High throughput in vivo functional validation of candidate congenital heart disease genes in Drosophila

  1. Jun-yi Zhu
  2. Yulong Fu
  3. Margaret Nettleton
  4. Adam Richman
  5. Zhe Han  Is a corresponding author
  1. Children's National Medical Center, United States

Abstract

Genomic sequencing has implicated large numbers of genes and de novo mutations as potential disease risk factors. A high throughput in vivo model system is needed to validate gene associations with pathology. We developed a Drosophila-based functional system to screen candidate disease genes identified from Congenital Heart Disease (CHD) patients. 134 genes were tested in the Drosophila heart using RNAi-based gene silencing. Quantitative analyses of multiple cardiac phenotypes demonstrated essential structural, functional, and developmental roles for more than 70 genes, including a subgroup encoding histone H3K4 modifying proteins. We also demonstrated the use of Drosophila to evaluate cardiac phenotypes resulting from specific, patient-derived alleles of candidate disease genes. We describe the first high throughput in vivo validation system to screen candidate disease genes identified from patients. This approach has the potential to facilitate development of precision medicine approaches for CHD and other diseases associated with genetic factors.

Article and author information

Author details

  1. Jun-yi Zhu

    Center for Cancer and Immunology Research, Children's National Medical Center, Washington, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Yulong Fu

    Center for Cancer and Immunology Research, Children's National Medical Center, Washington, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Margaret Nettleton

    Center for Cancer and Immunology Research, Children's National Medical Center, Washington, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Adam Richman

    Center for Cancer and Immunology Research, Children's National Medical Center, Washington, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Zhe Han

    Center for Cancer and Immunology Research, Children's National Medical Center, Washington, United States
    For correspondence
    zhan@childrensnational.org
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5177-7798

Funding

National Heart, Lung, and Blood Institute (NIH R01)

  • Zhe Han

National Institute of Diabetes and Digestive and Kidney Diseases (NIH R01)

  • Zhe Han

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

Reviewing Editor

  1. Richard P Harvey, Victor Chang Cardiac Research Institute, Australia

Publication history

  1. Received: October 23, 2016
  2. Accepted: January 11, 2017
  3. Accepted Manuscript published: January 13, 2017 (version 1)
  4. Accepted Manuscript updated: January 20, 2017 (version 2)
  5. Version of Record published: February 9, 2017 (version 3)
  6. Version of Record updated: August 8, 2017 (version 4)

Copyright

© 2017, Zhu 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

  • 3,336
    Page views
  • 756
    Downloads
  • 31
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, 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)

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

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

  1. Jun-yi Zhu
  2. Yulong Fu
  3. Margaret Nettleton
  4. Adam Richman
  5. Zhe Han
(2017)
High throughput in vivo functional validation of candidate congenital heart disease genes in Drosophila
eLife 6:e22617.
https://doi.org/10.7554/eLife.22617

Further reading

    1. Developmental Biology
    2. Genetics and Genomics
    Mahdi Moradi Marjaneh, Edwin P Kirk ... Richard P Harvey
    Research Article

    Unlike single-gene mutations leading to Mendelian conditions, common human diseases are likely to be emergent phenomena arising from multilayer, multiscale and highly interconnected interactions. Atrial and ventricular septal defects are the most common forms of cardiac congenital anomalies in humans. Atrial septal defects (ASD) show an open communication between left and right atria postnatally, potentially resulting in serious hemodynamic consequences if untreated. A milder form of atrial septal defect, patent foramen ovale (PFO), exists in about one quarter of the human population, strongly associated with ischaemic stroke and migraine. The anatomic liabilities and genetic and molecular basis of atrial septal defects remain unclear. Here, we advance our previous analysis of atrial septal variation through quantitative trait locus (QTL) mapping of an advanced intercross line (AIL) established between the inbred QSi5 and 129T2/SvEms mouse strains, that show extremes of septal phenotypes. Analysis resolved 37 unique septal QTL with high overlap between QTL for distinct septal traits and PFO as a binary trait. Whole genome sequencing of parental strains and filtering identified predicted functional variants, including in known human congenital heart disease genes. Transcriptome analysis of developing septa revealed downregulation of networks involving ribosome, nucleosome, mitochondrial and extracellular matrix biosynthesis in the 129T2/SvEms strain, potentially reflecting an essential role for growth and cellular maturation in septal development. Analysis of variant architecture across different gene features, including enhancers and promoters, provided evidence for involvement of non-coding as well as protein coding variants. Our study provides the first high resolution picture of genetic complexity and network liability underlying common congenital heart disease, with relevance to human ASD and PFO.

    1. Developmental Biology
    2. Stem Cells and Regenerative Medicine
    Virginia folgado-marco, Kristina Ames ... Nicholas E Baker
    Research Article

    Ribosomal protein (Rp) gene haploinsufficiency can result in Diamond-Blackfan Anemia (DBA), characterized by defective erythropoiesis and skeletal defects. Some mouse Rp mutations recapitulate DBA phenotypes, although others lack erythropoietic or skeletal defects. We generated a conditional knockout mouse to partially delete Rps12. Homozygous Rps12 deletion resulted in embryonic lethality. Mice inheriting the Rps12+/- genotype had growth and morphological defects, pancytopenia and impaired erythropoiesis. A striking reduction in hematopoietic stem cells (HSCs) and progenitors in the bone marrow (BM) was associated with decreased ability to repopulate the blood system after competitive and non-competitive BM transplantation. Rps12+/- mutants lost HSC quiescence, experienced ERK and MTOR activation and increased global translation in HSC and progenitors. Post-natal heterozygous deletion of Rps12 in hematopoietic cells using Tal1-Cre-ERT also resulted in pancytopenia with decreased HSC numbers. However, post-natal Cre-ERT induction led to reduced translation in HSCs and progenitors, suggesting that this is the most direct consequence of Rps12 haploinsufficiency in hematopoietic cells. Thus, RpS12 has a strong requirement in HSC function, in addition to erythropoiesis.