Insulin-stimulated disassociation of the SOS-Grb2 complex

S. B. Waters, K. Yamauchi, Jeffrey E. Pessin

Research output: Contribution to journalArticle

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Abstract

Insulin stimulation of differentiated 3T3-L1 adipocytes or Chinese hamster ovary cells expressing high levels of the insulin receptor resulted in a time-dependent decrease in the electrophoretic mobility of SOS on sodium dodecyl sulfate-polyacrylamide gels. The reduction in SOS mobility was completely reversed by alkaline phosphatase treatment, and the in vitro phosphorylation of SOS by mitogen-activated protein kinase resulted in a decrease of electrophoretic mobility identical to that following in vivo insulin stimulation. Immunoprecipitation of Grb2 followed by SOS immunoblotting demonstrated a disassociation of the SOS-Grb2 complex that paralleled the decrease in SOS electrophoretic mobility. Similarly, SOS immunoprecipitation followed by Grb2 immunoblotting also indicated an uncoupling of the SOS-Grb2 complex. Further, incubation of whole-cell extracts with glutathione-S-transferase-Grb2 fusion proteins demonstrated that insulin stimulation resulted in a decreased affinity of SOS for Grb2. In contrast, the disassociation of SOS from Grb2 did not affect the interactions between Grb2 and tyrosine-phosphorylated Shc. In addition to insulin, several other agents which activate the mitogen-activated protein kinase pathway (platelet-derived growth factor, serum, and phorbol ester) also resulted in the uncoupling of the SOS-Grb2 complex. Consistent with these results, expression of v-ras and v-raf resulted in a constitutive decrease in the association between SOS and Grb2. Together, these data suggest a molecular mechanism accounting for the transient activation of ras due to the uncoupling of the SOS-Grb2 complex following SOS phosphorylation.

Original languageEnglish (US)
Pages (from-to)2791-2799
Number of pages9
JournalMolecular and Cellular Biology
Volume15
Issue number5
StatePublished - 1995
Externally publishedYes

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Insulin
Mitogen-Activated Protein Kinases
Immunoprecipitation
Immunoblotting
Phosphorylation
Platelet-Derived Growth Factor
Insulin Receptor
Phorbol Esters
Cricetulus
Cell Extracts
Glutathione Transferase
Adipocytes
Sodium Dodecyl Sulfate
Alkaline Phosphatase
Tyrosine
Ovary
Serum
Proteins
In Vitro Techniques
polyacrylamide gels

ASJC Scopus subject areas

  • Cell Biology
  • Genetics
  • Molecular Biology

Cite this

Insulin-stimulated disassociation of the SOS-Grb2 complex. / Waters, S. B.; Yamauchi, K.; Pessin, Jeffrey E.

In: Molecular and Cellular Biology, Vol. 15, No. 5, 1995, p. 2791-2799.

Research output: Contribution to journalArticle

Waters, S. B. ; Yamauchi, K. ; Pessin, Jeffrey E. / Insulin-stimulated disassociation of the SOS-Grb2 complex. In: Molecular and Cellular Biology. 1995 ; Vol. 15, No. 5. pp. 2791-2799.
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