Evasion of Antiviral Innate Immunity by Theiler's Virus L* Protein through Direct Inhibition of RNase L

Abstract

Theiler's virus is a neurotropic picornavirus responsible for chronic infections of the central nervous system. The establishment of a persistent infection and the subsequent demyelinating disease triggered by the virus depend on the expression of L*, a viral accessory protein encoded by an alternative open reading frame of the virus. We discovered that L* potently inhibits the interferon-inducible OAS/RNase L pathway. The antagonism of RNase L by L* was particularly prominent in macrophages where baseline oligoadenylate synthetase (OAS) and RNase L expression levels are elevated, but was detectable in fibroblasts after IFN pretreatment. L* mutations significantly affected Theiler's virus replication in primary macrophages derived from wild-type but not from RNase L-deficient mice. L* counteracted the OAS/RNase L pathway through direct interaction with the ankyrin domain of RNase L, resulting in the inhibition of this enzyme. Interestingly, RNase L inhibition was species-specific as Theiler's virus L* protein blocked murine RNase L but not human RNase L or RNase L of other mammals or birds. Direct RNase L inhibition by L* and species specificity were confirmed in an in vitro assay performed with purified proteins. These results demonstrate a novel viral mechanism to elude the antiviral OAS/RNase L pathway. By targeting the effector enzyme of this antiviral pathway, L* potently inhibits RNase L, underscoring the importance of this enzyme in innate immunity against Theiler's virus.

Author Summary

Theiler's virus is a murine picornavirus (same family as poliovirus) which has a striking ability to establish persistent infections of the central nervous system. To do so, the virus has to counteract the immune response of the host and particularly the potent response mediated by interferon. We observed that a protein encoded by Theiler's virus, the L* protein, inhibited the RNase L pathway, one of the best-characterized pathways mediating the antiviral IFN response. In contrast to previously identified viral antagonists of this pathway, L* was found to act directly on RNase L, the effector enzyme of the pathway. L* activity was found to be species-specific as it inhibited murine but not human RNase L. We confirmed the species-specificity and the direct interaction between L* and RNase L in vitro, using purified proteins. Acting at the effector step in the pathway allows L* to block RNase L activity efficiently. This suggests that RNase L is particularly important to control Theiler's virus replication in vivo. Another virus, mouse hepatitis virus (MHV), was recently shown to interfere with RNase L activation. Theiler's virus and MHV share a marked tropism for macrophages which may suggest that the RNase L pathway is particularly important in this cell type.