In-cell structural dynamics of an EGF receptor during ligand-induced dimer–oligomer transition

The epidermal growth factor receptor (EGFR) is a membrane protein that regulates cell proliferation, differentiation and survival, and is a drug target for cancer therapy. Ligand-induced activation of the EGFR kinase is generally regarded to require ligand-bound-dimers, while phosphorylation and dow...

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Bibliographic Details
Published in:European biophysics journal 2020-01, Vol.49 (1), p.21-37
Main Authors: Kozer, Noga, Clayton, Andrew H. A.
Format: Article
Language:English
Subjects:
Activation
Binding
Biochemistry
Biological and Medical Physics
Biomedical and Life Sciences
Biophysics
Cell Biology
Cell differentiation
Cell proliferation
Cell survival
Correlation
Depolarization
Dimerization
Dimers
Dynamics
Epidermal growth factor
Epidermal growth factor receptors
Growth factors
Kinases
Life Sciences
Ligands
Membrane Biology
Membrane proteins
Membranes
Nanotechnology
Neurobiology
Oligomerization
Oligomers
Original Article
Phosphorylation
Stochasticity
Therapeutic targets
Variation
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Summary:The epidermal growth factor receptor (EGFR) is a membrane protein that regulates cell proliferation, differentiation and survival, and is a drug target for cancer therapy. Ligand-induced activation of the EGFR kinase is generally regarded to require ligand-bound-dimers, while phosphorylation and down-stream signalling is modulated by oligomers. Recent work has unveiled changes in EGFR dynamics from ligand-induced dimerization in membranes extracted from cells, however, less is known about the changes in EGFR dynamics that accompany the ligand-induced oligomerization in a live cell environment. Here, we determine the dynamics of a c-terminal GFP tag attached to EGFR in the unliganded dimer and in the liganded oligomers. By means of the single-frequency polarized phasor ellipse approach we extracted two correlation times on the sub-nanosecond and super-nanosecond timescales, respectively. EGF binding to the EGFR–GFP dimer lengthened the sub-nanosecond correlation time (from 0.1 to 1.3 ns) and shortened the super-nanosecond correlation time (from 210 to 56 ns) of the c-terminal GFP probe. The sub-nanosecond depolarization processes were assigned to electronic energy migration between proximal GFPs in the EGFR dimer or oligomer, while the super-nanosecond correlation times were assigned to nanosecond fluctuations of the GFP probe in the EGFR complex. Accordingly, these results show that ligand binding increased the average separation between the c-terminal tags and increased their rotational mobility. We propose that the dynamics are linked to an inhibitory function of the c-terminal tail in the un-liganded dimer and to the requirement of facile stochastic switching between kinase activation and cytoplasmic adaptor/effector binding in the active oligomers.
ISSN:0175-7571
1432-1017
DOI:10.1007/s00249-019-01410-2