Interplay Between Spindle Architecture and Function - International Review of Cell and Molecular Biology - page 91

spindles, to drive spatial distribution of spindle-associated factors or even the
MTs themselves.
Measurements of MT transport and lifetime through single molecule and
FSM imaging have provided insight into how MT dynamics may establish
and respond to the spindle environment. By tracking correlated movements
of pairs of fluxing tubulin dimers in a Xenopus egg extract spindle, the
arrangement of MTs was predicted to be a tiled-array in which many
MTs overlap to span the distance between the spindle pole and the meta-
phase plate ( Fig. 3.3 B; Yang et al., 2008 ). This suggests that spindle MTs
must be highly cross-linked to form a robust yet dynamic structure. How-
ever, chromosomes must be able to move within the spindle to align at the
metaphase plate. An investigation into the mechanical properties of the
Xenopus egg extract spindle indicates how MT cross-linking provides struc-
tural integrity to the spindle and yet allows chromosome movement
( Fig. 3.3 C). The viscoelastic properties were probed using force-calibrated
microneedles inserted into the spindle and were shown to depend on the
amount of MT cross-linking ( Shimamoto et al., 2011 ). Moreover, the vis-
coelastic behavior was timescale dependent and indicated that chromosome
movements occur on a timescale in which elastic strain can be dissipated
through remodeling of the spindle structure, possibly by MT depolymeri-
zation and repolymerization or flux ( Shimamoto et al., 2011 ; Fig. 3.3 C).
Thus, MT dynamics within the spindle drive the size, shape, and organiza-
tion of the spindle as well as determine its micromechanical properties.
Despite the highly cross-linked nature of the spindle, there is evidence
that the spindle environment is not uniquely stabilizing or destabilizing
for MTs ( Needleman et al., 2009 ). The stability of MTs within the spindle
was determined using single molecule lifetime measurements of labeled
tubulin in spindle MTs, similar to a FSM experiment. Instead of measuring
the displacement of the fluorescent speckle, these measurements assess how
long a labeled tubulin dimer resides in a spindle MT before a catastrophe
releases it. In Xenopus egg extracts, the residence lifetimes were very similar
within a spindle compared to much sparser MT arrays nucleated from Tet-
rahymena pellicles ( Needleman et al., 2009 ). This observation indicates that
the Xenopus cytoplasm is loaded with a variety of MT-stabilizing/
destabilizing proteins that can act on MTs, regardless of their density or
cross-linking. Perhaps more interestingly, tubulin lifetime measurements
did not vary across different regions of the spindle, suggesting that despite
specific localization of factors that alter MT dynamics within the spindle,
overall MT stability is uniform.