Research ArticleImmunology

Unphosphorylated ISGF3 drives constitutive expression of interferon-stimulated genes to protect against viral infections

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Sci. Signal.  25 Apr 2017:
Vol. 10, Issue 476, eaah4248
DOI: 10.1126/scisignal.aah4248

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An interferon-independent antiviral defense

Virally infected cells produce type I interferons (IFNs), which stimulate the phosphorylation and activation of the STAT1 and STAT2 transcription factors. When combined with the transcriptional regulator IRF9, phosphorylated STAT1 and STAT2 form the ISGF3 complex, which drives the expression of IFN-stimulated genes (ISGs) that are important for antiviral immunity. Wang et al. report the formation of an alternative form of ISGF3, U-ISGF3, which drove the expression of ISGs and protected cells from viral infection in the absence of detection of detectable IFN or IFN signaling. In addition to IRF9, U-ISGF3 contained unphosphorylated STAT1 and STAT2. Together, these data suggest that U-ISGF3 drives constitutive expression of ISGs as part of an IFN-independent, antiviral immune response.


Interferon (IFN)–stimulated genes (ISGs) are antiviral effectors that are induced by IFNs through the formation of a tripartite transcription factor ISGF3, which is composed of IRF9 and phosphorylated forms of STAT1 and STAT2. However, we found that IFN-independent ISG expression was detectable in immortalized cell lines, primary intestinal and liver organoids, and liver tissues. The constitutive expression of ISGs was mediated by the unphosphorylated ISGF3 (U-ISGF3) complex, consisting of IRF9 together with unphosphorylated STAT1 and STAT2. Under homeostatic conditions, STAT1, STAT2, and IRF9 were found in the nucleus. Analysis of a chromatin immunoprecipitation sequencing data set revealed that STAT1 specifically bound to the promoters of ISGs even in the absence of IFNs. Knockdown of STAT1, STAT2, or IRF9 by RNA interference led to the decreased expression of various ISGs in Huh7.5 human liver cells, which was confirmed in mouse embryonic fibroblasts (MEFs) from STAT1−/−, STAT2−/−, or IRF9−/− mice. Furthermore, decreased ISG expression was accompanied by increased replication of hepatitis C virus and hepatitis E virus. Conversely, simultaneous overexpression of all ISGF3 components, but not any single factor, induced the expression of ISGs and inhibited viral replication; however, no phosphorylated STAT1 and STAT2 were detected. A phosphorylation-deficient STAT1 mutant was comparable to the wild-type protein in mediating the IFN-independent expression of ISGs and antiviral activity, suggesting that ISGF3 works in a phosphorylation-independent manner. These data suggest that the U-ISGF3 complex is both necessary and sufficient for constitutive ISG expression and antiviral immunity under homeostatic conditions.

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