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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 sequestration and release cycle of condensin at the rDNA region during yeast meiosis. (A) Localization of condensin subunit Brn1 during yeast meiosis. Yeast cells (strain HY3528) were induced to undergo synchronous meiosis and then subjected to time-lapse live-cell fluorescence microscopy. Brn1 was tagged with GFP (green) and rDNA with tetO/tetR-RFP (red). Time zero was defined as the onset of anaphase I. Nuclear division shown by the residual tetR-RFP signal was used to determine the onset of anaphase I. Arrow indicates the point when condensin became undetectable at the rDNA region. (B) Representative images showing Brn1-GFP localization at early meiosis (strain HY3528), prophase I (Pro I, ndt80Δ, HY3584), metaphase I (Met I, PCLB2-CDC20, HY3628), and anaphase I (AnaI, strain HY3528). Fluorescence microscopy was performed as in A. Note that Brn1-GFP overlaps with rDNA-RFP during early meiosis (∼2.5 h after induction of meiosis), at prophase I (ndt80Δ), and at anaphase I, but not at metaphase I (PCLB2-CDC20). Right, graph showing the quantitative analysis of Brn1-GFP and Ycg1-GFP localization to the rDNA region at the indicated stages. (C) Localization of Brn1-GFP and Nop1-RFP at the indicated cell cycle stages as in B. Right, graph showing quantitative analysis. Strains HY4143, HY4144, HY4145, and HY4146 were used. (D) Western blot showing the protein level of Brn-6Myc in ndt80Δ (HY4193) and PCLB2-CDC20 cells (HY4206). (E) Cdc5 regulates condensin release from rDNA. PCUP1-CDC5 ndt80Δ (HY4008) and PCLB2-CDC5 (HY3995) cells were induced to undergo meiosis; 50 μM CuSO4 was added to the culture medium 4 h after. Representative images are shown. Bottom graph, quantitative analysis of these cells. Bars, 2 μm.
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Figure 1: The sequestration and release cycle of condensin at the rDNA region during yeast meiosis. (A) Localization of condensin subunit Brn1 during yeast meiosis. Yeast cells (strain HY3528) were induced to undergo synchronous meiosis and then subjected to time-lapse live-cell fluorescence microscopy. Brn1 was tagged with GFP (green) and rDNA with tetO/tetR-RFP (red). Time zero was defined as the onset of anaphase I. Nuclear division shown by the residual tetR-RFP signal was used to determine the onset of anaphase I. Arrow indicates the point when condensin became undetectable at the rDNA region. (B) Representative images showing Brn1-GFP localization at early meiosis (strain HY3528), prophase I (Pro I, ndt80Δ, HY3584), metaphase I (Met I, PCLB2-CDC20, HY3628), and anaphase I (AnaI, strain HY3528). Fluorescence microscopy was performed as in A. Note that Brn1-GFP overlaps with rDNA-RFP during early meiosis (∼2.5 h after induction of meiosis), at prophase I (ndt80Δ), and at anaphase I, but not at metaphase I (PCLB2-CDC20). Right, graph showing the quantitative analysis of Brn1-GFP and Ycg1-GFP localization to the rDNA region at the indicated stages. (C) Localization of Brn1-GFP and Nop1-RFP at the indicated cell cycle stages as in B. Right, graph showing quantitative analysis. Strains HY4143, HY4144, HY4145, and HY4146 were used. (D) Western blot showing the protein level of Brn-6Myc in ndt80Δ (HY4193) and PCLB2-CDC20 cells (HY4206). (E) Cdc5 regulates condensin release from rDNA. PCUP1-CDC5 ndt80Δ (HY4008) and PCLB2-CDC5 (HY3995) cells were induced to undergo meiosis; 50 μM CuSO4 was added to the culture medium 4 h after. Representative images are shown. Bottom graph, quantitative analysis of these cells. Bars, 2 μm.

Mentions: We showed previously that condensin is enriched at the rDNA array during yeast meiosis (Yu and Koshland, 2003). To determine condensin dynamics, we developed a live-cell fluorescence microscopy method to observe condensin and rDNA movement in yeast cells that were induced to undergo synchronous meiosis (Figure 1). Using green fluorescent protein (GFP)–tagged condensin subunits Brn1 and Ycg1 and a red fluorescent protein (RFP)–marked rDNA array (Li et al., 2011), we found that at the beginning of meiosis (∼5 h before anaphase I onset), Brn1-GFP was highly concentrated at the rDNA gene cluster (Figure 1A). Brn1-GFP became dispersed during prophase I (∼2 h before anaphase I), and its association with rDNA decreased precipitously at late prophase I. Brn1-GFP was undetectable at the rDNA at metaphase I (∼40 min before anaphase I, Shirk et al., 2011), but it reassociated with the rDNA array upon anaphase I onset (Figure 1A). These observations suggest that condensin is subject to relocation inside the yeast nucleus during meiosis.


