induced inflammatory cytokine responses by 24-hours post-infection. Exaggerated cytokine responses in females with impaired IFNAR signaling was associated with more effective control of viremia than their male counterparts. However, the trade-off was greater immune-mediated morbidity. The results of this study demonstrated a role for IFNAR signaling in the down-regulation of antiviral cytokine responses, which was strongly influenced by sex. Our findings suggested that the potential to mount toxic cytokine responses to a virus with concomitant disruption of IFNAR signaling was heavily biased towards females.

Partial Protection From Lupus-Like Disease by B-Cell Specific Type I Interferon Receptor Deficiency.

Systemic lupus erythematosus (SLE) is an autoimmune disease that can present with many different permutations of symptom presentation. A large subset of SLE patients have been shown to present with elevated interferon stimulated gene (ISG) expression, and Type I IFNs (IFNαß) have been shown to drive disease in murine models through global IFNα Receptor (IFNAR) knockouts. However, the disease contribution of distinct immune cell subsets in response to constitutively increased levels of IFNαß is not fully understood. We utilized a B-cell specific IFNAR knockout (BΔIFNAR) on the B6.Nba2 spontaneous-lupus background to determine the contribution of IFNαß stimulated B cells in disease. We found that IFNαß signaling in B cells is driving increased splenomegaly, increased populations of activated B cells, and increased populations of germinal center (GC) B cells, memory B cells, and plasma blasts/cells, but did not affect the development of glomerulonephritis and immune-complex deposition. IFNAR expression by B cells also drove production of anti-chromatin IgG, and anti-dsDNA and -nRNP IgG and IgG2C auto-antibody levels, as well as increased Bcl2 expression, affecting GC B cell survival in B6.Nba2 mice.

Type-Specific Crosstalk Modulates Interferon Signaling in Intestinal Epithelial Cells.

Intestinal epithelial cells (IECs) are the primary target of enteric viruses. Their infection by viruses leads to the upregulation of both type I and type III interferons (IFNs). These IFNs then act in an autocrine and paracrine manner to protect IECs from viral propagation. To date, whether both IFNs use similar signaling pathways and whether these 2 cytokines can act synergistically to protect against viral infection remain unclear. Using human IECs depleted of either the type I or type III IFN receptor, we found that both signal transduction pathways are interconnected and influence each other at the level of interferon-stimulated gene (ISG) expression and efficiency of antiviral protection. Precisely, in human IECs, the presence of a functional type III IFN receptor negatively regulates type I IFN signaling and activity, whereas the presence of type I IFN receptor positively reinforces type III IFN signaling and function. We propose that this complex crosstalk allows for a preferential type III IFN-mediated protection of human intestinal cells.

Decreased Methylation of IFNAR Gene Promoter from Peripheral Blood Mononuclear Cells Is Associated with Oxidative Stress in Chronic Hepatitis B.

Type I interferons (IFNs) play an antiviral effect by binding to type I interferon receptor (IFNAR). Oxidative stress might induce the gene promoter methylation. The purpose of our study was to evaluate the potential relationship between the methylation of IFNAR promoter and the status of oxidative stress in chronic hepatitis B (CHB). The methylation level of the IFNAR promoter in patients with CHB and healthy controls (HCs) was determined by methylation-specific polymerase chain reaction (MS-PCR). The quantitative real-time PCR (RT-qPCR) was used to evaluate the IFNAR mRNA status in peripheral blood mononuclear cells from CHB and HCs. Level of plasma-soluble IFNAR and oxidative stress parameters, including malondialdehyde (MDA) and glutathione (GSH) were determined by enzyme-linked immunosorbent assay (ELISA). The frequency of IFNAR promoter methylation in CHB patients was significantly lower than that of HCs. The IFNAR mRNA level of patients with CHB was higher than HCs. MDA level was higher in CHB patients, whereas GSH level was lower in CHB patients than that of HCs. In CHB patients, plasma MDA level was significantly higher with IFNAR promoter methylation than unmethylation, and soluble IFNAR in the circulation of methylated patients with CHB was decreased than unmethylated patients with CHB. Our results indicated that the IFNAR promoter methylation might have a potential relationship with the status of oxidative stress.

Characterization and signaling pathway analysis of interferon-kappa in bovine.

