The amino-terminal domain of bovine viral diarrhea virus Npro protein is necessary for alpha/beta interferon antagonism

The alpha/beta interferon (IFN-alpha/beta) system is the first line of defense against viral infection and a critical link between the innate and adaptive immune responses. IFN-alpha/beta secretion is the hallmark of cellular responses to acute RNA virus infections. As part of their survival strategy, many viruses have evolved mechanisms to counteract the host IFN-alpha/beta response. Bovine viral diarrhea virus (BVDV) (genus Pestivirus) was reported to trigger interferon production in infected cultured cells under certain circumstances or to suppress it under others. Our studies with various cultured fibroblasts and epithelial bovine cells indicated that cytopathic (cp) BVDV induces IFN-alpha/beta very inefficiently. Using a set of engineered cp BVDVs expressing mutant N-pro and appropriate controls, we found that the IFN-alpha/beta response to infection was dependent on N-pro expression and independent of viral replication efficiency. In order to investigate whether the protease activity of N-pro is required for IFN-alpha/beta antagonism, we engineered N-pro mutants lacking protease activity by replacement of amino acid E-22, H-49, or C-69. We found that E-22 and H-49 substitutions abolished the ability of N-pro to suppress IFN, whereas C-69 had no effect, suggesting that the structural integrity of the N terminus of N-pro was more important than its catalytic activity for IFN-alpha/beta suppression. A catalytically active mutant with a change at a conserved N-pro region near the N terminus (L8P) in both BVDV biotypes did not antagonize IFN-alpha/beta production, confirming its involvement in this process. Taken together, these results not only provide direct evidence for the role of N-pro in blocking IFN-alpha/beta induction, but also implicate the amino-terminal domain of the protein in this function.