Related Experiment Video
Updated: Feb 25, 2026
Methods to Study Mrp4-containing Macromolecular Complexes in the Regulation of Fibroblast Migration
Published on: May 19, 2016
Cells lay their own tracks - optogenetic Cdc42 activation stimulates fibronectin deposition supporting directed
Seth P Zimmerman1,2, Sreeja B Asokan1,3, Brian Kuhlman1,2
1UNC Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Abstract:
Rho GTPase family members are known regulators of directed migration and therefore play key roles in processes including development, the immune response and cancer metastasis. However, their individual contributions to these processes are complex. Here, we modify the activity of the two Rho GTPase family members Rac and Cdc42 by optogenetically recruiting specific guanine nucleotide exchange factor (GEF) DH or PH domains to defined regions of the cell membrane. We find that the localized activation of both GTPases produces lamellipodia in cells plated on a fibronectin substrate. By using a novel optotaxis assay, we show that biased activation can drive directional migration. Interestingly, in the absence of exogenous fibronectin, Rac activation is insufficient to produce stable lamellipodia or directional migration whereas Cdc42 activation is sufficient for these processes. We find that a remarkably small amount of fibronectin (<10 puncta per protrusion) is necessary to support stable GTPase-driven lamellipodia formation. Cdc42 bypasses the need for exogenous fibronectin by stimulating cellular fibronectin deposition under the newly formed lamellipodia.This article has an associated First Person interview with the first author of the paper.
Insights
Localized activation of Rac and Cdc42 GTPases drives cell migration. Cdc42, but not Rac, can promote directional movement and lamellipodia formation even with minimal fibronectin by stimulating cellular fibronectin deposition.
Area of Science:
- Cell biology
- Molecular biology
- Biophysics
Background:
- Rho GTPases (Rac, Cdc42) regulate cell migration, crucial for development, immunity, and cancer metastasis.
- Understanding individual Rho GTPase contributions to directed cell movement remains complex.
Purpose of the Study:
- To investigate the distinct roles of Rac and Cdc42 in directed cell migration using optogenetic control.
- To determine the role of fibronectin in GTPase-mediated lamellipodia formation and cell motility.
Main Methods:
- Optogenetic recruitment of guanine nucleotide exchange factor (GEF) domains to activate specific Rho GTPases (Rac, Cdc42) at the cell membrane.
- Development of a novel optotaxis assay to measure light-guided directional cell migration.
- Quantification of fibronectin puncta and analysis of cellular fibronectin deposition.
Main Results:
- Localized activation of both Rac and Cdc42 induced lamellipodia on fibronectin-coated substrates.
- Biased activation of these GTPases using optogenetics successfully drove directional cell migration.
- Cdc42 activation promoted stable lamellipodia and directional migration even without exogenous fibronectin, unlike Rac.
- Cdc42 bypassed the need for exogenous fibronectin by inducing cellular fibronectin deposition.
Conclusions:
- Optogenetic activation of Rho GTPases provides a powerful tool to dissect their roles in cell migration.
- Cdc42 plays a critical role in initiating and sustaining cell migration, potentially by modulating the extracellular matrix.
- Cellular fibronectin deposition is a key mechanism by which Cdc42 can drive migration in fibronectin-poor environments.
Related Concept Videos
01:19Cell Migration
01:09Cell Migration
01:39Mechanism of Filopodia Formation
Their main function is to guide migrating cells during normal tissue morphogenesis or cancer metastasis by recognizing and making initial contacts with the extracellular matrix. However, they can also act as stationary cell anchors or help to establish communication...
01:13Actin Polymerization and Cell Motility
Actin cytoskeleton dynamics can produce pushing, pulling, and resistance forces that help the cell to migrate....
01:32Cytoskeletal Coordination in Cell Migration
01:21Cell Polarization by Rho Proteins