Identification and characterization of the interferon-γ-inducible lysosomal thiol reductase gene in Chinese soft-shelled turtle,Pelodiscus sinensis

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The reduction of disulfide bonds of exogenous antigens is crucial to the MHC-II class antigen processing and presenting pathway and is catalysed by interferon-γ-inducible lysosomal thiol reductase (GILT). In this study, a reptile GILT gene from Chinese soft-shelled turtle, Pelodiscus sinensis (PsGILT), was identified. The full-length cDNA of PsGILT is 1631 nucleotides (nt), including a 5′-untranslated region (UTR) of 3 nt, a 3′-UTR of 860 nt and an open reading frame (ORF) of 768 nt encoding 255 amino acids (aa). The conserved features in known GILTs, such as signal peptide, CXXC motif, GILT signature sequence, N-glycosylation site and conserved cysteines, were all found in the putative PsGILT protein. Genomic analysis revealed that PsGILT kept the “7 exons and 6 introns” structure of vertebrate GILT genes. PsGILT was expressed in all examined organs/tissues and was mainly expressed in spleen and blood. Increased mRNA expression levels of PsIFN-γ and PsGILT in PBLs were observed after induction with LPS, PolyI:C and recombinant IFN-γ (rIFN-γ). We also tested the reductase activity of rGILT in vitro and found that it could reduce intact human IgG into H chains and L chains. These above results implied that PsGILT may play an important role in resisting bacterial and viral infections, like other vertebrate GILTs.HighlightsWe firstly identified a reptile GILT from Chinese soft-shelled turtle, Pelodiscus sinensis.PsGILT expression was induced in peripheral blood leucocytes by LPS, PolyI:C.Recombinant IFN-γ purified from E. coli (DE3) could induce the mRNA expression level of PsGILT in PBLs.In vitro, recombinant GILT purified from E. coli (DE3) had thiol reductase activity to reduce the intact human IgG into H and L chains.PsGILT might play an important role in immune defense possibly via a conserved functional mechanism throughout vertebrate evolution.

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