The classical model for the activation of the nucleotide exchange factor Son of sevenless (SOS) involves its recruitment to the membrane, where it engages Ras. The recent discovery that Ras*GTP is an allosteric activator of SOS indicated that the regulation of SOS is more complex than originally envisaged. We now present crystallographic and biochemical analyses of a construct of SOS that contains the Dbl homology-pleckstrin homology (DH-PH) and catalytic domains and show that the DH-PH unit blocks the allosteric binding site for Ras and suppresses the activity of SOS. SOS is dependent on Ras binding to the allosteric site for both a lower level of activity, which is a result of Ras*GDP binding, and maximal activity, which requires Ras*GTP. The action of the DH-PH unit gates a reciprocal interaction between Ras and SOS, in which Ras converts SOS from low to high activity forms as Ras*GDP is converted to Ras*GTP by SOS.

The kinetics of Ras activation by Son of sevenless (SOS) changes profoundly when Ras is tethered to membranes, instead of being in solution. SOS has two binding sites for Ras, one of which is an allosteric site that is distal to the active site. The activity of the SOS catalytic unit (SOS(cat)) is up to 500-fold higher when Ras is on membranes compared to rates in solution, because the allosteric Ras site anchors SOS(cat) to the membrane. This effect is blocked by the N-terminal segment of SOS, which occludes the allosteric site. We show that SOS responds to the membrane density of Ras molecules, to their state of GTP loading and to the membrane concentration of phosphatidylinositol-4,5-bisphosphate (PIP2), and that the integration of these signals potentiates the release of autoinhibition.

The RTK-Ras-ERK cascade is a central signaling module implicated in the control of diverse biological processes including cell proliferation, differentiation, and survival. The coupling of RTK to Ras is mediated by the Ras-specific nucleotide-exchange factor Son of Sevenless (Sos), which activates Ras by inducing the exchange of GDP for GTP . Considerable evidence indicates that the duration and amplitude of Ras signals are important determinants in controlling the biological outcome . However, the mechanisms that regulate the quantitative output of Ras signaling remain poorly understood. We define a previously unrecognized regulatory component of the machinery that specifies the kinetic properties of signals propagated through the RTK-Ras-ERK cascade. We demonstrate that the establishment of a positive feedback loop involving Ras.GTP and Sos leads to an increase in the amplitude and duration of Ras activation in response to EGF stimulation. This effect is propagated to downstream elements of the pathway as reflected by sustained EGF-induced ERK phosphorylation and enhanced SRE-dependent transcription. As a consequence, the physiological endpoint of EGF action is switched from proliferation to differentiation. We propose that the engagement of Ras/Sos positive feedback loop may contribute to the mechanism by which ligand stimulation is coupled to discrete biological responses.