Insulin and Epidermal Growth Factor Receptors Regulate Distinct Pools of Grb2-SOS in the Control of Ras Activation

doi:10.1074/jbc.271.30.18224

Insulin and Epidermal Growth Factor Receptors Regulate Distinct Pools of Grb2-SOS in the Control of Ras Activation*

From the Department of Physiology and Biophysics, The University of Iowa, Iowa City, Iowa 52242-1109

^§ To whom correspondence should be addressed.

↵^‡ Present address: Metabolex Inc., 3876 Bay Center Place, Hayward, CA 94545-3619.

Abstract

Insulin and epidermal growth factor (EGF) stimulate a rapid but transient increase in the amount of GTP bound to Ras that returns to the basal GDP-bound state within 10-30 min. Although insulin stimulation resulted in a dissociation of the Grb2·SOS complex, EGF did not affect the Grb2·SOS complex but instead induced dissociation of Grb2-SOS from tyrosine-phosphorylated Shc. The dissociation of Grb2-SOS from Shc was not due to dephosphorylation as Shc remained persistently tyrosine-phosphorylated during this time. Furthermore, there was no decrease in the extent of insulin receptor substrate 1, insulin receptor, or EGF receptor tyrosine phosphorylation. Surprisingly, however, despite the EGF-induced decrease in the amount of Grb2-SOS bound to Shc, the extent of Grb2 associated with Shc remained constant, and there was a concomitant increase in the amount of SOS associated with Grb2. In addition, after the insulin-stimulated dissociation of Grb2 from SOS, EGF treatment induced the reassociation of the Grb2·SOS complex. Quantitative immunoprecipitation demonstrated that only a small fraction of the total cellular pool of Grb2 was associated with SOS. Similarly, only a small fraction of SOS and Grb2 were co-immunoprecipitated with Shc. Together, these data suggest the presence of distinct Grb2-SOS pools that are independently utilized by insulin and EGF in their recruitment to tyrosine-phosphorylated Shc.

Footnotes

↵* This study was supported by Grants DK33823 and DK25925 from the National Institutes of Health. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

EGF

epidermal growth factor

SH2

src homology 2

SH3

src homology 3

IRS1

insulin receptor substrate 1

MEK

mitogen-activated and extracellular signal-regulated protein kinase kinase

ERK

extracellular signal-regulated protein kinase

CHO

Chinese hamster ovary

IR

insulin receptor

ER

EGF receptor.
- Received February 22, 1996.
- Revision received March 27, 1996.