Amphiregulin is a poor effector of ligand-induced EGF receptor dimerisation

Richter, A., Neelam, B., Chamberlin, S. and Davies, D. (1996) Amphiregulin is a poor effector of ligand-induced EGF receptor dimerisation. FASEB Journal, 10 (6), [A1377].

Abstract

The ability ofligand to stabilize epidermal growth factor receptor (EGFR) dimers is critical for subsequent activation of the intrinsic receptor tyrosine kinase. We have evaluated the capacity of the different EGFR ligands to promote dimer formation in a novel 96-well assay which exploits the higher ligand binding affinity of EGFR dimers compared with monomers. In this assay, the ability of the ligand to promote receptor dimerisation determines the proportion of dimeric receptor (but not total EGFR) captured by an immobilizing antibody (MoAb EGFR1). Subsequent measurement of 125/ - EGF binding reflects EGFR ligand binding affinity which in turn is indicative of the degree of dimer formation. When receptors are pretreated with equimolar concentrations of EGF, transforming growth factor alpha or betacellulin, 125/ - EGF binding is increased 5-fold compared to untreated EGFRs. Amphiregulin (ARJ, an EGFR ligand with a truncated C-tail that lacks Leu47, is ineffective in this assay, even when used at a 200x higher dose to compensate for its low receptor binding affinity. The inability of AR to promote EGFR dimerisation could not be explained by the presence of the basic N-terminal heparin-binding domain of AR, as a similar domain is also present in heparin binding-EGF which is equipotent with EGF in our assay. On the basis of these observations we propose that the C-terminal tail of EGF-R ligands is intimately involved in stabilising receptor dimers and that the interaction requires at least the conserved leucine residue. [Supported by the Cancer Research Campaign, UK].

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Contributors

Author: D. Davies

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