The Enhanced Tumorigenic Activity of a Mutant Epidermal Growth Factor Receptor Common in Human Cancers Is Mediated by Threshold Levels of Constitutive Tyrosine Phosphorylation and Unattenuated Signaling

doi:10.1074/jbc.272.5.2927

The Enhanced Tumorigenic Activity of a Mutant Epidermal Growth Factor Receptor Common in Human Cancers Is Mediated by Threshold Levels of Constitutive Tyrosine Phosphorylation and Unattenuated Signaling*

From the ^‡ Ludwig Institute for Cancer Research,
^§ Department of Medicine,
^∥ Department of Chemisty and Biochemistry and
^§§ Center for Molecular Genetics, University of California at San Diego, La Jolla, California 92093-0660,
^‡‡ Pharmingen, Inc., San Diego, California 92121, and the
^¶¶ Department of Pathology, University of Utah School of Medicine, Salt Lake City, Utah 84132

^¶ To whom correspondence should be addressed:
Ludwig Institute for Cancer Research, 9500 Gilman Dr., La Jolla, CA 92093-0660.
Tel.: 619-534-7814; Fax: 619-534-7750; E-mail: hhuang{at}ucsd.edu

↵** Present address: Dept. of Neurosurgery, Saitama Medical School, Saitama, 350-04 Japan.

Abstract

Deregulation of signaling by the epidermal growth factor receptor (EGFR) is common in human malignancy progression. One mutant EGFR (variously named ΔEGFR, de2-7 EGFR, or EGFRvIII), which occurs frequently in human cancers, lacks a portion of the extracellular ligand-binding domain due to genomic deletions that eliminate exons 2 to 7 and confers a dramatic enhancement of brain tumor cell tumorigenicity in vivo. In order to dissect the molecular mechanisms of this activity, we analyzed location, autophosphorylation, and attenuation of the mutant receptors. The mutant receptors were expressed on the cell surface and constitutively autophosphorylated at a significantly decreased level compared with wild-type EGFR activated by ligand treatment. Unlike wild-type EGFR, the constitutively active ΔEGFR were not down-regulated, suggesting that the altered conformation of the mutant did not result in exposure of receptor sequence motifs required for endocytosis and lysosomal sorting. Mutational analysis showed that the enhanced tumorigenicity was dependent on intrinsic tyrosine kinase activity and was mediated through the carboxyl terminus. In contrast with wild-type receptor, mutation of any major tyrosine autophosphorylation site abolished these activities suggesting that the biological functions of ΔEGFR are due to low constitutive activation with mitogenic effects amplified by failure to attenuate signaling by receptor down-regulation.

Footnotes

↵* These studies were partially supported by Japan Brain Foundation and YASUDA Medical Research Foundation (to M. N.) and by National Institutes of Health Grant DK13149 (to G. N. G.) and Grant DAMD17-94-J-4444 (to H. S. W.). 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:

wt

wild type

EGFR

epidermal growth factor receptor

PBS

phosphate-buffered saline

FACS

fluorescence-activated cell sorter

BSA

bovine serum albumin.
↵²H.-J. S. Huang, E. Stockert, and L. J. Old, unpublished data.
↵³ C. Klingbeil, unpublished observations.
↵⁴ H.-J. S. Huang, and W. K. Cavenee, unpublished observations.
↵⁵ L. K. Opresko, H.-J. S. Huang, M. Woolfe, and H. S. Wiley, manuscript in preparation.
- Received September 11, 1996.
- Revision received November 15, 1996.