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Condensin suppresses recombination and regulates double-strand break processing at the repetitive ribosomal DNA array to ensure proper chromosome segregation during meiosis in budding yeast.

Li P, Jin H, Yu HG - Mol. Biol. Cell (2014)

Bottom Line: Condensin is highly enriched at the rDNA region during prophase I, released at the prophase I/metaphase I transition, and reassociates with rDNA before anaphase I onset.We show that condensin plays a dual role in maintaining rDNA stability: it suppresses the formation of Spo11-mediated rDNA breaks, and it promotes DSB processing to ensure proper chromosome segregation.Our work reveals that condensin coordinates meiotic recombination with chromosome segregation at the repetitive rDNA sequence, thereby maintaining genome integrity.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, Florida State University, Tallahassee, FL 32306-4370.

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The relationship between Cdc14 release and condensin reloading at the rDNA. (A) Cdc14 dynamics in wild-type (HY2100) and PCLB2-BRN1 (HY2894) cells during yeast meiosis. Yeast cells were prepared for time-lapse live-cell microscopy as shown in Figure 1A. Cdc14 was tagged with GFP; Tub4, the γ-tubulin in yeast, was tagged with RFP. Time zero was defined as the onset of anaphase I. Note that the Cdc14-GFP focus disappeared at the onset of anaphase I and reemerged as two separated foci after meiosis I in the wild-type cell, but in the mutant cell it reemerged as a single focus. Arrows indicate the localization of Cdc14-GFP to one of the spindle pole bodies during anaphase I. Right, graphs showing SPB-to-SPB distance during metaphase I and anaphase I. (B) Cell progression in wild type and PCLB2-BRN1. Cell cycle stage was determined on the basis of spindle morphology. Both anaphase I and meiosis II cells are scored. (C) Reloading of Brn1-GFP at rDNA depends on Cdc14. Yeast cells (HY4056) were induced to undergo synchronous meiosis at 35°C, and time-lapse live-cell microscopy was performed at 35°C. Left, representative images. Right, quantitative analysis of Brn1-GFP localization. More than 200 cells were counted. Bars, 2 μm.
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Figure 4: The relationship between Cdc14 release and condensin reloading at the rDNA. (A) Cdc14 dynamics in wild-type (HY2100) and PCLB2-BRN1 (HY2894) cells during yeast meiosis. Yeast cells were prepared for time-lapse live-cell microscopy as shown in Figure 1A. Cdc14 was tagged with GFP; Tub4, the γ-tubulin in yeast, was tagged with RFP. Time zero was defined as the onset of anaphase I. Note that the Cdc14-GFP focus disappeared at the onset of anaphase I and reemerged as two separated foci after meiosis I in the wild-type cell, but in the mutant cell it reemerged as a single focus. Arrows indicate the localization of Cdc14-GFP to one of the spindle pole bodies during anaphase I. Right, graphs showing SPB-to-SPB distance during metaphase I and anaphase I. (B) Cell progression in wild type and PCLB2-BRN1. Cell cycle stage was determined on the basis of spindle morphology. Both anaphase I and meiosis II cells are scored. (C) Reloading of Brn1-GFP at rDNA depends on Cdc14. Yeast cells (HY4056) were induced to undergo synchronous meiosis at 35°C, and time-lapse live-cell microscopy was performed at 35°C. Left, representative images. Right, quantitative analysis of Brn1-GFP localization. More than 200 cells were counted. Bars, 2 μm.

Mentions: Failure of rDNA homologue segregation in the absence of condensin function resembles that of the cdc14 mutants (Buonomo et al., 2003; Marston et al., 2003). To determine the dynamics of Cdc14 release from the meiotic yeast nucleolus, we performed live-cell microscopy to observe Cdc14-GFP, which served as the only source of Cdc14 in experimental cells (Figure 4). In wild-type cells before anaphase I onset, Cdc14-GFP formed an intense focus (Figure 4A), which was colocalized with a nucleolus marker Nop1 (unpublished data), indicating its association with the yeast nucleolus. The focused Cdc14-GFP signals disappeared at the onset of anaphase I (t = 0 min) and then recovered as two separated Cdc14-GFP foci, representing two divided nucleoli (Figure 4A). On average, the interval from the disappearance, and therefore release of Cdc14 from the nucleolus, to the reformation of Cdc14-GFP foci lasted 12.2 ± 0.6 min (SD, n = 13). In PCLB2-BRN1 cells upon anaphase I onset, the Cdc14-GFP focus disappeared on time, as judged by the elongation of the meiosis I spindle (Figure 4A). The average duration of Cdc14-GFP dispersal was 12.0 ± 0.5 min (SD, n = 17) in the mutant, not significantly different from that of the wild type. Of note, Cdc14-GFP recovered as a single GFP focus after meiosis I (PCLB2-BRN1, t = 12 min), suggesting that the nucleolus failed to divide in PCLB2-BRN1 cells. This result supports our observation that rDNA homologues, which are tightly associated with the yeast nucleolus, remain linked in the absence of condensin function. We note that a small portion of Cdc14-GFP released from the nucleolus was concentrated at one of the spindle poles at anaphase I (wild type, t = 12, and PCLB2-BRN1, t = 9 min, arrows). The biological significance of Cdc14’s association with the spindle pole is unknown. Cdc14 release from the nucleolus and Cdc14 reloading appeared to be on time in PCLB2-BRN1 cells, because cells committed to anaphase I simultaneously with or without Brn1 (Figure 4B). These findings indicate that release of Cdc14 from the nucleolus is independent of condensin.


