Systematic characterization of RhoGEF/RhoGAP regulatory proteins reveals organization principles of Rho GTPase signaling

Rho GTPases control cell shape formation and thus fundamental physiological processes in all eukaryotes. Their functions are regulated by 145 RhoGEF and RhoGAP multi-domain proteins in humans. To provide the framework for a systems-level understanding of how these regulators orchestrate cellular morphogenesis, we comprehensively characterized their substrate specificities, localization and interactome. The resulting resource places the RhoGEFs/RhoGAPs in functional context, serving as a foundation for targeted and integrated studies. Our data reveals their critical role in the spatial organization of Rho signaling. They localize to multiple compartments to provide positional information, are extensively interconnected to jointly coordinate their signaling networks and are widely autoinhibited to remain sensitive to local activation. RhoGAPs exhibit lower substrate specificity than RhoGEFs and may contribute to preserving Rho activity gradients. We demonstrate the utility of our integrated data by detailing a multi-RhoGEF complex downstream of G-protein-coupled receptors in which the enzymes mutually regulate their activities.