THE ROLE OF THE PH DOMAIN IN THE SIGNAL-DEPENDENT MEMBRANE TARGETING OF SOS

The pleckstrin homology (PH) domain is a conserved protein module pres ent in diverse signal transducing proteins. To investigate the functio n of the PH domain of the Ras exchanger Sos, we have generated a recom binant (His)(6)-tagged PH domain from human Sos1 (PH-Sos). Here we sho w that PH-Sos binds with high affinity (1.5 mu M) to lipid vesicles co ntaining the negatively charged phospholipid phosphatidylinositol 4,5- bisphosphate (PIP2). When microinjected into serum-deprived rat embryo fibroblasts or COS cells, PH-Sos displays a homogenous subcellular di stribution. However, PH-Sos rapidly accumulates in the plasma membrane following serum stimulation and, under these conditions, is localized preferentially to the leading edge of motile cells, Surprisingly, the membrane localization of PH-Sos is not dependent on its ability to bi nd PIP2, Overexpression of the PH domain of Sos has a pronounced domin ant-negative effect on serum-induced activation of the Ras signaling p athway, These results suggest that the PH domain of Sos participates i n regulating the inducible association of Sos with the membrane, and i ndicate the presence of specific ligands that interact with this domai n to bring about the activation of Ras.