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Multiscale imaging and quantitative analysis of plasma membrane protein-cortical actin interplay
Aparajita Dasgupta1, Huong-Tra Ngo1, Deryl Tschoerner1
1Department of Biophysics, University of Texas Southwestern Medical Center, Dallas, Texas.
Biophysical Journal
|August 12, 2023
Summary
We developed a new imaging technique to directly link cortical actin (CA) structure and dynamics to cell surface protein organization without perturbing the cell. This method reveals how actin influences protein behavior, offering insights into cell signaling and processes.
Area of Science:
- Cell Biology
- Biophysics
- Biochemistry
Background:
- The spatiotemporal organization of cell surface receptors is crucial for effective cell signaling.
- Cortical actin (CA), beneath the plasma membrane (PM), significantly influences cell surface receptor organization.
- Previous studies using actin perturbation have limitations, including nonspecific effects and inability to quantify unperturbed actin architecture and dynamics.
Purpose of the Study:
- To establish direct relationships between CA architecture/dynamics and the spatiotemporal organization of PM proteins, including cell surface receptors.
- To overcome limitations of previous actin perturbation experiments by developing a non-disruptive imaging and analysis framework.
Main Methods:
- Developed a multiscale imaging and computational analysis framework integrating single-molecule imaging (SMI) of PM proteins and fluorescent speckle microscopy (FSM) of CA in live cells (SMI-FSM).
- Applied subcellular region analysis to investigate CA dynamics and PM protein mobility across different cellular locations.
- Examined the impact of global actin perturbation on local CA and PM protein properties.
Main Results:
- SMI-FSM demonstrated distinct relationships between PM proteins and CA, dependent on protein actin-binding ability, diffusion type, and local CA density.
- CA dynamics varied between cell edges and the cell center, predicting differences in PM protein mobility.
- Global actin perturbations did not always correlate with local CA changes near PM proteins, and PM protein responses varied with the local CA environment.
Conclusions:
- The integrated SMI-FSM framework provides a versatile tool for studying the influence of CA on PM protein organization under unperturbed conditions.
- Findings highlight the complex, context-dependent relationship between CA architecture/dynamics and PM protein spatiotemporal organization.
- This approach is expected to advance research on actin-dependent cellular processes and cell signaling.
