SARS-CoV-2 Nsp1 suppresses host but not viral translation through a bipartite mechanism

Abstract

SUMMARYThe Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a highly contagious virus that underlies the current COVID-19 pandemic. SARS-CoV-2 is thought to disable various features of host immunity and cellular defense. The SARS-CoV-2 nonstructural protein 1 (Nsp1) is known to inhibit host protein translation and could be a target for antiviral therapy against COVID-19. However, how SARS-CoV-2 circumvents this translational blockage for the production of its own proteins is an open question. Here, we report a bipartite mechanism of SARS-CoV-2 Nsp1 which operates by: (1) hijacking the host ribosome via direct interaction of its C-terminal domain (CT) with the 40S ribosomal subunit and (2) specifically lifting this inhibition for SARS-CoV-2 via a direct interaction of its N-terminal domain (NT) with the 5’ untranslated region (5’ UTR) of SARS-CoV-2 mRNA. We show that while Nsp1-CT is sufficient for binding to 40S and inhibition of host protein translation, the 5’ UTR of SARS-CoV-2 mRNA removes this inhibition by binding to Nsp1-NT, suggesting that the Nsp1-NT-UTR interaction is incompatible with the Nsp1-CT-40S interaction. Indeed, lengthening the linker between Nsp1-NT and Nsp1-CT of Nsp1 progressively reduced the ability of SARS-CoV-2 5’ UTR to escape the translational inhibition, supporting that the incompatibility is likely steric in nature. The short SL1 region of the 5’ UTR is required for viral mRNA translation in the presence of Nsp1. Thus, our data provide a comprehensive view on how Nsp1 switches infected cells from host mRNA translation to SARS-CoV-2 mRNA translation, and that Nsp1 and 5’ UTR may be targeted for anti-COVID-19 therapeutics.

SARS-CoV-2 utilizes a multipronged strategy to suppress host protein synthesis

“…By their nature, deep sequencing measurements provide relative values but not absolute quantification of RNA and translation levels. Since SARS-CoV-2 encoded protein, NSP1, was recently shown to interfere with translation by blocking the mRNA entry channel of ribosomes [25][26][27][28] , and since the extent to which SARS-CoV-2 interferes with the overall levels of cellular mRNA was not assessed, we next examined if SARS-CoV-2 infection affects global translation and RNA levels. To quantify absolute translation levels, we measured nascent protein synthesis levels using an analogue of puromycin, O-Propargyl Puromycin (OPP), which incorporates into elongating polypeptide chains 34 .…”

“…In CoVs, the most prominent and well characterized cellular shutoff protein is NSP1 23 . So far, studies on SARS-CoV-2 NSP1 demonstrated it restricts translation by directly binding to the ribosome 40S subunit [25][26][27][28] , thereby globally inhibiting translation initiation. For SARS-CoV, on top of this translation effect, NSP1 interactions with the 40S was also shown to induce cleavage of translated cellular mRNAs, thereby accelerating their turnover their translation we compiled a list of 14 long non-coding RNAs (lncRNAs) that localize to the cytoplasm and are well expressed in our data but, as expected, poorly translated ( Figure S4B).…”

“…Indeed, several studies focused on SARS-CoV-2 encoded NSP1, showing it binds to the 40S ribosomal subunit, leading to reduction in mRNA translation both in vitro and in cells [24][25][26][27][28] . Still, how the virus can overcome the NSP1-mediated block of translation and how NSP1 effects come into play during infection remained unclear.…”

“…NSP1 is the best characterized and most prominent coronavirus host shutoff factor 23 . Several recent studies showed SARS-CoV-2 NSP1 binds the 40s ribosome and inhibits translation [24][25][26][27][28] .…”

The viral protein NSP1 acts as a ribosome gatekeeper for shutting down host translation and fostering SARS-CoV-2 translation

“…Recently, it has been shown that the N-terminal domain of SARS-CoV-2 NSP1 is required for viral translation by NSP1-bound ribosomes (Shi et al, 2020). The linker length between the N-and C-terminal domains is critical for viral translation .…”