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The budding yeast Ipl1/Aurora protein kinase regulates mitotic spindle disassembly.

Buvelot S, Tatsutani SY, Vermaak D, Biggins S - J. Cell Biol. (2003)

Bottom Line: As the spindle disassembles, Ipl1p follows the plus ends of the depolymerizing spindle microtubules.Many Ipl1p substrates colocalize with Ipl1p to the spindle midzone, identifying additional proteins that may regulate spindle disassembly.We propose that Ipl1p regulates both the kinetochore and interpolar microtubule plus ends to regulate its various mitotic functions.

View Article: PubMed Central - PubMed

Affiliation: Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.

ABSTRACT
Ipl1p is the budding yeast member of the Aurora family of protein kinases, critical regulators of genomic stability that are required for chromosome segregation, the spindle checkpoint, and cytokinesis. Using time-lapse microscopy, we found that Ipl1p also has a function in mitotic spindle disassembly that is separable from its previously identified roles. Ipl1-GFP localizes to kinetochores from G1 to metaphase, transfers to the spindle after metaphase, and accumulates at the spindle midzone late in anaphase. Ipl1p kinase activity increases at anaphase, and ipl1 mutants can stabilize fragile spindles. As the spindle disassembles, Ipl1p follows the plus ends of the depolymerizing spindle microtubules. Many Ipl1p substrates colocalize with Ipl1p to the spindle midzone, identifying additional proteins that may regulate spindle disassembly. We propose that Ipl1p regulates both the kinetochore and interpolar microtubule plus ends to regulate its various mitotic functions.

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Ipl1 localization is similar to chromosomal passenger proteins. Live microscopy was performed on cells containing Ipl1–GFP (SBY556). DIC pictures are shown to the left of each corresponding fluorescence picture. In G1, S phase, and metaphase cells, Ipl1p localizes in a discrete dot corresponding to kinetochores. In anaphase and telophase cells, Ipl1p localizes along the whole spindle (top right), at the spindle midzone (middle right), or near the spindle poles (bottom right). Bar, 10 μm.
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fig1: Ipl1 localization is similar to chromosomal passenger proteins. Live microscopy was performed on cells containing Ipl1–GFP (SBY556). DIC pictures are shown to the left of each corresponding fluorescence picture. In G1, S phase, and metaphase cells, Ipl1p localizes in a discrete dot corresponding to kinetochores. In anaphase and telophase cells, Ipl1p localizes along the whole spindle (top right), at the spindle midzone (middle right), or near the spindle poles (bottom right). Bar, 10 μm.

Mentions: Ipl1p is required for chromosome segregation and the spindle checkpoint when kinetochores are not under tension. To learn more about Ipl1p functions, we examined the localization of an endogenous Ipl1p fusion to GFP at the COOH terminus in living cells using bud size as a marker for cell cycle stage (Fig. 1). In unbudded G1 cells, Ipl1–GFP localizes as a dot or small line. In small budded S phase cells, Ipl1p localizes as a distinct dot that separates into two discrete dots in medium budded metaphase cells and corresponds to precociously separated kinetochores (Goshima and Yanagida, 2000; He et al., 2000, 2001; Tanaka et al., 2000; Biggins and Murray, 2001; Pearson et al., 2001). In large budded cells, Ipl1–GFP exhibits dynamic localization patterns. In some cells, Ipl1–GFP is on the whole spindle but absent from the kinetochores that are clustered at the poles as reported previously (Tanaka et al., 2002). We also found two sites of Ipl1p localization that had not been seen before. Ipl1–GFP is found at the spindle midzone and in small tufts that vary in shape near the spindle poles at telophase. This likely represents Ipl1p bound to the remnants of depolymerized microtubules (Winey et al., 1995). Since the Ipl1–GFP fusion created a temperature-sensitive protein, we also localized endogenous Ipl1p. Immunofluorescence with anti-Ipl1p antibodies on chromosome spreads showed the same localization patterns as the Ipl1–GFP fusion protein (Loidl et al., 1998) (Fig. S1 available at http://www.jcb.org/cgi/content/full/jcb.200209018/DC1). Therefore, Ipl1p localization is similar to Aurora B and other “chromosomal passenger” proteins.


