• Tue. Feb 27th, 2024
Common mechanisms of translational shutdown of coronaviruses

Non-structural protein 1 (Nsp1) produced by coronaviruses appears to inhibit host protein synthesis in infected cells. Previous studies have shown that the C-terminal domain (CTD) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Nsp1 binds to the small ribosomal subunit and inhibits translation. However, whether this is a mechanism widely used by coronaviruses remains unknown.

In a recently published study bioRxiv* Preprint server, researchers investigate Nsp1 from SARS-CoV-2, Middle East respiratory syndrome coronavirus (MERS-CoV), and Bat-Hp-CoV using biophysical, structural, and biochemical assays.

Research: Universal features of Nsp1-mediated translational shutdown by coronaviruses. Image credit: Jerome-Cronenberger / Shutterstock.com

*Important Notice: bioRxiv Primary scientific reports are published that are not peer-reviewed and therefore should not be considered definitive, guide clinical practice/health-related behavior, or constitute established information.

background

Infection with members of the Betacoronavirus (β-CoV) genus causes severe respiratory disease in humans. A capped 5′ untranslated region (5’UTR) marks the beginning of the approximately 30 kilobase (kb) β-CoV genome, contains several protein-encoding open reading frames (ORFs), and ends with a polyadenylated 3’UTR.

Several NSPSs collectively facilitate viral infection through unclear mechanisms. Therefore, a better understanding of these mechanisms will accelerate the development of new therapeutics.

The SARS-CoV-2 Nsp1 CTD binds to the entry region of the messenger ribonucleic acid (mRNA) channel in the 40S subunit, where it strongly interacts with the mRNA and inhibits translation. Whether Nsp1 proteins from other β-CoVs share this mechanistic function is unclear.

About the study

In the present study, Nsp1 proteins from three representative β-CoVs were selected. These include SARS-CoV-2 subspecies Merbecovirus, MERS-CoV subspecies Merbecovirus, and Bat-Hp-CoV. Bat-HP-covi, the only member of the Hibecovirus subgenus, was chosen because it binds the human ribosome.

The main objective of this study was to obtain biochemical and structural evidence demonstrating that Nsp1 from all selected β-CoVs silences the translation of host mRNAs by binding to the mRNA channel of the 40S ribosomal subunit. To this end, paired structural and single-molecule analyzes were used to show that the N-terminal domain (NTD) of Bat-Hp-CoV Nsp1 binds to the decoding center of the 40S subunit.

Key findings

Binding of the Nsp1 CTD to the mRNA channel of the 40S subunit was shown to be a conserved mechanism. Furthermore, evasion of Nsp1-mediated translational inhibition of mRNAs was also documented. Furthermore, the elusive NTD binding of Nsp1 to the decoding center of the 40S subunit was visualized using the Bat-Hp-CoV protein as a model system.

Although only the NTD for Bat-Hp-CoV Nsp1 was visualized, biochemical data indicate that Nsps from all selected β-CoVs exert mechanistic effects on translation and binding mode. In vitro Translational experiments showed that NTD and CTD contribute significantly to translational inhibition in all viral systems tested. Furthermore, regions of Nsp1 responsible for ribosome interactions appear to be critical for selective translation of viral mRNAs.

Evaluation of the relative occupancy of the Nsp1 domains in the 40S subunit revealed that Nsp1 forms a bipartite interaction with the 40S subunit. The CTD of Nsp1 appears to have high affinity for the 40S mRNA entry channel. These findings are consistent with a previous model in which the CTD domain anchors the protein to the 40S subunit, and the Nsp1 NTD rapidly samples the 40S decoding center.

Nsp1 NTDs and viral mRNAs were first co-evolved and bypassed the translational barrier, matching Nsp1 proteins from the three viruses to their corresponding viral mRNAs.

The next step was loss of translational avoidance activity caused by mutation of either critical nucleotides in the stem-loop or conserved residues of the Nsp1 NTD in the viral 5’UTR. Thus, it is possible that Nsp1 immunoreactivity is reduced if viral mRNAs compete with the 40S subunit to interact with the Nsp1 NTD and prevent its localization to the decoding center.

Conclusions

A fundamental limitation of this study is that the interaction between viral mRNA and the ribosome-bound Nsp1 NTD has not yet been directly observed. However, the framework provided here rationalizes the substantial amount of scientific literature on the role of Nsp1.

These findings also provide the basis for further investigation into how coronaviruses balance the host immune response using viral protein synthesis. Future research is needed to expand on these findings to develop effective anti-viral therapeutics that target Nsp1 activity across betacoronaviruses.

*Important Notice: bioRxiv Primary scientific reports are published that are not peer-reviewed and therefore should not be considered definitive, guide clinical practice/health-related behavior, or constitute established information.

Journal Reference:

  • Preliminary Scientific Report. Schubert, K., Karosis, ED, Ban, I., etc. (2023) Universal features of Nsp1-mediated translational shutdown of coronaviruses. bioRxiv. doi:10.1101/2023.05.31.543022

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