A Rho exchange factor mediates thrombin and G alpha(12)-induced cytoskeletal responses

Thrombin induces astrocytoma cell rounding through a Rho-dependent pathway (Majumdar, M., Seasholtz, T. M., Goldstein, D., de Lanerolle, P,, and Brown , J. H. (1998) J. Biol. Chem. 273, 10099-10106), The involvement of the G(1 2) family of G proteins and the role of specific Rho exchange factors in tr ansducing signals from the thrombin receptor to Rho-dependent cytoskeletal responses was examined. Microinjection of cDNAs for activated G alpha(12) o r G alpha(13) induced cell rounding, and antibodies to G alpha(12) or G alp ha(13) blocked the response to thrombin, in contrast, activation or inhibit ion of G alpha(q) function had relatively little effect. The cytoskeletal r esponse to G alpha(12) was inhibited by microinjection of C3 exoenzyme, ind icating Rho dependence. Two Rho-specific guanine nucleotide exchange factor s (GEFs), oncogenic lbc and p115, increased the percentage of rounded cells 4-5-fold, and this was inhibited by C3. Mutant GEFs lacking the Dbl homolo gy (DH) domain required for exchange factor activity failed to induce cell rounding. However, the DH mutants of Ibc and p115 were efficacious inhibito rs off rounding induced by thrombin or G alpha(12). The effects of lbc were dependent on an intact pleckstrin homology domain, which may be required f or appropriate targeting of the Rho-GEF, These findings identify the G alph a(12) protein family as transducers of thrombin signaling to the cytoskelet on and provide the first evidence that a Rho-GEF transduces signals between G protein-coupled receptors and Rho-mediated cytoskeletal responses.