Early endonuclease-mediated evasion of RNA sensing ensures efficient coronavirus replication.

Eveline Kindler,Roland Züst,Yize Li,Frank J M Van Kuppeveld,Cornelia C Bergmann,John Ziebuhr,Nadja Karl,Julia Spanier,Hanspeter Stalder,Volker Thiel,Jochen Wilhelm,Burkhard Ludewig,Ruth Elliot,Sabrina Marti,Huib H Rabouw,Robert H Silverman,Philip V’Kovski,Christina Gaughan,Matthias Habjan,Susan R Weiss,Markus Keller,Ulrich Kalinke,Mi-Hyun Hwang ,Cristina Gil‐Cruz ,Luisa Cervantes-Barragán

doi:10.1371/journal.ppat.1006195

Abstract

Coronaviruses are of veterinary and medical importance and include highly pathogenic zoonotic viruses, such as SARS-CoV and MERS-CoV. They are known to efficiently evade early innate immune responses, manifesting in almost negligible expression of type-I interferons (IFN-I). This evasion strategy suggests an evolutionary conserved viral function that has evolved to prevent RNA-based sensing of infection in vertebrate hosts. Here we show that the coronavirus endonuclease (EndoU) activity is key to prevent early induction of double-stranded RNA (dsRNA) host cell responses. Replication of EndoU-deficient coronaviruses is greatly attenuated in vivo and severely restricted in primary cells even during the early phase of the infection. In macrophages we found immediate induction of IFN-I expression and RNase L-mediated breakdown of ribosomal RNA. Accordingly, EndoU-deficient viruses can retain replication only in cells that are deficient in IFN-I expression or sensing, and in cells lacking both RNase L and PKR. Collectively our results demonstrate that the coronavirus EndoU efficiently prevents simultaneous activation of host cell dsRNA sensors, such as Mda5, OAS and PKR. The localization of the EndoU activity at the site of viral RNA synthesis–within the replicase complex—suggests that coronaviruses have evolved a viral RNA decay pathway to evade early innate and intrinsic antiviral host cell responses.

Highlights

Host innate immune responses are of particular importance during the early phase of virus infection to restrict virus replication and spread
We found that coronaviruses that lack this enzymatic activity are readily visible to infected host cells that can mount a swift and potent host response restricting virus replication within hours
Our study provides a new paradigm of a first layer of RNA virus innate immune evasion during the early phase of infection, that takes place at the site of RNA synthesis, and is based on removal of double-stranded RNA (dsRNA) that would otherwise trigger the simultaneous activation of cytoplasmic host cell sensors

Summary

Introduction

Host innate immune responses are of particular importance during the early phase of virus infection to restrict virus replication and spread They rely on the ability to differentiate between immunological “self” and “non-self” in order to swiftly activate diverse antiviral effector mechanisms. Coronaviruses are positive-stranded RNA viruses that replicate in the host cell cytoplasm They are well known to evade innate immune activation, during the early phase of the infection [3,4,5,6]. The importance of functions encoded by the CoV replicase gene is further exemplified by non-structural protein (nsp) 1 that suppresses host gene expression by mediating host mRNA degradation [10, 11], and nsp that contains a papain-like proteinase with deubiquitination activity interfering with IFN-I host cell responses [12, 13]. A number of accessory gene functions, less conserved, have been described to target downstream events of innate immune activation, such as a phosphodiesterase (PDE) activity encoded by some

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Discussion

Conclusion