The RhoGTPases are characterized as membrane-associated molecular switches that cycle between active, GTP-bound and inactive, GDP-bound states. However, 90-95% of RhoGTPases are maintained in a soluble form by RhoGDI, which is generally viewed as a passive shuttle for inactive RhoGTPases. Our current understanding of RhoGTPase:RhoGDI dynamics has been limited by two experimental challenges: direct visualization of the RhoGTPases in vivo and reconstitution of the cycle in vitro. We developed methods to directly image vertebrate RhoGTPases in vivo or on lipid bilayers in vitro. Using these methods, we identified pools of active and inactive RhoGTPase associated with the membrane, found that RhoGDI can extract both inactive and active RhoGTPases, and found that extraction of active RhoGTPase contributes to their spatial regulation around cell wounds. These results indicate that RhoGDI directly contributes to the spatiotemporal patterning of RhoGTPases by removing active RhoGTPases from the plasma membrane.
All quantifications made in this study are included as source data files by figure.
- William M Bement
- Adriana E Golding
- Peter Bieling
- Ilaria Visco
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: The University of Wisconsin-Madison Animal Care and Use Committee has reviewed and approved all of the experiments performed for this study, outlined in protocol G005386-RO1.
- Jonathan Chernoff, Fox Chase Cancer Center, United States
© 2019, Golding et al.
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