Extraction of active RhoGTPases by RhoGDI regulates spatiotemporal patterning of RhoGTPases

Abstract

AbstractThe RhoGTPases are characterized as membrane-associated molecular switches cycling 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 RhoGTPasesin vivoand reconstitution of the cyclein vitro. We developed methods to directly image vertebrate RhoGTPasesin vivoor on lipid bilayersin vitro. Using these tools, we identified pools of active and inactive RhoGTPase associated with the membrane, showed that RhoGDI can actively extract both inactive and active RhoGTPases, and that the extraction of active RhoGTPase contributes to their spatial regulation around wounds. In contrast to the textbook model of the RhoGTPase cycle, these results indicate that RhoGDI actively contributes to spatiotemporal patterning by removing active RhoGTPases from the plasma membrane.