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Sue Biggins - One of the best experts on this subject based on the ideXlab platform.
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kinetochore associated stu2 promotes chromosome Biorientation in vivo
PLOS Genetics, 2019Co-Authors: Matthew P Miller, Rena K Evans, Alex Zelter, Elisabeth A Geyer, Michael J Maccoss, Luke M Rice, Trisha N Davis, Charles L Asbury, Sue BigginsAbstract:Accurate segregation of chromosomes to daughter cells is a critical aspect of cell division. It requires the kinetochores on duplicated chromosomes to biorient, attaching to microtubules from opposite poles of the cell. Bioriented attachments come under tension, while incorrect attachments lack tension and must be released to allow proper attachments to form. A well-studied error correction pathway is mediated by the Aurora B kinase, which destabilizes low tension-bearing attachments. We recently discovered that in vitro, kinetochores display an additional intrinsic tension-sensing pathway that utilizes Stu2. The contribution of kinetochore-associated Stu2 to error correction in cells, however, was unknown. Here, we identify a Stu2 mutant that abolishes its kinetochore function and show that it causes Biorientation defects in vivo. We also show that this Stu2-mediated pathway functions together with the Aurora B-mediated pathway. Altogether, our work indicates that cells employ multiple pathways to ensure Biorientation and the accuracy of chromosome segregation.
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a stu2 mediated intrinsic tension sensing pathway promotes chromosome Biorientation in vivo
bioRxiv, 2018Co-Authors: Matthew P Miller, Rena K Evans, Alex Zelter, Elisabeth A Geyer, Michael J Maccoss, Luke M Rice, Trisha N Davis, Charles L Asbury, Sue BigginsAbstract:Accurate segregation of chromosomes to daughter cells is a critical aspect of cell division. It requires the kinetochores on duplicated chromosomes to biorient, attaching to microtubules from opposite poles of the cell. Bioriented attachments come under tension, while incorrect attachments lack tension and must be destabilized. A well-studied error correction pathway is mediated by the Aurora B kinase, which destabilizes low tension-bearing attachments. We recently discovered that in vitro, kinetochores display an additional intrinsic tension-sensing pathway that utilizes Stu2. This pathway9s contribution to error correction in cells, however, was unknown. Here, we identify a Stu2 mutant that abolishes its kinetochore function and show that it causes error correction defects in vivo. We also show that this intrinsic tension-sensing pathway functions in concert with the Aurora B-mediated pathway. Together, our work indicates that cells employ at least two pathways to ensure Biorientation and the accuracy of chromosome segregation.
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Kinetochore Function and Chromosome Segregation Rely on Critical Residues in Histones H3 and H4 in Budding Yeast
Genetics, 2013Co-Authors: Tessie M Ng, Tineke L. Lenstra, Nicole Duggan, Shuangying Jiang, Steven Ceto, Frank C. P. Holstege, Jef D. Boeke, Sue BigginsAbstract:Accurate chromosome segregation requires that sister kinetochores biorient and attach to microtubules from opposite poles. Kinetochore Biorientation relies on the underlying centromeric chromatin, which provides a platform to assemble the kinetochore and to recruit the regulatory factors that ensure the high fidelity of this process. To identify the centromeric chromatin determinants that contribute to chromosome segregation, we performed two complementary unbiased genetic screens using a library of budding yeast mutants in every residue of histone H3 and H4. In one screen, we identified mutants that lead to increased loss of a nonessential chromosome. In the second screen, we isolated mutants whose viability depends on a key regulator of Biorientation, the Aurora B protein kinase. Nine mutants were common to both screens and exhibited kinetochore Biorientation defects. Four of the mutants map near the unstructured nucleosome entry site, and their genetic interaction with reduced IPL1 can be suppressed by increasing the dosage of SGO1, a key regulator of Biorientation. In addition, the composition of purified kinetochores was altered in six of the mutants. Together, this work identifies previously unknown histone residues involved in chromosome segregation and lays the foundation for future studies on the role of the underlying chromatin structure in chromosome segregation.
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pericentromeric sister chromatid cohesion promotes kinetochore Biorientation
Molecular Biology of the Cell, 2009Co-Authors: Tessie M Ng, William G Waples, Brigitte D Lavoie, Sue BigginsAbstract:Accurate chromosome segregation depends on sister kinetochores making bioriented attachments to microtubules from opposite poles. An essential regulator of Biorientation is the Ipl1/Aurora B protein kinase that destabilizes improper microtubule–kinetochore attachments. To identify additional Biorientation pathways, we performed a systematic genetic analysis between the ipl1-321 allele and all nonessential budding yeast genes. One of the mutants, mcm21Δ, precociously separates pericentromeres and this is associated with a defect in the binding of the Scc2 cohesin-loading factor at the centromere. Strikingly, Mcm21 becomes essential for Biorientation when Ipl1 function is reduced, and this appears to be related to its role in pericentromeric cohesion. When pericentromeres are artificially tethered, Mcm21 is no longer needed for Biorientation despite decreased Ipl1 activity. Taken together, these data reveal a specific role for pericentromeric linkage in ensuring kinetochore Biorientation.
