The nsp3 Macrodomain Promotes Virulence in Mice with Coronavirus-Induced Encephalitis

Abstract

All coronaviruses encode for a macrodomain containing ADP-ribose-1”-phosphatase (ADRP) activity within the N-terminus of non-structural protein 3 (nsp3). Previous work showed that mouse hepatitis virus strain A59 (MHV-A59) with a mutated catalytic site (N1348A) replicated similarly to wild-type virus but was unable to cause acute hepatitis in mice. To determine whether this attenuated phenotype is applicable to multiple disease models, we mutated the catalytic residue in the JHM strain of MHV (JHMV), which causes acute and chronic encephalomyelitis, using a newly developed bacterial artificial chromosome (BAC)-based MHV reverse genetics system. Infection of mice with the macrodomain catalytic point mutant virus (N1347A) resulted in reduced lethality, weight loss, viral titers, pro-inflammatory cytokine and chemokine expression, and immune cell infiltration in the brain when compared to mice infected with wild-type virus. Specifically, macrophages were most affected, with approximately 2.5-fold fewer macrophages at day 5 post-infection in N1347A-infected brains. Tumor necrosis factor (TNF) and interferon (IFN) signaling were not required for effective host control of mutant virus, as all N1347A-infected mice survived the infection. However, the adaptive immune system was required for protection, since N1347A virus was able to cause lethal encephalitis in RAG1 (recombination activation gene1)^-/- knockout mice, although disease onset was modestly delayed. Overall, these results indicate that the BAC-based MHV reverse genetics system will be useful for studies of JHMV, and expand upon previous studies, showing that the macrodomain is critical for the ability of coronaviruses to evade the immune system and promote viral pathogenesis.