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1.
Methods Mol Biol ; 2818: 161-169, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-39126473

RESUMEN

For over a century, major advances in understanding meiosis have come from the use of microscopy-based methods. Studies using the budding yeast, Saccharomyces cerevisiae, have made important contributions to our understanding of meiosis because of the facility with which budding yeast can be manipulated as a genetic model organism. In contrast, imaging-based approaches with budding yeast have been constrained by the small size of its chromosomes. The advent of advances in fluorescent chromosome tagging techniques has made it possible to use yeast more effectively for imaging-based approaches as well. This protocol describes live cell imaging methods that can be used to monitor chromosome movements throughout meiosis in living yeast cells.


Asunto(s)
Meiosis , Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/citología , Cromosomas Fúngicos/genética , Microscopía Fluorescente/métodos , Saccharomycetales/genética , Saccharomycetales/citología
2.
Curr Biol ; 34(10): 2085-2093.e6, 2024 05 20.
Artículo en Inglés | MEDLINE | ID: mdl-38670094

RESUMEN

Proper chromosome segregation in meiosis I relies on the formation of connections between homologous chromosomes. Crossovers between homologs provide a connection that allows them to attach correctly to the meiosis I spindle. Tension is transmitted across the crossover when the partners attach to microtubules from opposing poles of the spindle. Tension stabilizes microtubule attachments that will pull the partners toward opposite poles at anaphase. Paradoxically, in many organisms, non-crossover partners segregate correctly. The mechanism by which non-crossover partners become bioriented on the meiotic spindle is unknown. Both crossover and non-crossover partners pair their centromeres early in meiosis (prophase). In budding yeast, centromere pairing is correlated with subsequent correct segregation of the partners. The mechanism by which centromere pairing, in prophase, promotes later correct attachment of the partners to the metaphase spindle is unknown. We used live cell imaging to track the biorientation process of non-crossover chromosomes. We find that centromere pairing allows the establishment of connections between the partners that allows their later interdependent attachment to the meiotic spindle using tension-sensing biorientation machinery. Because all chromosome pairs experience centromere pairing, our findings suggest that crossover chromosomes also utilize this mechanism to achieve maximal segregation fidelity.


Asunto(s)
Centrómero , Segregación Cromosómica , Meiosis , Saccharomyces cerevisiae , Centrómero/metabolismo , Segregación Cromosómica/fisiología , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Huso Acromático/metabolismo , Huso Acromático/fisiología , Emparejamiento Cromosómico , Cromosomas Fúngicos/genética , Microtúbulos/metabolismo
3.
bioRxiv ; 2023 Jan 10.
Artículo en Inglés | MEDLINE | ID: mdl-36712123

RESUMEN

Tumor cell lines with elevated chromosome numbers frequently have correlated elevations of Mps1 expression and these tumors are more dependent on Mps1 activity for their survival than control cell lines. Mps1 is a conserved kinase involved in controlling aspects of chromosome segregation in mitosis and meiosis. The mechanistic explanation for the Mps1-addiction of aneuploid cells is unknown. To address this question, we explored Mps1-dependence in yeast cells with increased sets of chromosomes. These experiments revealed that in yeast, increasing ploidy leads to delays and failures in orienting chromosomes on the mitotic spindle. Yeast cells with elevated numbers of chromosomes proved vulnerable to reductions of Mps1 activity. Cells with reduced Mps1 activity exhibit an extended prometaphase with longer spindles and delays in orienting the chromosomes. One known role of Mps1 is in recruiting Bub1 to the kinetochore in meiosis. We found that the Mps1-addiction of polyploid yeast cells is due in part to its role in Bub1 recruitment. Together, the experiments presented here demonstrate that increased ploidy renders cells more dependent on Mps1 for orienting chromosomes on the spindle. The phenomenon described here may be relevant in understanding why hyper-diploid cancer cells exhibit elevated reliance on Mps1 expression for successful chromosome segregation.

4.
Chromosoma ; 128(3): 355-367, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-31165256

RESUMEN

In meiosis, crossovers between homologous chromosomes link them together. This enables them to attach to microtubules of the meiotic spindle as a unit, such that the homologs will be pulled away from one another at anaphase I. Homologous pairs can sometimes fail to become linked by crossovers. In some organisms, these non-exchange partners are still able to segregate properly. In several organisms, associations between the centromeres of non-exchange partners occur in meiotic prophase. These associations have been proposed to promote segregation in meiosis I. But it is unclear how centromere pairing could promote subsequent proper segregation. Here we report that meiotic centromere pairing of chromosomes in mouse spermatocytes allows the formation of an association between chromosome pairs. We find that heterochromatin regions of homologous centromeres remain associated even after centromere-pairing dissolves. Our results suggest the model that, in mouse spermatocytes, heterochromatin maintains the association of homologous centromeres in the absence crossing-over.


