Functional domains and amino acid residues of Japanese eel IRF1, AjIRF1, regulate its nuclear import and IFN/Mx promoter activation.

Interferon regulatory factors (IRFs) are a family of transcriptional factors capable of regulating the expression of distinct subsets of interferon (IFN)-stimulated genes by binding to their promoters. IRF1 was the first member identified for its ability to regulate the IFNβ gene and has now been revealed to exhibit remarkable functional diversity in the regulation of different cellular responses. In the present study, the IRF1 gene was identified and characterized in Japanese eel, Anguilla japonica (AjIRF1). The open reading frame of AjIRF1 was 804 bp in length, encoding a protein of 267 amino acids (aa) that encompasses a conserved N-terminal DNA binding domain (DBD). Sequence alignment shows the presence of six highly conserved tryptophan (W) residues in the DBD of IRF1, IRF2 and IRF11, while other IRF members have only five tryptophans. Expression analysis showed that AjIRF1 was significantly upregulated in all tested organs/tissues in response to Poly I:C stimulation or Edwardsiella tarda infection. Furthermore, the functional activity of AjIRF1 was confirmed in driving the transcription of AjIFN promoters, which depends on the highly conserved residues within DBD. Subcellular distribution analysis revealed that AjIRF1 was localized exclusively in the nucleus, which is cooperatively regulated by a bipartite NLS embedded within the DBD and a monopartite NLS located immediately downstream of the DBD. Taken together, this study presents the expression profile of AjIRF1 and defines the functional motifs required for its nuclear import and its role in activating IFN promoters, thus providing helpful information for further research on the regulatory mechanisms of teleost IRF1.