Characterization,Function And Regulation Of DIGIRR Gene In Miiuy Croaker

Members of the TLR-IL-1R(Toll-like receptor-interleukin 1 receptor signaling,TIR)superfamily play an important role in the mediated immune responses,and their family members share a common feature: all of them contain a conserved Toll-IL-1R(TIR)domain,which is closely related to the signal transduction process.DIGIRR(Double-Ig-IL-1R Related Molecule),one of the members of the TIL-IL-1R receptor superfamily,plays an important role in the immune system and is also a negative regulator of IL-1 signaling pathway.In this study,the cDNA sequence of the miiuy croaker DIGIRR(mmi-DIGIRR)gene was identified and characterized.The mmi-DIGIRR cDNA is 1837 bp in length and includes a 211 bp 5’-untranslated region(UTR),a 1575 bp open reading frame(ORF)and a 51 bp 3’UTR.The mmi-DIGIRR gene encodes a polypeptide of 524 amino acids with a molecular weight of 59.43 KDa and a PI of 6.16.Mmi-DIGIRR has the common characteristics of IL-IR receptor family.Its protein domain is mainly composed of intracellular conserved TIR domain and two extracellular immune protein(immunoglobulin,Ig)domain.In addition,the extracellular domain of mmi-DIGIRR has a signal peptide sequence of 16 amino acids and a transmembrane region of 33 amino acids.Phylogenetic tree analysis showed that the DIGIRR gene of mackerel was homologous to SIGIRR and might come from the same ancestor molecule.The comparison analysis of multiple sequences and key amino acids of signal transduction showed that mmi-DIGIRR had the same conserved sequence in the intracellular region as the SIGIRR/DIGIRR of other vertebrates,and its evolution was relatively conservative.Only one of the two key amino acid sites(Ser and Arg-Tyr)of the TIR family signal transduction was mutated in mmi-DIGIRR(Arg-Tyr mutated to Arg-Leu),while in mammalian SIGIRR,both amino acid sites were mutated(Ser mutated to Cys,Arg-Tyr mutated to Gln-Leu).qPCR analysis showed that mmi-DIGIRR was significantly expressed in liver,skin and muscle,but lower in heart.Functional and regulatory studies showed that overexpression of mmi-DIGIRR could significantly inhibit NF-κB activation induced by LPS.This study comprehensively analyzed the evolution and function of the mmi-DIGIRR,which provided a theoretical basis for the future study of fish DIGIRR.