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Budding yeast chromosome structure and dynamics during mitosis.

Pearson CG, Maddox PS, Salmon ED, Bloom K - J. Cell Biol. (2001)

Bottom Line: Centromeres are in a metaphase-like conformation, whereas chromosome arms are neither aligned nor separated before anaphase.The stretched chromatin was observed to segregate to the spindle pole bodies at rates greater than centromere to pole movement, indicative of rapid elastic recoil between the chromosome arm and the centromere.These results indicate that the elastic properties of DNA play an as of yet undiscovered role in the poleward movement of chromosome arms.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, USA. cgpearso@email.unc.edu

ABSTRACT
Using green fluorescent protein probes and rapid acquisition of high-resolution fluorescence images, sister centromeres in budding yeast are found to be separated and oscillate between spindle poles before anaphase B spindle elongation. The rates of movement during these oscillations are similar to those of microtubule plus end dynamics. The degree of preanaphase separation varies widely, with infrequent centromere reassociations observed before anaphase. Centromeres are in a metaphase-like conformation, whereas chromosome arms are neither aligned nor separated before anaphase. Upon spindle elongation, centromere to pole movement (anaphase A) was synchronous for all centromeres and occurred coincident with or immediately after spindle pole separation (anaphase B). Chromatin proximal to the centromere is stretched poleward before and during anaphase onset. The stretched chromatin was observed to segregate to the spindle pole bodies at rates greater than centromere to pole movement, indicative of rapid elastic recoil between the chromosome arm and the centromere. These results indicate that the elastic properties of DNA play an as of yet undiscovered role in the poleward movement of chromosome arms.

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Centromere proximal spots exhibited dynamic separation and oscillations along the preanaphase mitotic spindle. The lacO marker was integrated ∼1.1 kb from CEN11 with Spc72–GFP-labeled spindle pole bodies. (A) Selected frames of an ∼7.5-min single-plane time-lapse. (B) A kymograph sequence of the entire time-lapse collected at ∼0.9-s intervals. Narrow tick marks indicate corresponding time points to the above selected frames. (C) Graphical plot of the above time course. In both A and B, arrowheads denote the spindle pole bodies, and arrows define the centromere proximal chromosome spots. Elapsed time provided in seconds. Bar, 2 μm.
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Figure 2: Centromere proximal spots exhibited dynamic separation and oscillations along the preanaphase mitotic spindle. The lacO marker was integrated ∼1.1 kb from CEN11 with Spc72–GFP-labeled spindle pole bodies. (A) Selected frames of an ∼7.5-min single-plane time-lapse. (B) A kymograph sequence of the entire time-lapse collected at ∼0.9-s intervals. Narrow tick marks indicate corresponding time points to the above selected frames. (C) Graphical plot of the above time course. In both A and B, arrowheads denote the spindle pole bodies, and arrows define the centromere proximal chromosome spots. Elapsed time provided in seconds. Bar, 2 μm.

Mentions: To measure the dynamic movements of centromeres in preanaphase cells, we recorded the ∼1.1-kb centromere proximal marker movements relative to GFP-labeled spindle pole bodies using rapid frame acquisition rates (∼1 frame/s) (Fig. 2). Fig. 2 shows that, in addition to dynamic movements between the separated chromosome spots, there were also distinctive oscillations of the centromere proximal spots along the mitotic spindle with Spc72–GFP-labeled spindle pole bodies. Two classes of centromere proximal spot movements are evident in Fig. 2: major oscillatory movements relative to the spindle pole body (Fig. 2 C, directed oscillation) and minor movements of <0.25 μm that may be due to diffusional motion. Analyses were repeated three times for each time-lapse sequence, and the mean was displayed, indicating that the smaller movements are not a product of tracking errors (Fig. 2 C). The centromere proximal markers separated ≤1.26 μm (Fig. 2 C, time point = 70.3 s) and reassociated with each other before reseparation (Fig. 2 C, time point = 199.0 s). There was no apparent strong coordination of the separated centromere spot oscillations relative to the spindle pole bodies, as determined by the independent movement of each spot relative to its sister (Fig. 2 C, directed oscillation). However, we did occasionally observe both separated spots to move toward a single pole in what appeared to be coordinated movements toward the spindle pole body (data not shown).


