Triggering Unfolded Protein Response By 2-Deoxy-D-Glucose Inhibits Porcine Epidemic Diarrhea Virus Propagation

Porcine epidemic diarrhea (PED) is a devastating swine disease that is characterized by acute enteritis and lethal watery diarrhea followed by dehydration. The virus infection results in a high mortality rate in piglets. PED is caused by porcine epidemic diarrhea virus (PEDV) that manifests as sporadic outbreaks during the winter. Although it was first identified in Europe, it now becomes an increasing problem in many Asian countries, causing to huge economic losses on breeding farms. PEDV outbroke among U.S. farms in 2013. Although the application of bivalent inactivated vaccine or attenuated PEDV & TGEV vaccine has greatly decreased the incidence of two types of viral disease, the occurrence of PED is still increasing annually in our country. No specific antiviral drugs can be used when PED outbreaks.The unfolded protein response (UPR) is cyto-protective machinery elicited towards an influx of large amount of protein synthesis in the endoplasmic reticulum (ER). Extensive studies suggest that the UPR can also be activated during virus infection. In the present study, we use transmission electron microscopy to analyze the morphology changes of ER post-PEDV infection. We observed the ER morphology was impressively deranged with hyper-swollen membranes throughout the majority of PEDV-infected cells. When viewed in sagittal section, multiple large vesicles appeared to be surrounded by the swollen ER, some of which were also full of virions. Western Blot and real-time PCR identified the differences of UPR genes in response to PEDV infection. The results suggested that PEDV infection induced UPR in Vero cells. Meanwhile, we silenced the expression of PKR-like ER kinase (PERK) by shRNA, we found that the knockdown of PERK increased virus loads in the cells, which was consistent with the result on 4-phenylbutyrate (4-PBA) treatment.We next determined whether 2-Deoxy-D-glucose (2-DG), an ER stress inducer, possesses inhibitory activity against PEDV infection. Plaque formation assay, RT-PCR and Western Blot analysis suggested that 2-DG might inhibit virus infection by affecting viral protein translation during the early stage of virus infection. Interestingly, we also found that 2-DG treatment could affect virus assembly, which is similar to previous studies on influenza virus. All these results support the therapeutic potential of using 2-DG or glucose/mannose analogs to induce the UPR to block virus replication.