Role of Nuclear Factor-κB in the Antiviral Action of Interferon and Interferon-regulated Gene Expression

Abstract

Interferons (IFNs)1 are a family of multifunctional cytokines that block viral infection, inhibit cell proliferation, and modulate cell differentiation. Whereas type I IFNs (IFN-␣, IFN-␤, and IFN-) bind to a common cell-surface receptor, the receptor for type II IFN (IFN-␥) is a distinct entity (1). IFNs transduce signals from the cell surface, resulting in selective gene induction through the activation of JAK tyrosine kinases and STAT transcription factors (1-3). Upon JAK-mediated tyrosine phosphorylation, the STAT proteins (STAT1, STAT2, and STAT3) dimerize and translocate to the nucleus to activate transcription of IFN-stimulated genes (3). IFN also activates the nuclear factor-B (NF-B) transcription factor in a serine/threonine kinase-dependent signaling pathway that protects cells against apoptosis (4, 5).NF-B regulates the expression of genes involved in cell signaling, stress responses, growth, survival, and apoptosis by binding to cis-acting B sites in the promoters and enhancers of these genes. Viruses, cytokines, lipopolysaccharides, and other stimulating agents promote NF-B translocation to the nucleus and DNA binding. Active DNA-binding forms of NF-B are dimeric combinations of Rel proteins (p50, p52, c-Rel, v-Rel, RelA/p65, and RelB). In most cell types, the predominant form of NF-B is the p50⅐p65 heterodimer. NF-B dimers are retained in the cytoplasm of unstimulated cells in an inactive state by the binding of a family of inhibitory IB proteins.Whereas STAT proteins play crucial roles in the transcriptional response to IFN-␣/␤ and in the induction of antiviral activity, the role of NF-B in IFN signaling and action has not been studied extensively. To identify the functional role of NF-B in IFN action, sensitivity to the antiviral effect of IFN was examined in fibroblasts that either had normal NF-B function or had a functional deletion of the IFN-induced NF-B pathway by germ line disruption of the Rel p50 and p65 proteins. Antiviral assays demonstrated that NF-B knockout (KO) fibroblasts were sensitized to the antiviral action of IFN-␤. To determine the relationship between gene regulation by IFN-␤ and antiviral activity, microarray analysis was performed on RNA samples collected from IFN-treated murine fibroblasts. Microarray analysis identified several classical IFN-stimulated genes (ISGs) involved in the antiviral action of IFN. Quantitative real-time PCR demonstrated that, whereas the IFN-induced expression of some ISGs was enhanced in NF-B KO cells relative to mouse wild-type fibroblasts, the IFN-induced expression of other ISGs was lower in NF-B KO cells. Our results demonstrate the distinctive role of NF-B in the regulation of ISGs and in the induction of antiviral activity. Thus, the IFN-activated NF-B pathway not only counterbalances apoptosis, but also regulates the expression of ISGs and the induction of antiviral activity. EXPERIMENTAL PROCEDURESBiological Reagents and Cell Culture-Recombinant Chinese hamster ovary cell-expressed rat IFN-␤ was obtained from Biogen Idec, Inc. (6). The b...

NFκB Negatively Regulates Interferon-induced Gene Expression and Anti-influenza Activity

“…For example, tumor necrosis factor-mediated JNK signaling is enhanced in NFB-KO mice (23,24). Moreover, we previously showed that the IFN-induced expression of some ISGs (Mx1 and Nmi) is negatively regulated by NFB, while the expression of other ISGs, such as Ifit1 and Isg15, is enhanced by NFB (6). In the present study, we show that NFB has opposing effects on different ISGs that map to a single genetic locus.…”

“…1) were induced by IFN␤ in a dose-dependent manner to higher expression levels in NFB-KO cells than in WT cells. Similarly, we showed previously that Mx1 and Nmi were also induced more in NFB-KO cells than in WT cells by IFN in a dose-dependent manner (6).…”

“…Are NFB-dependent-We previously reported that NFB plays a role in regulating ISG expression (6). Subsequently, a functional genomics approach was used to monitor the role of NFB in gene expression changes induced by IFN.…”

“…Control cultures received no IFN but were treated in parallel. Expression values were determined using Affymetrix Microarray Suite (MAS) 5.0 software, and the data were filtered and analyzed using GeneSpring software 7.0 (Silicon Genetics, Inc.) as described previously (6).…”

“…All data analysis was performed using GeneSpring software 7.0 (Silicon Genetics, Inc.). The MAS 5.0 gene expression values for each gene were normalized as described previously (6). Fold induction by IFN was calculated in matched pairs of WT and NFB-KO MEFs, respectively.…”