ZNFX1 functions as a compensatory dsRNA recognition receptor to exert antiviral effect in orange-spotted grouper.

Zinc finger NFX1-type containing 1 (ZNFX1) has been established as a critical mediator of the antiviral response in mammals, functioning through dsRNA recognition and priority activation of the retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) signaling pathway. However, the role of its fish ortholog, particularly in relation to aquatic virus interactions, remains elusive. The absence of the RIG-I homolog, a key pattern recognition receptor, in multiple Actinopterygii may compromise their innate antiviral immune responses. Here, ZNFX1 in Epinephelus coioides (EcZNFX1) is identified as an evolutionarily primitive, interferon (IFN)-stimulated dsRNA sensor that compensates for the absence of RIG-I in Actinopterygii. EcZNFX1 is rapidly upregulated by orange-spotted grouper nervous necrosis virus (OGNNV) infection and restricts viral replication in grouper brain-derived (GB) and spleen-derived (GS) cells after binding to viral dsRNA intermediates via its conserved P-loop NTPase domain. Notably, EcZNFX1 exerts a dual immunoregulatory role in modulating virus-induced inflammatory responses in diverse cellular contexts. In GB cells that are highly susceptible to OGNNV and have attenuated regenerative capacity, EcZNFX1 suppresses IFN-I/ISGs production and pyroptosis mediated by viral infection, thereby limiting neurotoxicity by precise tuning of the antiviral response. Conversely, in GS cells with stronger resistance to OGNNV, EcZNFX1 directly interacts with TBK1 to promote its phosphorylation and subsequent nuclear translocation of IRF3, activating a robust IFN-I signaling. Overall, this study elucidates that ZNFX1 is a compensatory receptor for dsRNA sensing in RIG-I-deficient teleost, which exerts context-dependent antiviral effects in cell-type-specific microenvironments, providing mechanistic insights for aquatic virus countermeasures.