SARS-CoV-2 impairs interferon production via NSP2-induced repression of mRNA translation.

Viruses evade the innate immune response by suppressing the production or activity of cytokines such as type I interferons (IFNs). Here we report the discovery of a mechanism by which the SARS-CoV-2 virus coopts an intrinsic cellular machinery to suppress the production of the key immunostimulatory cytokine IFN-ß. We reveal that the SARS-CoV-2 encoded nonstructural protein 2 (NSP2) directly interacts with the cellular GIGYF2 protein. This interaction enhances the binding of GIGYF2 to the mRNA cap-binding protein 4EHP, thereby repressing the translation of the Ifnb1 mRNA. Depletion of GIGYF2 or 4EHP significantly enhances IFN-ß production, which inhibits SARS-CoV-2 replication. Our findings reveal a target for rescuing the antiviral innate immune response to SARS-CoV-2 and other RNA viruses.

SARS-CoV-2 viral protein Nsp2 stimulates translation under normal and hypoxic conditions.

When viruses like SARS-CoV-2 infect cells, they reprogram the repertoire of cellular and viral transcripts that are being translated to optimize their strategy of replication, often targeting host translation initiation factors, particularly eIF4F complex consisting of eIF4E, eIF4G and eIF4A. A proteomic analysis of SARS-CoV-2/human proteins interaction revealed viral Nsp2 and initiation factor eIF4E2, but a role of Nsp2 in regulating translation is still controversial. HEK293T cells stably expressing Nsp2 were tested for protein synthesis rates of synthetic and endogenous mRNAs known to be translated via cap- or IRES-dependent mechanism under normal and hypoxic conditions. Both cap- and IRES-dependent translation were increased in Nsp2-expressing cells under normal and hypoxic conditions, especially mRNAs that require high levels of eIF4F. This could be exploited by the virus to maintain high translation rates of both viral and cellular proteins, particularly in hypoxic conditions as may arise in SARS-CoV-2 patients with poor lung functioning.

Proteomic screening identifies RPLp2 as a specific regulator for the translation of coronavirus.

Viral mRNA of coronavirus translates in an eIF4E-dependent manner, and the phosphorylation of eIF4E can modulate this process, but the role of p-eIF4E in coronavirus infection is not yet entirely evident. p-eIF4E favors the translation of selected mRNAs, specifically the mRNAs that encode proteins associated with cell proliferation, inflammation, the extracellular matrix, and tumor formation and metastasis. In the present work, two rounds of TMT relative quantitative proteomics were used to screen 77 cellular factors that are upregulated upon infection by coronavirus PEDV and are potentially susceptible to a high level of p-eIF4E. PEDV infection increased the translation level of ribosomal protein lateral stalk subunit RPLp2 (but not subunit RPLp0/1) in a p-eIF4E-dependent manner. The bicistronic dual-reporter assay and polysome profile showed that RPLp2 is essential for translating the viral mRNA of PEDV. RNA binding protein and immunoprecipitation assay showed that RPLp2 interacted with PEDV 5'UTR via association with eIF4E. Moreover, the cap pull-down assay showed that the viral nucleocapsid protein is recruited in m7GTP-precipitated complexes with the assistance of RPLp2. The heterogeneous ribosomes, which are different in composition, regulate the selective translation of specific mRNAs. Our study proves that viral mRNA and protein utilize translation factors and heterogeneous ribosomes for preferential translation initiation. This previously uncharacterized process may be involved in the selective translation of coronavirus.

Emetine suppresses SARS-CoV-2 replication by inhibiting interaction of viral mRNA with eIF4E.

Emetine is a FDA-approved drug for the treatment of amebiasis. Previously we demonstrated the antiviral efficacy of emetine against some RNA and DNA viruses. In this study, we evaluated the in vitro antiviral efficacy of emetine against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and found it to be a low nanomolar (nM) inhibitor. Interestingly, emetine exhibited protective efficacy against lethal challenge with infectious bronchitis virus (IBV; a chicken coronavirus) in the embryonated chicken egg infection model. Emetine treatment led to a decrease in viral RNA and protein synthesis without affecting other steps of viral life cycle such as attachment, entry and budding. In a chromatin immunoprecipitation (CHIP) assay, emetine was shown to disrupt the binding of SARS-CoV-2 mRNA with eIF4E (eukaryotic translation initiation factor 4E, a cellular cap-binding protein required for initiation of protein translation). Further, molecular docking and molecular dynamics simulation studies suggested that emetine may bind to the cap-binding pocket of eIF4E, in a similar conformation as m7-GTP binds. Additionally, SARS-CoV-2 was shown to exploit ERK/MNK1/eIF4E signalling pathway for its effective replication in the target cells. Collectively our results suggest that further detailed evaluation of emetine as a potential treatment for COVID-19 may be warranted.