The D614G mutation in the SARS-CoV2 Spike protein increases infectivity in an ACE2 receptor dependent manner

  1. Junko Ogawa
  2. Wei Zhu
  3. Nina Tonnu
  4. Oded Singer
  5. Tony Hunter
  6. Amy L Ryan (Firth)
  7. Gerald M Pao

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Abstract

The SARS-CoV2 coronavirus responsible for the current COVID19 pandemic has been reported to have a relatively low mutation rate. Nevertheless, a few prevalent variants have arisen that give the appearance of undergoing positive selection as they are becoming increasingly widespread over time. Most prominent among these is the D614G amino acid substitution in the SARS-CoV2 Spike protein, which mediates viral entry. The D614G substitution, however, is in linkage disequilibrium with the ORF1b P314L mutation where both mutations almost invariably co-occur, making functional inferences problematic. In addition, the possibility of repeated new introductions of the mutant strain does not allow one to distinguish between a founder effect and an intrinsic genetic property of the virus. Here, we synthesized and expressed the WT and D614G variant SARS-Cov2 Spike protein, and report that using a SARS-CoV2 Spike protein pseudotyped lentiviral vector we observe that the D614G variant Spike has >1/2 log 10 increased infectivity in human cells expressing the human ACE2 protein as the viral receptor. The increased binding/fusion activity of the D614G Spike protein was corroborated in a cell fusion assay using Spike and ACE2 proteins expressed in different cells. These results are consistent with the possibility that the Spike D614G mutant increases the infectivity of SARS-CoV2.

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  1. ScreenIT

    SciScore for 10.1101/2020.07.21.214932: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Primary antibodies were GeneTex 632604 anti-Spike antibody (mouse monoclonal) and anti-calnexin (C5C9) rabbit mAb (CST #2679) were detected with 2nd anti-Mouse IgG Alexa 488 (Life tech Cat# A11029) and anti-rabbit IgG Dylight 633 (Thermo Fisher #35563) antibodies.
    anti-Spike
    suggested: None
    anti-calnexin
    suggested: (Thermo Fisher Scientific Cat# MA3027A488, RRID:AB_2633337)
    anti-Mouse IgG
    suggested: (Molecular Probes Cat# A-11029, RRID:AB_138404)
    anti-rabbit IgG Dylight 633
    suggested: (Thermo Fisher Scientific Cat# 35563, RRID:AB_1965953)
    Experimental Models: Cell Lines
    SentencesResources
    A stable Spike expressing 293T cell line was generated by infection with pBOB-CAG-SARS-CoV2-Spike-HA lentiviral vector https://www.addgene.org/141347/ followed by single cell cloning and screening by cell extract immunoblotting of individual clones with HA tag antibody (CST #3724).
    293T
    suggested: None
    Software and Algorithms
    SentencesResources
    Immunofluorescence was performed following the Cell Signaling Technology protocol (https://www.cellsignal.com/).
    https://www.cellsignal.com/
    suggested: (Cell Signaling Technology, RRID:SCR_004431)

    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 found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

    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.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2020.07.21.214932 on bioRxiv