Feedback activation of neurofibromin terminates growth factor-induced Ras activation.

Anne Hennig,Robby Markwart,Katharina Wolff,Graham Ladds,Katja Schubert,Ignacio Rubio,Yan Cui,Manuel A Esparza-Franco,Ian A Prior

doi:10.1186/s12964-016-0128-z

Abstract

BackgroundGrowth factors induce a characteristically short-lived Ras activation in cells emerging from quiescence. Extensive work has shown that transient as opposed to sustained Ras activation is critical for the induction of mitogenic programs. Mitogen-induced accumulation of active Ras-GTP results from increased nucleotide exchange driven by the nucleotide exchange factor Sos. In contrast, the mechanism accounting for signal termination and prompt restoration of basal Ras-GTP levels is unclear, but has been inferred to involve feedback inhibition of Sos. Remarkably, how GTP-hydrolase activating proteins (GAPs) participate in controlling the rise and fall of Ras-GTP levels is unknown.ResultsMonitoring nucleotide exchange of Ras in permeabilized cells we find, unexpectedly, that the decline of growth factor-induced Ras-GTP levels proceeds in the presence of unabated high nucleotide exchange, pointing to GAP activation as a major mechanism of signal termination. Experiments with non-hydrolysable GTP analogues and mathematical modeling confirmed and rationalized the presence of high GAP activity as Ras-GTP levels decline in a background of high nucleotide exchange. Using pharmacological and genetic approaches we document a raised activity of the neurofibromatosis type I tumor suppressor Ras-GAP neurofibromin and an involvement of Rsk1 and Rsk2 in the down-regulation of Ras-GTP levels.ConclusionsOur findings show that, in addition to feedback inhibition of Sos, feedback stimulation of the RasGAP neurofibromin enforces termination of the Ras signal in the context of growth-factor signaling. These findings ascribe a precise role to neurofibromin in growth factor-dependent control of Ras activity and illustrate how, by engaging Ras-GAP activity, mitogen-challenged cells play safe to ensure a timely termination of the Ras signal irrespectively of the reigning rate of nucleotide exchange.Electronic supplementary materialThe online version of this article (doi:10.1186/s12964-016-0128-z) contains supplementary material, which is available to authorized users.

Highlights

Growth factors induce a characteristically short-lived Ras activation in cells emerging from quiescence
Extensive work has shown that the duration of the Ras signal is a key determinant of cell fate in the sense that growth factor activation of Ras must be transient to promote a proper proliferative response
We provide evidence that neurofibromin, the product of the tumor suppressor of neurofibromatosis type 1 (NF1), is the RasGAP species mediating the deactivation of Ras

Summary

Introduction

Growth factors induce a characteristically short-lived Ras activation in cells emerging from quiescence. Mitogen-induced accumulation of active Ras-GTP results from increased nucleotide exchange driven by the nucleotide exchange factor Sos. In contrast, the mechanism accounting for signal termination and prompt restoration of basal Ras-GTP levels is unclear, but has been inferred to involve feedback inhibition of Sos. Remarkably, how GTP-hydrolase activating proteins (GAPs) participate in controlling the rise and fall of Ras-GTP levels is unknown. Cells emerging from quiescence upon growth factor encounter feature a pronounced activation of Ras which is characteristically short-lived. Erk phosphorylates multiple sites at the Sos C-terminus, promoting the dissociation of Sos from the adapter protein Grb-2 [16,17,18,19] This reaction is inferred to down-modulate Sos activity by removing Sos from the vicinity of Ras, not all studies are in support of this model [19,20,21,22]

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