scholarly journals The N-terminal and central domains of CoV-2 nsp1 play key functional roles in suppression of cellular gene expression and preservation of viral gene expression

2021 ◽  
Author(s):  
Aaron Stephen Mendez ◽  
Michael Ly ◽  
Angélica M. González-Sánchez ◽  
Ella Hartenian ◽  
Nicholas Ingolia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mutational Analysis ◽  
Nonstructural Protein ◽  
Ribosomal Subunit ◽  
Viral Gene ◽  
Viral Mrna ◽  
Cellular Gene ◽  
Cellular Gene Expression ◽  
Nonstructural Protein 1

Nonstructural protein 1 (nsp1) is the first viral protein synthesized during coronavirus (CoV) infection and is a key virulence factor that dampens the innate immune response. It restricts cellular gene expression through a combination of inhibiting translation by blocking the mRNA entry channel of the 40S ribosomal subunit and by promoting mRNA degradation. We performed a detailed structure-guided mutational analysis of CoV-2 nsp1 coupled with in vitro and cell-based functional assays, revealing insight into how it coordinates these activities against host but not viral mRNA. We found that residues in the N-terminal and central regions of nsp1 not involved in docking into the 40S mRNA entry channel nonetheless stabilize its association with the ribosome and mRNA, thereby enhancing its restriction of host gene expression. These residues are also critical for the ability of mRNA containing the CoV-2 leader sequence to escape translational repression. Notably, we identify CoV-2 nsp1 mutants that gain the ability to repress translation of viral leader-containing transcripts. These data support a model in which viral mRNA binding functionally alters the association of nsp1 with the ribosome, which has implications for drug targeting and understanding how engineered or emerging mutations in CoV-2 nsp1 could attenuate the virus.

scholarly journals Lysine 164 is critical for SARS-CoV-2 Nsp1 inhibition of host gene expression

2020 ◽  
Author(s):  
Zhou Shen ◽  
Guangxu Zhang ◽  
Yilin Yang ◽  
Mengxia Li ◽  
Siqi Yang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nonstructural Protein ◽  
Antiviral Drug ◽  
Ribosomal Subunit ◽  
Virus Production ◽  
Host Gene ◽  
Host Protein ◽  
Renilla Luciferase ◽  
Host Gene Expression ◽  
Nonstructural Protein 1

The emerging pathogen severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused social and economic disruption worldwide, infecting over 9.0 million people and killing over 469 000 by 24 June 2020. Unfortunately, no vaccine or antiviral drug that completely eliminates the transmissible disease coronavirus disease 2019 (COVID-19) has been developed to date. Given that coronavirus nonstructural protein 1 (nsp1) is a good target for attenuated vaccines, it is of great significance to explore the detailed characteristics of SARS-CoV-2 nsp1. Here, we first confirmed that SARS-CoV-2 nsp1 had a conserved function similar to that of SARS-CoV nsp1 in inhibiting host-protein synthesis and showed greater inhibition efficiency, as revealed by ribopuromycylation and Renilla luciferase (Rluc) reporter assays. Specifically, bioinformatics and biochemical experiments showed that by interacting with 40S ribosomal subunit, the lysine located at amino acid 164 (K164) was the key residue that enabled SARS-CoV-2 nsp1 to suppress host gene expression. Furthermore, as an inhibitor of host-protein expression, SARS-CoV-2 nsp1 contributed to cell-cycle arrest in G0/G1 phase, which might provide a favourable environment for virus production. Taken together, this research uncovered the detailed mechanism by which SARS-CoV-2 nsp1 K164 inhibited host gene expression, laying the foundation for the development of attenuated vaccines based on nsp1 modification.

scholarly journals Rat Cytomegalovirus Gene Expression in Cardiac Allograft Recipients Is Tissue Specific and Does Not Parallel the Profiles Detected In Vitro

2007 ◽  
Vol 81 (8) ◽  
pp. 3816-3826 ◽  
Author(s):  
Daniel N. Streblow ◽  
Koen W. R. van Cleef ◽  
Craig N. Kreklywich ◽  
Christine Meyer ◽  
Patricia Smith ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rat Heart ◽  
Cultured Cells ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Viral Mrna ◽  
Tissue Specific ◽  
Viral Genes

ABSTRACT Rat cytomegalovirus (RCMV) is a β-herpesvirus with a 230-kbp genome containing over 167 open reading frames (ORFs). RCMV gene expression is tightly regulated in cultured cells, occurring in three distinct kinetic classes (immediate early, early, and late). However, the extent of viral-gene expression in vivo and its relationship to the in vitro expression are unknown. In this study, we used RCMV-specific DNA microarrays to investigate the viral transcriptional profiles in cultured, RCMV-infected endothelial cells, fibroblasts, and aortic smooth muscle cells and to compare these profiles to those found in tissues from RCMV-infected rat heart transplant recipients. In cultured cells, RCMV expresses approximately 95% of the known viral ORFs with few differences between cell types. By contrast, in vivo viral-gene expression in tissues from rat heart allograft recipients is highly restricted. In the tissues studied, a total of 80 viral genes expressing levels twice above background (5,000 to 10,000 copies per μg total RNA) were detected. In each tissue type, there were a number of genes expressed exclusively in that tissue. Although viral mRNA and genomic DNA levels were lower in the spleen than in submandibular glands, the number of individual viral genes expressed was higher in the spleen (60 versus 41). This finding suggests that the number of viral genes expressed is specific to a given tissue and is not dependent upon the viral load or viral mRNA levels. Our results demonstrate that the profiles, as well as the amplitude, of viral-gene expression are tissue specific and are dramatically different from those in infected cultured cells, indicating that RCMV gene expression in vitro does not reflect viral-gene expression in vivo.

