Molecular characterization and expression analysis of interferon-gamma-inducible lysosomal thiol reductase (GILT) gene from pearl oyster Pinctada fucata

Abstract

Interferon-gamma-inducible lysosomal thiol reductase (GILT) is an important thiol reductase, involved in class, MHC-restricted antigen processing by catalyzing disulfide bond reduction in mammals. Herein, we describe the identification and characterization of pearl oyster Pinctada fucata GILT (designated as poGILT). The poGILT cDNA was 1273bp long and consisted of a 5'-untranslated region (UTR) of 24bp, a 3'-UTR of 484bp with two cytokine RNA instability motifs (ATTTA), and an open reading frame (ORF) of 765bp encoding a polypeptide of 254 amino acids with an estimated molecular mass of 28.9kDa and a theoretical isoelectric point of 7.4. The N-terminus of the poGILT was found to have a putative signal peptide with a cleavage site amino acid position at 19-20. SMART analysis showed that the poGILT contained a GILT active-site C(69)PDC(72) motif and a GILT signature motif C(115)QHGKEECIGNLIETC(130). Homology analysis of the deduced amino acid sequence of the poGILT with other known GILT sequences by MatGAT software revealed that the poGILT shared 42.9-67.3% similarity and 22.9-49.8% identity to the other known GILT sequences. The expression level of poGILT mRNA was higher in digestive gland, moderate in adductor muscle, gills, gonad, intestine and mantle, and lower in hemocytes. The poGILT mRNA expression was significantly up-regulated in gill and digestive gland after LPS or V. alginolyticus stimulation, respectively. These results suggested that the poGILT was a constitutively expressed acute-phase protein, the expression of which can be enhanced after LPS or V. algrinolyticus stimulation, perhaps involved in the innate immune response of pearl oyster.