Structural basis for translational shutdown and immune evasion by the Nsp1 protein of SARS-CoV-2

Thoms, Matthias; Buschauer, Robert; Ameismeier, Michael; Koepke, Lennart; Denk, T.; Hirschenberger, Maximilian; Kratzat, Hanna; Hayn, Manuel; Mackens-Kiani, Timur; Cheng, Jingdong; Stürzel, Christina M.; Fröhlich, Thomas; Berninghausen, Otto; Becker, Thomas; Kirchhoff, Frank; Sparrer, Konstantin M. J.; Beckmann, Roland

doi:10.1101/2020.05.18.102467

Cited by 151 publications

(269 citation statements)

References 63 publications

(58 reference statements)

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“…While this work was in progress, multiple groups reported structures of NSP1 bound to the human ribosome 31,32 . Despite its apparent flexibility, the N-terminal globular domain of NSP1 was localized to the solvent-exposed surface of the 40S subunit, near the entrance to the mRNA entry channel (Supp.…”

Section: Mrna Within the Entry Channel Of The 40s Subunit Inhibited Nmentioning

confidence: 99%

Dynamic competition between SARS-CoV-2 NSP1 and mRNA on the human ribosome inhibits translation initiation

Lapointe¹,

Grosely²,

Johnson

et al. 2020

Preprint

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SARS-CoV-2 recently emerged as a human pathogen and is the causative agent of the COVID-19 pandemic. A molecular framework of how the virus manipulates host cellular machinery to facilitate infection remains unclear. Here, we focus on SARS-CoV-2 NSP1, which is proposed to be a virulence factor that inhibits protein synthesis by directly binding the human ribosome. Using extract-based and reconstitution experiments, we demonstrate that NSP1 inhibits translation initiation on model human and SARS-CoV-2 mRNAs. NSP1 also specifically binds to the small (40S) ribosomal subunit, which is required for translation inhibition. Using single-molecule fluorescence assays to monitor NSP1-40S subunit binding in real time, we demonstrate that eukaryotic translation initiation factors (eIFs) modulate the interaction: NSP1 rapidly and stably associates with most ribosomal pre-initiation complexes in the absence of mRNA, with particular enhancement and inhibition by eIF1 and eIF3j, respectively. Using model mRNAs and an inter-ribosomal-subunit FRET signal, we elucidate that NSP1 competes with RNA segments downstream of the start codon to bind the 40S subunit and that the protein is unable to associate rapidly with 80S ribosomes assembled on an mRNA. Collectively, our findings support a model where NSP1 associates with the open head conformation of the 40S subunit to inhibit an early step of translation, by preventing accommodation of mRNA within the entry channel.

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Section: Mrna Within the Entry Channel Of The 40s Subunit Inhibited Nmentioning

confidence: 99%

Dynamic competition between SARS-CoV-2 NSP1 and mRNA on the human ribosome inhibits translation initiation

Lapointe¹,

Grosely²,

Johnson

et al. 2020

Preprint

View full text Add to dashboard Cite

show abstract

“…This alters the positive charge of the protein and may help improve its role in the assembly of the virus genome in the capsid, as discussed below in relation to the appearance of blooms (36). After this triple modification, we see several reversals of the tendency to lose C in the genome.…”

Section: Reversal Of the Tendency Of The Viral Genome To Lose Its Cytmentioning

confidence: 93%

“…In fact, the second branch comes from the same descent, to which is added the U490A (Asp75Glu) mutation in the Nsp1 protein, which controls the specific translation of viral RNA [36], systematically associated with the mutation C3177U (Pro971Leu) in the acidic domain, without any clearly identified function, of the multifunctional protease Nsp3 [37], and finally the U18736C (Phe1757Leu) doublet of the exonuclease, N7-methyltransferase Nsp14, and A29700G in the 3'UTR region of the virus. The Phe1757Leu modification is located in the middle of a zinc binding site at the interface between the two domains of the Nsp3 protein.…”

Section: Reversal Of the Tendency Of The Viral Genome To Lose Its Cytmentioning

confidence: 99%

Biochemical and mathematical lessons from the evolution of the SARS-CoV-2 virus: paths for novel antiviral warfare

