Human Epidermal Growth Factor Receptor-1 Expression Renders Chinese Hamster Ovary Cells Sensitive to Alternative Aldosterone Signaling

doi:10.1074/jbc.M208851200

Human Epidermal Growth Factor Receptor-1 Expression Renders Chinese Hamster Ovary Cells Sensitive to Alternative Aldosterone Signaling*

From the ^‡Physiologisches Institut and the ^§Institut für Medizinische Strahlenkunde und Zellforschung, Universität Würzburg, 97070 Würzburg, Germany

Abstract

The epidermal growth factor (EGF) regulates cell proliferation, differentiation, and ion transport using ERK1/2 as a downstream effector. Furthermore, the EGF receptor (EGFR) is involved in signaling by G-protein-coupled receptors, growth hormone, and cytokines via transactivation. It has been suggested that steroids interact with peptide hormones. Previously, we have shown that aldosterone modulates EGF responses in Madin-Darby canine kidney cells (Gekle, M., Freudinger, R., Mildenberger, S., and Silbernagl, S. (2002) Am. J. Physiol. 282, F669–F679). Here, we tested the hypothesis that human EGFR-1 can confer alternative aldosterone responsiveness with respect to ERK1/2 phosphorylation to Chinese hamster ovary cells, which do not express EGFR. Wild-type Chinese hamster ovary cells did not respond to EGF or aldosterone. After transfection of human EGFR-1, the cells responded to EGF, but not to aldosterone. However, when submaximal concentrations of EGF were used, nanomolar concentrations of aldosterone potentiated the action of EGF within minutes, resulting in a leftward shift of the EGF dose-response curve. This was not the case in mock-transfected cells. The EGFR kinase inhibitor tyrphostin AG1478 or the MEK1/2 inhibitor U0126 completely prevented the effect. Furthermore, aldosterone enhanced Tyr phosphorylation of c-Src and EGFR, and an inhibitor of cytosolic tyrosine kinases (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyriociaine) prevented the action of aldosterone. Our data show that aldosterone uses the EGF-EGFR-MEK1/2-ERK1/2 signaling cascade to elicit its alternative effects. In the presence of EGF, aldosterone leads to EGFR transactivation via cytosolic tyrosine kinases of the Src family.

Footnotes

↵* This work was supported by Deutsche Forschungsgemeinschaft Grants Ge 905/4-2, Ge 905/4-3, and SFB487/A6.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.
↵¶ To whom correspondence should be addressed: Physiologisches Inst., Universität Würzburg, Röntgenring 9, 97070 Würzburg, Germany. Tel.: 49-931-312739; Fax: 49-931-312741; E-mail: michael.gekle@mail.uni-wuerzburg.de.
Published, JBC Papers in Press, September 19, 2002, DOI 10.1074/jbc.M208851200
Abbreviations:

PKC

protein kinase C

ERK

extracellular signal-regulated kinase

EGF

epidermal growth factor

EGFR

epidermal growth factor receptor

MDCK

Madin-Darby canine kidney

HER1

human epidermal growth factor receptor-1

CHO

Chinese hamster ovary

MEK

mitogen-activated protein kinase/extracellular signal-regulated kinase kinase

pERK1/2

phosphorylated ERK1/2

ELISA

enzyme-linked immunosorbent assay

PBS

phosphate-buffered saline

PMA

phorbol 12-myristate 13-acetate

PP2

4-amino-5-(4-chloro- phenyl)-7-(t-butyl)pyrazolo[3,4-d]pyriociaine
- Received August 29, 2002.
The American Society for Biochemistry and Molecular Biology, Inc.