The budding yeast Ipl1/Aurora protein kinase regulates mitotic spindle disassembly.

Buvelot S, Tatsutani SY, Vermaak D, Biggins S - J. Cell Biol. (2003)

Ipl1 localization is similar to chromosomal passenger proteins. Live microscopy was performed on cells containing Ipl1–GFP (SBY556). DIC pictures are shown to the left of each corresponding fluorescence picture. In G1, S phase, and metaphase cells, Ipl1p localizes in a discrete dot corresponding to kinetochores. In anaphase and telophase cells, Ipl1p localizes along the whole spindle (top right), at the spindle midzone (middle right), or near the spindle poles (bottom right). Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172676&req=5

fig1: Ipl1 localization is similar to chromosomal passenger proteins. Live microscopy was performed on cells containing Ipl1–GFP (SBY556). DIC pictures are shown to the left of each corresponding fluorescence picture. In G1, S phase, and metaphase cells, Ipl1p localizes in a discrete dot corresponding to kinetochores. In anaphase and telophase cells, Ipl1p localizes along the whole spindle (top right), at the spindle midzone (middle right), or near the spindle poles (bottom right). Bar, 10 μm.
Mentions: Ipl1p is required for chromosome segregation and the spindle checkpoint when kinetochores are not under tension. To learn more about Ipl1p functions, we examined the localization of an endogenous Ipl1p fusion to GFP at the COOH terminus in living cells using bud size as a marker for cell cycle stage (Fig. 1). In unbudded G1 cells, Ipl1–GFP localizes as a dot or small line. In small budded S phase cells, Ipl1p localizes as a distinct dot that separates into two discrete dots in medium budded metaphase cells and corresponds to precociously separated kinetochores (Goshima and Yanagida, 2000; He et al., 2000, 2001; Tanaka et al., 2000; Biggins and Murray, 2001; Pearson et al., 2001). In large budded cells, Ipl1–GFP exhibits dynamic localization patterns. In some cells, Ipl1–GFP is on the whole spindle but absent from the kinetochores that are clustered at the poles as reported previously (Tanaka et al., 2002). We also found two sites of Ipl1p localization that had not been seen before. Ipl1–GFP is found at the spindle midzone and in small tufts that vary in shape near the spindle poles at telophase. This likely represents Ipl1p bound to the remnants of depolymerized microtubules (Winey et al., 1995). Since the Ipl1–GFP fusion created a temperature-sensitive protein, we also localized endogenous Ipl1p. Immunofluorescence with anti-Ipl1p antibodies on chromosome spreads showed the same localization patterns as the Ipl1–GFP fusion protein (Loidl et al., 1998) (Fig. S1 available at http://www.jcb.org/cgi/content/full/jcb.200209018/DC1). Therefore, Ipl1p localization is similar to Aurora B and other “chromosomal passenger” proteins.

Bottom Line: As the spindle disassembles, Ipl1p follows the plus ends of the depolymerizing spindle microtubules.Many Ipl1p substrates colocalize with Ipl1p to the spindle midzone, identifying additional proteins that may regulate spindle disassembly.We propose that Ipl1p regulates both the kinetochore and interpolar microtubule plus ends to regulate its various mitotic functions.

View Article: PubMed Central - PubMed

Affiliation: Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.

ABSTRACT
Ipl1p is the budding yeast member of the Aurora family of protein kinases, critical regulators of genomic stability that are required for chromosome segregation, the spindle checkpoint, and cytokinesis. Using time-lapse microscopy, we found that Ipl1p also has a function in mitotic spindle disassembly that is separable from its previously identified roles. Ipl1-GFP localizes to kinetochores from G1 to metaphase, transfers to the spindle after metaphase, and accumulates at the spindle midzone late in anaphase. Ipl1p kinase activity increases at anaphase, and ipl1 mutants can stabilize fragile spindles. As the spindle disassembles, Ipl1p follows the plus ends of the depolymerizing spindle microtubules. Many Ipl1p substrates colocalize with Ipl1p to the spindle midzone, identifying additional proteins that may regulate spindle disassembly. We propose that Ipl1p regulates both the kinetochore and interpolar microtubule plus ends to regulate its various mitotic functions.

Show MeSH