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 sequestration and release cycle of condensin at the rDNA region during yeast meiosis. (A) Localization of condensin subunit Brn1 during yeast meiosis. Yeast cells (strain HY3528) were induced to undergo synchronous meiosis and then subjected to time-lapse live-cell fluorescence microscopy. Brn1 was tagged with GFP (green) and rDNA with tetO/tetR-RFP (red). Time zero was defined as the onset of anaphase I. Nuclear division shown by the residual tetR-RFP signal was used to determine the onset of anaphase I. Arrow indicates the point when condensin became undetectable at the rDNA region. (B) Representative images showing Brn1-GFP localization at early meiosis (strain HY3528), prophase I (Pro I, ndt80Δ, HY3584), metaphase I (Met I, PCLB2-CDC20, HY3628), and anaphase I (AnaI, strain HY3528). Fluorescence microscopy was performed as in A. Note that Brn1-GFP overlaps with rDNA-RFP during early meiosis (∼2.5 h after induction of meiosis), at prophase I (ndt80Δ), and at anaphase I, but not at metaphase I (PCLB2-CDC20). Right, graph showing the quantitative analysis of Brn1-GFP and Ycg1-GFP localization to the rDNA region at the indicated stages. (C) Localization of Brn1-GFP and Nop1-RFP at the indicated cell cycle stages as in B. Right, graph showing quantitative analysis. Strains HY4143, HY4144, HY4145, and HY4146 were used. (D) Western blot showing the protein level of Brn-6Myc in ndt80Δ (HY4193) and PCLB2-CDC20 cells (HY4206). (E) Cdc5 regulates condensin release from rDNA. PCUP1-CDC5 ndt80Δ (HY4008) and PCLB2-CDC5 (HY3995) cells were induced to undergo meiosis; 50 μM CuSO4 was added to the culture medium 4 h after. Representative images are shown. Bottom graph, quantitative analysis of these cells. Bars, 2 μm.
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Figure 1: The sequestration and release cycle of condensin at the rDNA region during yeast meiosis. (A) Localization of condensin subunit Brn1 during yeast meiosis. Yeast cells (strain HY3528) were induced to undergo synchronous meiosis and then subjected to time-lapse live-cell fluorescence microscopy. Brn1 was tagged with GFP (green) and rDNA with tetO/tetR-RFP (red). Time zero was defined as the onset of anaphase I. Nuclear division shown by the residual tetR-RFP signal was used to determine the onset of anaphase I. Arrow indicates the point when condensin became undetectable at the rDNA region. (B) Representative images showing Brn1-GFP localization at early meiosis (strain HY3528), prophase I (Pro I, ndt80Δ, HY3584), metaphase I (Met I, PCLB2-CDC20, HY3628), and anaphase I (AnaI, strain HY3528). Fluorescence microscopy was performed as in A. Note that Brn1-GFP overlaps with rDNA-RFP during early meiosis (∼2.5 h after induction of meiosis), at prophase I (ndt80Δ), and at anaphase I, but not at metaphase I (PCLB2-CDC20). Right, graph showing the quantitative analysis of Brn1-GFP and Ycg1-GFP localization to the rDNA region at the indicated stages. (C) Localization of Brn1-GFP and Nop1-RFP at the indicated cell cycle stages as in B. Right, graph showing quantitative analysis. Strains HY4143, HY4144, HY4145, and HY4146 were used. (D) Western blot showing the protein level of Brn-6Myc in ndt80Δ (HY4193) and PCLB2-CDC20 cells (HY4206). (E) Cdc5 regulates condensin release from rDNA. PCUP1-CDC5 ndt80Δ (HY4008) and PCLB2-CDC5 (HY3995) cells were induced to undergo meiosis; 50 μM CuSO4 was added to the culture medium 4 h after. Representative images are shown. Bottom graph, quantitative analysis of these cells. Bars, 2 μm.
Mentions: We showed previously that condensin is enriched at the rDNA array during yeast meiosis (Yu and Koshland, 2003). To determine condensin dynamics, we developed a live-cell fluorescence microscopy method to observe condensin and rDNA movement in yeast cells that were induced to undergo synchronous meiosis (Figure 1). Using green fluorescent protein (GFP)–tagged condensin subunits Brn1 and Ycg1 and a red fluorescent protein (RFP)–marked rDNA array (Li et al., 2011), we found that at the beginning of meiosis (∼5 h before anaphase I onset), Brn1-GFP was highly concentrated at the rDNA gene cluster (Figure 1A). Brn1-GFP became dispersed during prophase I (∼2 h before anaphase I), and its association with rDNA decreased precipitously at late prophase I. Brn1-GFP was undetectable at the rDNA at metaphase I (∼40 min before anaphase I, Shirk et al., 2011), but it reassociated with the rDNA array upon anaphase I onset (Figure 1A). These observations suggest that condensin is subject to relocation inside the yeast nucleus during meiosis.

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