Asunto(s)
Centrómero , Emparejamiento Cromosómico , Segregación Cromosómica , Heterocromatina , Meiosis , Espermatocitos , Animales , Masculino , Ratones , Profase , Recombinación Genética
5.
Proc Natl Acad Sci U S A ; 116(19): 9417-9422, 2019 05 07.
Artículo en Inglés | MEDLINE | ID: mdl-31019073

RESUMEN

Faithful chromosome segregation during meiosis I depends upon the formation of connections between homologous chromosomes. Crossovers between homologs connect the partners, allowing them to attach to the meiotic spindle as a unit, such that they migrate away from one another at anaphase I. Homologous partners also become connected by pairing of their centromeres in meiotic prophase. This centromere pairing can promote proper segregation at anaphase I of partners that have failed to become joined by a crossover. Centromere pairing is mediated by synaptonemal complex (SC) proteins that persist at the centromere when the SC disassembles. Here, using mouse spermatocyte and yeast model systems, we tested the role of shugoshin in promoting meiotic centromere pairing by protecting centromeric synaptonemal components from disassembly. The results show that shugoshin protects the centromeric SC in meiotic prophase and, in anaphase, promotes the proper segregation of partner chromosomes that are not linked by a crossover.


Asunto(s)
Anafase/fisiología , Proteínas de Ciclo Celular/metabolismo , Centrómero/metabolismo , Segregación Cromosómica/fisiología , Profase/fisiología , Espermatocitos/metabolismo , Animales , Proteínas de Ciclo Celular/genética , Centrómero/genética , Masculino , Ratones , Ratones Noqueados , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Espermatocitos/citología , Huso Acromático/genética , Huso Acromático/metabolismo , Complejo Sinaptonémico/genética , Complejo Sinaptonémico/metabolismo
6.
PLoS Genet ; 14(8): e1007513, 2018 08.
Artículo en Inglés | MEDLINE | ID: mdl-30091974

RESUMEN

In meiosis I, homologous chromosomes segregate away from each other-the first of two rounds of chromosome segregation that allow the formation of haploid gametes. In prophase I, homologous partners become joined along their length by the synaptonemal complex (SC) and crossovers form between the homologs to generate links called chiasmata. The chiasmata allow the homologs to act as a single unit, called a bivalent, as the chromosomes attach to the microtubules that will ultimately pull them away from each other at anaphase I. Recent studies, in several organisms, have shown that when the SC disassembles at the end of prophase, residual SC proteins remain at the homologous centromeres providing an additional link between the homologs. In budding yeast, this centromere pairing is correlated with improved segregation of the paired partners in anaphase. However, the causal relationship of prophase centromere pairing and subsequent disjunction in anaphase has been difficult to demonstrate as has been the relationship between SC assembly and the assembly of the centromere pairing apparatus. Here, a series of in-frame deletion mutants of the SC component Zip1 were used to address these questions. The identification of a separation-of-function allele that disrupts centromere pairing, but not SC assembly, has made it possible to demonstrate that centromere pairing and SC assembly have mechanistically distinct features and that the centromere pairing function of Zip1 drives disjunction of the paired partners in anaphase I.


Asunto(s)
Centrómero/metabolismo , Proteínas Nucleares/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomycetales/genética , Alelos , Anafase/genética , Emparejamiento Cromosómico , Segregación Cromosómica , Meiosis , Proteínas Nucleares/metabolismo , Recombinación Genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomycetales/metabolismo , Complejo Sinaptonémico/metabolismo
7.
Mol Biol Cell ; 26(17): 2986-3000, 2015 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-26157162

RESUMEN

In mitosis, the centromeres of sister chromosomes are pulled toward opposite poles of the spindle. In meiosis I, the opposite is true: the sister centromeres move together to the same pole, and the homologous chromosomes are pulled apart. This change in segregation patterns demands that between the final mitosis preceding meiosis and the first meiotic division, the kinetochores must be restructured. In budding yeast, unlike mammals, kinetochores are largely stable throughout the mitotic cycle. In contrast, previous work with budding and fission yeast showed that some outer kinetochore proteins are lost in early meiosis. We use quantitative mass spectrometry methods and imaging approaches to explore the kinetochore restructuring process that occurs in meiosis I in budding yeast. The Ndc80 outer kinetochore complex, but not other subcomplexes, is shed upon meiotic entry. This shedding is regulated by the conserved protein kinase Ipl1/Aurora-B and promotes the subsequent assembly of a kinetochore that will confer meiosis-specific segregation patterns on the chromosome.


Asunto(s)
Aurora Quinasas/metabolismo , Cinetocoros/fisiología , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/enzimología , Aurora Quinasas/genética , Ciclo Celular/fisiología , Puntos de Control del Ciclo Celular/genética , Centrómero/metabolismo , Segregación Cromosómica , Cinetocoros/enzimología , Cinetocoros/metabolismo , Meiosis/fisiología , Proteínas Nucleares/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomycetales/genética , Saccharomycetales/metabolismo
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