scholarly journals Coronavirus nonstructural protein 1: Common and distinct functions in the regulation of host and viral gene expression

2015 ◽  
Vol 202 ◽  
pp. 89-100 ◽  
Author(s):  
Krishna Narayanan ◽  
Sydney I. Ramirez ◽  
Kumari G. Lokugamage ◽  
Shinji Makino
Keyword(s):  
Gene Expression ◽  
Nonstructural Protein ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Nonstructural Protein 1

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

Science ◽  
2020 ◽  
Vol 369 (6508) ◽  
pp. 1249-1255 ◽  
Author(s):  
Matthias Thoms ◽  
Robert Buschauer ◽  
Michael Ameismeier ◽  
Lennart Koepke ◽  
Timo Denk ◽  
...  
Keyword(s):  
Messenger Rna ◽  
Nonstructural Protein ◽  
Ribosomal Subunit ◽  
Mrna Translation ◽  
Structural Basis ◽  
Innate Immune Responses ◽  
Structure Based Drug Design ◽  
Nonstructural Protein 1 ◽  
C Terminus

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the current coronavirus disease 2019 (COVID-19) pandemic. A major virulence factor of SARS-CoVs is the nonstructural protein 1 (Nsp1), which suppresses host gene expression by ribosome association. Here, we show that Nsp1 from SARS-CoV-2 binds to the 40S ribosomal subunit, resulting in shutdown of messenger RNA (mRNA) translation both in vitro and in cells. Structural analysis by cryo–electron microscopy of in vitro–reconstituted Nsp1-40S and various native Nsp1-40S and -80S complexes revealed that the Nsp1 C terminus binds to and obstructs the mRNA entry tunnel. Thereby, Nsp1 effectively blocks retinoic acid–inducible gene I–dependent innate immune responses that would otherwise facilitate clearance of the infection. Thus, the structural characterization of the inhibitory mechanism of Nsp1 may aid structure-based drug design against SARS-CoV-2.

scholarly journals The N-terminal domain of SARS-CoV-2 nsp1 plays key roles in suppression of cellular gene expression and preservation of viral gene expression

Cell Reports ◽  
2021 ◽  
pp. 109841
Author(s):  
Aaron S. Mendez ◽  
Michael Ly ◽  
Angélica M. González-Sánchez ◽  
Ella Hartenian ◽  
Nicholas T. Ingolia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Cellular Gene ◽  
Cellular Gene Expression ◽  
Terminal Domain

scholarly journals Mechanisms of Coronavirus Nsp1-Mediated Control of Host and Viral Gene Expression

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 300
Author(s):  
Keisuke Nakagawa ◽  
Shinji Makino
Keyword(s):  
Gene Expression ◽  
Virulence Factor ◽  
Nonstructural Protein ◽  
Host Gene ◽  
Viral Gene ◽  
Host Gene Expression ◽  
Viral Mrnas ◽  
Nonstructural Protein 1 ◽  
Cellular Environment ◽  
High Amino Acid

Many viruses disrupt host gene expression by degrading host mRNAs and/or manipulating translation activities to create a cellular environment favorable for viral replication. Often, virus-induced suppression of host gene expression, including those involved in antiviral responses, contributes to viral pathogenicity. Accordingly, clarifying the mechanisms of virus-induced disruption of host gene expression is important for understanding virus–host cell interactions and virus pathogenesis. Three highly pathogenic human coronaviruses (CoVs), including severe acute respiratory syndrome (SARS)-CoV, Middle East respiratory syndrome (MERS)-CoV, and SARS-CoV-2, have emerged in the past two decades. All of them encode nonstructural protein 1 (nsp1) in their genomes. Nsp1 of SARS-CoV and MERS-CoV exhibit common biological functions for inducing endonucleolytic cleavage of host mRNAs and inhibition of host translation, while viral mRNAs evade the nsp1-induced mRNA cleavage. SARS-CoV nsp1 is a major pathogenic determinant for this virus, supporting the notion that a viral protein that suppresses host gene expression can be a virulence factor, and further suggesting the possibility that SARS-CoV-2 nsp1, which has high amino acid identity with SARS-CoV nsp1, may serve as a major virulence factor. This review summarizes the gene expression suppression functions of nsp1 of CoVs, with a primary focus on SARS-CoV nsp1 and MERS-CoV nsp1.

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