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the overexpression of a saccharomyces cerevisiae centromeric histone h3 variant mutant protein leads to a defect in kinetochore Biorientation
Genetics, 2007Co-Authors: Kimberly A Collins, Jennifer L. Gerton, Raymond Camahort, Chris Seidel, Sue BigginsAbstract:Chromosomes segregate using their kinetochores, the specialized protein structures that are assembled on centromeric DNA and mediate attachment to the mitotic spindle. Because centromeric sequences are not conserved, centromere identity is propagated by an epigenetic mechanism. All eukaryotes contain an essential histone H3 variant (CenH3) that localizes exclusively to centromeres. Because CenH3 is required for kinetochore assembly and is likely to be the epigenetic mark that specifies centromere identity, it is critical to elucidate the mechanisms that assemble and maintain CenH3 exclusively at centromeres. To learn more about the functions and regulation of CenH3, we isolated mutants in the budding yeast CenH3 that are lethal when overexpressed. These CenH3 mutants fall into three unique classes: (I) those that localize to euchromatin but do not alter kinetochore function, (II) those that localize to the centromere and disrupt kinetochore function, and (III) those that no longer target to the centromere but still disrupt chromosome segregation. We found that a class III mutant is specifically defective in the ability of sister kinetochores to biorient and attach to microtubules from opposite spindle poles, indicating that CenH3 mutants defective in kinetochore Biorientation can be obtained.
Adele L Marston - One of the best experts on this subject based on the ideXlab platform.
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tension dependent removal of pericentromeric shugoshin is an indicator of sister chromosome Biorientation
Genes & Development, 2014Co-Authors: Olga O Nerusheva, David A Kelly, Stefan Galander, Josefin Fernius, Adele L MarstonAbstract:During mitosis and meiosis, sister chromatid cohesion resists the pulling forces of microtubules, enabling the generation of tension at kinetochores upon chromosome Biorientation. How tension is read to signal the bioriented state remains unclear. Shugoshins form a pericentromeric platform that integrates multiple functions to ensure proper chromosome Biorientation. Here we show that budding yeast shugoshin Sgo1 dissociates from the pericentromere reversibly in response to tension. The antagonistic activities of the kinetochore-associated Bub1 kinase and the Sgo1-bound phosphatase protein phosphatase 2A (PP2A)-Rts1 underlie a tension-dependent circuitry that enables Sgo1 removal upon sister kinetochore Biorientation. Sgo1 dissociation from the pericentromere triggers dissociation of condensin and Aurora B from the centromere, thereby stabilizing the bioriented state. Conversely, forcing sister kinetochores to be under tension during meiosis I leads to premature Sgo1 removal and precocious loss of pericentromeric cohesion. Overall, we show that the pivotal role of shugoshin is to build a platform at the pericentromere that attracts activities that respond to the absence of tension between sister kinetochores. Disassembly of this platform in response to intersister kinetochore tension signals the bioriented state. Therefore, tension sensing by shugoshin is a central mechanism by which the bioriented state is read.
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shugoshin biases chromosomes for Biorientation through condensin recruitment to the pericentromere
eLife, 2014Co-Authors: Kitty F Verzijlbergen, Dean Clift, Olga O Nerusheva, David A Kelly, Alastair R W Kerr, Flavia De Lima Alves, Juri Rappsilber, Adele L MarstonAbstract:When a cell divides to create two new daughter cells, it must produce a copy of each of its chromosomes. It is important that each daughter cell gets just one copy of each chromosome. If an error occurs and one cell gets two copies of a single chromosome, it can lead to cancer or birth defects. Fortunately, there are multiple checks to ensure that this does not happen. During cell division the chromosomes line up in a way that increases the likelihood that each daughter cell will have one copy of each chromosome. After this process—which is called Biorientation—is completed, microtubules pull the chromosomes to opposite ends of the cell, which then divides. Proteins called shugoshin proteins are known to be involved in Biorientation in many organisms. These proteins are found in a region called the pericentromere, which surrounds the area on the chromosomes that the microtubules attach to, but the details of their involvement in Biorientation are not fully understood. Now Verzijlbergen et al. have exploited sophisticated genetic techniques in yeast to explore how shugoshin proteins work. These experiments showed that the shugoshin protein helps to recruit condensin—a protein that keeps the DNA organized within the chromosome—to the pericentromere to assist with Biorientation. It also keeps aurora B kinase—one of the enzymes that helps to correct errors during cell division—in the pericentromere when a microtubule attaches to the wrong chromosome. These results help us understand how a ‘hub’ in the pericentromere ensures Biorientation. The next challenge will be to understand how this hub is disassembled after Biorientation to allow error-free cell division to proceed. As shugoshins have been found to be damaged in some cancers, understanding the workings of this hub could also shed new light on how they contribute to disease.
