Biomedical Engineering Reference
In-Depth Information
of the lamellipodium in the cell frame of reference was nearly equal to cell
speed with only a small retrograde flow of the actin meshwork relative to the
substrate [31, 62, 63]. These results show that in keratocytes the actin cy-
toskeleton is firmly anchored to the substrate so that actin polymerization is
directly coupled to protrusion at the leading edge. Note the coherence of the
observed flow across the lamellipodium, indicating that the actin meshwork
moved as a single, relatively rigid, mechanical unit. The actin network at the
rear of the cell displayed a rapid inward flow toward the cell body, with most
of the motion in the cell frame of reference perpendicular to the direction of
motion (see Figure 2.4c). This inward flow was probably due to forces gener-
ated by myosin contraction pulling the actin meshwork (see below), whereas
the retrograde flow in the central lamellipodium in keratocytes was largely
caused by actin protrusion at the leading edge, although some contribution
from myosin mediated contraction is possible.
The observed kinetics of actin assembly and disassembly within a cell are
dramatically accelerated compared to the rates measured for pure actin
in
vitro
, due to the action of a large number of actin-associated proteins, many
of which have been shown to be required for the protrusion process. These
include actin capping proteins, profilin, and ADF/cofilin. Capping proteins
bind to barbed ends and terminates their growth. This activity regulates the
length and the number of active filaments and contributes to the formation of
the brushlike zone in the actin network at the leading edge (see Figure 2.3c).
Profilin binds actin monomers with a one-to-one stoichiometry and acts as a
monomer sequestering protein, to allow higher actin monomer concentrations.
At the same time, profilin-actin complexes can readily be incorporated at the
barbed ends of growing filaments [64, 65]. ADF/cofilin is required for rapid
filament disassembly [66]. Together, these proteins allow the cell to maintain
a high concentration of actin monomers that can rapidly add on to the barbed
ends of actin filaments. These components, together with actin and Arp2/3,
were originally shown to be necessary and sucient for reconstituting the
formation of actin “comet tails” and motility of
Listeria monocytogenes
[9],
and have since been found to be required for the formation and protrusion of
lamellipodia in tissue culture cells [67]. Protrusion at the leading edge is fur-
ther regulated by a number of other auxiliary proteins, such as the Ena/VASP
protein family [68, 69, 70]. Further details regarding the biochemistry of actin
protrusion are described elsewhere [5, 6, 7, 45].
The spatially regulated dynamics of the actin network are responsible for
protrusion at the leading edge. The local rate of protrusion will depend on
the load force acting on a growing filament, as well as other force-independent
factors. The load force is determined by the pressure imposed by the mem-
brane tension and the differences in osmotic/hydrostatic pressures across the
cell membrane. The load force on an individual filament is inversely propor-
tional to the local filament density, because the load is distributed among
the filaments. Force-independent factors affecting protrusion include the local
concentration of actin monomers (as well as the local concentrations of acces-
