2 SH2 DOMAINS OF P120 RAS GTPASE-ACTIVATING PROTEIN BIND SYNERGISTICALLY TO TYROSINE-PHOSPHORYLATED P190 RHO-GTPASE-ACTIVATING PROTEIN

p120 GTPase-activating protein (GAP) is a negative regulator of Pas th at functions at a key relay point in signal transduction pathways that control cell proliferation. Among other proteins, p120 GAP associates with p190, a GAP for the Pas-related protein, Rho. To characterize th e p120 . p190 interaction further, we used bacterially expressed gluta thione S-transferase fusion polypeptides to map the regions of p120 ne cessary for its interactions with p190. Our results show that both the N-terminal and the C-terminal SH2 domains of p120 are individually ca pable of binding p190 expressed in a baculovirus/insect cell system. M oreover, the two SH2 domains together on one polypeptide bind synergis tically to p190, and this interaction is dependent on tyrosine phospho rylation of p190. In addition, mutation of the highly conserved Arg re sidues in the critical FLVR sequences of both SH2 domains of full-leng th p120 reduces binding to tyrosine-phosphorylated p190. The dependenc e on p190 phosphorylation for complex formation with p120 . SH2 domain s observed in vitro is consistent with analysis of the native p120 . p 190 complexes formed in vivo. These findings suggest that SH2-phosphot yrosine interaction is one mechanism by which the cell regulates p120 p190 association and thus may be a means for coordinating the Ras- and Rho-mediated signaling pathways.