Interplay Between Spindle Architecture and Function - International Review of Cell and Molecular Biology - page 92

Further support for a model of spindle organization with homogenous
MT stability in all regions stems from using a femtosecond laser ablation tech-
nique to cut a subset of MTs within a metaphase Xenopus egg extract spindle
( Brugues et al., 2012 ; Fig. 3.3 D). The cut MTs depolymerized from their
newly generated plus-ends at the site of laser irradiation, generating waves
of depolymerization moving toward each spindle pole, which could be
observed by tracking the changes in MT intensity over time. Quantitative
information was derived about the MT population from the wave decay, such
as the density of minus-ends. By comparing the two waves, information about
the polarity of MTs could be obtained, and irradiating different regions of the
spindle showed that MTs were longer near the center of the spindle and
shorter toward the poles. Interestingly, inhibition of poleward MT flux
homogenized spindleMT lengths, supporting a model in which spindle archi-
tecture depends on both the position of MT nucleation near the center of the
spindle and poleward MT transport ( Brugues et al., 2012 ).
Together, these studies provide clues as to how global features such as
spindle size and shape are established and maintained over timescales much
longer than the MT lifetime and over distances much longer than the aver-
age spindle MT. Due to its large size and accessibility for perturbation, the
Xenopus egg extract spindle has been investigated most intensively, and in the
future, it will be critical to elucidate howMT nucleation, dynamics, and flux
operate to produce a functional spindle and how these properties are mod-
ulated in different spindle types.
3. NUCLEATION AND ORIGIN OF MICROTUBULES IN
THE SPINDLE
There are two recognized spindle assembly pathways, “search-and--
capture” and “spindle self-organization,” which are not mutually exclusive
and are thought to operate simultaneously in most cases to promote robust
spindle formation. These two pathways are broadly defined by where MTs
nucleate. Search-and-capture occurs by selective stabilization of MTs nucle-
ated at the MTOC or centrosome and self-organization describes any number
of modes of nucleation followed by incorporation of the MTs into a bipolar
array by the activity of associated proteins including MT-based motors. Less
well understood, however, is what determines the relative role of a pathway or
the preferred types of MT nucleation in a particular cell type and how these
pathways are interregulated and contribute to spindle architecture. Depending