Coronavirus Non-Structural Protein 1 Is a Major Pathogenicity Factor: Implications for the Rational Design of Coronavirus Vaccines

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Abstract
Attenuated viral vaccines can be generated by targeting essential pathogenicity factors. We report here the rational design of an attenuated recombinant coronavirus vaccine based on a deletion in the coding sequence of the non-structural protein 1 (nsp1). In cell culture, nsp1 of mouse hepatitis virus (MHV), like its SARS-coronavirus homolog, strongly reduced cellular gene expression. The effect of nsp1 on MHV replication in vitro and in vivo was analyzed using a recombinant MHV encoding a deletion in the nsp1-coding sequence. The recombinant MHV nsp1 mutant grew normally in tissue culture, but was severely attenuated in vivo. Replication and spread of the nsp1 mutant virus was restored almost to wild-type levels in type I interferon (IFN) receptor-deficient mice, indicating that nsp1 interferes efficiently with the type I IFN system. Importantly, replication of nsp1 mutant virus in professional antigen-presenting cells such as conventional dendritic cells and macrophages, and induction of type I IFN in plasmacytoid dendritic cells, was not impaired. Furthermore, even low doses of nsp1 mutant MHV elicited potent cytotoxic T cell responses and protected mice against homologous and heterologous virus challenge. Taken together, the presented attenuation strategy provides a paradigm for the development of highly efficient coronavirus vaccines. Prevention of viral diseases by vaccination aims for controlled induction of protective immune responses against viral pathogens. Live viral vaccines consist of attenuated, replication-competent viruses that are believed to be superior in the induction of broad immune responses, including cell-mediated immunity. The recent proceedings in the area of virus reverse genetics allows for the rational design of recombinant vaccines by targeting, i.e., inactivating, viral pathogenicity factors. For coronaviruses, a major pathogenicity factor has now been identified. The effect of coronavirus non-structural protein 1 on pathogenicity has been analyzed in a murine model of coronavirus infection. By deleting a part of this protein, a recombinant virus has been generated that is greatly attenuated in vivo, while retaining immunogenicity. In particular, the mutant virus retained the ability to replicate in professional antigen-presenting cells and fulfilled an important requirement of a promising vaccine candidate: the induction of a protective long-lasting, antigen-specific cellular immune response. This study has implications for the rational design of live attenuated coronavirus vaccines aimed at preventing coronavirus-induced diseases of veterinary and medical importance, including the potentially lethal severe acute respiratory syndrome.