Chemistry Reference
In-Depth Information
Figure 6.2 Reconstructed, sectioned image from the
programmable array microscope (PAM) showing distribution
of EGF-bound EGFR on A431 cells. The cells stably express
EGFR
EGF (red) bind to the EGFR on the
filopodia and the cell membrane. The image is a maximum
intensity projection of 19 focal planes acquired using an exposure
of
eGFP (green). QD655
-
-
16ms per slice with a spacing of 0.5
m
m.
an activated EGF receptor dimer was necessary to maintain the transport process
(Figure 6.3, taken from Figure 4 in [3]).
The actin bundles in
filopodia undergo growth and exchange by addition of
monomers to the plus ends and depolymerization from the minus ends, a process
referred to as treadmilling. In combination with the active pulling of actin
filaments
by myosin, treadmilling results in a net
flow of F-actin towards the interior of the cell.
Association of macromolecules with an actin
filament leads to a translocation of the
cargo towards the cell body. Speci
c inhibitors of the EGFR RTK as well as
cytochalasin D, a disruptor of the actin cytoskeleton, abolish transport but not free
diffusion of the receptor
-
ligand complex. The coupling of the QD
-
EGF
-
EGFR
complex to this retrograde
flow was further corroborated by photobleaching of
actin
-
EGFP bundles during transport and correlation of the movement of the QDs
and the bleached actin segment (see Figure 3 in [3]). Fromdata such as these we were
able to calculate the velocity of retrograde transport to be 20
8 nm/s.
In the same studies, it was demonstrated that retrograde transport precedes
receptor internalization, which occurs at the base of the
filopodia. The fact that
initiation of transport requires the cooperative interaction of two or more activated
