AEN Suppresses the Replication of Porcine Epidemic Diarrhea Virus by Inducing the Expression of Type I IFN and ISGs in MARC-145 Cells.

Apoptosis-enhancing nuclease (AEN), which shares close evolutionary relationships with the interferon-stimulated gene 20 protein (ISG20) homologs in humans, is a member of the DEDDh exonuclease family. Numerous studies on various pathogens have identified the essential roles of ISG20 in inhibiting virus replication. However, the fundamental functions of AEN during viral infection remain largely unknown. This study discovered that AEN expression was significantly upregulated in MARC-145 cells infected with Porcine epidemic diarrhea virus (PEDV) strain 85-7. In contrast, the amount of AEN protein decreased as viral replication increased. It was found that PEDV nsp1 and nsp5 mediated the decrease in AEN production, suggesting that an increase in AEN was not conducive to virus replication. By comparing AEN and its exonuclease-inactive mutant AEN-4A, we determined that the antiviral activity of AEN was independent of its exonuclease function. qPCR analyses revealed that AEN and AEN-4A could induce a significant increase in the transcription levels of IFN-α, IFN-β, and ISGs (OASL, IFI44, IFIT2, ISG15, Mx1, Mx2), and that AEN-4A has a higher induction ability. Overexpression of AEN and AEN-4A in MARC-145 cells targeting IFN-β knockdown or IFN-deficient Vero cells showed reduced or a complete loss of antiviral activity of both, suggesting that AEN may activate the type I IFN immune response and promote the expression of ISGs, thereby inhibiting PEDV replication. Taken together, our data prove the novel mechanism of AEN-mediated virus restriction.