SGIV and RGNNV regulate fish innate immune responses by mediating ubiquitinated modifications of proteasome and ribosomal proteins.

Singapore grouper iridovirus (SGIV) and red-spotted grouper nervous necrosis virus (RGNNV) have been identified as the predominant DNA and RNA viral pathogens, respectively, affecting grouper populations. Protein ubiquitination, a crucial post-translational modification, plays pivotal regulatory roles in both host innate immune responses and viral replication processes. In the present study, we employed quantitative proteomics approaches to systematically identify and characterize alterations in ubiquitination-modified proteins in grouper spleen (Epinephelus akaara, GS) cells following SGIV and RGNNV infection. Proteomic analysis revealed that differentially expressed proteins were predominantly enriched in ribosome- and proteasome-related signaling pathways and associated biological processes. STRING analysis and correlation analysis revealed ubiquitin-40S ribosomal protein S27a (RPS27a) as a key regulatory molecule demonstrating significant interactions and functional correlations with proteins involved in ribosome biogenesis and proteasome-mediated degradation during both SGIV and RGNNV infection. Mechanistic investigations demonstrated that the replication processes of both SGIV and RGNNV are critically dependent on the host proteasome system, with viral regulation of the NF-κB signaling pathway being mediated through the ubiquitin-proteasome system (UPS). Functional validation through overexpression and RNA interference approaches demonstrated that grouper ub-RPS27a (Ecub-RPS27a) significantly inhibits viral replication while modulating innate immune responses in virus-infected GS cells. Collectively, these findings provide novel insights into the molecular mechanisms governing viral pathogenesis and host-pathogen interactions during SGIV and RGNNV infection.