TrioGEF1 controls Rac- and Cdc42-dependent cell structures through the direct activation of RhoG

Rho GTPases regulate the morphology of cells stimulated by extracellular li gands. Their activation is controlled by guanine exchange factors (GEF) tha t catalyze their binding to GTP. The multidomain Trio protein represents an emerging class of Rho regulators that contain two GEF domains of distinct specificities. We report here the characterization of Rho signaling pathway s activated by the N-terminal GEF domain of Trio (TrioD1). In fibroblasts, TrioD1 triggers the formation of particular cell structures, similar to tho se elicited by RhoG, a GTPase known to activate both Rac1 and Cdc42Hs. In a ddition, the activity of TrioD1 requires the microtubule network and reloca lizes RhoG at the active sites of the plasma membrane. Using a classical in vitro exchange assay, TrioD1 displays a higher GEF activity on RhoG than o n Rac1. In fibroblasts, expression of dominant negative RhoG mutants totall y abolished TrioD1 signaling, whereas dominant negative Rac1 and Cdc42Hs on ly led to partial and complementary inhibitions. Finally, expression of a R ho Binding Domain that specifically binds RhoG(GTP) led to the complete abo lition of TrioD1 signaling, which strongly supports Rac1 not being activate d by TrioD1 in vivo. These data demonstrate that Trio controls a signaling cascade that activates RhoG, which in turn activates Rad and Cdc42Hs.