A vital ubiquitin-conjugating enzyme CgUbe2g1 participated in regulation of immune response of Pacific oyster Crassostrea gigas.

As an important post-translational protein modification, ubiquitination has been demonstrated to play a vital role in immune response of vertebrates. Ubiquitin (Ub)-conjugating enzyme E2 is the "heart" of ubiquitination, which is responsible for Ub cellular signaling and substrate modification. In the present study, an Ub-conjugating enzyme E2 (designed as CgUbe2g1) was identified from oyster Crassostrea gigas, and its regulation in the immune response against lipopolysaccharide (LPS) stimulation was investigated. CgUbe2g1 encoded a polypeptide of 168 amino acids with the predicted molecular mass of 19.20 kDa and contained conserved catalytic 'Ubc' domains. It shared a higher similarity with the known UBC2G1 type E2s and was closely clustered with the type E2s identified from invertebrates in the phylogenetic assay. The mRNA transcripts of CgUbe2g1 were mainly distributed in hemocyte, mantle, hepatopancreas and male gonad of C. gigas. CgUbe2g1 protein was found to be colocalized with Ub around the nucleus of oyster hemocyte. The recombinant CgUbe2g1 protein (rCgUbe2g1) could activate the ubiquitination in vitro by binding both activated and un-activated Ub. The expressions of inflammation-related factors TNF-α and NF-κB in CgUbe2g1 transfected cells were both significantly up-regulated after LPS stimulation, which were 12.9-fold at 3 h (p < 0.01) and 2.3-fold at 6 h (p < 0.01) of that in negative control group, respectively. The phagocytic rate of hemocyte and the ROS level in hemocyte were both significantly decreased (p < 0.01), while the apoptosis rate was significantly increased (p < 0.01) after CgUbe2g1 mRNA was interfered. These results demonstrated that Ub-conjugating enzyme CgUbe2g1 was involved in the innate immune response of oyster against invading pathogen, which might play important roles in the activation of inflammatory response and regulation of cellular immune response.