Pea peptide depressed the replication of porcine epidemic diarrhea virus through ROS-PERK mediated autophagy

This study examined the anti-PEDV capability of PP in Vero cells. PP, at a concentration of 200 μM, was found to inhibit the replication of both PEDV hubei/2016/China strain and CV777 strain in Vero cells with concentration of 200 μM. Further investigations indicated that PP depressed the replication of virus life cycle, but not attachment and invasion into cells. In addition, PP can inhibit the generation of reactive oxygen species (ROS) induced by PEDV in Vero cells, which depressed autophagy caused by PEDV through PERK-eIF2α signal pathway. Our study expanded knowledge of bio-active PP antiviral capacity and deepened understanding of PEDV pathogenicity, which would be valuable for the development of control and prevention for PEDV. Fig 1showed that there was no clear toxic response to Vero cells when the PP concentration was 12.5–200 µM. Fig 2 revealed that the expression of N protein notably declined following PP treatment after PEDV CH/hubei/2016 strain (Fig.2 A and 2 B) and CV777 strain (Fig.2 C and 2 D) infection, indicating that PP was able to depress the replication of PEDV. Consistent with the western blot results, TCID50 assay indicated that the viral tiers of PP treated group decreased significantly (figure 2 E and figure 2 F). Compared to the control group, PP significantly decreased viral load (figure 2 G. figure 2 H). Fig 3 showed the addition of PP was performed under different treatment conditions, 18 hpi with PEDV hubei/2016 strain, cells were harvested and subject to western blot (Fig.3 A) , and TCID50 for release. The results showed that the PP had no effect on the viral adsorption (Fig.3 B and Fig.3 C), viral internalization (Fig.3 D and Fig.3 E) and viral release (Fig.3 H), but significantly inhibited the replication stage of viral protein (Fig.3F, G). Fig 4showed that the PEDV infection increased the fluorescence intensity compared to mocks, indicating that PEDV infection elevated the ROS production, while PP decreased the elevation of ROS caused by PEDV infection (Fig.4A, B), which was similar to that observed by NAC treatment. Fig 5 showed that PEDV infection significant increased KEAP 1 protein expression (Fig.5A, B), enhanced the LC 3II / LC3 B I ratio (Fig.5A, F) and phosphorylation of PERK and eIF2α (Fig.5A, C, D). However, addition of PP lowered KEAP 1 up-regulation, ratio of LC 3II / LC3 I (Fig.5A, B), phosphorylation of PERK and eIF2α induced by PEDV infection, which was consistent with NAC treatment (Fig.5A, B, C, and F). In line with NAC treatment, PP significantly depressed the replication of PEDV (Fig.5A and E).