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shugoshin promotes sister kinetochore Biorientation in saccharomyces cerevisiae
Molecular Biology of the Cell, 2007Co-Authors: Brendan M Kiburz, Angelika Amon, Adele L MarstonAbstract:Chromosome segregation must be executed accurately during both mitotic and meiotic cell divisions. Sgo1 plays a key role in ensuring faithful chromosome segregation in at least two ways. During meiosis this protein regulates the removal of cohesins, the proteins that hold sister chromatids together, from chromosomes. During mitosis, Sgo1 is required for sensing the absence of tension caused by sister kinetochores not being attached to microtubules emanating from opposite poles. Here we describe a differential requirement for Sgo1 in the segregation of homologous chromosomes and sister chromatids. Sgo1 plays only a minor role in segregating homologous chromosomes at meiosis I. In contrast, Sgo1 is important to bias sister kinetochores toward Biorientation. We suggest that Sgo1 acts at sister kinetochores to promote their Biorientation.
Geert J P L Kops - One of the best experts on this subject based on the ideXlab platform.
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Arrayed BUB recruitment modules in the kinetochore scaffold KNL1 promote accurate chromosome segregation
Journal of Cell Biology, 2013Co-Authors: Mathijs Vleugel, Eelco Tromer, Manja Omerzu, Vincent Groenewold, Wilco Nijenhuis, Berend Snel, Geert J P L KopsAbstract:Fidelity of chromosome segregation relies on coordination of chromosome Biorientation and the spindle checkpoint. Central to this is the kinetochore scaffold KNL1 that integrates the functions of various mitotic regulators including BUB1 and BUBR1. We show that KNL1 contains an extensive array of short linear sequence modules that encompass TxxΩ and MELT motifs and that can independently localize BUB1. Engineered KNL1 variants with few modules recruit low levels of BUB1 to kinetochores but support a robust checkpoint. Increasing numbers of modules concomitantly increase kinetochore BUB1 levels and progressively enhance efficiency of chromosome Biorientation. Remarkably, normal KNL1 function is maintained by replacing all modules with a short array of naturally occurring or identical, artificially designed ones. A minimal array of generic BUB recruitment modules in KNL1 thus suffices for accurate chromosome segregation. Widespread divergence in the amount and sequence of these modules in KNL1 homologues may represent flexibility in adapting regulation of mitotic processes to altered requirements for chromosome segregation during evolution.
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Connecting up and clearing out: how kinetochore attachment silences the spindle assembly checkpoint
Chromosoma, 2012Co-Authors: Geert J P L Kops, Jagesh V. ShahAbstract:With the goal of creating two genetically identical daughter cells, cell division culminates in the equal segregation of sister chromatids. This phase of cell division is monitored by a cell cycle checkpoint known as the spindle assembly checkpoint (SAC). The SAC actively prevents chromosome segregation while one or more chromosomes, or more accurately kinetochores, remain unattached to the mitotic spindle. Such unattached kinetochores recruit SAC proteins to assemble a diffusible anaphase inhibitor. Kinetochores stop production of this inhibitor once microtubules (MTs) of the mitotic spindle are bound, but productive attachment of all kinetochores is required to satisfy the SAC, initiate anaphase, and exit from mitosis. Although mechanisms of kinetochore signaling and SAC inhibitor assembly and function have received the bulk of attention in the past two decades, recent work has focused on the principles of SAC silencing. Here, we review the mechanisms that silence SAC signaling at the kinetochore, and in particular, how attachment to spindle MTs and Biorientation on the mitotic spindle may turn off inhibitor generation. Future challenges in this area are highlighted towards the goal of building a comprehensive molecular model of this process.
