Biomedical Engineering Reference
In-Depth Information
Why may pushing forces be used in certain systems and pulling forces in
others? One important aspect is probably the size of the system. It is simply
not possible to eciently generate pushing forces when microtubules become
too long. A growing microtubule can move a microtubule-organizing center
through the cell, and withstand counteracting forces up to its stall force, as
long as the counteracting force is less than the critical buckling force of the
microtubule. When a microtubule buckles, the pushing force on the organizing
center is decreased and continued growth of the microtubule no longer nec-
essarily leads to forward motion of the organizing center. Long microtubules
buckle more easily than short microtubules, which is probably why small fis-
sion yeast cells can use microtubule pushing to position the nucleus, whereas
spindles in the much larger
Celegans
embryos need to rely on pulling forces.
One remark to make, however, is that long microtubules in large cells do
not necessarily buckle at the critical buckling force associated with their full
length. Microtubules in cells are embedded in a visco-elastic environment due
to the presence of (for example) the actin cytoskeleton. Elastic deformation
of this network may prevent large amplitude bending of microtubules leading
to shorter wavelength buckling and consequent increased resistance to force
[41].
Recent modeling efforts have shown that positioning processes based on
pulling forces cannot be stable in the absence of restoring (pushing) forces [40,
42]. The intuitive reason is that as soon as an organizing center moves closer
to one side of the cell, more microtubule-cortex interactions become possible,
leading to larger forces in that same direction (assuming that an equal number
of microtubules are nucleated in all directions and that every microtubule-
cortex contact, or a fixed percentage thereof, generates only a pulling force). In
contrast, in the case of pushing, more interaction automatically leads to larger
forces in the opposite direction, and a stable situation is reached when the
organizing center sits (on average) in the middle [37]. In
Celegans
embryos,
it appears that not all microtubules interact with cortical force generating
sites. It is therefore likely that antagonistic pushing forces are generated by
the other microtubules and that the balance between them, combined with
regulation of the actual number of pulling-force generators on the anterior and
posterior sides controls the final (asymmetric) position of the spindle [43].
To study the antagonistic effect of pulling and pushing forces in a simpli-
fied experimental setting, we recently modified our microfabricated chamber
experiments to be able to introduce localized pulling activities at the cham-
ber boundaries (see Figure 4.11b). Instead of using chambers made from pure
silicon monoxide (SiO), as we normally do, we etch these chambers into de-
posited multilayers of SiO (1.5 micron thick), chromium (thin layer to help
adhere the gold), gold (between 100 and 800 nm), chromium, and SiO again.
This provides us with chambers with a rim of gold along the edges to which
we can specifically react biotin tags after assembling a monolayer of thiol-
terminated reactive groups to the gold. Using streptavidin as an intermediate
layer, we then attach biotinylated dynein molecules (a gift from Sam Reck-
