Linking the production of reactive oxygen species to the expression of antimicrobial peptides through aquaporin in an arthropod

In arthropods, bacterial infection activates dual oxidase 2 (Duox2) to produce extracellular reactive oxygen species (ROS), while stimulating the expression of antimicrobial peptides (AMPs) through immune deficiency (Imd)/Relish pathway. Although both strategies contribute significantly to antibacterial immunity, the Duox2/ROS system and Relish/AMPs pathway are generally considered separate branches. In the present study, we report that these two branches are connected by an aquaporin 4 (Aqp4) in a crustacean, the red swamp crayfish <i>Procambarus clarkii</i>. Aqp4 is found to be critical for the crayfish antibacterial response. The significance of Aqp4 is achieved by transporting extracellular ROS into the intracellular compartment after bacterial infection. The increase in intracellular ROS levels, mediated by the c-Jun N-terminal kinase, promotes the nuclear translocation and thereby the transcriptional activity of FoxO. As the key transcription factor of Relish, FoxO regulates the upregulation of Relish and ultimately the expression of Relish-targeting AMPs. By linking ROS production to AMP expression, the sequential ROS/Aqp4/FoxO/Relish/AMPs axis makes a relatively significant, yet previously unnoticed, contribution to the antibacterial response. Therefore, this study reveals the intriguing association between ROS production and AMP expression, the two central antibacterial strategies and provides insights into the regulatory mechanisms of arthropod immunity.