A bovine interferon-kappa (BoIFN-κ) gene was amplified, which encodes a protein of 215 amino acids sharing 63% identity with human IFN-κ. BoIFN-κ was demonstrated to have antiviral and antiproliferative activities. Moreover, BoIFN-κ was shown to be highly sensitive to trypsin, however, it remained stable despite changes in pH and temperature. Result showed that BoIFN-κ can bind with bovine type I IFN receptors, and the antiviral activity can be blocked by antibodies against type I IFN receptors or BoIFN-κ. Additionally, BoIFN-κ can induce the transcription of Mx1, ISG15 and ISG56 gene, as well as the expression of Mx1 protein. The NF-κB, ISRE, and BoIFN-ß promoter can all be activated by BoIFN-κ. This study revealed that BoIFN-κ exhibits the typical characteristics of type I IFNs and exerts antiviral activity via activation of the JAK-STAT signaling pathway. Overall, these findings will enrich the current knowledge about IFN-κ and facilitate further research on the role of type I IFN in antiviral defense responses in bovine.

Receptor dimer stabilization by hierarchical plasma membrane microcompartments regulates cytokine signaling.

The interaction dynamics of signaling complexes is emerging as a key determinant that regulates the specificity of cellular responses. We present a combined experimental and computational study that quantifies the consequences of plasma membrane microcompartmentalization for the dynamics of type I interferon receptor complexes. By using long-term dual-color quantum dot (QD) tracking, we found that the lifetime of individual ligand-induced receptor heterodimers depends on the integrity of the membrane skeleton (MSK), which also proved important for efficient downstream signaling. By pair correlation tracking and localization microscopy as well as by fast QD tracking, we identified a secondary confinement within ~300-nm-sized zones. A quantitative spatial stochastic diffusion-reaction model, entirely parameterized on the basis of experimental data, predicts that transient receptor confinement by the MSK meshwork allows for rapid reassociation of dissociated receptor dimers. Moreover, the experimentally observed apparent stabilization of receptor dimers in the plasma membrane was reproduced by simulations of a refined, hierarchical compartment model. Our simulations further revealed that the two-dimensional association rate constant is a key parameter for controlling the extent of MSK-mediated stabilization of protein complexes, thus ensuring the specificity of this effect. Together, experimental evidence and simulations support the hypothesis that passive receptor confinement by MSK-based microcompartmentalization promotes maintenance of signaling complexes in the plasma membrane.

Luteolin sensitizes the antiproliferative effect of interferon α/ß by activation of Janus kinase/signal transducer and activator of transcription pathway signaling through protein kinase A-mediated inhibition of protein tyrosine phosphatase SHP-2 in cancer cells.

New negative regulators of interferon (IFN) signaling, preferably with tissue specificity, are needed to develop therapeutic means to enhance the efficacy of type I IFNs (IFN-α/ß) and reduce their side effects. We conducted cell-based screening for IFN signaling enhancer and discovered that luteolin, a natural flavonoid, sensitized the antiproliferative effect of IFN-α in hepatoma HepG2 cells and cervical carcinoma HeLa cells. Luteolin promoted IFN-ß-induced Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway activation by enhancing the phosphorylation of Jak1, Tyk2, and STAT1/2, thereby promoting STAT1 accumulation in the nucleus and endogenous IFN-α-regulated gene expression. Of interest, inhibition of phosphodiesterase (PDE) abolished the effect of IFN-ß and luteolin on STAT1 phosphorylation. Luteolin also increased the cAMP-degrading activity of PDE bound with type I interferon receptor 2 (IFNAR2) and decreased the intracellular cAMP level, indicating that luteolin may act on the JAK/STAT pathway via PDE. Protein kinase A (PKA) was found to negatively regulate IFN-ß-induced JAK/STAT signaling, and its inhibitory effect was counteracted by luteolin. Pull-down and immunoprecipitation assays revealed that type II PKA interacted with IFNAR2 via the receptor for activated C-kinase 1 (RACK-1), and such interaction was inhibited by luteolin. Src homology domain 2 containing tyrosine phosphatase-2 (SHP-2) was further found to mediate the inhibitory effect of PKA on the JAK/STAT pathway. These data suggest that PKA/PDE-mediated cAMP signaling, integrated by RACK-1 to IFNAR2, may negatively regulate IFN signaling through SHP-2. Inhibition of this signaling may provide a new way to sensitize the efficacy of IFN-α/ß.

A report from the American college of rheumatology/association of rheumatology health professionals (ACR/ARHP) - 2012 annual meeting (November 9-14, 2012 - Washington, D.C., USA).

The annual meeting of the American College of Rheumatology (ACR), jointly held with the Association of Rheumatology Health Professionals (ARHP), brought together attendees focused on all aspects of rheumatology, including researchers looking into treatment options and various services around the care of rheumatologic conditions. As well as networking opportunities at the meeting, there were a wide range of symposia and posters available covering various conditions and levels of research. There were also educational and meet-the-professor sessions. This report will cover a selection of interesting talks from poster and oral sessions on the latest preclinical and clinical research.