Generation of a versatile BiFC ORFeome library for analyzing protein-protein interactions in live Drosophila

  1. Johannes Bischof
  2. Marilyne Duffraisse
  3. Edy Furger
  4. Leiore Ajuria
  5. Guillaume Giraud
  6. Solene Vanderperre
  7. Rachel Paul
  8. Mikael Björklund
  9. Damien Ahr
  10. Alexis W Ahmed
  11. Lionel Spinelli
  12. Christine Brun
  13. Konrad Basler
  14. Samir Merabet  Is a corresponding author
  1. University of Zurich, Switzerland
  2. Institut de Génomique Fonctionnelle de Lyon, France
  3. Zhejiang University, China
  4. Aix Marseille University, France

Abstract

Transcription factors achieve specificity by establishing intricate interaction networks that will change depending on the cell context. Capturing these interactions in live condition is however a challenging issue that requires sensitive and non-invasive methods. We present a set of fly lines, called 'multicolor BiFC library', which covers most of the Drosophila transcription factors for performing Bimolecular Fluorescence Complementation (BiFC). The multicolor BiFC library can be used to probe two different binary interactions simultaneously and is compatible for large-scale interaction screens. The library can also be coupled with established Drosophila genetic resources to analyze interactions in the developmentally relevant expression domain of each protein partner. We provide proof of principle experiments of these various applications, using Hox proteins in the live Drosophila embryo as a case study. Overall this novel collection of ready-to-use fly lines constitutes an unprecedented genetic toolbox for the identification and analysis of protein-protein interactions in vivo.

Data availability

Fly lines generated for the project have been deposited to the FlyORF library and are available upon request to FlyORF (https://flyorf.ch/index.php/orf-collection). The numerical, processed data used for this study is provided in the manuscript, figures and supplementary files.

Article and author information

Author details

  1. Johannes Bischof

    Institute of Molecular Life Sciences, University of Zurich, Zürich, Switzerland
    Competing interests
    Johannes Bischof, involved in maintaining and distributing the fly lines via the not-for-profit FlyORF project. There are no other competing interests to declare.
  2. Marilyne Duffraisse

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  3. Edy Furger

    Institute of Molecular Life Sciences, University of Zurich, Zürich, Switzerland
    Competing interests
    No competing interests declared.
  4. Leiore Ajuria

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  5. Guillaume Giraud

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  6. Solene Vanderperre

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  7. Rachel Paul

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  8. Mikael Björklund

    Zhejiang University-University of Edinburgh Institute, Zhejiang University, Haining, China
    Competing interests
    Mikael Björklund, involved in the development of the FlyORF resource. There are no other competing interests to declare.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2176-681X
  9. Damien Ahr

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  10. Alexis W Ahmed

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    Competing interests
    No competing interests declared.
  11. Lionel Spinelli

    TAGC U1090, Aix Marseille University, Marseille, France
    Competing interests
    No competing interests declared.
  12. Christine Brun

    TAGC U1090, Aix Marseille University, Marseille, France
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5563-6765
  13. Konrad Basler

    Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland
    Competing interests
    Konrad Basler, involved in maintaining and distributing the fly lines via the not-for-profit FlyORF project. There are no other competing interests to declare.
  14. Samir Merabet

    ENS Lyon UMR5242, Institut de Génomique Fonctionnelle de Lyon, Lyon, France
    For correspondence
    samir.merabet@ens-lyon.fr
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7629-703X

Funding

Fondation pour la Recherche Médicale (1122556)

  • Johannes Bischof
  • Marilyne Duffraisse
  • Edy Furger
  • Leiore Ajuria
  • Guillaume Giraud
  • Solene Vanderperre
  • Rachel Paul
  • Samir Merabet

Cefipra

  • Johannes Bischof
  • Marilyne Duffraisse
  • Edy Furger
  • Leiore Ajuria
  • Guillaume Giraud
  • Solene Vanderperre
  • Rachel Paul
  • Samir Merabet

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

Copyright

© 2018, Bischof 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

  • 5,664
    views
  • 853
    downloads
  • 41
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