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Mps1 promotes rapid centromere accumulation of Aurora B
EMBO Reports, 2012Co-Authors: Maike S. Van Der Waal, Geert J P L Kops, Mathijs Vleugel, Adrian T. Saurin, Daniel W. Gerlich, Martijn J. M. Vromans, Claudia Wurzenberger, René H. Medema, Susanne M. A. LensAbstract:Aurora B localization to mitotic centromeres, which is required for proper chromosome alignment during mitosis, relies on Haspin-dependent histone H3 phosphorylation and on Bub1-dependent histone H2A phosphorylation—which interacts with Borealin through a Shugoshin (Sgo) intermediate. We demonstrate that Mps1 stimulates the latter recruitment axis. Mps1 activity enhances H2A-T120ph and is critical for Sgo1 recruitment to centromeres, thereby promoting Aurora B centromere recruitment in early mitosis. Importantly, chromosome Biorientation defects caused by Mps1 inhibition are improved by restoring Aurora B centromere recruitment. As Mps1 kinetochore localization reciprocally depends on Aurora B, we propose that this Aurora B-Mps1 recruitment circuitry cooperates with the Aurora B-Haspin feedback loop to ensure rapid centromere accumulation of Aurora B at the onset of mitosis.
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Finding the middle ground: how kinetochores power chromosome congression
Cellular and Molecular Life Sciences, 2010Co-Authors: Geert J P L Kops, Adrian T. Saurin, Patrick MeraldiAbstract:Genomic stability requires error-free chromosome segregation during mitosis. Chromosome congression to the spindle equator precedes chromosome segregation in anaphase and is a hallmark of metazoan mitosis. Here we review the current knowledge and concepts on the processes that underlie chromosome congression, including initial attachment to spindle microtubules, Biorientation, and movements, from the perspective of the kinetochore.
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dividing the goods co ordination of chromosome Biorientation and mitotic checkpoint signalling by mitotic kinases
Biochemical Society Transactions, 2009Co-Authors: Geert J P L KopsAbstract:Error-free chromosome segregation during cell division relies on chromosome Biorientation and mitotic checkpoint activity. A group of unrelated kinases controls various aspects of both processes. The present short review outlines our current understanding of the roles of these kinases in maintaining chromosomal stability.
Richard J Mcintosh - One of the best experts on this subject based on the ideXlab platform.
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mechanisms of chromosome Biorientation and bipolar spindle assembly analyzed by computational modeling
eLife, 2020Co-Authors: Christopher Edelmaier, Adam Lamson, Zachary R Gergely, Saad Ansari, Robert Blackwell, Richard J Mcintosh, Matthew A Glaser, M D BettertonAbstract:The essential functions required for mitotic spindle assembly and chromosome Biorientation and segregation are not fully understood, despite extensive study. To illuminate the combinations of ingredients most important to align and segregate chromosomes and simultaneously assemble a bipolar spindle, we developed a computational model of fission-yeast mitosis. Robust chromosome Biorientation requires progressive restriction of attachment geometry, destabilization of misaligned attachments, and attachment force dependence. Large spindle length fluctuations can occur when the kinetochore-microtubule attachment lifetime is long. The primary spindle force generators are kinesin-5 motors and crosslinkers in early mitosis, while interkinetochore stretch becomes important after Biorientation. The same mechanisms that contribute to persistent Biorientation lead to segregation of chromosomes to the poles after anaphase onset. This model therefore provides a framework to interrogate key requirements for robust chromosome Biorientation, spindle length regulation, and force generation in the spindle.
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mechanisms of chromosome Biorientation and bipolar spindle assembly analyzed by computational modeling
bioRxiv, 2019Co-Authors: Christopher Edelmaier, Adam Lamson, Zachary R Gergely, Saad Ansari, Robert Blackwell, Richard J Mcintosh, Matthew A Glaser, M D BettertonAbstract:The essential functions required for mitotic spindle assembly and chromosome Biorientation and segregation are not fully understood, despite extensive study. To illuminate the combinations of ingredients most important to align and segregate chromosomes in mitosis and simultaneously assemble a bipolar spindle, we developed a computational model of fission-yeast mitosis. Starting from an initial condition that mimics the onset of mitosis, the model assembles a bipolar spindle, attaches to and aligns chromosomes, corrects attachment errors, and segregates the chromosomes to the poles. The model exhibits quantitative agreement with data from fission yeast. By examining model requirements for long-lived Biorientation, we find that progressive restriction of attachment geometry, destabilization of misaligned attachments, and attachment force dependence are important for robust chromosome alignment. Spindle length changes consistent with the force-balance model, and large fluctuations in spindle length can occur when the kinetochore-microtubule attachment lifetime is long. During spindle assembly, the primary spindle force generators are kinesin-5 motors and crosslinkers, while interkinetochore stretch becomes important after Biorientation occurs. The same mechanisms that contribute to persistent Biorientation lead to segregation of chromosomes to the poles after anaphase onset. This model therefore provides a framework to interrogate key requirements for robust chromosome Biorientation, spindle length regulation, and force generation in the spindle.
