In this study, we described the structure, mRNA tissue distribution and regulation of an IRF-3 gene from turbot, Scophthalmus maximus (SmIRF-3). The gene sequence of SmIRF-3 is 6077 bp long, composed of 11 exons and 10 introns similar to known IRF-3 genes of fish, and encodes a peptide of 466 amino acids. The deduced protein sequence shares the highest identity of 56.0–81.2% with fish IRF-3 and possesses a DNA-binding domain (DBD), an IRF association domain (IAD) and a serine-rich domain (SRD) known to be important for the functions of IRF-3 in vertebrates. Phylogenetic analysis grouped SmIRF-3 with other IRF3s of vertebrates. SmIRF-3 transcripts were detectable in limited tissue types of healthy fish, with higher expression observed in head, kidney, spleen and kidney,. The SmIRF-3 was transcriptionally up-regulated by turbot reddish body iridovirus (TRBIV) and polyinosinic:polycytidylic acid (poly I:C) in the head kidney, spleen and gills, with showing a two wave induced expression during a 7-day time course in all cases. The highest inducibility and the likely earliest increase of SmIRF-3 expression were observed in the spleen, and poly I:C was a stronger inducer. In addition, the maximal expression level of SmIRF-3 arose prior to that of the Mx in all the cases.