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Individual pericentromeres display coordinated motion and stretching in the yeast spindle.

Stephens AD, Snider CE, Haase J, Haggerty RA, Vasquez PA, Forest MG, Bloom K - J. Cell Biol. (2013)

Bottom Line: By labeling several chromosomes, we find that pericentromeres display coordinated motion and stretching in metaphase.The pericentromeres of different chromosomes exhibit physical linkage dependent on centromere function and structural maintenance of chromosomes complexes.Coordinated motion is dependent on condensin and the kinesin motor Cin8, whereas coordinated stretching is dependent on pericentric cohesin and Cin8.

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Affiliation: Department of Biology, 2 Department of Mathematics, and 3 Department Biomedical Engineering, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599.

ABSTRACT
The mitotic segregation apparatus composed of microtubules and chromatin functions to faithfully partition a duplicated genome into two daughter cells. Microtubules exert extensional pulling force on sister chromatids toward opposite poles, whereas pericentric chromatin resists with contractile springlike properties. Tension generated from these opposing forces silences the spindle checkpoint to ensure accurate chromosome segregation. It is unknown how the cell senses tension across multiple microtubule attachment sites, considering the stochastic dynamics of microtubule growth and shortening. In budding yeast, there is one microtubule attachment site per chromosome. By labeling several chromosomes, we find that pericentromeres display coordinated motion and stretching in metaphase. The pericentromeres of different chromosomes exhibit physical linkage dependent on centromere function and structural maintenance of chromosomes complexes. Coordinated motion is dependent on condensin and the kinesin motor Cin8, whereas coordinated stretching is dependent on pericentric cohesin and Cin8. Linking of pericentric chromatin through cohesin, condensin, and kinetochore microtubules functions to coordinate dynamics across multiple attachment sites.

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Coordinated stretching of different pericentromeres is pericentric cohesin and Cin8 dependent. (A and B) Trans-labeled pericentromeres were categorized as no stretch (both arrays compact foci), uncoordinated (stretching in only one of the labeled CEN arrays), or coordinated (both CEN nonsister arrays display stretching). Bar, 1 µm. (C) WT pericentromere stretching frequency for each CEN11 and CEN15. (D) Coordinated stretching occurs in 40 ± 1% of cells that show stretching, higher than predicted by independent stretching frequencies (11% dotted line). (E) Graph of coordinated stretching events that occur on the same or opposite side of the spindle. (F–H) Graphs of categorized stretching (F), coordinated stretching (G), and same versus opposite side coordinated stretching (H) for chromatin (GalH3, mcm21Δ, and brn1-9) and microtubule motor mutants (cin8Δ and kip1Δ). Asterisks denote mutants in which single pericentromere stretching (black lines) and coordinated stretching frequency (red bars) are statistically similar (G, χ2 > 0.4), and thus, stretching is independent. Values are listed in Table 1 and Table S1. Error bars represent standard deviations.
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fig2: Coordinated stretching of different pericentromeres is pericentric cohesin and Cin8 dependent. (A and B) Trans-labeled pericentromeres were categorized as no stretch (both arrays compact foci), uncoordinated (stretching in only one of the labeled CEN arrays), or coordinated (both CEN nonsister arrays display stretching). Bar, 1 µm. (C) WT pericentromere stretching frequency for each CEN11 and CEN15. (D) Coordinated stretching occurs in 40 ± 1% of cells that show stretching, higher than predicted by independent stretching frequencies (11% dotted line). (E) Graph of coordinated stretching events that occur on the same or opposite side of the spindle. (F–H) Graphs of categorized stretching (F), coordinated stretching (G), and same versus opposite side coordinated stretching (H) for chromatin (GalH3, mcm21Δ, and brn1-9) and microtubule motor mutants (cin8Δ and kip1Δ). Asterisks denote mutants in which single pericentromere stretching (black lines) and coordinated stretching frequency (red bars) are statistically similar (G, χ2 > 0.4), and thus, stretching is independent. Values are listed in Table 1 and Table S1. Error bars represent standard deviations.

Mentions: LacO arrays in the pericentromere are observed as compact foci or decompacted linear filaments reflecting the pericentromere chromatin response to force (Bachant et al., 2002; Stephens et al., 2011). Using pericentromeres labeled in trans (CEN11 and CEN15), we investigated the occurrence of coordinated stretching (Fig. 2, A–E). Each pericentromere LacO/TetO displayed similar stretching (CEN11, 12%; CEN15, 11%, n = 267; Fig. 2 C). In cells with a stretched pericentromere, 40% ± 1% of cells exhibit a second stretched pericentromere (n = 37; Fig. 2 D), significantly more than predicted by independent probabilities (11% single stretching dotted line; χ2 < 1 × 10−8; Fig. 2 D). Of the coordinated stretching, events greater than 2/3 displayed stretching on the same side of the spindle (70% same side; Fig. 2 E). Furthermore, both coordinated stretching and bias for the same side of the spindle are reproducible for CEN11 and CEN3 (Fig. S1, B–E). Correlated motion and stretching dynamics between multiple pairs of pericentromeres of different chromosomes is indicative of a cross-linked network.


