No evidence for basigin/CD147 as a direct SARS-CoV-2 spike binding receptor

  1. Jarrod Shilts
  2. Thomas W. M. Crozier
  3. Edward J. D. Greenwood
  4. Paul J. Lehner
  5. Gavin J. Wright

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Abstract

The spike protein of SARS-CoV-2 is known to enable viral invasion into human cells through direct binding to host receptors including ACE2. An alternate entry receptor for the virus was recently proposed to be basigin/CD147. These early studies have already prompted a clinical trial and multiple published hypotheses speculating on the role of this host receptor in viral infection and pathogenesis. Here, we report that we are unable to find evidence supporting the role of basigin as a putative spike binding receptor. Recombinant forms of the SARS-CoV-2 spike do not interact with basigin expressed on the surface of human cells, and by using specialized assays tailored to detect receptor interactions as weak or weaker than the proposed basigin-spike binding, we report no evidence for a direct interaction between the viral spike protein to either of the two common isoforms of basigin. Finally, removing basigin from the surface of human lung epithelial cells by CRISPR/Cas9 results in no change in their susceptibility to SARS-CoV-2 infection. Given the pressing need for clarity on which viral targets may lead to promising therapeutics, we present these findings to allow more informed decisions about the translational relevance of this putative mechanism in the race to understand and treat COVID-19.

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  1. Version published to 10.1038/s41598-020-80464-1
  2. ScreenIT

    SciScore for 10.1101/2020.07.25.221036: (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
    Antibody staining was performed with a similar procedure, except during the first 30 minutes on ice cells were incubated with 30 μg/mL of monoclonal Ab-1 anti-BSG antibody12, then resuspended in 1:500 anti-human IgG antibody conjugated to Cy3 (Sigma
    anti-BSG
    suggested: None
    anti-human IgG
    suggested: None
    Anti-human basigin monoclonal antibodies were added at the following concentrations: 1.7 μg/mL Ab-1 (Zenonos et al., formerly known as ch6D9)12, 2.2 μg/mL MEM-M6/1 (Abcam #ab666), and 1.3 μg/mL MEM-M6/6 (Abcam #ab119114)25.
    Anti-human basigin
    suggested: None
    Both secondary antibodies were conjugated to alkaline phosphatase.
    alkaline phosphatase.
    suggested: None
    A control mouse anti-rat Cd4 domain 3+4 monoclonal antibody (OX68) against the tags of our recombinant proteins was used at a 1.6 μg/mL concentration.
    anti-rat Cd4 domain
    suggested: None
    After 1 hour of incubation with the primary antibody and three HBS-T washes, secondary antibody was added as 1:7000 donkey anti-human IgG (Abcam #ab102407) for ch6D9 and for all other antibodies as 1:3500 goat anti-mouse IgG (Sigma #A9316).
    anti-mouse IgG
    suggested: (Sigma-Aldrich Cat# A9316, RRID:AB_258446)
    Experimental Models: Cell Lines
    SentencesResources
    Recombinant protein expression and purification: Human embryonic kidney (HEK)-293E cells were transiently transfected with polyethylenimine as previously described46,47.
    HEK)-293E
    suggested: None
    Software and Algorithms
    SentencesResources
    Data processing: For flow cytometry experiments, measurement events for analysis were gated on live singlet cells (based on DAPI and forward and side scatter profiles) using FlowJo version 9.
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Cytometry data was visualized using the CytoML package50 in R version 3.6.1.
    CytoML
    suggested: None

    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.
    • No funding statement was detected.
    • 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.

  3. Version published to 10.1101/2020.07.25.221036v1 on bioRxiv