Irg1 mediates inhibition of A20-triggered NF-κB signaling cascade in tilapia macrophages during early immune response

Abstract

Immune response gene 1 (Irg1) is prominently upregulated during the early phase of macrophage immune response to lipopolysaccharide (LPS) stimulation, concomitant with pronounced inflammatory activity. Prior investigations have identified Irg1 as one of the most rapidly and significantly upregulated immune-related genes in the teleost fish model following pathogenic challenge, suggesting its pivotal role in teleost immune responses. Herein, we investigated how Irg1 modulates early (12 h) inflammatory responses in head kidney–derived macrophages of Oreochromis niloticus (Nile tilapia). Our results reveal widespread distribution of Irg1 across 11 healthy O. niloticus tissues, with the highest expression observed in the spleen, followed by the head kidney and gills. Furthermore, dynamic upregulation of Irg1 expression was evident within immune-related tissues (head kidney and spleen) during a 72 h period following in vivo stress. We successfully expressed and purified the recombinant OnIRG1 protein ((r)OnIRG1) and established a macrophage model with OnIrg1 knockdown. At the onset of the cellular immune response (12 h post-LPS stimulation) the up-regulated Irg1 was found to inhibit A20 (an NF-κB negative regulatory), which activated the NF-κB signaling pathway and facilitating the expression of subsequent downstream inflammatory factors. Overall, these findings not only affirm the highly conserved structure and function of Irg1, but also broaden our comprehension of its role in teleost cellular immune regulation, emphasizing its significance in immune modulation.