RhoG is a member of the Rho family of GTPases that shares 72% and 62% sequence identity with Rac1 and Cdc42Hs, respectively. We have expressed mutant RhoG proteins fused to the green fluorescent protein and analyzed subsequent changes in cell surface morphology and modifications of cytoskeletal structures. In rat and mouse fibroblasts, green fluorescent protein chimera and endogenous RhoG proteins colocalize according to a tubular cytoplasmic pattern, with perinuclear accumulation and local concentration at the plasma membrane. Constitutively active RhoG proteins produce morphological and cytoskeletal changes similar to those elicited by a simultaneous activation of Rac1 and Cdc42Hs, i.e., the formation of ruffles, lamellipodia, filopodia, and partial loss of stress fibers. In addition, RhoG and Cdc42Hs promote the formation of microvilli at the cell apical membrane. RhoG-dependent events are not mediated through a direct interaction with Rac1 and Cdc42Hs targets such as PAK-1, POR1, or WASP proteins but require endogenous Rac1 and Cdc42Hs activities: coexpression of a dominant negative Rac1 impairs membrane ruffling and lamellipodia but not filopodia or microvilli formation. Conversely, coexpression of a dominant negative Cdc42Hs only blocks microvilli and filopodia, but not membrane ruffling and lamellipodia. Microtubule depolymerization upon nocodazole treatment leads to a loss of RhoG protein from the cell periphery associated with a reversal of the RhoG phenotype, whereas PDGF or bradykinin stimulation of nocodazole-treated cells could still promote Rac1- and Cdc42Hs-dependent cytoskeletal reorganization. Therefore, our data demonstrate that RhoG controls a pathway that requires the microtubule network and activates Rac1 and Cdc42Hs independently of their growth factor signaling pathways.
RhoG GTPase controls a pathway that independently activates Rac1 and Cdc42Hs
Gauthier-Rouviere, C.; Vignal, E.; Meriane, M.; Roux, P.; Montcourier, P.; Fort, P.
Mol Biol Cell
1998-06 / vol 9 / pages 1379-94
Animals; Mice; Cell Line; Recombinant Fusion Proteins/genetics/metabolism; Rats; Transcription Factors/genetics/*metabolism; Actins/metabolism; GTP-Binding Proteins/*metabolism; Green Fluorescent Proteins; 3T3 Cells; cdc42 GTP-Binding Protein; Cell Cycle Proteins/*metabolism; GTP Phosphohydrolases/genetics/*metabolism; Bradykinin/pharmacology; Cytoskeleton/physiology; Luminescent Proteins/metabolism; Microtubules/metabolism; Platelet-Derived Growth Factor/pharmacology; rac GTP-Binding Proteins