Black carp Nup93 negatively regulates IRF3- and IRF7-mediated antiviral immune response

Nucleoporin 93 (Nup93), a key component of the nuclear pore complex, is involved in various cellular processes, such as immune signaling pathway. In mammals, Nup93 positively regulates the RLR signaling pathway by targeting TBK1 and IRF3. However, the role of Nup93 in teleost immunity remains unexplored. In this study, we have cloned and characterized the Nup93 homolog (bcNup93) in black carp. bcNup93 consists of 821 amino acids, composed of a coiled-coil (CC) domain and a NIC96 domain. Quantitative RT-PCR analysis revealed fluctuations in bcNup93 mRNA levels in response to spring viremia of carp virus, grass carp reovirus, and poly(I:C) stimulation. bcNup93 migrated around 93 kDa in an immunoblotting assay, and immunofluorescence and nucleo-cytoplasmic fractionation assays identified its predominant localization in the nuclear envelope. Knockdown of bcNup93 in host cells significantly enhanced the expression of interferons and interferon-stimulated genes, as well as the antiviral capacity, suggesting a suppressive role of bcNup93 in antiviral immunity. In addition, bcNup93 knockdown in black carp juveniles led to reduced viral gene expression in multiple tissues postinfection, as well as an enhanced survival ratio. Mechanistically, bcNup93 interacts with bcIRF3/7 and attenuates their protein levels by lysosomal degradation pathways and decreases their protein stability or activity by degrading K63-linked ubiquitination of bcIRF3/7. Further analysis of bcNup93 functional domains shows that both CC and NIC96 domains are critical for its inhibitory effects on bcIRF3/7-mediated antiviral signaling. In conclusion, these findings demonstrate that bcNup93 negatively modulates the antiviral immune response in black carp by attenuating bcIRF3/7 protein levels.