Negative regulation of RIG-I by STAT1a and STAT1b in the innate immunity of black carp.

Signal transducer and activator of transcription 1 (STAT1) is a key component of the interferon (IFN) signaling pathway, in which retinoic acid-inducible gene I (RIG-I) functions as a key cytosolic viral RNA sensor. However, the role of STAT1 in regulating the upstream components of the IFN pathway remains much unknown in teleost. In this study, we investigated the interaction and functional roles of STAT1a and STAT1b in regulating the RIG-I-mediated antiviral signaling pathway in black carp (Mylopharyngodon piceus). Co-immunoprecipitation (Co-IP) and immunofluorescence (IF) assays showed that STAT1a and STAT1b interact with RIG-I in the cytoplasm. Dual-luciferase reporter assays revealed that STAT1a and STAT1b inhibit RIG-I-induced IFN promoter activity. Further analysis in EPC cells infected with spring viremia of carp virus (SVCV) showed that both STAT1 isoforms suppressed RIG-I's antiviral activity, particularly STAT1b. Mechanistically, STAT1a degraded RIG-I through the ubiquitin-proteasome pathway and selectively inhibited K63-linked ubiquitination, while STAT1b impaired K63-linked ubiquitination without affecting oligomerization. Our data conclude that STAT1a and STAT1b negatively regulate RIG-I by reducing K63-linked polyubiquitination, providing a novel mechanism for post-translational regulation of the RIG-I signaling pathway in teleosts.