Grass carp Hnf4β orchestrates antibacterial defense via the AIF/Hnf4α/caspase 3 signaling axis

Abstract

Hepatocyte nuclear factor 4 (HNF4), a transcription factor family critical for hepatic development and metabolic homeostasis, consists of three isoforms (HNF4α, HNF4β, HNF4γ). While HNF4α and HNF4γ are evolutionarily conserved across vertebrates, HNF4β is restricted to teleosts, amphibians and birds, with its antimicrobial function poorly characterized. Here, we investigate the role of grass carp Hnf4β (gcHnf4β) in response to Aeromonas salmonicida infection, uncovering a novel antibacterial signaling axis. Overexpression of gcHnf4β significantly attenuates bacterial proliferation and augments Ctenopharyngodon idella kidney (CIK) cell viability through transcriptional upregulation of caspase 3, caspase 9 and apoptosis-inducing factor (AIF). Mechanistically, gcHnf4β acts as a transcriptional hub, directly interacting with AIF and forming a ternary complex with gcHnf4α to indirectly engage caspase 3. Subcellular dynamics show nuclear retention of gcHnf4β during infection, concurrent with cytoplasmic-to-nuclear translocation of AIF/caspase 3 and their co-localization in the nucleus. Functional validation reveals that AIF knockdown or caspase 3 inhibition abolishes antibacterial activity mediated by gcHnf4β, whereas caspase 9 inhibition does not. Our findings establish gcHnf4β as a critical regulator of antimicrobial immunity, providing novel insights into host-pathogen interactions and potential targets for aquaculture disease control.