The mRNA and protein of IL-8 oppositely regulate PRRSV replication via nucleoprotein and 14-3-3γ

Porcine reproductive and respiratory syndrome virus (PRRSV) has posed a significant threat to the global swine industry for over 3 decades, resulting in substantial economic losses. In this study, we identified the cellular protein 14-3-3γ as an antiviral factor that suppresses PRRSV replication by downregulating the mRNA levels of interleukin-8 (IL-8). Intriguingly, while IL-8 mRNA levels were positively correlated with PRRSV genome replication, the IL-8 protein itself exhibited a potent inhibitory effect on viral replication. Mechanistic studies revealed that 14-3-3γ enhances IL-8 transcription. In contrast, the viral nucleocapsid (N) protein upregulates IL-8 transcription. A critical nuclear localization signal (NLS) domain (amino acid residues 11-13) within the N protein, particularly the lysine residues at positions 10 and 12, was identified as essential for this regulatory function. The nuclear distribution of the N protein was found to be indispensable for its activity. Furthermore, we demonstrated that 14-3-3γ and IL-8 competitively bind to the N protein, modulating its nuclear distribution. This study elucidates the intricate regulation of IL-8 at both transcriptional and protein levels in modulating PRRSV replication. It uncovers a novel function of IL-8 in viral pathogenesis and highlights its potential as a promising candidate for the development of anti-PRRSV therapeutics.IMPORTANCEPorcine reproductive and respiratory syndrome virus (PRRSV) has severely impacted swine production worldwide for decades. This study reveals that the cellular protein 14-3-3γ suppresses PRRSV replication by downregulating interleukin-8 mRNA, while IL-8 protein itself inhibits viral replication. Mechanistically, 14-3-3γ upregulates IL-8 transcription, whereas the viral nucleocapsid (N) protein enhances IL-8 transcription, which is dependent on its nuclear localization signal (amino acid residues 11-13, particularly K10/K12). Nuclear distribution of the N protein is essential for this function. Additionally, 14-3-3γ and IL-8 competitively bind N protein, modulating its nuclear trafficking. This dual regulation of IL-8 at transcriptional and protein levels highlights its antiviral role and potential as a therapeutic target against PRRSV.