Dynamic competition between SARS-CoV-2 NSP1 and mRNA on the human ribosome inhibits translation initiation
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Abstract
SARS-CoV-2 is the causative agent of the COVID-19 pandemic. A molecular framework for how the virus manipulates host cellular machinery to facilitate infection is needed. Here, we integrate biochemical and single-molecule strategies to reveal molecular insight into how NSP1 from SARS-CoV-2 inhibits translation initiation. NSP1 directly binds to the small (40S) subunit of the human ribosome, which is modulated by human initiation factors. Further, NSP1 and mRNA compete with each other to bind the ribosome. Our findings suggest that the presence of NSP1 on the small ribosomal subunit prevents proper accommodation of the mRNA. How this competition disrupts the many steps of translation initiation is an important target for future studies.
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SciScore for 10.1101/2020.08.20.259770: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources nLuc in vitro translation assays: HeLa cell-free translation (ThermoFisher, #88884) reactions setup according to manufacturer’s protocol were programed with a final mRNA concentration of 200 nM (endpoint) or 80 nM (real-time). HeLasuggested: NoneHAP1 cells were grown at 37 °C with 5% CO2 in Iscove’s Modified Dulbecco’s medium (IMDM) (Gibco, #sh30228) supplemented with 10% (v/v) HAP1suggested: NoneWild-type HEK293T cells were purchased … SciScore for 10.1101/2020.08.20.259770: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources nLuc in vitro translation assays: HeLa cell-free translation (ThermoFisher, #88884) reactions setup according to manufacturer’s protocol were programed with a final mRNA concentration of 200 nM (endpoint) or 80 nM (real-time). HeLasuggested: NoneHAP1 cells were grown at 37 °C with 5% CO2 in Iscove’s Modified Dulbecco’s medium (IMDM) (Gibco, #sh30228) supplemented with 10% (v/v) HAP1suggested: NoneWild-type HEK293T cells were purchased from ATCC (CRL-3216) and engineered HEK293T cells are described here. HEK293Tsuggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)Software and Algorithms Sentences Resources Co-localized molecules were identified and analyzed using custom MATLAB scripts. MATLABsuggested: (MATLAB, RRID:SCR_001622)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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