Cluzel¹,

Lambert

Maday³

et al. 2020

Preprint

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In the fight against the spread of COVID-19 the emphasis is on vaccination or on reactivating existing drugs used for other purposes. The tight links that necessarily exist between the virus as it multiplies and the metabolism of its host are systematically ignored. Here we show that the metabolism of all cells is coordinated by the availability of a core building block of the cell’s genome, cytidine triphosphate (CTP). This metabolite is also the key to the synthesis of the viral envelope and to the translation of its genome into proteins. This unique role explains why evolution has led to the early emergence in animals of an antiviral immunity enzyme, viperin, that synthesizes a toxic analogue of CTP. The constraints arising from this dependency guide the evolution of the virus. With this in mind, we explored the real-time experiment taking place before our eyes using probabilistic modelling approaches to the molecular evolution of the virus. We have thus followed, almost on a daily basis, the evolution of the composition of the viral genome to link it to the progeny produced over time, particularly in the form of blooms that sparked a firework of viral mutations. Some of those certainly increase the propagation of the virus. This led us to make out the critical role in this evolution of several proteins of the virus, such as its nucleocapsid N, and more generally to begin to understand how the virus ties up the host metabolism to its own benefit. A way for the virus to escape CTP-dependent control in cells would be to infect cells that are not expected to grow, such as neurons. This may account for unexpected body sites of viral development in the present epidemic.

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“…Further indicative of its preserved biological function, nsp1 from alpha-and beta-CoVs have different size, but show comparable biological activities in their ability to reduce host gene expression, even though the mechanism seems different [15,[20][21][22]. SARS-CoV nsp1 almost completely blocks host protein translation by binding the 40S ribosome of the host cell, which stops canonical mRNA translation at different steps during the initiation process [23][24][25].…”

Section: Introductionmentioning

confidence: 99%

“…For example, in the case of SARS-CoV several residues have been identi ed that differentially inhibit host gene expression, like interferon alpha, responsible for antiviral activity [18]. More recently, a region in the C-terminal domain of nsp1 of SARS-CoV-2 has been demonstrated to interfere with host expression factors [25].…”

Section: Introductionmentioning

confidence: 99%

Emerging of a SARS-CoV-2 viral strain with a deletion in nsp1

Benedetti

Snyder²,

Giovanetti

et al. 2020

Preprint

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Background:The new Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), which was ﬁrst detected in Wuhan (China) in December of 2019 is responsible for the current global pandemic.Phylogenetic analysis revealed that it is similar to other betacoronaviruses, such as SARS-CoV and Middle-Eastern Respiratory Syndrome, MERS-CoV. Its genome is ∼30 kb in length and contains two large overlapping polyproteins, ORF1a and ORF1ab that encode for several structural and non-structural proteins. The non-structural protein 1 (nsp1) is arguably the most important pathogenic determinant, and previous studies on SARS-CoV indicate that it is both involved in viral replication and hampering the innate immune system response. Detailed experiments of site-specific mutagenesis and in vitro reconstitution studies determined that the mechanisms of action are mediated by i) the presence of specific amino acid residues of nsp1 and b) the interaction between the protein and the host’s small ribosomal unit. In fact, substitution of certain amino acids resulted in reduction of its negative effects.Methods: A total of 17928 genome sequences were obtained from the GISAID database (December 2019 to July 2020) from patients infected by SARS-CoV-2 from different areas around the world. Genomes alignment was performed using MAFFT (REFF) and the nsp1 genomic regions were identified using BioEdit and verified using BLAST. Nsp1 protein of SARS-CoV-2 with and without deletion have been subsequently modelled using I-TASSER.Results: We identified SARS-CoV-2 genome sequences, from several Countries, carrying a previously unknown deletion of 9 nucleotides in position 686-694, corresponding to the AA position 241-243 (KSF). This deletion was found in different geographical areas. Structural prediction modelling suggests an effect on the C-terminal tail structure.Conclusions: Modelling analysis of a newly identified deletion of 3 amino acids (KSF) of SARS-CoV-2 nsp1 suggests that this deletion could affect the structure of the C-terminal region of the protein, important for regulation of viral replication and negative effect on host’s gene expression. In addition, substitution of the two amino acids (KS) from nsp1 of SARS-CoV was previously reported to revert loss of interferon-alpha expression. The deletion that we describe indicates that SARS-CoV-2 is undergoing profound genomic changes. It is important to: i) confirm the spreading of this particular viral strain, and potentially of strains with other deletions in the nsp1 protein, both in the population of asymptomatic and pauci-symptomatic subjects, and ii) correlate these changes in nsp1 with potential decreased viral pathogenicity.

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Structural basis for translational shutdown and immune evasion by the Nsp1 protein of SARS-CoV-2

Cited by 151 publications

References 63 publications

Dynamic competition between SARS-CoV-2 NSP1 and mRNA on the human ribosome inhibits translation initiation

Dynamic competition between SARS-CoV-2 NSP1 and mRNA on the human ribosome inhibits translation initiation

Biochemical and mathematical lessons from the evolution of the SARS-CoV-2 virus: paths for novel antiviral warfare

Emerging of a SARS-CoV-2 viral strain with a deletion in nsp1

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