The landscape of antibody binding in SARS-CoV-2 infection

  1. Anna S. Heffron
  2. Sean J. McIlwain
  3. Maya F. Amjadi
  4. David A. Baker
  5. Saniya Khullar
  6. Ajay K. Sethi
  7. Ann C. Palmenberg
  8. Miriam A. Shelef
  9. David H. O’Connor
  10. Irene M. Ong

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Abstract

The search for potential antibody-based diagnostics, vaccines, and therapeutics for pandemic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has focused almost exclusively on the spike (S) and nucleocapsid (N) proteins. Coronavirus membrane (M), ORF3a, and ORF8 proteins are humoral immunogens in other coronaviruses (CoVs) but remain largely uninvestigated for SARS-CoV-2. Here we use ultradense peptide microarray mapping to show that SARS-CoV-2 infection induces robust antibody responses to epitopes throughout the SARS-CoV-2 proteome, particularly in M, in which one epitope achieved excellent diagnostic accuracy. We map 79 B cell epitopes throughout the SARS-CoV-2 proteome and demonstrate that antibodies that develop in response to SARS-CoV-2 infection bind homologous peptide sequences in the six other known human CoVs. We also confirm reactivity against four of our top-ranking epitopes by enzyme-linked immunosorbent assay (ELISA). Illness severity correlated with increased reactivity to nine SARS-CoV-2 epitopes in S, M, N, and ORF3a in our population. Our results demonstrate previously unknown, highly reactive B cell epitopes throughout the full proteome of SARS-CoV-2 and other CoV proteins.

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    SciScore for 10.1101/2020.10.10.334292: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: Human subjects: The study was conducted in accordance with the Declaration of Helsinki and approved by the Institutional Review Board of the University of Wisconsin-Madison.
    Consent: All subjects were 18 years of age or older at the time of recruitment and provided informed consent.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    647-conjugated goat antihuman IgG secondary antibody (Jackson ImmunoResearch).
    antihuman IgG
    suggested: (GeneTex Cat# GTX28798, RRID:AB_374523)
    To determine whether the peptide was in an epitope (in SARS-CoV-2 proteins) or cross-reactive for anti-SARS-CoV-2 antibodies (in non-SARS-CoV-2 proteins), we used an adjusted p-value cutoff of <0.1 (based on multiple hypothesis testing correction for all 119,487 unique sequences on the array) and a fold-change of greater than or equal to 2 and grouped consecutive peptides as a represented epitope.
    anti-SARS-CoV-2
    suggested: None
    Software and Algorithms
    SentencesResources
    Additional unmodeled regions were generated using Modeller [59].
    Modeller
    suggested: (MODELLER, RRID:SCR_008395)
    C-proximal HR2 regions were modeled as single helices (Phe1148-Leu1211) in Coot [60].
    Coot
    suggested: (Coot, RRID:SCR_014222)
    The data2bfactor Python script written by Robert L.
    Python
    suggested: (IPython, RRID:SCR_001658)
    Campbell, Thomas Holder, and Suguru Asai (downloaded from http://pldserver1.biochem.queensu.ca/~rlc/work/pymol/) was used to substitute peptide array data onto this structure in place of the B factor in PyMol (The PyMOL Molecular Graphics System, Version 2.0 Schrödinger, LLC) using a dark blue (low) to red (high) color scale.
    PyMol
    suggested: (PyMOL, RRID:SCR_000305)
    Alignments for heatmaps were created using MUSCLE [63].
    MUSCLE
    suggested: (MUSCLE, RRID:SCR_011812)

    Results from OddPub: Thank you for sharing your code and data.

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    A caveat is that our methods cannot discern whether the increased IgG binding to CCCoVs in COVID-19 convalescent sera is due to newly developed cross-reactive antibodies or due to the stimulation of a memory response against the original CCCoV antigens. However, cross-reactivity of anti-SARS-CoV-2 antibodies with SARS-CoV or MERS-CoV is likely real, since our population was very unlikely to have been exposed to those viruses. A more stringent assessment of cross-reactivity as well as functional investigations into these cross-reactive antibodies will be vital in determining their capacity for cross-protection. Further, our methods efficiently detect antibody binding to linear epitopes [55], but their sensitivity for detecting parts of conformational epitopes is unknown, and additional analyses will be required to determine whether epitopes identified induce neutralizing or otherwise protective antibodies. Finally, we demonstrated that more severely ill patients have significantly greater reactivity to certain epitopes in S, M, N, and ORF3a. The nine epitopes with significantly higher magnitude reactivity in intubated patients may play a role in the overaggressive immune response known to characterize severe COVID-19 [7, 56], suggesting that they may be targets for treatment in or prevention of severe disease. Alternatively, the antibody response in general may be higher in very sick patients, expanding the repertoire of antibody reactivity. Future studies should investigate w...

    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: Please consider improving the rainbow (“jet”) colormap(s) used on page 14. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

    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

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  2. Version published to 10.1101/2020.10.10.334292 on bioRxiv