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Vol. 10, Issue 6, 2017-2031, June 1999
and
*Boulder Laboratory for Three-dimensional Fine Structure and
The spindle pole body (SPB) is the major microtubule-organizing
center of budding yeast and is the functional equivalent of the
centrosome in higher eukaryotic cells. We used fast-frozen, freeze-substituted cells in conjunction with high-voltage electron tomography to study the fine structure of the SPB and the events of
early spindle formation. Individual structures were imaged at 5-10 nm
resolution in three dimensions, significantly better than can be
achieved by serial section electron microscopy. The SPB is organized in
distinct but coupled layers, two of which show ordered two-dimensional
packing. The SPB central plaque is anchored in the nuclear envelope
with hook-like structures. The minus ends of nuclear microtubules (MTs)
are capped and are tethered to the SPB inner plaque, whereas the
majority of MT plus ends show a distinct flaring. Unbudded cells
containing a single SPB retain 16 MTs, enough to attach to each of the
expected 16 chromosomes. Their median length is ~150 nm. MTs growing
from duplicated but not separated SPBs have a median length of ~130
nm and interdigitate over the bridge that connects the SPBs. As a
bipolar spindle is formed, the median MT length increases to ~300 nm
and then decreases to ~30 nm in late anaphase. Three-dimensional
models confirm that there is no conventional metaphase and that
anaphase A occurs. These studies complement and extend what is known
about the three-dimensional structure of the yeast mitotic spindle and
further our understanding of the organization of the SPB in intact cells.
Department of Molecular, Cellular, and Developmental
Biology, University of Colorado, Boulder, Colorado 80309-0347
Online version of this
article contains video material for Figures 2 and 10. Online version
available at www.molbiolcell.org.
Corresponding author. E-mail address:
eileen{at}bio3d.colorado.edu.
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