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mitotic chromosome Biorientation in fission yeast is enhanced by dynein and a minus end directed kinesin like protein
Molecular Biology of the Cell, 2007Co-Authors: Ekaterina L Grishchuk, Ilia S Spiridonov, Richard J McintoshAbstract:Chromosome Biorientation, the attachment of sister kinetochores to sister spindle poles, is vitally important for accurate chromosome segregation. We have studied this process by following the congression of pole-proximal kinetochores and their subsequent anaphase segregation in fission yeast cells that carry deletions in any or all of this organism's minus end–directed, microtubule-dependent motors: two related kinesin 14s (Pkl1p and Klp2p) and dynein. None of these deletions abolished Biorientation, but fewer chromosomes segregated normally without Pkl1p, and to a lesser degree without dynein, than in wild-type cells. In the absence of Pkl1p, which normally localizes to the spindle and its poles, the checkpoint that monitors chromosome Biorientation was defective, leading to frequent precocious anaphase. Ultrastructural analysis of mutant mitotic spindles suggests that Pkl1p contributes to error-free Biorientation by promoting normal spindle pole organization, whereas dynein helps to anchor a focused bundle of spindle microtubules at the pole.
Christopher S Campbell - One of the best experts on this subject based on the ideXlab platform.
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an engineered minimal chromosomal passenger complex reveals a role for incenp sli15 spindle association in chromosome Biorientation
Journal of Cell Biology, 2017Co-Authors: Sarah Fink, Kira Turnbull, Arshad Desai, Christopher S CampbellAbstract:The four-subunit chromosomal passenger complex (CPC), whose enzymatic subunit is Aurora B kinase, promotes chromosome Biorientation by detaching incorrect kinetochore–microtubule attachments. In this study, we use a combination of truncations and artificial dimerization in budding yeast to define the minimal CPC elements essential for its Biorientation function. We engineered a minimal CPC comprised of the dimerized last third of the kinase-activating Sli15/INCENP scaffold and the catalytic subunit Ipl1/Aurora B. Although native Sli15 is not oligomeric, artificial dimerization suppressed the Biorientation defect and lethality associated with deletion of a majority of its microtubule-binding domain. Dimerization did not act through a physical clustering-based kinase activation mechanism but instead promoted spindle association, likely via a putative helical domain in Sli15 that is essential even when dimerized and is required to target kinetochore substrates. Based on the engineering and characterization of a minimal CPC, we suggest that spindle association is important for active Ipl1/Aurora B complexes to preferentially destabilize misattached kinetochores.
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An engineered minimal chromosomal passenger complex reveals a role for INCENP/Sli15 spindle association in chromosome Biorientation.
Journal of Cell Biology, 2017Co-Authors: Sarah Fink, Kira Turnbull, Arshad Desai, Christopher S CampbellAbstract:The four-subunit chromosomal passenger complex (CPC), whose enzymatic subunit is Aurora B kinase, promotes chromosome Biorientation by detaching incorrect kinetochore–microtubule attachments. In this study, we use a combination of truncations and artificial dimerization in budding yeast to define the minimal CPC elements essential for its Biorientation function. We engineered a minimal CPC comprised of the dimerized last third of the kinase-activating Sli15/INCENP scaffold and the catalytic subunit Ipl1/Aurora B. Although native Sli15 is not oligomeric, artificial dimerization suppressed the Biorientation defect and lethality associated with deletion of a majority of its microtubule-binding domain. Dimerization did not act through a physical clustering-based kinase activation mechanism but instead promoted spindle association, likely via a putative helical domain in Sli15 that is essential even when dimerized and is required to target kinetochore substrates. Based on the engineering and characterization of a minimal CPC, we suggest that spindle association is important for active Ipl1/Aurora B complexes to preferentially destabilize misattached kinetochores.
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Tension sensing by Aurora B kinase is independent of survivin-based centromere localization
Nature, 2013Co-Authors: Christopher S Campbell, Arshad DesaiAbstract:The current model to explain accurate chromosome segregation after DNA replication holds that kinetochore–microtubule attachments exert tension across the centromere and are stabilized by spatial separation from inner centromere-localized Aurora B; here an alternative model is presented, wherein active Aurora B produced by clustering is sufficient to ensure Biorientation through a mechanism that is intrinsic to the kinetochore.
