Functionally Antagonistic Transcription Factors IRF1 and IRF2 Regulate the Transcription of the Dopamine Receptor D2 Gene Associated with Aggressive Behavior of Weaned Pigs

Aggressive behavior has negative effects on animal welfare and growth performance in pigs. The dopamine receptor D2 (DRD2) has a critical neuromodulator role in the dopamine signal pathway within the brain to control behavior. A functional single-nucleotide polymorphism (SNP), rs1110730503, in the promoter region of the porcine DRD2 gene was identified, which affects aggressive behavior in pigs. A chromatin immunoprecipitation (ChIP) assay was used to identify the interactions between interferon regulatory factor 1 (IRF1) and IRF2 with the DRD2 gene. The overexpression or knockdown of these two transcription factors in porcine kidney-15 (PK15) and porcine neuronal cells (PNCs) indicate that the binding of IRF1 to DRD2 promotes the transcription of the DRD2 gene, but the binding of IRF2 to the DRD2 gene inhibits its transcription. Furthermore, IRF1 and IRF2 are functionally antagonistic to each other. The downregulation of DRD2 or upregulation of IRF2 increased the apoptosis rate of porcine neuroglial cells. Taken together, we found that transcriptional factors IRF1 and IRF2 have vital roles in regulating the transcription of the DRD2 gene, and rs1110730503 (−915A/T) is a functional SNP that influences IRF2 binding to the promoter of the DRD2 gene. These findings will provide further insight towards controlling aggressive behavior in pigs.

Chromatin bound by survivin regulates the glycolytic switch in interferon-γ producing CD4+ T cells

Upon activation, CD4+ T cells adapt metabolically to fulfill their effector function in autoimmunity. Here we show that nuclear survivin is essential for transcriptional regulation of glucose utilization. We found that the glycolytic switch in interferon (IFN) g–producing CD4+ cells is dependent on a complex of survivin with interferon regulatory factor 1 (IRF1), and Smad3 and was reversed by survivin inhibition. Transcriptome analysis of CD4+ cells and sequencing of survivin-bound chromatin identified a hub of metabolism regulating genes whose transcription depended on survivin. Direct binding of survivin to IRF1 and SMAD3 promoted IRF1-mediated transcription, repressed SMAD3 activity, and lowered PFKFB3 production. Inhibiting survivin upregulated PFKFB3, restored glycolysis, and reduced glucose uptake, improving control over IFNg-dependent T-cell functions. Thus, IRF1-survivin-SMAD3 interactions are important for metabolic adaptation of CD4+ cells and provide an attractive strategy to counteract IFNg-dependent inflammation.

Stem cell specific interferon stimulated gene expression is regulated by the formative pluripotency network through IRF1

Stem cells intrinsically express a subset of genes which are normally associated with interferon stimulation, and thus the innate immunity response. Expression of these interferon stimulated genes (ISG) in stem cells is independent of external stimuli such as viral infection. Here we show that the interferon regulatory factor 1, Irf1, is directly controlled by the murine formative pluripotency gene regulatory network and therefore upregulated in the transition from naive to formative pluripotency. IRF1 in turn binds to regulatory regions of a conserved set of ISGs and is required for their faithful expression in formative pluripotent cells. IRF1 also binds to an enhancer of the formative pluripotency transcription factor Oct6 and is partially required for upregulation of Oct6. IRF1 therefore acts as a link between the formative pluripotency network and the regulation of innate immunity genes in formative pluripotency.

Ataxin-10 Inhibits TNF-α-Induced Endothelial Inflammation via Suppressing Interferon Regulatory Factor-1

Endothelial inflammation is a crucial event in the initiation of atherosclerosis. Here, we identify Ataxin-10 protein as a novel negative modulator of endothelial activation by suppressing IRF-1 transcription activity. The protein level of Ataxin-10 is relatively higher in human vascular endothelial cells, which can be significantly suppressed by TNF-α in both HUVECs and HLMECs. Overexpression of Ataxin-10 markedly inhibited the mRNA expressions of VCAM-1 and several cytokines including MCP-1, CXCL-1, CCL-5, and TNF-α; thus, it can also suppress monocyte adhesion to endothelial cells. Accordingly, Ataxin-10 silencing promoted endothelial inflammation. However, Ataxin-10 did not affect the MAPK/NF-κB signaling pathway stimulated by TNF-α in HUVECs. Using the yeast two-hybrid assay, we found that Ataxin-10 can directly bind to interferon regulatory factor-1 (IRF-1). Upon TNF-α stimulation, Ataxin-10 promoted the cytoplasmic localization of IRF-1, which inhibited the transcription of VCAM-1. Moreover, knockdown of IRF-1 can eliminate the effect of Ataxin-10 on the expression of VCAM-1 in HUVECs induced by TNF-α. Taken together, these results indicate that Ataxin-10 inhibits endothelial cell activation and may serve as a promising therapeutic target for some vascular inflammatory-related diseases such as atherosclerosis.