Condensin suppresses recombination and regulates double-strand break processing at the repetitive ribosomal DNA array to ensure proper chromosome segregation during meiosis in budding yeast.

Li P, Jin H, Yu HG - Mol. Biol. Cell (2014)

The relationship between Cdc14 release and condensin reloading at the rDNA. (A) Cdc14 dynamics in wild-type (HY2100) and PCLB2-BRN1 (HY2894) cells during yeast meiosis. Yeast cells were prepared for time-lapse live-cell microscopy as shown in Figure 1A. Cdc14 was tagged with GFP; Tub4, the γ-tubulin in yeast, was tagged with RFP. Time zero was defined as the onset of anaphase I. Note that the Cdc14-GFP focus disappeared at the onset of anaphase I and reemerged as two separated foci after meiosis I in the wild-type cell, but in the mutant cell it reemerged as a single focus. Arrows indicate the localization of Cdc14-GFP to one of the spindle pole bodies during anaphase I. Right, graphs showing SPB-to-SPB distance during metaphase I and anaphase I. (B) Cell progression in wild type and PCLB2-BRN1. Cell cycle stage was determined on the basis of spindle morphology. Both anaphase I and meiosis II cells are scored. (C) Reloading of Brn1-GFP at rDNA depends on Cdc14. Yeast cells (HY4056) were induced to undergo synchronous meiosis at 35°C, and time-lapse live-cell microscopy was performed at 35°C. Left, representative images. Right, quantitative analysis of Brn1-GFP localization. More than 200 cells were counted. Bars, 2 μm.
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Related In: Results  -  Collection

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Figure 4: The relationship between Cdc14 release and condensin reloading at the rDNA. (A) Cdc14 dynamics in wild-type (HY2100) and PCLB2-BRN1 (HY2894) cells during yeast meiosis. Yeast cells were prepared for time-lapse live-cell microscopy as shown in Figure 1A. Cdc14 was tagged with GFP; Tub4, the γ-tubulin in yeast, was tagged with RFP. Time zero was defined as the onset of anaphase I. Note that the Cdc14-GFP focus disappeared at the onset of anaphase I and reemerged as two separated foci after meiosis I in the wild-type cell, but in the mutant cell it reemerged as a single focus. Arrows indicate the localization of Cdc14-GFP to one of the spindle pole bodies during anaphase I. Right, graphs showing SPB-to-SPB distance during metaphase I and anaphase I. (B) Cell progression in wild type and PCLB2-BRN1. Cell cycle stage was determined on the basis of spindle morphology. Both anaphase I and meiosis II cells are scored. (C) Reloading of Brn1-GFP at rDNA depends on Cdc14. Yeast cells (HY4056) were induced to undergo synchronous meiosis at 35°C, and time-lapse live-cell microscopy was performed at 35°C. Left, representative images. Right, quantitative analysis of Brn1-GFP localization. More than 200 cells were counted. Bars, 2 μm.
Mentions: Failure of rDNA homologue segregation in the absence of condensin function resembles that of the cdc14 mutants (Buonomo et al., 2003; Marston et al., 2003). To determine the dynamics of Cdc14 release from the meiotic yeast nucleolus, we performed live-cell microscopy to observe Cdc14-GFP, which served as the only source of Cdc14 in experimental cells (Figure 4). In wild-type cells before anaphase I onset, Cdc14-GFP formed an intense focus (Figure 4A), which was colocalized with a nucleolus marker Nop1 (unpublished data), indicating its association with the yeast nucleolus. The focused Cdc14-GFP signals disappeared at the onset of anaphase I (t = 0 min) and then recovered as two separated Cdc14-GFP foci, representing two divided nucleoli (Figure 4A). On average, the interval from the disappearance, and therefore release of Cdc14 from the nucleolus, to the reformation of Cdc14-GFP foci lasted 12.2 ± 0.6 min (SD, n = 13). In PCLB2-BRN1 cells upon anaphase I onset, the Cdc14-GFP focus disappeared on time, as judged by the elongation of the meiosis I spindle (Figure 4A). The average duration of Cdc14-GFP dispersal was 12.0 ± 0.5 min (SD, n = 17) in the mutant, not significantly different from that of the wild type. Of note, Cdc14-GFP recovered as a single GFP focus after meiosis I (PCLB2-BRN1, t = 12 min), suggesting that the nucleolus failed to divide in PCLB2-BRN1 cells. This result supports our observation that rDNA homologues, which are tightly associated with the yeast nucleolus, remain linked in the absence of condensin function. We note that a small portion of Cdc14-GFP released from the nucleolus was concentrated at one of the spindle poles at anaphase I (wild type, t = 12, and PCLB2-BRN1, t = 9 min, arrows). The biological significance of Cdc14’s association with the spindle pole is unknown. Cdc14 release from the nucleolus and Cdc14 reloading appeared to be on time in PCLB2-BRN1 cells, because cells committed to anaphase I simultaneously with or without Brn1 (Figure 4B). These findings indicate that release of Cdc14 from the nucleolus is independent of condensin.

Bottom Line: Condensin is highly enriched at the rDNA region during prophase I, released at the prophase I/metaphase I transition, and reassociates with rDNA before anaphase I onset.We show that condensin plays a dual role in maintaining rDNA stability: it suppresses the formation of Spo11-mediated rDNA breaks, and it promotes DSB processing to ensure proper chromosome segregation.Our work reveals that condensin coordinates meiotic recombination with chromosome segregation at the repetitive rDNA sequence, thereby maintaining genome integrity.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, Florida State University, Tallahassee, FL 32306-4370.

Show MeSH
Related in: MedlinePlus