In vivo structure and dynamics of the SARS-CoV-2 RNA genome

  1. Yan Zhang
  2. Kun Huang
  3. Dejian Xie
  4. Jian You Lau
  5. Wenlong Shen
  6. Ping Li
  7. Dong Wang
  8. Zhong Zou
  9. Shu Shi
  10. Hongguang Ren
  11. Youliang Wang
  12. Youzhi Mao
  13. Meilin Jin
  14. Grzegorz Kudla
  15. Zhihu Zhao

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Abstract

The dynamics of SARS-CoV-2 RNA structure and their functional relevance are largely unknown. Here we develop a simplified SPLASH assay and comprehensively map the in vivo RNA-RNA interactome of SARS-CoV-2 genome across viral life cycle. We report canonical and alternative structures including 5′-UTR and 3′-UTR, frameshifting element (FSE) pseudoknot and genome cyclization in both cells and virions. We provide direct evidence of interactions between Transcription Regulating Sequences, which facilitate discontinuous transcription. In addition, we reveal alternative short and long distance arches around FSE. More importantly, we find that within virions, while SARS-CoV-2 genome RNA undergoes intensive compaction, genome domains remain stable but with strengthened demarcation of local domains and weakened global cyclization. Taken together, our analysis reveals the structural basis for the regulation of replication, discontinuous transcription and translational frameshifting, the alternative conformations and the maintenance of global genome organization during the whole life cycle of SARS-CoV-2, which we anticipate will help develop better antiviral strategies.

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  1. Version published to 10.1038/s41467-021-25999-1
  2. Version published to 10.1101/2021.01.15.426526v2 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2021.01.15.426526: (What is this?)

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

    Table 1: Rigor

    NIH rigor criteria are not applicable to paper type.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    Cell lines: Chlorocebus sabaeus (Green monkey) VeroE6 (female, RRID:CVCL_YQ49) were purchased from American Type Culture Collection (ATCC, id: ATCC CRL-1586).
    detected: (JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
    Vero E6 and VeroE6/TMPRSS2 cells were cultured in Dulbecco’s modified Eagles medium (DMEM) supplemented with 10% fetal bovine serum at 37 °C in a humidified CO2 incubator.
    VeroE6/TMPRSS2
    suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
    VeroE6 cells were inoculated with SARS-CoV-2 strain Wuhan-Hu-1 at MOI=0.01 pfu/cell for 24 hours.
    VeroE6
    suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
    Experimental Models: Organisms/Strains
    SentencesResources
    SARS-CoV-2 virus strain Wuhan-Hu-1 was kindly provided by (Wuhan institute of viology).
    Wuhan-Hu-1
    suggested: None
    Software and Algorithms
    SentencesResources
    Viewpoint histograms and arc plots were plotted with ggplot2 R package [43].
    ggplot2
    suggested: (ggplot2, RRID:SCR_014601)
    For the spatial location of particular gene loci, we used 1 point/20 balls to calculate the position of specific genes in the whole 3D simulation, and then modify the pymol results by a python script.
    pymol
    suggested: (PyMOL, RRID:SCR_000305)
    Differential interaction identifying: Differential gene expression analysis was performed using DESeq2 [32].
    DESeq2
    suggested: (DESeq, RRID:SCR_000154)

    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:
    • 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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  4. Version published to 10.1101/2021.01.15.426526v1 on bioRxiv