Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction

  1. Alex Cornean
  2. Jakob Gierten
  3. Bettina Welz
  4. Juan Luis Mateo
  5. Thomas Thumberger
  6. Joachim Wittbrodt  Is a corresponding author
  1. Heidelberg University, Germany
  2. University of Oviedo, Spain

Abstract

Single nucleotide variants (SNVs) are prevalent genetic factors shaping individual trait profiles and disease susceptibility. The recent development and optimizations of base editors, rubber and pencil genome editing tools now promise to enable direct functional assessment of SNVs in model organisms. However, the lack of bioinformatic tools aiding target prediction limits the application of base editing in vivo. Here, we provide a framework for adenine and cytosine base editing in medaka (Oryzias latipes) and zebrafish (Danio rerio), ideal for scalable validation studies. We developed an online base editing tool ACEofBASEs (a careful evaluation of base-edits), to facilitate decision-making by streamlining sgRNA design and performing off-target evaluation. We used state-of-the-art adenine (ABE) and cytosine base editors (CBE) in medaka and zebrafish to edit eye pigmentation genes and transgenic GFP function with high efficiencies. Base editing in the genes encoding troponin T and the potassium channel ERG faithfully recreated known cardiac phenotypes. Deep-sequencing of alleles revealed the abundance of intended edits in comparison to low levels of insertion or deletion (indel) events for ABE8e and evoBE4max. We finally validated missense mutations in novel candidate genes of congenital heart disease (CHD) dapk3, ube2b, usp44, and ptpn11 in F0 and F1 for a subset of these target genes with genotype-phenotype correlation. This base editing framework applies to a wide range of SNV-susceptible traits accessible in fish, facilitating straight-forward candidate validation and prioritization for detailed mechanistic downstream studies.

Data availability

Source code for ACEofBASEs has been provided.

Article and author information

Author details

  1. Alex Cornean

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3727-7057
  2. Jakob Gierten

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  3. Bettina Welz

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  4. Juan Luis Mateo

    Deparment of Computer Science, University of Oviedo, Oviedo, Spain
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9902-6048
  5. Thomas Thumberger

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8485-457X
  6. Joachim Wittbrodt

    Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany
    For correspondence
    jochen.wittbrodt@cos.uni-heidelberg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8550-7377

Funding

Deutsche Forschungsgemeinschaft (WI 1824/9-1)

  • Joachim Wittbrodt

H2020 European Research Council (810172)

  • Joachim Wittbrodt

Deutsches Zentrum für Herz-Kreislaufforschung

  • Joachim Wittbrodt

Deutsche Herzstiftung (S/02/17)

  • Jakob Gierten

Joachim Herz Stiftung

  • Jakob Gierten

Deutsche Forschungsgemeinschaft (3DMM2O)

  • Joachim Wittbrodt

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Stephen C Ekker, Mayo Clinic, United States

Version history

  1. Received: July 12, 2021
  2. Preprint posted: July 26, 2021 (view preprint)
  3. Accepted: March 21, 2022
  4. Accepted Manuscript published: April 4, 2022 (version 1)
  5. Version of Record published: April 22, 2022 (version 2)

Copyright

© 2022, Cornean et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Alex Cornean
  2. Jakob Gierten
  3. Bettina Welz
  4. Juan Luis Mateo
  5. Thomas Thumberger
  6. Joachim Wittbrodt
(2022)
Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction
eLife 11:e72124.
https://doi.org/10.7554/eLife.72124

Share this article

https://doi.org/10.7554/eLife.72124

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