TNF-Mediated Inhibition of Classical Swine Fever Virus Replication Is IRF1-, NF-κB- and JAK/STAT Signaling-Dependent

The sera from pigs infected with virulent classical swine fever virus (CSFV) contain substantial amounts of tumor necrosis factor (TNF), a prototype proinflammatory cytokine with pleiotropic activities. TNF limits the replication of CSFV in cell culture. In order to investigate the signaling involved in the antiviral activity of TNF, we employed small-molecule inhibitors to interfere specifically with JAK/STAT and NF-κB signaling pathways in near-to-primary endothelial PEDSV.15 cells. In addition, we knocked out selected factors of the interferon (IFN) induction and signaling pathways using CRISPR/Cas9. We found that the anti-CSFV effect of TNF was sensitive to JAK/STAT inhibitors, suggesting that TNF induces IFN signaling. Accordingly, we observed that the antiviral effect of TNF was dependent on intact type I IFN signaling as PEDSV.15 cells with the disrupted type I IFN receptor lost their capacity to limit the replication of CSFV after TNF treatment. Consequently, we examined whether TNF activates the type I IFN induction pathway. With genetically modified PEDSV.15 cells deficient in functional interferon regulatory factor 1 or 3 (IRF1 or IRF3), we observed that the anti-CSFV activity exhibited by TNF was dependent on IRF1, whereas IRF3 was dispensable. This was distinct from the lipopolysaccharide (LPS)-driven antiviral effect that relied on both IRF1 and IRF3. In agreement with the requirement of IRF1 to induce TNF- and LPS-mediated antiviral effects, intact IRF1 was also essential for TNF- and LPS-mediated induction of IFN-β mRNA, while the activation of NF-κB was not dependent on IRF1. Nevertheless, NF-κB activation was essential for the TNF-mediated antiviral effect. Finally, we observed that CSFV failed to counteract the TNF-mediated induction of the IFN-β mRNA in PEDSV.15 cells, suggesting that CSFV does not interfere with IRF1-dependent signaling. In summary, we report that the proinflammatory cytokine TNF limits the replication of CSFV in PEDSV.15 cells by specific induction of an IRF1-dependent antiviral type I IFN response.

Interferon Regulatory Factor-1 Regulates Cisplatin-Induced Apoptosis and Autophagy in A549 Lung Cancer Cells

Purpose: This study aimed to investigate the expression and function of interferon regulatory factor-1 (IRF-1) in non-small cell lung cancer (NSCLC). Methods: IRF-1 expression and its prognostic value were investigated through bioinformatic analysis. The protein expression levels of IRF-1, cleaved caspase-3, and LC3-I/II were analyzed by western blotting. A lentiviral vector was used to overexpress or knockdown IRF-1 in vitro. Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) were analyzed by JC-1 and DCFH-DA staining, respectively. ATP, SOD, MDA, cell viability, LDH release, and caspase-3 activity were evaluated using commercial kits. Results: Compared to the levels in normal tissues, IRF-1 expression was significantly lower in lung cancer tissues and was a prognostic factor for NSCLC. Cisplatin treatment induced IRF-1 activation, ROS production, ATP depletion, SOD consumption, and MDA accumulation in A549 lung cancer cells. IRF-1 overexpression promoted mitochondrial depolarization, oxidative stress, and apoptotic cell death and inhibited autophagy in A549 cells, and these effects could be reversed by IRF-1 knockdown. Conclusion: These data suggest that IRF-1 regulates apoptosis, autophagy and oxidative stress, which might be served as a potential target for increasing chemotherapy sensitivity of lung cancer.

T cell-intrinsic Interferon Regulatory Factor-1 expression suppresses differentiation of CD4 + T cell populations that support chronic gammaherpesvirus infection.

Gammaherpesviruses are ubiquitous pathogens that establish life-long infection and are associated with B cell lymphomas. To establish chronic infection, these viruses usurp B cell differentiation and drive a robust germinal center response to expand the latent viral reservoir and gain access to memory B cells. Germinal center B cells, while important for the establishment of latent infection, are also thought to be the target of viral transformation. The host and viral factors that impact the gammaherpesvirus-driven germinal center response are not clearly defined. We showed that global expression of the antiviral and tumor-suppressor interferon regulatory factor 1 (IRF-1) selectively attenuates the murine gammaherpesvirus 68 (MHV68)-driven germinal center response and restricts expansion of the latent viral reservoir. In this study we found that T cell intrinsic IRF-1 expression recapitulates some aspects of antiviral state imposed by IRF-1 during chronic MHV68 infection, including attenuation of the germinal center response and viral latency in the spleen. We also discovered that global and T cell-intrinsic IRF-1 deficiency leads to unhindered rise of IL-17A-expressing and follicular helper T cell populations, two CD4 + T cell subsets that support chronic MHV68 infection. Thus, this study unveils a novel aspect of antiviral activity of IRF-1 by demonstrating IRF-1-mediated suppression of specific CD4 + T cell subsets that support chronic gammaherpesvirus infection. Importance Gammaherpesviruses infect over 95% of the adult population, last the lifetime of the host, and are associated with multiple cancers. These viruses usurp the germinal center response to establish lifelong infection in memory B cells. This manipulation of B cell differentiation by the virus is thought to contribute to lymphomagenesis, though exactly how the virus precipitates malignant transformation in vivo is unclear. IRF-1, a host transcription factor and a known tumor suppressor, restricts the MHV68-driven germinal center response in a B cell-extrinsic manner. We found that T cell intrinsic IRF-1 expression attenuates the MHV68-driven germinal center response by restricting the CD4 + T follicular helper population. Further, our study identified IRF-1 as a novel negative regulator of IL-17-driven immune responses, highlighting the multifaceted role of IRF-1 in gammaherpesvirus infection.