Identification and functional characterization of the goldfish (Carassius auratus L.) high mobility group box 1 (HMGB1) chromatin-binding protein

• HMGB1 mRNA levels were highest in the brain, spleen, intestine and kidney.
• HMGB1 expression was up-regulated in tissues of goldfish infected with M. marinum.
• Recombinant HMGB1 induced respiratory burst and nitric oxide response in phagocytes.
• Macrophages treated with HMGB1 produced TNFα-2 and IL1-β1.
• HMGB1 induced activation of NFκB signaling pathway.

We report on the identification and functional characterization of HMGB1 of the goldfish. Quantitative analysis indicated the highest expression of goldfish HMGB1 in the brain, with lower mRNA levels in spleen, intestine, kidney, gill and heart. HMGB1 was also differentially expressed in goldfish immune cell populations with highest mRNA levels present in splenocytes and neutrophils. We generated and functionally characterized the recombinant HMGB1 (rgHMGB1). The rgHMGB1 primed the respiratory burst response in monocytes and induced nitric oxide production of primary goldfish macrophages. Treatment of goldfish macrophages with heat-killed Mycobacterium marinum and Aeromonas salmonicida elevated the expression of HMGB1 and resulted in higher HMGB1 protein levels. The rgHMGB1 induced a dose-dependent production of TNFα-2 and IL-1β1 of goldfish macrophages. Furthermore, the dual luciferase reporter assay revealed that goldfish HMGB1 induced the activation of the NF-κB signaling pathway. Our results indicate that goldfish HMGB1 is a critical regulatory cytokine of inflammatory and antimicrobial response of the goldfish.