Inactivation of Glycogen Synthase Kinase-3 by Epidermal Growth Factor Is Mediated by Mitogen-activated Protein Kinase/p90 Ribosomal Protein S6 Kinase Signaling Pathway in NIH/3T3 Cells

doi:10.1074/jbc.270.3.987

Inactivation of Glycogen Synthase Kinase-3 by Epidermal Growth Factor Is Mediated by Mitogen-activated Protein Kinase/p90 Ribosomal Protein S6 Kinase Signaling Pathway in NIH/3T3 Cells (*)

From the (1)Departments of Pharmacology and
(2)Biochemistry and
(3)Howard Hughes Medical Institute, University of Washington, Seattle, Washington 98195, the
(4)Department of Membrane Research and Biophysics, The Weizmann Institute of Science, Rehovot 76100, Israel, and
(5)Afdeling Biochemie Facultei der Geneeskunde, Katholieke Universiteit Leuven, B 300 Leuven, Belgium

** To whom correspondence should be addressed:
Dept. of Pharmacology, SL-15, University of Washington, Seattle, WA 98195.
Tel.: 206-543-8500; Fax: 206-543-0858.

Abstract

The role of the p90 ribosomal protein S6 kinase/mitogen-activated protein kinase (RSK/MAPK) signaling pathway in regulating glycogen synthase kinase-3 (GSK-3) activity was investigated. In vitro studies showed that GSK-3 was inactivated by 50% upon incubation with RSK purified from epidermal growth factor (EGF)-stimulated NIH/3T3 cells. Subsequently, the effect of EGF on GSK-3 activity was measured in NIH/3T3 cells that stably overexpressed mutated forms of MAPK kinase (MAPKK). The activation of RSK by EGF was markedly decreased in cell lines expressing the dominant negative MAPKK mutants S222A and K97A and was increased in cells expressing the S222E mutant as compared with control cell lines. EGF induced a rapid decrease in GSK-3β activity (50%) in control and S222E cells; however, only 25 and 10% inhibition in GSK-3β activity was observed in cell lines expressing the dominant negative mutants K97A and S222A, respectively, suggesting that inhibition of GSK-3 was partially blocked in these cells. Taken together, these results suggest that the action of EGF on GSK-3 inactivation is mediated by the RSK/MAPK signaling pathway in NIH/3T3 cells and provide evidence for a mechanism regulating GSK-3 activity in intact cells.

Footnotes

↵^§ Supported by an American Heart Association training fellowship.
↵^¶ Research Director of the Belgian “National Fonds voor Wetenschappelijk Onderzoek.”
↵* This work was supported by Grant DK 42528 from the National Institutes of Health and by a grant from the Muscular Dystrophy Association. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

GSK-3

glycogen synthase kinase-3

EGF

epidermal growth factor

MAPK

mitogen-activated protein kinase

MAPKK

MAPK kinase

RSK

p90 ribosomal protein S6 kinase

I2

inhibitor-2.
- Received October 18, 1994.
- Revision received November 18, 1994.