An Incomplete Program of Cellular Tyrosine Phosphorylations Induced by Kinase-defective Epidermal Growth Factor Receptors

doi:10.1074/jbc.270.20.12085

An Incomplete Program of Cellular Tyrosine Phosphorylations Induced by Kinase-defective Epidermal Growth Factor Receptors (*)

From the (1) Department of Microbiology and
(2) Pharmacology, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908

^¶To whom correspondence should be addressed:
Dept. of Microbiology, Box 441, 2-16 Jordan Hall, University of Virginia Health Sciences Center, Charlottesville, VA 22908.
Tel.: 804-924-5022; Fax: 804-982-0689; E-mail: mjw{at}virginia.edu.

Abstract

Although signaling by the epidermal growth factor (EGF) receptor is thought to be dependent on receptor tyrosine kinase activity, it is clear that mitogen-activated protein (MAP) kinase can be activated by receptors lacking kinase activity. Since analysis of the signaling pathways used by kinase-defective receptors could reveal otherwise masked capabilities, we examined in detail the tyrosine phosphorylations and enzymes of the MAP kinase pathway induced by kinase-defective EGF receptors. Following EGF stimulation of B82L cells expressing a kinase-defective EGF receptor mutant (K721M), we found that ERK2 and ERK1 MAP kinases, as well as MEK1 and MEK2 were all activated, and SHC became prominently tyrosine-phosphorylated. By contrast, kinase-defective receptors failed to induce detectable phosphorylations of GAP (GTPase-activating protein), p62, JAK1, or p91STAT1, all of which were robustly phosphorylated by wild-type receptors. These data demonstrate that kinase-defective receptors induce several protein tyrosine phosphorylations, but that these represent only a subset of those seen with wild-type receptors. This suggests that kinase-defective receptors activate a heterologous tyrosine kinase with a specificity different from the EGF receptor. We found that kinase-defective receptors induced ErbB2/c-Neu enzymatic activation and ErbB2/c-Neu binding to SHC at a level even greater than that induced by wild-type receptors. Thus, heterodimerization with and activation of endogenous ErbB2/c-Neu is a possible mechanism by which kinase-defective receptors stimulate the MAP kinase pathway.

Footnotes

↵^§ Supported by a fellowship from the Deutsche Forschungsgemeinschaft (RE-864/2-2).
↵* This work was supported by National Institutes of Health Grants CA 39076, CA 40042, GM 47332, GM 49772, and 5-T32-GM 07055. 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:

EGF-R

epidermal growth factor receptor

EGF

epidermal growth factor

K-

kinase-defective

K+

wild-type (kinase-active)

MAP kinase

mitogen-activated protein kinase

ERK

extracellular signal-regulated kinase

MEK

MAPK or Erk kinase

GAP

GTPase-activating protein

PAGE

polyacrylamide gel electrophoresis

DMEM

Dulbecco's modified Eagle's medium

PBS

phosphate-buffered saline.