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Imaging and quantifying organelle contact sites using a reversible splitFAST complementation system.

Jason A Weesner1, Chi-Lun Chang1

  • 1Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, TN, United States.

Methods in Enzymology
|March 1, 2026
PubMed
Summary
This summary is machine-generated.

The FABCON toolkit enables visualization and quantification of organelle contact sites in living cells. This method allows researchers to study the dynamic regulation of these crucial cellular communication hubs.

Keywords:
Bimolecular fluorescence complementationLipid dropletsMitochondriaOrganelle contact sitesSplitFAST

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Area of Science:

  • Cell Biology
  • Molecular Imaging
  • Biophysics

Background:

  • Organelle contact sites are vital for cellular functions but challenging to visualize in living cells.
  • Existing methods struggle to dynamically track these sub-micro scale interactions.
  • Understanding organelle communication is key to deciphering cellular logistics.

Purpose of the Study:

  • To introduce the FABCON (Fluorogen-Activated Bimolecular complementation at CONtact sites) toolkit for detecting and quantifying organelle contact sites.
  • To provide a customizable and robust platform for studying dynamic organelle interactions.
  • To enable detailed analysis of specific contact sites, such as mitochondria-lipid droplet interactions.

Main Methods:

  • FABCON utilizes reversible, fluorogen-dependent complementation of the splitFAST system to label contact sites.
  • The protocol details the engineering principles for customizing FABCON to different model systems.
  • Methods include intensity-based measurement and line scanning for quantification in mammalian cells.

Main Results:

  • FABCON successfully visualizes and quantifies organelle contact sites, demonstrated with mitochondria-lipid droplet interactions.
  • The toolkit allows for dynamic monitoring of these interactions in living cells.
  • Quantitative measurements of contact site levels are achievable through established imaging techniques.

Conclusions:

  • FABCON offers a versatile and robust platform for the study of organelle contact sites.
  • This toolkit can be broadly applied to various contact site types with appropriate validation.
  • FABCON facilitates the investigation of the dynamic regulation of inter-organelle communication.