process of lamellipodium. The source of the fluctuation has not been elucidated in
that study. Inhibition of the polymerization by cytochalasin or by depletion of ATP
significantly lowered the degree of fluctuation. These results are consistent with the
energy-requiring actin turnover is involved in the protrusive process, but it may
simply indicate the requirement of the intact network structure.
Cell is a multi-scale entity and fluctuations must occur on a wide range of spatial
and temporal scales. This is indeed the case, because fluctuations measured by
optical microscopy exhibit various spectral characteristics [170–172]. Some fluctuations that have been revealed in other studies is an order of magnitude lower than
that observed with the optical trapping technique, but the fluctuation detected on the
apical (top) surface of the cell in one study [171] seems to have similar frequency
range (a few Hz). Thus, although the location of the fluctuation was different
between two studies, these fluctuations may share a common mechanism.
7.18.3 What Fluctuates?
One may attribute the difference between the results obtained with AFM technique
and the optical trapping technique to the different cell type (ie.,
keratocyte vs. fibroblast). But the commonality of the basic construction of
lamellipodia of these cells (actin network wrapped in the cell membrane) and
necessity of actin polymerization for the protrusion make this notion unlikely.
Suppose the fluctuation-aided lamellipdial protrusion did occur, it will be natural
to assume that both the fluctuation of lipid membrane and the fluctuation of actin
filament were involved in the protrusive process. The lipid bilayer membrane is
highly flexible: its out-of-plane amplitude of thermal fluctuation [85, 173] is estimated to be ~7 nm for 1 μm  1 μm membrane (~size of the probe-bead used in the
optical trapping measurement) in the presence of a lateral membrane tension
(0.01 pN/nm; [174]). Hence, the thermal fluctuation of the membrane will create
the gap sufficiently large for the insertion of an actin monomer. However, since the
membrane tension is on the same or only slightly higher than the trap stiffness, the
membrane fluctuation will be readily suppressed by the bead in a stiffer trap (k is up
to 0.1 pN/nm).
On the other hand, the cantilever of the atomic force microscope is much stiffer
(10–100 pN/nm). Thus, the membrane will be unable to fluctuate under the AFM
cantilever. In this case, however, the fluctuation of actin filament will allow the
growth of the actin filament as has been proposed. If a single 30 nm actin filament
bends, the stiffness will be 0.01 pN/nm [175, 176]. The bending creates the ~1.8 nm
gap between the filament tip and the cell membrane (assumed to be flat). This is
comparable to the gap created by the fluctuating membrane. Of course, the stiffness
of the cantilever is much larger than that of a single actin filament, but as mentioned
earlier, the load can be borne by many filaments (% 100 filaments per μm edge of
lamellipodium [95, 127]. Hence, the growth of the will be possible. This notion is
consistent with the result of an experiment using purified actin and ActA [177].
7.18 Polymerization Force Developed by Lamellipodium
139
that study. Inhibition of the polymerization by cytochalasin or by depletion of ATP
significantly lowered the degree of fluctuation. These results are consistent with the
energy-requiring actin turnover is involved in the protrusive process, but it may
simply indicate the requirement of the intact network structure.
Cell is a multi-scale entity and fluctuations must occur on a wide range of spatial
and temporal scales. This is indeed the case, because fluctuations measured by
optical microscopy exhibit various spectral characteristics [170–172]. Some fluctuations that have been revealed in other studies is an order of magnitude lower than
that observed with the optical trapping technique, but the fluctuation detected on the
apical (top) surface of the cell in one study [171] seems to have similar frequency
range (a few Hz). Thus, although the location of the fluctuation was different
between two studies, these fluctuations may share a common mechanism.
7.18.3 What Fluctuates?
One may attribute the difference between the results obtained with AFM technique
and the optical trapping technique to the different cell type (ie.,
keratocyte vs. fibroblast). But the commonality of the basic construction of
lamellipodia of these cells (actin network wrapped in the cell membrane) and
necessity of actin polymerization for the protrusion make this notion unlikely.
Suppose the fluctuation-aided lamellipdial protrusion did occur, it will be natural
to assume that both the fluctuation of lipid membrane and the fluctuation of actin
filament were involved in the protrusive process. The lipid bilayer membrane is
highly flexible: its out-of-plane amplitude of thermal fluctuation [85, 173] is estimated to be ~7 nm for 1 μm  1 μm membrane (~size of the probe-bead used in the
optical trapping measurement) in the presence of a lateral membrane tension
(0.01 pN/nm; [174]). Hence, the thermal fluctuation of the membrane will create
the gap sufficiently large for the insertion of an actin monomer. However, since the
membrane tension is on the same or only slightly higher than the trap stiffness, the
membrane fluctuation will be readily suppressed by the bead in a stiffer trap (k is up
to 0.1 pN/nm).
On the other hand, the cantilever of the atomic force microscope is much stiffer
(10–100 pN/nm). Thus, the membrane will be unable to fluctuate under the AFM
cantilever. In this case, however, the fluctuation of actin filament will allow the
growth of the actin filament as has been proposed. If a single 30 nm actin filament
bends, the stiffness will be 0.01 pN/nm [175, 176]. The bending creates the ~1.8 nm
gap between the filament tip and the cell membrane (assumed to be flat). This is
comparable to the gap created by the fluctuating membrane. Of course, the stiffness
of the cantilever is much larger than that of a single actin filament, but as mentioned
earlier, the load can be borne by many filaments (% 100 filaments per μm edge of
lamellipodium [95, 127]. Hence, the growth of the will be possible. This notion is
consistent with the result of an experiment using purified actin and ActA [177].
7.18 Polymerization Force Developed by Lamellipodium
139