1. Developmental Biology
Download icon

NKX2-5 mutations causative for congenital heart disease retain functionality and are directed to hundreds of targets

  1. Romaric Bouveret  Is a corresponding author
  2. Ashley J Waardenberg
  3. Nicole Schonrock
  4. Mirana Ramialison
  5. Tram Doan
  6. Danielle de Jong
  7. Antoine Bondue
  8. Gurpreet Kaur
  9. Stephanie Mohamed
  10. Hananeh Fonoudi
  11. Chiann-mun Chen
  12. Merridee Wouters
  13. Shoumo Bhattacharya
  14. Nicolas Plachta
  15. Sally L Dunwoodie
  16. Gavin Chapman
  17. Cédric Blanpain
  18. Richard P Harvey
  1. Victor Chang Cardiac Research Institute, Australia
  2. Université Libre de Bruxelles, Belgium
  3. Monash University, Australia
  4. University of Oxford, United Kingdom
Research Article
  • Cited 32
  • Views 2,934
  • Annotations
Cite this article as: eLife 2015;4:e06942 doi: 10.7554/eLife.06942

Abstract

To model cardiac gene regulatory networks in health and disease we used DamID to establish robust target gene sets for the cardiac homeodomain factor NKX2-5 and two congenital heart disease-associated mutants carrying a crippled homeodomain, which normally functions as DNA- and protein-binding interface. Despite compromised direct DNA-binding, NKX2-5 mutants retained partial functionality and bound hundreds of targets, including NKX2-5 wild type targets and unique sets of 'off-targets'. NKX2-5∆HD, which lacks the entire homeodomain, could still dimerise with wild type NKX2-5 and its cofactors, including newly-discovered cofactors of the ETS family, through the conserved tyrosine-rich domain (YRD). NKX2-5∆HD off-targets showed overrepresentation of many binding motifs, including ETS motifs, the majority co-occupied by ETS proteins as determined by DamID. Off-targets of an NKX2-5 YRD mutant were not enriched in ETS targets. Our study reveals off-target binding and transcriptional activity for NKX2-5 mutations driven in part by cofactor interactions, suggesting a novel type of gain-of-function in congenital heart disease.

Article and author information

Author details

  1. Romaric Bouveret

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    For correspondence
    r.bouveret@victorchang.edu.au
    Competing interests
    The authors declare that no competing interests exist.

  2. Ashley J Waardenberg

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  3. Nicole Schonrock

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  4. Mirana Ramialison

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  5. Tram Doan

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  6. Danielle de Jong

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  7. Antoine Bondue

    Institut de Recherche Interdisciplinaire en Biologie Humaine et Moléculaire, Université Libre de Bruxelles, Brussels, Belgium
    Competing interests
    The authors declare that no competing interests exist.

  8. Gurpreet Kaur

    European Molecular Biology Laboratory, Australian Regenerative Medicine Institute, Monash University, Clayton, Australia
    Competing interests
    The authors declare that no competing interests exist.

  9. Stephanie Mohamed

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  10. Hananeh Fonoudi

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  11. Chiann-mun Chen

    Department of Cardiovascular Medicine, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  12. Merridee Wouters

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  13. Shoumo Bhattacharya

    Department of Cardiovascular Medicine, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  14. Nicolas Plachta

    European Molecular Biology Laboratory, Australian Regenerative Medicine Institute, Monash University, Clayton, Australia
    Competing interests
    The authors declare that no competing interests exist.

  15. Sally L Dunwoodie

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  16. Gavin Chapman

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

  17. Cédric Blanpain

    Institut de Recherche Interdisciplinaire en Biologie Humaine et Moléculaire, Université Libre de Bruxelles, Brussels, Belgium
    Competing interests
    The authors declare that no competing interests exist.

  18. Richard P Harvey

    Victor Chang Cardiac Research Institute, Darlinghurst, Australia
    Competing interests
    The authors declare that no competing interests exist.

Ethics

Animal experimentation: Animal experimentation was performed with approval of the Garvan Institute/St Vincent's Hospital Animal Ethics Committee (Project numbers 10/19 and 10/01).

Reviewing Editor

  1. Margaret Buckingham, Institut Pasteur, France

Publication history

  1. Received: February 10, 2015
  2. Accepted: July 5, 2015
  3. Accepted Manuscript published: July 6, 2015 (version 1)
  4. Version of Record published: August 25, 2015 (version 2)

Copyright

© 2015, Bouveret 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.

Metrics

  • 2,934
    Page views
  • 629
    Downloads
  • 32
    Citations

Article citation count generated by polling the highest count across the following sources: Scopus, Crossref, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Categories and tags

Research organism

    1. Related to

    Genetic Diseases: Many ways to break a heart

    Megan Rowton, Ivan P Moskowitz
  2. Further reading

Further reading

    1. Developmental Biology

    Genetic Diseases: Many ways to break a heart

    Megan Rowton, Ivan P Moskowitz
    Insight

    A mutant transcription factor that has been linked to congenital heart disease has wider effects than previously thought.

    1. Developmental Biology
    2. Genetics and Genomics

    Cell competition removes segmental aneuploid cells from Drosophila imaginal disc-derived tissues based on ribosomal protein gene dose

    Zhejun Ji et al.
    Research Article

    Aneuploidy causes birth defects and miscarriages, occurs in nearly all cancers and is a hallmark of aging. Individual aneuploid cells can be eliminated from developing tissues by unknown mechanisms. Cells with ribosomal protein (Rp) gene mutations are also eliminated, by cell competition with normal cells. Because Rp genes are spread across the genome, their copy number is a potential marker for aneuploidy. We found that elimination of imaginal disc cells with irradiation-induced genome damage often required cell competition genes. Segmentally aneuploid cells derived from targeted chromosome excisions were eliminated by the RpS12-Xrp1 cell competition pathway if they differed from neighboring cells in Rp gene dose, whereas cells with normal doses of the Rp and eIF2γ genes survived and differentiated adult tissues. Thus, cell competition, triggered by differences in Rp gene dose between cells, is a significant mechanism for the elimination of aneuploid somatic cells, likely to contribute to preventing cancer.

    1. Developmental Biology
    2. Neuroscience

    Robo recruitment of the Wave Regulatory Complex plays an essential and conserved role in midline repulsion

    Karina Chaudhari et al.
    Research Article

    The Roundabout (Robo) guidance receptor family induces axon repulsion in response to its ligand Slit by inducing local cytoskeletal changes; however, the link to the cytoskeleton and the nature of these cytoskeletal changes are poorly understood. Here, we show that the heteropentameric Scar/Wave Regulatory Complex (WRC) which drives Arp2/3-induced branched actin polymerization, is a direct effector of Robo signaling. Biochemical evidence shows that Slit triggers WRC recruitment to the Robo receptor's WIRS motif. In Drosophila embryos, mutants of the WRC enhance Robo1-dependent midline crossing defects. Additionally, mutating Robo1's WIRS motif significantly reduces receptor activity in rescue assays in vivo, and CRISPR-Cas9 mutagenesis shows that the WIRS motif is essential for endogenous Robo1 function. Finally, axon guidance assays in mouse dorsal spinal commissural axons and gain-of-function experiments in chick embryos demonstrate that the WIRS motif is also required for Robo1 repulsion in mammals. Together, our data support an essential conserved role for the WIRS-WRC interaction in Robo1-mediated axon repulsion.