Analysis of signal transduction mediated by Ras and Rac family GTPases

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The role of guanine nucleotide exchange factors (GEFs) for Ras and Rho family GTP-binding proteins was explored. Dbl is a prototype of Dbl family GEFs, containing tandem DH and PH domains, which are responsible for GEF activity toward Rho family GTP-binding proteins. G protein βγ subunits (Gβγ) bind to the N-terminal region of Dbl although the role in the regulation of GEF activity remains elusive. Furthermore, the tyrosine kinase ACK-1, an effector of Cdc42, phosphorylates Dbl thereby stimulating GEF activity. In fact, the epidermal growth factor receptor activates ACK-1 in a Grb-2 and Cdc42-dependent manner, which in turn phosphorylates Dbl, leading to actin cytoskeletal reorganization. Ras-GRF-1 was originally isolated as a Ras GEF that is expressed exclusively in the brain. We found that the Gβγ signal induced GEF activity of Ras-GRF-1 toward Rac that is latent without stimulation. As expected, DH and PH domains were required for this activity. Gβγ-dependent tyrosine phosphorylation of Ras-GRF-1 was critical for the induction of GEF activity toward Rac. The non-receptor tyrosine kinase Src phosphorylates Ras-GRF-1, and induces its Rac GEF activity in vivo and in vitro. Thus, Src may be involved in Gβγ-dependent activation of Ras-GRF-1. On the other hand, ACK-1 as well stimulates tyrosine phosphorylation of Ras-GRF-1, thereby enhancing GEF activity toward Ras.