Characterization of DNA Binding and Nuclear Retention Identifies Zebrafish IRF11 as a Positive Regulator of IFN Antiviral Response

In mammals, transcription factors of IFN-regulatory factors (IRFs) family translate viral recognition into IFN antiviral responses through translocating to nucleus and subsequently binding to the promoters of IFN and IFN-stimulated genes (ISGs). In addition to IRF1-9 conserved across vertebrates and IRF10 in teleost fish and bird, teleost fish has another novel member, IRF11; however, little is known about its role in IFN response. In this study, we provide evidence that IRF11 is present only in Osteichthyes (bony fish) but lost in tetrapods and subsequently characterize the stimulatory potential of zebrafish IRF11 to IFN antiviral response relevant to its subcellular localization and promoter binding. Overexpression of zebrafish IRF11 restricts virus replication through induction of IFN and ISGs. Zebrafish IRF11 is constitutively localized to nucleus, which is driven by a tripartite NLS motif, consisting of three interdependent basic clusters, two in DNA binding domain (DBD) and one in the region immediately C-terminal to DBD. Nuclear IRF11 binds to the IRF-binding element/IFN-stimulated response element motifs of zebrafish IFN promoters depending on the two conserved amino acids (K78, R82) within DBD helix a3. K78 and R82 also benefit zebrafish IRF11 nuclear import as two key residues positioned at the first basic cluster of the tripartite NLS motif. Such features enable zebrafish IRF11 to function as a positive transcription factor for fish IFN antiviral response. Our results identify a unique tripartite NLS motif that integrates DNA-binding activity and nuclear import ability, allowing zebrafish IRF11 to initiate IFN and ISG expression.