FADD cooperates with Caspase-8 to positively regulate the innate immune response and promote apoptosis following bacterial infection in Japanese eel.

Fas-associated protein with Death Domain (FADD) is a crucial signaling component of apoptosis and a vital immunomodulator on inflammatory signaling pathways. However, information on FADD-mediated apoptosis and immune regulation is limited in teleost. We herein cloned a FADD homolog, AjFADD, from Japanese eel (Anguilla japonica). Expression analysis revealed that AjFADD was significantly induced by LPS, poly I:C, and Aeromonas hydrophila infection in vivo and in vitro. The expression of IFNs and IRFs, c-Rel and c-Fos, IL1 and TNF-α, and the essential antimicrobial peptide LEAP-2 in Japanese eel liver cells was enhanced by overexpressing AjFADD, with a significant decrease of those genes following knockdown AjFADD. Luciferase activity assay, flow cytometry, and wound healing results showed that AjFADD cooperated with AjCaspase-8 to promote apoptosis of HEK293 cells and Japanese eel liver cells infected with A. hydrophila. Furthermore, AjFADD and AjCaspase-8 co-localized in the cytoplasm and displayed a direct protein-protein interaction by immunoprecipitation. Our results collectively showed that FADD cooperated with Caspase-8 to positively regulate the innate immune response and promote apoptosis in response to the A. hydrophila challenge in Japanese eel.