Characterisation of the transcriptome and proteome of SARS-CoV-2 reveals a cell passage induced in-frame deletion of the furin-like cleavage site from the spike glycoprotein

  1. Andrew D. Davidson
  2. Maia Kavanagh Williamson
  3. Sebastian Lewis
  4. Deborah Shoemark
  5. Miles W. Carroll
  6. Kate J. Heesom
  7. Maria Zambon
  8. Joanna Ellis
  9. Philip A. Lewis
  10. Julian A. Hiscox
  11. David A. Matthews

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Abstract

Background

SARS-CoV-2 is a recently emerged respiratory pathogen that has significantly impacted global human health. We wanted to rapidly characterise the transcriptomic, proteomic and phosphoproteomic landscape of this novel coronavirus to provide a fundamental description of the virus’s genomic and proteomic potential.

Methods

We used direct RNA sequencing to determine the transcriptome of SARS-CoV-2 grown in Vero E6 cells which is widely used to propagate the novel coronavirus. The viral transcriptome was analysed using a recently developed ORF-centric pipeline. Allied to this, we used tandem mass spectrometry to investigate the proteome and phosphoproteome of the same virally infected cells.

Results

Our integrated analysis revealed that the viral transcripts (i.e. subgenomic mRNAs) generally fitted the expected transcription model for coronaviruses. Importantly, a 24 nt in-frame deletion was detected in over half of the subgenomic mRNAs encoding the spike (S) glycoprotein and was predicted to remove a proposed furin cleavage site from the S glycoprotein. Tandem mass spectrometry identified over 500 viral peptides and 44 phosphopeptides in virus-infected cells, covering almost all proteins predicted to be encoded by the SARS-CoV-2 genome, including peptides unique to the deleted variant of the S glycoprotein.

Conclusions

Detection of an apparently viable deletion in the furin cleavage site of the S glycoprotein, a leading vaccine target, shows that this and other regions of SARS-CoV-2 proteins may readily mutate. The furin site directs cleavage of the S glycoprotein into functional subunits during virus entry or exit and likely contributes strongly to the pathogenesis and zoonosis of this virus. Our data emphasises that the viral genome sequence should be carefully monitored during the growth of viral stocks for research, animal challenge models and, potentially, in clinical samples. Such variations may result in different levels of virulence, morbidity and mortality.

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  1. Version published to 10.1186/s13073-020-00763-0
  2. ScreenIT

    SciScore for 10.1101/2020.03.22.002204: (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

    Experimental Models: Cell Lines
    SentencesResources
    Aliquots of each dilution were added to 1 × 104 Vero E6 cells in the same medium in each of 12 wells of a 96-well plate.
    Vero E6
    suggested: None
    Software and Algorithms
    SentencesResources
    All spectra were acquired using an Orbitrap Fusion Lumos mass spectrometer controlled by Xcalibur 4.1 software (Thermo Scientific) and operated in data-dependent acquisition mode.
    Xcalibur
    suggested: (Thermo Xcalibur, RRID:SCR_014593)
    Data Analysis: The raw data files were processed using Proteome Discoverer software v2.1
    Proteome Discoverer
    suggested: (Proteome Discoverer, RRID:SCR_014477)

    Results from OddPub: Thank you for sharing your data.

    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:
    • No conflict of interest statement was detected. If there are no conflicts, we encourage authors to explicit state so.
    • 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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  3. Version published to 10.1101/2020.03.22.002204v1 on bioRxiv