Porcine reproductive and respiratory syndrome virus (PRRSV), the most economically important infectious disease of pigs, elicits poor innate and adaptive immune responses. Soluble CD83 (sCD83), a secretion from various immune cell populations, especially MoDCs, is involved in negatively regulating the immune response. We speculate sCD83 may be a critical factor in the process of PRRSV-coordinated macrophage polarization. In this study, we found that PAMs co-cultured with PRRSV-infected MoDCs inhibited the M1 macrophage while enhancing the M2 macrophage. This was accompanied by a decrease in the pro-inflammatory cytokine TNF-α and iNOS and an increase in the anti-inflammatory cytokine IL-10 and Arg1. Meanwhile, sCD83 incubation causes the same specific effects lead to a switch in macrophage from M1 to M2. Neutralization of sCD83 removes the inhibitory effects of PRRSV on PAMs. Using reverse genetics, we generated recombinant PRRSVs with mutations in N protein, nsp1α, and nsp10 (knockout sCD83-concerned key amino acid site). Four mutant viruses lost the suppression of M1 macrophage markers, in contrast to the restriction of the upregulation of M2 macrophage markers. These findings suggest that PRRSV modulates the switch of macrophage polarization from M1 to M2 by upregulating the MoDC-induced secretion of CD83, providing new insights into the mechanism by which PRRSV regulates host immunity.

Heat shock protein 27 (HSP27) is a multifunctional protein and belongs to the small HSP family. It has been shown that HSP27 is involved in viral replication as a cellular chaperone, but the function of HSP27 during porcine reproductive and respiratory syndrome virus (PRRSV) infections remains unexplored. Here, we found that PRRSV replication can induce HSP27 expression and phosphorylation in vitro. HSP27 overexpression promoted PRRSV replication, whereas its knockdown reduced PRRSV proliferation. Additionally, suppressing HSP27 phosphorylation reduced PRRSV replication and the level of viral double-stranded RNA (dsRNA), a marker of the viral replication and transcription complexes (RTCs). Furthermore, HSP27 can interact with multiple viral nonstructural proteins (nsps), including nsp1α, nsp1β, nsp5, nsp9, nsp11 and nsp12. Suppressing the phosphorylation of HSP27 almost completely disrupted its interaction with nsp1β and nsp12. Altogether, our study revealed that HSP27 plays an important role in PRRSV replication.

Porcine reproductive and respiratory syndrome virus (PRRSV) seriously affects the world pig industry, and existing vaccines are not enough to protect against PRRSV efficiently. Identifying host factors that inhibit PRRSV infection may provide potential targets for antiviral therapies. Galectin 3 binding protein (LGALS3BP), also referred to as 90K, is a multifunctional protein that plays significant roles in viral infections, but its role in PRRSV infection is not clearly defined. In this study, we found that 90K expression increased during PRRSV infection. Additionally, 90K inhibits PRRSV replication by restricting both viral RNA synthesis and viral assembly. Further studies showed that 90K can interact with various PRRSV structural proteins, such as GP2, GP3, and GP5. Moreover, 90K targets K217 in GP3 for K48-linked ubiquitination, facilitating GP3 degradation through the proteasomal pathway. Meanwhile, we found that 90K induces the expression of type I interferon (IFN) and inflammatory cytokines by activating NF-κB and IRF3. Overall, our findings suggest that 90K plays a negative role in PRRSV replication by inducing PRRSV-GP3 degradation and activating innate immune responses.