Caillaud A, Hovanessian AG, Levy DE, Marie IJ.. Regulatory serine residues mediate phosphorylation-dependent and phosphorylation-independent activation of interferon regulatory factor 7. J Biol Chem 280: 17671-17677

ArticleinJournal of Biological Chemistry 280(18):17671-7 · June 2005with13 Reads
DOI: 10.1074/jbc.M411389200 · Source: PubMed
Interferon regulatory factor (IRF)7 is a key transcription factor required for establishment of antiviral resistance. In response to infection, IRF7 is activated by phosphorylation through the action of the non-canonical IkappaB kinases, IkappaB kinase-epsilon and TANK-binding kinase 1. Activation leads to nuclear retention, DNA binding, and derepression of transactivation ability. Clusters of serine residues located in the carboxyl-terminal regulatory domain of IRF7 are putative targets of virus-activated kinases. However, the exact sites of phosphorylation have not yet been established. Here, we report a comprehensive structure-activity examination of potential IRF7 phosphorylation sites through analysis of mutant proteins in which specific serine residues were altered to alanine or aspartate. Phosphorylation patterns of these mutants were analyzed by two-dimensional gel electrophoresis, and their transcriptional activity was monitored by reporter assays. Essential phosphorylation events were mapped to amino acids 437-438 and a redundant set of sites at either amino acids 429-431 or 441. IRF7 recovered from infected cells was heterogeneously phosphorylated at these sites, and greater phosphorylation correlated with increased transactivation. Interestingly, a distinct serine cluster conserved in the related protein IRF3 was also essential for IRF7 activation and distal phosphorylation. However, the essential role of this motif did not appear to be fulfilled by phosphorylation. Rather, these serine residues and an adjacent leucine were required for phosphorylation at distal sites and may determine a conformational element required for function.
    • "In mice, cells that are deficient in both TBK1 and IKKε usually fail to produce type-I IFN in response to viral infection [97, 98]. Following activation of TLRs and RLRs by viral nucleic acids, TBK1 and IKKε assemble with TNF receptor-associated factor 3 (TRAF3) and TANK to phosphorylate IRF3 and IRF7 at multiple serine and threonine residues99100101102. The phosphorylation leads to the dimerization and nuclear transfer of IRFs, and also induces the expression of pro-inflammatory and antiviral genes [103, 104]. "
    [Show abstract] [Hide abstract] ABSTRACT: Aquacultured fish are threatened by many pathogens, often with serious consequences. Vaccination is one of the most effective tools for enhancing host immunity and protecting fish from infections. Vaccination with DNA vaccine is based on the administration of the gene encoding a vaccine antigen. Several effective DNA vaccines that encode viral or bacterial antigenic proteins have already been shown to be effective for cultured fish. This review summarizes current knowledge on fish DNA vaccines, and the mechanism of interaction between the DNA vaccines and host immunity, especially focusing on the enhancement of innate immunity mediated through direct-recognition of DNA vaccine by pattern recognition receptors (PRRs). To date, numerous fish PRR genes have been identified, and the primordial functions of PRRs involved in the innate immune response to viral and bacterial nucleic acids have been increasingly clarified. The evolutionary conservations and divergences in the PRR mechanisms of teleosts and mammals are focused on their molecular features and the recognition of DNA vaccine mediated by TANK binding kinase 1. In addition, the mechanism of type I interferon production in teleosts, which is enhanced after the recognition of cytosolic nucleic acids and current topics on DNA sensing by PRRs are also introduced.
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    • "However, the presence of five tryptophan residues in AaIRF7, a character as observed in mammalian IRF7, may indicate a rather conserved feature in AaIRF7 and the requirement for further functional study on the fourtryptophan DBD in fish IRF7. In addition, AaIRF3 and AaIRF7 have all serines in the C terminus, as observed in their mammalian counterparts, which have been confirmed to be the essential phosphoacceptor sites for dimerization following virus infection and in human counterparts [19,22,32,33], suggesting that the activation of AaIRF3 and AaIRF7 may be similar to human IRF3 and IRF7, respectively [23,34]. In mammals, IRF3 and IRF7 have been experimentally identified as the key regulators of type I IFN gene expression in response to viral infection [1,9e12,18]. "
    [Show abstract] [Hide abstract] ABSTRACT: Interferon regulatory factor (IRF) 3 and IRF7 have been identified as regulators of type I interferon (IFN) gene expression in mammals. In the present study, the two genes were cloned and characterized in the European eel, Anguilla anguilla. The full-length cDNA sequence of IRF3 and IRF7 in the European eel, named as AaIRF3 and AaIRF7 consists of 2879 and 2419 bp respectively. Multiple alignments showed that the two IRFs have a highly conserved DNA binding domain (DBD) in the N terminus, with the characteristic motif containing five tryptophan residues, which is a feature present in their mammalian homologues. But, IRF7 has only four of the five residues in other species of fish. The expression of AaIRF3 and AaIRF7 both displayed an obvious dose-dependent manner following polyinosinic:polycytidylic acid (PolyI:C) challenge. In vivo expression analysis showed that the mRNA level of AaIRF3 and AaIRF7 was significantly up-regulated in response to PolyI:C stimulation in all examined tissues/organs except in muscle, with a lower level of increase observed in response to lipopolysaccharide (LPS) challenge and Edwardsiella tarda infection, indicating that AaIRF3 and AaIRF7 may be more likely involved in antiviral immune response. In addition, some pattern recognition receptors genes related with the production of type I IFNs and those genes in response to type I IFNs were identified in the European eel genome database, indicating a relatively conserved system in the production of type I IFN and its signalling in the European eel.
    Full-text · Article · Apr 2014
    • "Upon KSHV infection, IRF-7 competes with RTA for viral gene promoters to modulate viral transactivation (Wang et al., 2005). The function of IRF-7 can in turn be regulated by post-translational modifications including phosphorylation (Caillaud et al., 2005; Marie et al., 2000), acetylation (Caillaud et al., 2002), ubiquitination (Kawai et al., 2004; Ning et al., 2008; Yu et al., 2005) and SUMOylation (Chang et al., 2009; Kubota et al., 2008). Lysine residues are potential sites for acetylation, ubiquitination and SUMOylation, and may serve as targets to modulate the functions of IRF-7. "
    [Show abstract] [Hide abstract] ABSTRACT: Kaposi's sarcoma-associated herpesvirus (KSHV) infection goes through latent and lytic phases, which are controlled by the viral replication and transcription activator (RTA). Upon KSHV infection, the host responds by suppressing RTA-activated lytic gene expression through interferon regulatory factor 7 (IRF-7), a key regulator of host innate immune response. Lysine residues are potential sites for post-translational modification of IRF-7, and were suggested to be critical for its activity. In this study, we analysed the 15 lysine residues for their effects on IRF-7 function by site-directed mutagenesis. We found that some mutations affect the ability of IRF-7 to activate interferon (IFN)-α1 and IFN-β promoters, to suppress RTA-mediated lytic gene expression and to repress KSHV reactivation and lytic replication. However, other mutations affect only a subset of these four functions. These findings demonstrate that the lysine residues of IRF-7 play important roles in mediating IFN synthesis and modulating viral lytic replication.
    Full-text · Article · Jan 2011
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