Salvianolic acid A inhibits PRRSV replication via binding to Keap1 to activate the MKRN1–Nrf2–NQO1 pathway

Porcine reproductive and respiratory syndrome (PRRS) has caused significant economic losses to the global pig industry, and there are currently no safe, effective, or commercially available vaccines. Traditional Chinese medicine may serve as a beneficial supplement to vaccines and provide a feasible solution for preventing and controlling PRRS. The present study explored the effects and molecular mechanisms of the traditional Chinese medicine Salviae miltiorrhizae Bunge extract salvianolic acid A (SalA) on porcine reproductive and respiratory syndrome virus (PRRSV) replication. SalA effectively inhibited PRRSV replication in vitro and in vivo without affecting the adsorption, entry, or release stages of the viral replication cycle. SalA directly binds to the Thr560 site of Kelch-like ECH-associated protein 1 (Keap1) via a hydrogen bond, promoting the recruitment of Keap1 to a newly identified E3 ubiquitin ligase, makorin RING finger protein 1 (MKRN1). This led to K48-linked ubiquitination of Keap1 at the K615 amino acid residue, subsequent proteasomal degradation, and eventual activation of the nuclear factor E2-related factor 2 (Nrf2)-NADPH quinone oxidoreductase 1 (NQO1) pathway. Knockdown of MKRN1 blocked SalA-induced Keap1 ubiquitination, degradation, and activation of the Nrf2-NQO1 pathway. Further viral infection experiments revealed that SalA inhibited PRRSV replication by activating the MKRN1-Nrf2-NQO1 pathway. In addition to its anti-PRRSV activity, SalA effectively inhibited PRRSV- and lipopolysaccharide (LPS)-induced expression of inflammatory cytokines, activation of inflammatory pathways and inflammasomes, and inhibition of cellular pyroptosis by activating the Nrf2 pathway. These results suggest that SalA can inhibit PRRSV replication and alleviate the inflammatory response during PRRS and secondary bacterial infections, providing a novel candidate strategy for PRRS treatment.