Ligand-independent phosphorylation of Y869 (Y845) links mutant EGFR signaling to stat-mediated gene expression

Exp Cell Res. 2008 Jan 15;314(2):413-9. doi: 10.1016/j.yexcr.2007.09.002. Epub 2007 Sep 8.

Abstract

Activating mutants of EGFR have been identified in a subset of non-small-cell lung cancers. To investigate mutant-driven signaling, we focused on Y869, a residue in the same activation loop where the L858R and L861Q mutations are located. We observed ligand-independent phosphorylation of Y869 in 32D cells EGFR(L858R) and EGFR(L861Q). The EGFR tyrosine kinase inhibitor (TKI) erlotinib inhibited Y869 P-EGFR in intact cells as well as in a cell-free kinase reaction. Expression of kinase domain of EGFR(L858R) and EGFR(L861Q) exhibited auto-phosphorylation of Y869; this was inhibited by EGFR TKIs but not by Src kinase inhibitor. P-Y859 of EGFR-mediated downstream component, STAT5, was also analyzed. Y694 P-STAT5 was eliminated by erlotinib treatment. Analysis of immune-complexes showed constitutive association of mutant EGFRs with STAT5 and Src which was unaffected by erlotinib or PP1. On the other hand, 32D-EGFR(WT) exhibited constitutive STAT5 phosphorylation and association of EGFR with JAK2. In these cells, a JAK2 inhibitor abrogated P-STAT5 whereas mutant EGFRs did not associate with JAK2. Expression of c-myc was regulated by EGFR/STAT5 signaling in cells expressing EGFR(L858R) and EGFR(L861Q). Our results suggest that ligand-independent and Src activity-independent phosphorylation of Y869 in mutant EGFR regulates STAT5 activation and c-myc expression.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • ErbB Receptors / antagonists & inhibitors
  • ErbB Receptors / genetics*
  • ErbB Receptors / metabolism*
  • Erlotinib Hydrochloride
  • Genes, myc
  • Humans
  • Ligands
  • Mutation
  • Phosphorylation
  • Proto-Oncogene Proteins pp60(c-src) / metabolism
  • Quinazolines / pharmacology
  • STAT5 Transcription Factor / metabolism*
  • Signal Transduction*
  • Tumor Cells, Cultured

Substances

  • Ligands
  • Quinazolines
  • STAT5 Transcription Factor
  • Erlotinib Hydrochloride
  • ErbB Receptors
  • Proto-Oncogene Proteins pp60(c-src)