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Human interferon-? and interferon-? exhibit low potency and low affinity for cell-surface IFNAR and the poxvirus antagonist B18R.

ABSTRACT: IFN? and IFN? are interferons that induce microbial immunity at mucosal surfaces and in the skin. They are members of the type-I interferon (IFN) family, which consists of 16 different IFNs, that all signal through the common IFNAR1/IFNAR2 receptor complex. Although IFN? and IFN? have unique expression and functional properties, their biophysical properties have not been extensively studied. In this report, we describe the expression, purification, and characterization of recombinant human IFN? and IFN?. In cellular assays, IFN? and IFN? exhibit ?1000-fold lower potency than IFN?2 and IFN?. The reduced potency of IFN? and IFN? are consistent with their weak affinity for the IFNAR2 receptor chain. Despite reduced IFNAR2-binding affinities, IFN? and IFN? exhibit affinities for the IFNAR1 chain that are similar to other IFN subtypes. As observed for cellular IFNAR2 receptor, the poxvirus antagonist, B18R, also exhibits reduced affinity for IFN? and IFN?, relative to the other IFNs. Taken together, our data suggest IFN? and IFN? are specialized IFNs that have evolved to weakly bind to the IFNAR2 chain, which allows innate protection of the mucosa and skin and limits neutralization of IFN? and IFN? biological activities by viral IFN antagonists.

SUBMITTER: Harris BD

PROVIDER: S-EPMC6187621 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Human interferon-ϵ and interferon-κ exhibit low potency and low affinity for cell-surface IFNAR and the poxvirus antagonist B18R.

Harris Bethany D BD Schreiter Jessica J Chevrier Marc M Jordan Jarrat L JL Walter Mark R MR

The Journal of biological chemistry 20180831 41

IFNϵ and IFNκ are interferons that induce microbial immunity at mucosal surfaces and in the skin. They are members of the type-I interferon (IFN) family, which consists of 16 different IFNs, that all signal through the common IFNAR1/IFNAR2 receptor complex. Although IFNϵ and IFNκ have unique expression and functional properties, their biophysical properties have not been extensively studied. In this report, we describe the expression, purification, and characterization of recombinant human IFNϵ ...[more]

PMID: 30171073

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Project description:We previously identified a subset of interferon-stimulated genes (ISGs) upregulated by West Nile virus (WNV) infection in wild-type mouse embryo fibroblasts (MEFs) after viral proteins had inhibited type I interferon (IFN)-mediated JAK-STAT signaling and also in WNV-infected RIG-I-/-, MDA5-/-, STAT1-/-, STAT2-/-, IFNAR-/-, IRF3-/-, IRF7-/-, and IRF3/7-/- MEFs. In this study, ISG upregulation by WNV infection in IFNAR-/- MEFs was confirmed by transcriptome sequencing (RNA-seq). ISG upregulation by WNV infection was inhibited in RIG-I/MDA5-/- MEFs. ISGs were upregulated in IRF1-/- and IRF5-/- MEFs but only minimally upregulated in IRF3/5/7-/- MEFs, suggesting redundant IRF involvement. We previously showed that a single proximal interferon-stimulated response element (ISRE) in the Oas1a and Oas1b promoters bound the ISGF3 complex after type I IFN treatment. In this study, we used wild-type and mutant promoter luciferase reporter constructs to identify critical regions in the Oas1b and Ifit1 promoters for gene activation in infected IFNAR-/- MEFs. Two ISREs were required in both promoters. Mutation of these ISREs in an Ifit1 promoter DNA probe reduced in vitro complex formation with infected nuclear extracts. An NF-κB inhibitor decreased Ifit1 promoter activity in cells and in vitro complex formation. IRF3 and p50 promoter binding was detected by chromatin immunoprecipitation (ChIP) for upregulated ISGs with two proximal ISREs. The data indicate that ISREs function cooperatively to upregulate the expression of some ISGs when type I IFN signaling is absent, with the binding complex consisting of IRF3, IRF5, and/or IRF7 and an NF-κB component(s) as well as other, as-yet-unknown factors. IMPORTANCE Type I IFN signaling in mammalian cells induces formation of the ISGF3 transcription factor complex, which binds to interferon stimulated response elements (ISREs) in the promoters of interferon-stimulated genes (ISGs) in the cell nucleus. Flavivirus proteins counteract type I IFN signaling by preventing either the formation or nuclear localization of ISGF3. A subset of ISRE-regulated ISGs was still induced in West Nile virus (WNV)-infected mouse embryo fibroblasts (MEFs), indicating that cells have an alternative mechanism for activating these ISGs. In this study, cellular components involved in this ISG upregulation mechanism were identified using gene knockout MEFs and ChIP, and critical promoter regions for gene activation were mapped using reporter assays. The data indicate a cooperative function between two ISREs and required binding of IRF3, IRF5, and/or IRF7 and an NF-κB component(s). Moreover, type I IFN signaling-independent ISG activation requires different additional promoter activation regions than type I IFN-dependent activation.

Dataset Information

Human interferon-? and interferon-? exhibit low potency and low affinity for cell-surface IFNAR and the poxvirus antagonist B18R.

Publications

Human interferon-ϵ and interferon-κ exhibit low potency and low affinity for cell-surface IFNAR and the poxvirus antagonist B18R.

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