Budding yeast chromosome structure and dynamics during mitosis.

Pearson CG, Maddox PS, Salmon ED, Bloom K - J. Cell Biol. (2001)

Centromere proximal spots exhibited dynamic separation and oscillations along the preanaphase mitotic spindle. The lacO marker was integrated ∼1.1 kb from CEN11 with Spc72–GFP-labeled spindle pole bodies. (A) Selected frames of an ∼7.5-min single-plane time-lapse. (B) A kymograph sequence of the entire time-lapse collected at ∼0.9-s intervals. Narrow tick marks indicate corresponding time points to the above selected frames. (C) Graphical plot of the above time course. In both A and B, arrowheads denote the spindle pole bodies, and arrows define the centromere proximal chromosome spots. Elapsed time provided in seconds. Bar, 2 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2199205&req=5

Figure 2: Centromere proximal spots exhibited dynamic separation and oscillations along the preanaphase mitotic spindle. The lacO marker was integrated ∼1.1 kb from CEN11 with Spc72–GFP-labeled spindle pole bodies. (A) Selected frames of an ∼7.5-min single-plane time-lapse. (B) A kymograph sequence of the entire time-lapse collected at ∼0.9-s intervals. Narrow tick marks indicate corresponding time points to the above selected frames. (C) Graphical plot of the above time course. In both A and B, arrowheads denote the spindle pole bodies, and arrows define the centromere proximal chromosome spots. Elapsed time provided in seconds. Bar, 2 μm.
Mentions: To measure the dynamic movements of centromeres in preanaphase cells, we recorded the ∼1.1-kb centromere proximal marker movements relative to GFP-labeled spindle pole bodies using rapid frame acquisition rates (∼1 frame/s) (Fig. 2). Fig. 2 shows that, in addition to dynamic movements between the separated chromosome spots, there were also distinctive oscillations of the centromere proximal spots along the mitotic spindle with Spc72–GFP-labeled spindle pole bodies. Two classes of centromere proximal spot movements are evident in Fig. 2: major oscillatory movements relative to the spindle pole body (Fig. 2 C, directed oscillation) and minor movements of <0.25 μm that may be due to diffusional motion. Analyses were repeated three times for each time-lapse sequence, and the mean was displayed, indicating that the smaller movements are not a product of tracking errors (Fig. 2 C). The centromere proximal markers separated ≤1.26 μm (Fig. 2 C, time point = 70.3 s) and reassociated with each other before reseparation (Fig. 2 C, time point = 199.0 s). There was no apparent strong coordination of the separated centromere spot oscillations relative to the spindle pole bodies, as determined by the independent movement of each spot relative to its sister (Fig. 2 C, directed oscillation). However, we did occasionally observe both separated spots to move toward a single pole in what appeared to be coordinated movements toward the spindle pole body (data not shown).

Bottom Line: Centromeres are in a metaphase-like conformation, whereas chromosome arms are neither aligned nor separated before anaphase.The stretched chromatin was observed to segregate to the spindle pole bodies at rates greater than centromere to pole movement, indicative of rapid elastic recoil between the chromosome arm and the centromere.These results indicate that the elastic properties of DNA play an as of yet undiscovered role in the poleward movement of chromosome arms.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, USA. cgpearso@email.unc.edu

ABSTRACT
Using green fluorescent protein probes and rapid acquisition of high-resolution fluorescence images, sister centromeres in budding yeast are found to be separated and oscillate between spindle poles before anaphase B spindle elongation. The rates of movement during these oscillations are similar to those of microtubule plus end dynamics. The degree of preanaphase separation varies widely, with infrequent centromere reassociations observed before anaphase. Centromeres are in a metaphase-like conformation, whereas chromosome arms are neither aligned nor separated before anaphase. Upon spindle elongation, centromere to pole movement (anaphase A) was synchronous for all centromeres and occurred coincident with or immediately after spindle pole separation (anaphase B). Chromatin proximal to the centromere is stretched poleward before and during anaphase onset. The stretched chromatin was observed to segregate to the spindle pole bodies at rates greater than centromere to pole movement, indicative of rapid elastic recoil between the chromosome arm and the centromere. These results indicate that the elastic properties of DNA play an as of yet undiscovered role in the poleward movement of chromosome arms.

Show MeSH
Related in: MedlinePlus