Grouper deubiquitinating enzyme ovarian tumor domain-containing protein 5 (OTUD5) stabilizes stimulator of interferon genes (STING) via the autophagy-lysosomal pathway to modulate antiviral innate immunity in fish.

Abstract

The deubiquitinating enzyme ovarian tumor domain-containing protein 5 (OTUD5) in fish modulates antiviral immunity through a novel, non-canonical mechanism, as demonstrated by comprehensive functional and mechanistic studies. This work demonstrates that grouper OTUD5 (EcOTUD5) dichotomously regulates viral replication in fish cells, suppressing DNA virus (Singapore grouper iridovirus, SGIV) while enhancing RNA virus (Red-spotted grouper nervous necrosis virus, RGNNV) proliferation. Notably, this dual regulation is independent of EcOTUD5's catalytic activity, evidenced by persistent effects following mutations in its OTU domain or catalytic site (C224S). Subcellular localization analysis reveals nucleocytoplasmic distribution of EcOTUD5 post-infection, while the C224S mutation compromises nuclear localization stability. Mechanistically, EcOTUD5 modulates innate immunity by stabilizing stimulator of interferon genes (STING) through autophagy-lysosomal pathway activation. Overexpression of EcOTUD5 extends STING half-life via autophagy inhibition, while lysosomal inhibition further suppresses STING degradation. Importantly, EcOTUD5 selectively targets K63-linked polyubiquitin chains on STING, a modification critical for antiviral signaling. These findings uncover a unique regulatory axis where EcOTUD5 functions as a molecular switch, differentially controlling DNA and RNA virus replication through STING stabilization and non-enzymatic mechanisms. This work provides critical insights into the role of deubiquitinases in innate immune signaling and highlights the complexity of host-virus interactions in aquatic species.