Individual pericentromeres display coordinated motion and stretching in the yeast spindle.

Stephens AD, Snider CE, Haase J, Haggerty RA, Vasquez PA, Forest MG, Bloom K - J. Cell Biol. (2013)

Coordinated stretching of different pericentromeres is pericentric cohesin and Cin8 dependent. (A and B) Trans-labeled pericentromeres were categorized as no stretch (both arrays compact foci), uncoordinated (stretching in only one of the labeled CEN arrays), or coordinated (both CEN nonsister arrays display stretching). Bar, 1 µm. (C) WT pericentromere stretching frequency for each CEN11 and CEN15. (D) Coordinated stretching occurs in 40 ± 1% of cells that show stretching, higher than predicted by independent stretching frequencies (11% dotted line). (E) Graph of coordinated stretching events that occur on the same or opposite side of the spindle. (F–H) Graphs of categorized stretching (F), coordinated stretching (G), and same versus opposite side coordinated stretching (H) for chromatin (GalH3, mcm21Δ, and brn1-9) and microtubule motor mutants (cin8Δ and kip1Δ). Asterisks denote mutants in which single pericentromere stretching (black lines) and coordinated stretching frequency (red bars) are statistically similar (G, χ2 > 0.4), and thus, stretching is independent. Values are listed in Table 1 and Table S1. Error bars represent standard deviations.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3824013&req=5

fig2: Coordinated stretching of different pericentromeres is pericentric cohesin and Cin8 dependent. (A and B) Trans-labeled pericentromeres were categorized as no stretch (both arrays compact foci), uncoordinated (stretching in only one of the labeled CEN arrays), or coordinated (both CEN nonsister arrays display stretching). Bar, 1 µm. (C) WT pericentromere stretching frequency for each CEN11 and CEN15. (D) Coordinated stretching occurs in 40 ± 1% of cells that show stretching, higher than predicted by independent stretching frequencies (11% dotted line). (E) Graph of coordinated stretching events that occur on the same or opposite side of the spindle. (F–H) Graphs of categorized stretching (F), coordinated stretching (G), and same versus opposite side coordinated stretching (H) for chromatin (GalH3, mcm21Δ, and brn1-9) and microtubule motor mutants (cin8Δ and kip1Δ). Asterisks denote mutants in which single pericentromere stretching (black lines) and coordinated stretching frequency (red bars) are statistically similar (G, χ2 > 0.4), and thus, stretching is independent. Values are listed in Table 1 and Table S1. Error bars represent standard deviations.
Mentions: LacO arrays in the pericentromere are observed as compact foci or decompacted linear filaments reflecting the pericentromere chromatin response to force (Bachant et al., 2002; Stephens et al., 2011). Using pericentromeres labeled in trans (CEN11 and CEN15), we investigated the occurrence of coordinated stretching (Fig. 2, A–E). Each pericentromere LacO/TetO displayed similar stretching (CEN11, 12%; CEN15, 11%, n = 267; Fig. 2 C). In cells with a stretched pericentromere, 40% ± 1% of cells exhibit a second stretched pericentromere (n = 37; Fig. 2 D), significantly more than predicted by independent probabilities (11% single stretching dotted line; χ2 < 1 × 10−8; Fig. 2 D). Of the coordinated stretching, events greater than 2/3 displayed stretching on the same side of the spindle (70% same side; Fig. 2 E). Furthermore, both coordinated stretching and bias for the same side of the spindle are reproducible for CEN11 and CEN3 (Fig. S1, B–E). Correlated motion and stretching dynamics between multiple pairs of pericentromeres of different chromosomes is indicative of a cross-linked network.

Bottom Line: By labeling several chromosomes, we find that pericentromeres display coordinated motion and stretching in metaphase.The pericentromeres of different chromosomes exhibit physical linkage dependent on centromere function and structural maintenance of chromosomes complexes.Coordinated motion is dependent on condensin and the kinesin motor Cin8, whereas coordinated stretching is dependent on pericentric cohesin and Cin8.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, 2 Department of Mathematics, and 3 Department Biomedical Engineering, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599.

ABSTRACT
The mitotic segregation apparatus composed of microtubules and chromatin functions to faithfully partition a duplicated genome into two daughter cells. Microtubules exert extensional pulling force on sister chromatids toward opposite poles, whereas pericentric chromatin resists with contractile springlike properties. Tension generated from these opposing forces silences the spindle checkpoint to ensure accurate chromosome segregation. It is unknown how the cell senses tension across multiple microtubule attachment sites, considering the stochastic dynamics of microtubule growth and shortening. In budding yeast, there is one microtubule attachment site per chromosome. By labeling several chromosomes, we find that pericentromeres display coordinated motion and stretching in metaphase. The pericentromeres of different chromosomes exhibit physical linkage dependent on centromere function and structural maintenance of chromosomes complexes. Coordinated motion is dependent on condensin and the kinesin motor Cin8, whereas coordinated stretching is dependent on pericentric cohesin and Cin8. Linking of pericentric chromatin through cohesin, condensin, and kinetochore microtubules functions to coordinate dynamics across multiple attachment sites.

Show MeSH
Related in: MedlinePlus