RICH, a rho GTPase-activating protein domain-containing protein involved in signaling by Cdc42 and Rac1

A previously unidentified Rho GTPase-activating protein (GAP) domain-contai ning protein was found in a yeast two-hybrid screen for cDNAs encoding prot eins binding to the Src homology 3 domain of Cdc42-interacting protein 4 (C IP4). The protein was named RICH-1 (RhoGAP interacting with CIP4 homologues ), and, in addition to the RhoGAP domain, it contained an N-terminal domain with endophilin homology and a C-terminal proline-rich domain. Transient t ransfections of RICH-1 indicated that it bound to CIP4 in vivo, as shown by co-immunoprecipitation experiments, as well as colocalization assays. In v itro assays demonstrated that the RhoGAP domain of RICH-1 catalyzed GTP hyd rolysis on Cdc42 and Rac1, but not on RhoA. Ectopic expression of the RhoGA P domain as well as the full-length protein interfered with platelet-derive d growth factor BB-induced membrane ruffling, but not with serum-induced st ress fiber formation, further emphasizing the notion that, in vivo, RICH-1 is a GAP for Cdc42 and Rac1.