Black carp STAT2 enhances IRF3-mediated antiviral signaling by regulating its ubiquitination and improving its nuclear translocation

Interferon (IFN) regulatory factor 3 (IRF3) is a critical transcription factor involved in inducing IFN production. However, the regulatory mechanisms of piscine IRF3 remain inadequately explored. Here, we report that the signal transducer and activator of transcription 2 (STAT2) of black carp (Mylopharyngodon piceus) functions as a positive regulator in the black carp IRF3 (bcIRF3)-mediated IFN signaling pathway. Knockdown of bcSTAT2 in vivo facilitates viral replications, and the mRNA levels of bcSTAT2 gene increase after IFN stimulation in host cells. Additionally, overexpression of bcSTAT2 promotes the activation of interferon stimulated response element (ISRE) ex vivo. Co-immunoprecipitation assays identify the interaction between bcSTAT2 and bcIRF3, and both molecules exhibit similar subcellular distributions in immunofluorescent staining assay. When co-expressed, bcSTAT2 improves the protein level of bcIRF3, and enhances the bcIRF3-mediated IFN production and antiviral capabilities. Mechanistically, we demonstrate that bcSTAT2 significantly enhances the nuclear translocation of bcIRF3. Besides, bcSTAT2 enhances K29-linked ubiquitination, while reduces K33- and K48-linked ubiquitination of bcIRF3. Taken together, our data concludes that bcSTAT2 positively regulates bcIRF3-mediated antiviral activity by regulating its ubiquitination and facilitating its nuclear translocation, which has shed a light on the regulation of IFN signaling.