The tethering arm of the EGF receptor is required for negative cooperativity and signal transduction

Sangeeta Adak, Diana DeAndrade, Linda J. Pike

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

The EGF receptor is a classical receptor-tyrosine kinase. In the absence of ligand, the receptor adopts a closed conformation in which the dimerization arm of subdomain II interacts with the tethering arm in subdomain IV. Following the binding of EGF, the receptor opens to form a symmetric, back-toback dimer. Although it is clear that the dimerization arm of subdomain II is central to the formation of receptor dimers, the role of the tethering arm of subdomain IV (residues 561-585) in this configuration is not known. Here we use 125I-EGF binding studies to assess the functional role of the tethering arm in the EGF receptor dimer. Mutation of the three major residues that contribute to tethering (D563A,H566A,K585AEGF receptor) did not significantly alter either the ligand binding properties or the signaling properties of the EGF receptor. By contrast, breaking the Cys 558-Cys567 disulfide bond through double alanine replacements or deleting the loop entirely led to a decrease in the negative cooperativity in EGF binding and was associated with small changes in downstream signaling. Deletion of the Cys571-Cys593 disulfide bond abrogated cooperativity, resulting in a high affinity receptor and increased sensitivity of downstream signaling pathways to EGF. Releasing the Cys 571-Cys593 disulfide bond resulted in extreme negative cooperativity, ligand-independent kinase activity, and impaired downstream signaling. These data demonstrate that the tethering arm plays an important role in supporting cooperativity in ligand binding. Because cooperativity implies subunitsubunit interactions, these results also suggest that the tethering arm contributes to intersubunit interactions within the EGF receptor dimer.

Original languageEnglish (US)
Pages (from-to)1545-1555
Number of pages11
JournalJournal of Biological Chemistry
Volume286
Issue number2
DOIs
StatePublished - Jan 14 2011
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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