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2015 ; 109
(9
): 1925-36
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Plasma Membrane Organization of Epidermal Growth Factor Receptor in Resting and
Ligand-Bound States
#MMPMID26536269
Bag N
; Huang S
; Wohland T
Biophys J
2015[Nov]; 109
(9
): 1925-36
PMID26536269
show ga
The spatial arrangement of the epidermal growth factor receptor (EGFR) on the
cellular plasma membrane is one of the prime factors that control its downstream
signaling pathways and related functions. However, the molecular organization,
which spans the scale from nanometers to micrometer-size clusters, has not been
resolved in detail, mainly due to a lack of techniques with the required
spatiotemporal resolution. Therefore, we used imaging total internal
reflection-fluorescence correlation spectroscopy to investigate EGFR dynamics on
live CHO-K1 plasma membranes in resting and ligand-bound states. In combination
with the fluorescence correlation spectroscopy diffusion law, this provides
information on the subresolution organization of EGFR on cell membranes. We found
that overall EGFR organization is sensitive to both cholesterol and the actin
cytoskeleton. EGFR in the resting state is partly trapped in
cholesterol-containing domains, whereas another fraction exhibits cholesterol
independent trapping on the membrane. Disruption of the cytoskeleton leads to a
broader range of EGFR diffusion coefficients and a reduction of hop diffusion. In
the ligand-bound state we found a dose-dependent behavior. At 10 ng/mL EGF the
EGFR is endocytosed and recycled to the membrane, whereas diffusion and
organization do not change significantly. At 100 ng/mL EGF the EGFR forms
clusters, which are subsequently internalized, whereas outside the clusters
diffusivity increases and the organization of the receptor remains unchanged.
After disruption of cholesterol-containing domains or actin cytoskeleton, EGF
induces microscopic EGFR clusters on the membrane and endocytosis is inhibited.