Serum amyloid P component suppresses porcine epidemic diarrhea virus replication through TLR4-mediated IFN-β signaling pathway

Abstract

Porcine epidemic diarrhea virus (PEDV) is a porcine enteropathogenic coronavirus that causes significant economic losses in many Asian and European countries. It is characterized by lethal watery diarrhea and high mortality rate in piglets. Serum amyloid P component (SAP), a member of acute response phase protein (APP) family, has been reported to play a crucial role in innate immune response against various microbial pathogens. However, its antiviral activities are little known. In this study, the antiviral activity of SAP during PEDV infection was investigated. In virus-infected IPEC cells, it was found that SAP expression was significantly upregulated. To study the role of SAP in PEDV replication, the expression of SAP was regulated in cells using eukaryotic expression plasmids expressing the SAP protein and sgRNA. PEDV replication was then assessed through real-time PCR, Western blotting, and TCID₅₀ assays. The result showed that PEDV replication was inhibited in cells overexpressing SAP and promoted in cells with SAP knocking out. To further investigate the mechanism by which SAP inhibits PEDV replication, Interferon Beta (IFN-β) and its related signaling pathway proteins were detected. The results demonstrated that SAP activates the promoter of (IFN-β) and IFN regulatory factor 3 (IRF3) mediated by Toll-Like Receptor 4 (TLR4) signaling. During PEDV infection, SAP enhances TLR4-mediated IFN-β signaling, leading to increased IFN-β expression, which subsequently suppresses PEDV replication. By using TBK1/IKBKE inhibitor MRT67307 in PEDV-infected cells, the antiviral activity of SAP was inhibited. This suggests that the antiviral effect of SAP may rely on the activation of the TBK1/IKBKE signaling pathway, which is critical for the induction of type I interferons and other antiviral responses. Moreover, the interaction between SAP and PEDV N protein and the functional domain of SAP were investigated. From the results of this study, it can be concluded that the interaction between SAP and PEDV N protein activates the TLR4-mediated IFN signaling pathway, thereby inhibiting PEDV replication.