PSMB4 Degrades the Porcine Reproductive and Respiratory Syndrome Virus Nsp1α Protein via the Autolysosome Pathway and Induces the Production of Type I Interferon

Porcine reproductive and respiratory syndrome virus (PRRSV) causes respiratory disease in pigs of all ages and reproductive failure in sows, resulting in great economic losses to the swine industry. In this work, we identified the interaction between PSMB4 and PRRSV Nsp1 alpha by yeast two-hybrid screening. The PSMB4-Nsp1 alpha interaction was further confirmed by coimmunoprecipitation, glutathione S-transferase (GST) pulldown, and laser confocal experiments. The PCP alpha domain (amino acids 66 to 166) of Nsp1 alpha and the C-terminal domain (amino acids 250 to 264) of PSMB4 were shown to be critical for the PSMB4-Nsp1 alpha interaction. PSMB4 overexpression reduced PRRSV replication, whereas PSMB4 knockdown elicited opposing effects. Mechanistically, PSMB4 targeted K169 in Nsp1 alpha for K63-linked ubiquitination and targeted Nsp1 alpha for autolysosomal degradation by interacting with LC3 to enhance the activation of the lysosomal pathway. Meanwhile, we found that PSMB4 activated the NF-kappa B signaling pathway to produce type I interferons by downregulating the expression of I kappa B alpha and p-I kappa B alpha. In conclusion, our data revealed a new mechanism of PSMB4-mediated restriction of PRRSV replication, whereby PSMB4 was found to induce Nsp1 alpha degradation and type I interferon expression, in order to impede the replication of PRRSV.IMPORTANCE In the swine industry, PRRSV is a continuous threat, and the current vaccines are not effective enough to block it. This study determined that PSMB4 plays an antiviral role against PRRSV. PSMB4 was found to interact with PRRSV Nsp1 alpha, mediate K63-linked ubiquitination of Nsp1 alpha at K169, and thus trigger its degradation via the lysosomal pathway. Additionally, PSMB4 activated the NF-kappa B signaling pathway to produce type I interferons by downregulating the expression of I kappa B alpha and p-I kappa B alpha. This study extends our understanding of the proteasome subunit PSMB4 against PRRSV replication and will contribute to the development of new antiviral strategies. In the swine industry, PRRSV is a continuous threat, and the current vaccines are not effective enough to block it. This study determined that PSMB4 plays an antiviral role against PRRSV.