Effect of phosphorylation on EGFR dimer stability probed by single-molecule dynamics and FRET/FLIM

Biophysical Journal
Oana CobanTony Ng

Abstract

Deregulation of epidermal growth factor receptor (EGFR) signaling has been correlated with the development of a variety of human carcinomas. EGF-induced receptor dimerization and consequent trans- auto-phosphorylation are among the earliest events in signal transduction. Binding of EGF is thought to induce a conformational change that consequently unfolds an ectodomain loop required for dimerization indirectly. It may also induce important allosteric changes in the cytoplasmic domain. Despite extensive knowledge on the physiological activation of EGFR, the effect of targeted therapies on receptor conformation is not known and this particular aspect of receptor function, which can potentially be influenced by drug treatment, may in part explain the heterogeneous clinical response among cancer patients. Here, we used Förster resonance energy transfer/fluorescence lifetime imaging microscopy (FRET/FLIM) combined with two-color single-molecule tracking to study the effect of ATP-competitive small molecule tyrosine kinase inhibitors (TKIs) and phosphatase-based manipulation of EGFR phosphorylation on live cells. The distribution of dimer on-times was fitted to a monoexponential to extract dimer off-rates (koff). Our data show that p...Continue Reading

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Citations

Dec 19, 2019·Journal of the National Cancer Institute·Paul R BarberTony Ng
Jul 22, 2020·Annual Review of Cell and Developmental Biology·Aneta Koseska, Philippe I H Bastiaens
Oct 14, 2020·Journal of Cellular and Molecular Medicine·Zhiyuan LuBaobing Zhao
Jun 18, 2021·Nano-micro Letters·Yuzhen OuyangMinghua Wu
Mar 20, 2018·Biochemistry·Steven D QuinnGabriela S Schlau-Cohen
Oct 4, 2019·Analytical Chemistry·Rong ZhaoXiaohong Fang
Oct 24, 2020·Biophysical Journal·Ovia M ThirukkumaranHideaki Mizuno

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Methods Mentioned

BETA
x-ray crystallography
fluorescence imaging
FRET
transfection
imaging techniques

Software Mentioned

Elements
spFRET
LabVIEW
NIS
TRI2

Related Concepts

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