Characteristics And Inductive Expression Analysis Of IRF-8in Turbot, Scophthalmus Maximus, IRF-2and IRF-8in Japanese Flounder, Paralichthys Olivaceus

Turbot (Scophthalmus maximus) is an important fish cultured widely inNorth China’s coastal areas, but highly vulnerable to the infection of turbotreddish body irrdovirus (TRBIV). As the same, flounder farming industry in Chinahas been suffered a growing damage from various viral epidemics includinglymphocystis disease caused by the infection of lymphocystis disease virus Chinastrain (LCDV-cn) during the past decades. Interferon regulatory factor (IRF) familytranscription factors play essential roles in various life processes in vertebrates, suchas early immune responses to pathogens, antiviral defense, immuno-regulation andgrowth control. An understanding of the IFN system in turbot and flounder will behelpful to the control of viral diseases of this commercially important species. So far,IRF-1, IRF-3, IRF-5, IRF-7and IRF-10have been reported in flounder, but IRF-2hasn’t been reported. In addition, little data are available presently about IRF-8in fish.The objects of this study are turbot and Japanese flounder, we used thehomologous PCR and Rapid Amplification of cDNA Ends（RACE）method to isolatethe cDNA and genomic sequences of SmIRF-8, PoIRF-2and PoIRF-8respectively.The full-length PoIRF-2cDNA (GenBank accession no. JF312911) is1819bp long,with an open reading frame (ORF) of993bp that encodes331amino acid residues.Similar to IRF-2in other vertebrates, PoIRF-2contains three conserved domains: anN-terminal DBD domain, a C-terminal IRF association domain2(IAD2) and aC-terminal transcriptional repression domain (TRD). The cDNA sequence of SmIRF8(GenBank accession number: JQ219663) is2317bp long, PoIRF-8full-length cDNA(GenBank accession number: FJ828965) is1854bp in length, they both contain a1263bp coding region encoding a putative protein of420amino acid residues. Besides, they possess an N-terminal DBD domain, a C-terminal IAD domain and aNLS domain. The SmIRF8gene sequence (GenBank accession number: JQ219665) is4363bp long, the PoIRF-8gene has a length of approximately5.7kb (GenBankaceesion no. FJ556995) which both consisting of9exons and8introns.The tissue distribution of SmIRF8, PoIRF-2and PoIRF-8mRNA was examinedby qPCR in twelve tissue types including brain, gills, stomach, intestine, heart, headkidney, kidney, liver, spleen, gonad, muscle and skin of healthy turbots and flounders,respectively. The constitutive expression of SmIRF8, PoIRF-2and PoIRF-8wereobserved in all tissues examined, but have a high level in the lymphomyeloid-richtissues, such as the spleen, kidney and head kidney. SmIRF-8was less abundant in theskin, muscle and liver, PoIRF-8was less abundant in the mucule. Thus, it suggeststhat SmIRF8, PoIRF-2and PoIRF-8play a role predominantly in the immune system.In this study, although the SmIRF8was found to be up-regulated by thestimulation of both poly I:C and TRBIV, the induction by poly I:C was earlier and abit stronger and the spleen exhibited the highest and earliest increase. The expressionprofile of PoIRF-2can be up-regulated by the stimulation of both poly I:C and LCDV,and the spleen exhibited the earliest expression peak. The expression profile ofPoIRF-8can be up-regulated by the stimulation of both poly I:C and LCDV during a7-day time course, but the induction by poly I:C was a bit earlier. Upon poly I:Cchallenge, the expression peak of PoIRF-8arose at hour9post-stimulation in the gills,head kidney and muscle and hour3in the spleen, with an approximate3.0-,2.0-,3.0-and4.0-fold increase, respectively. In the case of LCDV challenge, exposure ofJapanese flounder to LCDV up-regulated PoIRF-8expression, with maximum4.0-,3.0-,3.0-and3.0-fold increase observed at hour12in the spleen, head kidney, gillsand muscle, respectively. Collectively, these results provide a possibility that SmIRF8,PoIRF-2and PoIRF-8play a role in antiviral IFN response.Our findings contribute to the reserch of the IFN system JAK-STAT signalpathway in turbot and Japanese flouder, and help to develop effective immunestrateges to protect turbot and Japanese flouder from viral diseases.