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individual mitotic microtubule, due to its short length and numerous adjacent con-
founding neighbor microtubules, is relatively difficult to track. Indeed, most of the
mitotic microtubules cannot be tracked with precision. However, infrequently, there
are individual mitotic microtubules which grow longer and away from the dense
monopolar spindle core. These longer mitotic microtubules can be tracked with pre-
cision. This does imply that individual mitotic measurements will tend to bias toward
the longer microtubules. Nevertheless, useful measurements can still be made from
individual mitotic microtubule dynamics as a comparison to other mutant mitotic
microtubules or to interphase microtubules.
The microtubule length versus time plot provides information to derive all
parameters of microtubule dynamics, including velocities of growth and shrink-
age, and frequencies of catastrophe and rescue (
Walker et al., 1988
). Statistical
significance and inference will require many measurements. Here, we choose
to present one example to make a point.
Figure 24.2
B highlights the contrast in
length and life time of an interphase and mitotic microtubule. Qualitatively, the
velocities of growth and shrinkage (i.e., similar growth and shrinkage slopes)
are similar between interphase and mitotic microtubules, and their time to catas-
trophe (i.e., inverse of catastrophe frequency) is very different.
Figure 24.2
C
shows comparative length differences between an individual mitotic microtubule
of the control cut7.24
ts
and an individual mitotic microtubule of the mutant
cut7.24
ts
:csi2
. Qualitatively, the mutant mitotic microtubule appears longer.
Figure 24.2
D shows that the velocities of growth and shrinkage are similar be-
tween mitotic microtubules of the control and mutant cell. However, the mutant
mitotic microtubule grows a longer time before catastrophe. This suggests that
csi2p regulates mitotic microtubule dynamics. Specifically, csi2p positively reg-
ulates microtubule frequency of catastrophe.
D
CONCLUSION
We presented a simple method to image and analyze individual spindle microtubule
dynamics in the fission yeast. This method overcomes the spatial limitation of having
a dense microtubule structure within the diffraction-limited spindle width and spin-
dle poles. In principle, this method could be applied generally to other yeasts and
fungi having similar spindle structures, provided that they have a thermosensitive
kinesin-5 as the motor controlling spindle bipolarity.
Acknowledgments
We thank Dr. Andrea Stout of the Penn CDB Imaging Core for technical support. J. C. is sup-
ported by a predoctoral fellowship from the FCT-Portugal. This work is supported by grants
from the NIH and ANR.