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. Johannes Bischof
  2. Marilyne Duffraisse
  3. Edy Furger
  4. Leiore Ajuria
  5. Guillaume Giraud
  6. Solene Vanderperre
  7. Rachel Paul
  8. Mikael Björklund
  9. Damien Ahr
  10. Alexis W Ahmed
  11. Lionel Spinelli
  12. Christine Brun
  13. Konrad Basler
  14. Samir Merabet
(2018)
Generation of a versatile BiFC ORFeome library for analyzing protein-protein interactions in live Drosophila
eLife 7:e38853.
https://doi.org/10.7554/eLife.38853

Share this article

https://doi.org/10.7554/eLife.38853

Further reading

    1. Chromosomes and Gene Expression
    2. Genetics and Genomics
    Omid Gholamalamdari, Tom van Schaik ... Andrew S Belmont
    Research Article

    Models of nuclear genome organization often propose a binary division into active versus inactive compartments yet typically overlook nuclear bodies. Here, we integrated analysis of sequencing and image-based data to compare genome organization in four human cell types relative to three different nuclear locales: the nuclear lamina, nuclear speckles, and nucleoli. Although gene expression correlates mostly with nuclear speckle proximity, DNA replication timing correlates with proximity to multiple nuclear locales. Speckle attachment regions emerge as DNA replication initiation zones whose replication timing and gene composition vary with their attachment frequency. Most facultative LADs retain a partially repressed state as iLADs, despite their positioning in the nuclear interior. Knock out of two lamina proteins, Lamin A and LBR, causes a shift of H3K9me3-enriched LADs from lamina to nucleolus, and a reciprocal relocation of H3K27me3-enriched partially repressed iLADs from nucleolus to lamina. Thus, these partially repressed iLADs appear to compete with LADs for nuclear lamina attachment with consequences for replication timing. The nuclear organization in adherent cells is polarized with nuclear bodies and genomic regions segregating both radially and relative to the equatorial plane. Together, our results underscore the importance of considering genome organization relative to nuclear locales for a more complete understanding of the spatial and functional organization of the human genome.

    1. Cell Biology
    2. Genetics and Genomics
    Keva Li, Nicholas Tolman ... UK Biobank Eye and Vision Consortium
    Research Article

    A glaucoma polygenic risk score (PRS) can effectively identify disease risk, but some individuals with high PRS do not develop glaucoma. Factors contributing to this resilience remain unclear. Using 4,658 glaucoma cases and 113,040 controls in a cross-sectional study of the UK Biobank, we investigated whether plasma metabolites enhanced glaucoma prediction and if a metabolomic signature of resilience in high-genetic-risk individuals existed. Logistic regression models incorporating 168 NMR-based metabolites into PRS-based glaucoma assessments were developed, with multiple comparison corrections applied. While metabolites weakly predicted glaucoma (Area Under the Curve = 0.579), they offered marginal prediction improvement in PRS-only-based models (p=0.004). We identified a metabolomic signature associated with resilience in the top glaucoma PRS decile, with elevated glycolysis-related metabolites—lactate (p=8.8E-12), pyruvate (p=1.9E-10), and citrate (p=0.02)—linked to reduced glaucoma prevalence. These metabolites combined significantly modified the PRS-glaucoma relationship (Pinteraction = 0.011). Higher total resilience metabolite levels within the highest PRS quartile corresponded to lower glaucoma prevalence (Odds Ratiohighest vs. lowest total resilience metabolite quartile=0.71, 95% Confidence Interval = 0.64–0.80). As pyruvate is a foundational metabolite linking glycolysis to tricarboxylic acid cycle metabolism and ATP generation, we pursued experimental validation for this putative resilience biomarker in a human-relevant Mus musculus glaucoma model. Dietary pyruvate mitigated elevated intraocular pressure (p=0.002) and optic nerve damage (p<0.0003) in Lmx1bV265D mice. These findings highlight the protective role of pyruvate-related metabolism against glaucoma and suggest potential avenues for therapeutic intervention.