High mobility group box 1 can enhance NF-κB activation and act as a pro-inflammatory molecule in the Pacific oyster, Crassostrea gigas.

High-mobility group box 1 (HMGB1), a highly conserved DNA-binding protein, is involved in nucleosome formation and transcriptional regulation, and can also act as an extracellular cytokine to trigger inflammation and immune responses. In this study, we identified a HMGB1 gene (hereafter designated as CgHMGB1) in the Pacific oyster Crassostrea gigas. The full-length CgHMGB1 cDNA is 833 bp including 5' and 3'-untranslated regions (UTRs) of 145 and 79 bp, respectively, and an open reading frame (ORF) of 609 bp. The gene encodes a 202 amino acid polypeptide with an estimated molecular mass of 23.3 kDa. Sequence alignment shows that CgHMGB1 contains two basic HMG boxes and a highly acidic C-terminal domain. Recombinant CgHMGB1 proteins can enhance the mRNA level of various inflammatory cytokines in vivo. Typically, CgHMGB1 is localized in the nucleus, though lipopolysaccharide can induce its release to cytoplasm. Moreover, luciferase reporter assays reveal that CgHMGB1 cannot stimulate Nuclear Factor-κB reporter activity alone, but it can enhance Rel-dependent NF-κB activation in a dose-dependent manner. CgHMGB1 is highly expressed in hemocytes and its transcripts are significantly more abundant following bacterial challenge. Our results suggest that CgHMGB1 plays an essential role in innate defense by enhancing Rel-activated NF-κB activity and inducing the expression of inflammatory cytokines.