Related Experiment Video
Updated: Dec 21, 2025
Methods to Study Mrp4-containing Macromolecular Complexes in the Regulation of Fibroblast Migration
Published on: May 19, 2016
Imaging the fibroblast growth factor receptor network on the plasma membrane with DNA-assisted single-molecule
Mark S Schröder1, Marie-Lena I E Harwardt1, Johanna V Rahm1
1Institute of Physical and Theoretical Chemistry, Goethe-University Frankfurt, Max-von-Laue-Str. 7, 60438 Frankfurt, Germany.
Abstract:
Fibroblast growth factor receptors (FGFRs) are a subfamily of receptor tyrosine kinases and central players in health and disease. Following ligand binding and the formation of homo- and heteromeric complexes, FGFRs initiate a cellular response. Challenges in studying FGFR activation are inner-subfamily interactions and a complex heterogeneity of these in the cell membrane, which demand for observation techniques that can resolve individual protein complexes and that are compatible with endogenous protein levels. Here, we established an imaging and analysis pipeline for multiplexed single-molecule localization microscopy (SMLM) of the FGFR network at the plasma membrane. Using DNA-labeled primary antibodies, we visualize all four FGFRs in the same cell with near-molecular spatial resolution. From the super-resolution imaging data, we extract information on FGFR density, spatial distribution, and inner-subfamily colocalization. Our approach is straightforward and easily adaptable to other multiplexed SMLM data of membrane proteins.
Insights
We developed a new imaging technique to study Fibroblast Growth Factor Receptors (FGFRs) in cells. This method visualizes individual FGFRs and their interactions, offering insights into cell signaling pathways.
Area of Science:
- Cell Biology
- Molecular Biology
- Biophysics
Background:
- Fibroblast Growth Factor Receptors (FGFRs) are crucial receptor tyrosine kinases involved in cellular processes and disease.
- Studying FGFR activation is challenging due to complex interactions and membrane heterogeneity.
- Existing methods lack the resolution to observe individual FGFR complexes at endogenous levels.
Purpose of the Study:
- To establish a novel imaging and analysis pipeline for multiplexed single-molecule localization microscopy (SMLM) of the FGFR network.
- To visualize and quantify FGFR density, distribution, and inner-subfamily colocalization at the plasma membrane.
- To provide a versatile tool for studying other membrane protein networks.
Main Methods:
- Developed a multiplexed SMLM pipeline using DNA-labeled primary antibodies.
- Visualized all four FGFRs within the same cell with near-molecular spatial resolution.
- Analyzed super-resolution imaging data to extract FGFR network characteristics.
Main Results:
- Successfully visualized individual FGFRs and their complexes on the cell membrane.
- Quantified FGFR density, spatial distribution, and colocalization patterns within the FGFR subfamily.
- Demonstrated the pipeline's compatibility with endogenous protein levels.
Conclusions:
- The established SMLM pipeline enables high-resolution visualization of the FGFR network.
- This approach provides novel insights into FGFR interactions and cellular signaling.
- The method is adaptable for studying diverse membrane protein systems.
Related Concept Videos
01:37Super-resolution Fluorescence Microscopy
01:19Protein Dynamics in Living Cells
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...