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A GFP Complementation-based Dual-expression System for Assessing Cell-Cell Contact Mediated by Cytonemes in Live Drosophila Wing Imaginal Discs
Published on: August 22, 2025
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A GFP Complementation-based Dual-expression System for Assessing Cell-Cell Contact Mediated by Cytonemes in Live
Mathieu Joseph1, David R Hipfner2
1Institut de recherches cliniques de Montréal (IRCM); Programmes de biologie moléculaire, Université de Montréal.
Journal of Visualized Experiments : Jove
|September 8, 2025
Summary
Researchers developed a novel method to quantify cytoneme-mediated cell contacts in Drosophila wing imaginal discs. This technique uses a split-GFP system to visualize and measure signaling filopodia interactions between distinct epithelial layers.
Area of Science:
- Developmental Biology
- Cell Biology
- Molecular Biology
Background:
- Embryonic tissue development relies on local cell-to-cell signaling.
- Cytonemes are specialized filopodia facilitating signal exchange between cells.
- Understanding cytoneme function is crucial for deciphering developmental patterning.
Purpose of the Study:
- To present a protocol for quantifying cytoneme-mediated contacts between Drosophila wing disc epithelial cells.
- To enable the study of factors influencing cytoneme formation and function.
- To visualize signaling filopodia interactions across the disc lumen.
Main Methods:
- Utilized a split-GFP reconstitution assay in live Drosophila wing imaginal discs.
- Employed GAL4-UAS and LexA-LexAop systems to express complementary GFP fragments.
- Quantified reconstituted GFP fluorescence using confocal microscopy on live wing disc preparations.
Main Results:
- Successfully quantified cytoneme-mediated cell contacts between disc proper (DP) and peripodial membrane (PerM) epithelia.
- Established a system for live imaging and quantification of signaling filopodia.
- Demonstrated the ability to manipulate gene expression in specific cell populations to assess effects on cell contacts.
Conclusions:
- The developed GFP reconstitution protocol provides a robust method for studying cytoneme-mediated cell interactions.
- This system facilitates the identification of molecular regulators of cytoneme formation and function.
- The adaptable protocol can be applied to other tissues and developmental contexts.
