Abstract
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.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 2017-2031 |
| Number of pages | 15 |
| Journal | Molecular Biology of the Cell |
| Volume | 10 |
| Issue number | 6 |
| DOIs | |
| State | Published - Jan 1 1999 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Molecular Biology
- Cell Biology
Cite this
High-voltage electron tomography of spindle pole bodies and early mitotic spindles in the yeast Saccharomyces cerevisiae. / O'Toole, Eileen T.; Winey, Mark; McIntosh, J. Richard.
In: Molecular Biology of the Cell, Vol. 10, No. 6, 01.01.1999, p. 2017-2031.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - High-voltage electron tomography of spindle pole bodies and early mitotic spindles in the yeast Saccharomyces cerevisiae
AU - O'Toole, Eileen T.
AU - Winey, Mark
AU - McIntosh, J. Richard
PY - 1999/1/1
Y1 - 1999/1/1
N2 - 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.
AB - 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.
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U2 - 10.1091/mbc.10.6.2017
DO - 10.1091/mbc.10.6.2017
M3 - Article
VL - 10
SP - 2017
EP - 2031
JO - Molecular Biology of the Cell
JF - Molecular Biology of the Cell
SN - 1059-1524
IS - 6
ER -