Genome-wide CRISPR screening identifies TMEM106B as a proviral host factor for SARS-CoV-2

  1. Jim Baggen
  2. Leentje Persoons
  3. Els Vanstreels
  4. Sander Jansen
  5. Dominique Van Looveren
  6. Bram Boeckx
  7. Vincent Geudens
  8. Julie De Man
  9. Dirk Jochmans
  10. Joost Wauters
  11. Els Wauters
  12. Bart M. Vanaudenaerde
  13. Diether Lambrechts
  14. Johan Neyts
  15. Kai Dallmeier
  16. Hendrik Jan Thibaut
  17. Maarten Jacquemyn
  18. Piet Maes
  19. Dirk Daelemans

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

No abstract available

Article activity feed

  1. Version published to 10.1038/s41588-021-00805-2
  2. ScreenIT

    SciScore for 10.1101/2020.09.28.316281: (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
    Employed primary antibodies were rabbit anti-LC3B (L7543, Sigma) at a 1:200 dilution and mouse anti-dsRNA (J2, Scicons) at a 1:1000 dilution.
    anti-LC3B
    suggested: (Sigma-Aldrich Cat# L7543, RRID:AB_796155)
    anti-dsRNA
    suggested: (Millipore Cat# MABE1134, RRID:AB_2819101)
    Secondary antibodies Alexa Fluor® 568 goat anti-rabbit (A11011, Invitrogen, ThermoFisher Scientific) and Alexa Fluor® 488 goat anti-mouse (A11029, Invitrogen, ThermoFisher Scientific) were diluted 1:500.
    anti-rabbit
    suggested: (Molecular Probes Cat# A-11011, RRID:AB_143157)
    A11011
    suggested: (Molecular Probes Cat# A-11011, RRID:AB_143157)
    anti-mouse
    suggested: (Molecular Probes Cat# A-11029, RRID:AB_138404)
    A11029
    suggested: (Molecular Probes Cat# A-11029, RRID:AB_138404)
    Proteins were separated by size (12-230 kDa) and visualized on a Wes system (ProteinSimple, San Jose, CA, USA) with an anti-mouse or anti-goat IgG-HRP antibody (R&D systems, HAF109) detecting the primary antibody against GAPDH (Santa Cruz Biotechnology, sc-47724) or anti-hACE2 (R&D systems, AF933), respectively.
    anti-goat
    suggested: (R and D Systems Cat# HAF109, RRID:AB_357236)
    GAPDH
    suggested: (Santa Cruz Biotechnology Cat# sc-47724, RRID:AB_627678)
    anti-hACE2
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Cell culture: HEK293T (received from prof
    HEK293T
    suggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)
    Vero E6, Huh-7 (CLS - 300156; human hepatoblastoma), Hep3B (ATCC HB-8064; human hepatocellular carcinoma) and HRT-18G (ATCC CRL-11663; human colorectal adenocarcinoma) were maintained in Dulbecco’s Modified Eagle Medium (DMEM, Gibco Life Technologies) supplemented with 8% heat-inactivated fetal bovine serum (HyClone, GE Healthcare Life Sciences), 0.075% sodium bicarbonate (Gibco Life Technologies) and 1mM sodium pyruvate (Gibco Life Technologies)
    Huh-7
    suggested: None
    Hep3B
    suggested: None
    A549 cells were maintained in F-12K medium supplemented with 10% heat-inactivated fetal bovine serum.
    A549
    suggested: None
    NCI-H1975 (ATCC-CRL-5908) and NCI-H2110 (ATCC-CRL-5924) cells were maintained in RPMI medium supplemented with 10% heat-inactivated fetal bovine serum.
    NCI-H1975
    suggested: ATCC Cat# CRL-5908, RRID:CVCL_1511)
    NCI-H2110
    suggested: ATCC Cat# CRL-5924, RRID:CVCL_1530)
    The HCoV-229E (ATCC VR-740) and HCoV-OC43 (ATCC VR-1558) virus stocks were obtained by inoculating a confluent monolayer of Huh7 or HRT-18G cells, respectively.
    HCoV-229E
    suggested: None
    HCoV-OC43
    suggested: None
    For the SARS-CoV-2 low stringency screen, 1.5 x 108 Huh7 cells for each of two replicates were transduced at a MOI of ~0.2 with lentivirus containing the Brunello library and selected with puromycin for three days.
    Huh7
    suggested: CLS Cat# 300156/p7178_HuH7, RRID:CVCL_0336)
    Virus inhibition assays: The antiviral activity of PI3K type 3 inhibitors on SARS-CoV-2 in Vero E6 cells was evaluated as follows: on day −1, the test compounds were serially diluted in DMEM (Gibco cat no 41965-039) supplemented with 2% v/v heat-inactivated FCS and sodium bicarbonate (Gibco 25080-060)
    Vero E6
    suggested: None
    Software and Algorithms
    SentencesResources
    Protein signals were visualized and quantified with the Compass software, v4.0.0
    Compass
    suggested: (COMPASS, RRID:SCR_015874)

    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.

  3. Version published to 10.1101/2020.09.28.316281v1 on bioRxiv