scholarly journals Premature Sister Chromatid Separation Is Poorly Detected by the Spindle Assembly Checkpoint as a Result of System-Level Feedback

Cell Reports ◽  
2015 ◽  
Vol 13 (3) ◽  
pp. 469-478 ◽  
Author(s):  
Mihailo Mirkovic ◽  
Lukas H. Hutter ◽  
Béla Novák ◽  
Raquel A. Oliveira
Keyword(s):  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
System Level ◽  
Sister Chromatid Separation

scholarly journals Cdc14 Inhibition by the Spindle Assembly Checkpoint Prevents Unscheduled Centrosome Separation in Budding Yeast

2009 ◽  
Vol 20 (10) ◽  
pp. 2626-2637 ◽  
Author(s):  
Elena Chiroli ◽  
Giulia Rancati ◽  
Ilaria Catusi ◽  
Giovanna Lucchini ◽  
Simonetta Piatti
Keyword(s):  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cytoplasmic Dynein ◽  
Spindle Pole ◽  
Mitotic Exit ◽  
Centrosome Separation ◽  
Spindle Pole Bodies ◽  
Early Anaphase ◽  
Sister Chromatid Separation

The spindle assembly checkpoint (SAC) is an evolutionarily conserved surveillance mechanism that delays anaphase onset and mitotic exit in response to the lack of kinetochore attachment. The target of the SAC is the E3 ubiquitin ligase anaphase-promoting complex (APC) bound to its Cdc20 activator. The Cdc20/APC complex is in turn required for sister chromatid separation and mitotic exit through ubiquitin-mediated proteolysis of securin, thus relieving inhibition of separase that unties sister chromatids. Separase is also involved in the Cdc-fourteen early anaphase release (FEAR) pathway of nucleolar release and activation of the Cdc14 phosphatase, which regulates several microtubule-linked processes at the metaphase/anaphase transition and also drives mitotic exit. Here, we report that the SAC prevents separation of microtubule-organizing centers (spindle pole bodies [SPBs]) when spindle assembly is defective. Under these circumstances, failure of SAC activation causes unscheduled SPB separation, which requires Cdc20/APC, the FEAR pathway, cytoplasmic dynein, and the actin cytoskeleton. We propose that, besides inhibiting sister chromatid separation, the SAC preserves the accurate transmission of chromosomes also by preventing SPBs to migrate far apart until the conditions to assemble a bipolar spindle are satisfied.

scholarly journals Absence of the Spindle Assembly Checkpoint restores mitotic fidelity upon loss of sister chromatid cohesion

2018 ◽  
Author(s):  
Rui D. Silva ◽  
Mihailo Mirkovic ◽  
Leonardo G. Guilgur ◽  
Om S. Rathore ◽  
Rui Gonçalo Martinho ◽  
...  
Keyword(s):  
Cell Survival ◽  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Genome Shuffling ◽  
Spindle Assembly ◽  
Sister Chromatid Cohesion ◽  
Abnormal Development ◽  
Wing Development ◽  
Cohesin Complex ◽  
Chromatid Cohesion

AbstractSister chromatid cohesion is essential for faithful mitosis, as premature cohesion loss leads to random chromosome segregation and aneuploidy, resulting in abnormal development. To identify specific conditions capable of restoring defects associated with cohesion loss, we screened for genes whose depletion modulates Drosophila wing development when sister chromatid cohesion is impaired. Cohesion deficiency was induced by knock-down of the acetyltransferase Separation anxiety (San)/Naa50, a cohesin complex stabilizer. Several genes whose function impacts wing development upon cohesion loss were identified. Surprisingly, knockdown of key Spindle Assembly Checkpoint (SAC) proteins, Mad2 and Mps1, suppressed developmental defects associated with San depletion. SAC impairment upon cohesin removal, triggered by San depletion or artificial removal of the cohesin complex, prevented extensive genome shuffling, reduced segregation defects and restored cell survival. This counterintuitive phenotypic suppression was caused by an intrinsic bias for efficient chromosome bi-orientation at mitotic entry, coupled with slow engagement of error-correction reactions. We conclude that mitotic timing determines the severity of defects associated with cohesion deficiency. Therefore, although divisions are still error-prone, SAC inactivation enhances cell survival and tissue homeostasis upon cohesion loss.

Unscheduled Sister-Chromatid-Separation in the Presence of Spindle Disruption in Acute Myelogenous Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3114-3114
Author(s):  
Dominik Schnerch ◽  
Julia Felthaus ◽  
Monika Engelhardt ◽  
Ralph M. Waesch
Keyword(s):  
Cell Cycle ◽  
Cell Lines ◽  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Burkitt’S Lymphoma ◽  
Lymphoma Cell ◽  
Myelogenous Leukemia ◽  
Burkitt's Lymphoma ◽  
Cell Cycle Regulators ◽  
Sister Chromatid Separation

Abstract Mitosis is known to be one of the most critical events in the cell cycle. The spindle assembly checkpoint (SAC) is required for proper chromosome segregation during mitosis. The SAC serves as a mitotic surveillance mechanism responsible for detection of misassembly of chromosomes to the mitotic spindle. Lack of chromosome attachment and spindle tension generate a specific „wait-anaphase-signal“. This particular signal interferes with proteolysis, depending on the ubiquitin-ligase APCCdc20, thereby inhibiting mitotic progression through stabilization of mitotic regulators. We found several AML cell lines to be incapable of properly accumulating in mitosis upon nocodazole-induced spindle disruption when compared to a set of Burkitt’s lymphoma cell lines. This result was further supported by the degradation of the mitotic regulators Cyclin B and Securin in synchronized Kasumi-1 cells in the presence of nocodazole shortly after entering mitosis. Interestingly, the SAC proteins BubR1 and Bub1 were found at low expression levels in AML cell lines in comparison to Burkitt’s lymphoma cell lines. We established a shRNA-based model to evaluate the effects of BubR1- and/or Bub1-repression to levels found in AML cell lines to directly compare the Bub1/BubR1 knockdown phenotype with the investigated AML cell lines. Our findings support the view that BubR1 repression alone is sufficient to confer SAC deficiency. To determine the frequency of BubR1 repression in patient-derived primary cells, AML blasts were cytokine-stimulated to enter the cell cycle. Flow cytometry-based G2/M-specific expression analysis of BubR1 in primary AML blasts revealed lower expression in most analyzed cell populations. To further test the hypothesis that AML cells override the metaphase-to-anaphase-transition despite spindle damage, we performed Giemsa staining of cells that were incubated in nocodazole containing growth medium. In AML cell lines, unlike the analyzed Burkitt’s lymphoma cell lines, a significant number of metaphase-like cells contained single chromatids, suggesting premature sister chromatid separation in the presence of spindle damage. Premature sister-chromatid-separation in the presence of chromosomal misalignment would lead to aneuploidy and favor the onset of genomic instability. Our recent efforts focus on high-throughput automated live cell scanning, promising a better understanding of cell division and chromosome separation in the context of different challenges, such as spindle damage. This powerful tool allows a more precise characterization of our knockdown phenotypes in the double-knockdown system, which is a prerequisite for comparison of our model system with AML cell lines. Finally, this new technique might also prove useful to extend our analyses to patient derived AML blasts. As we observed deregulation of SAC protein levels in AML cell lines and primary AML blasts, our findings of premature degradation of cell cycle regulators and unscheduled sister-chromatid-separation suggest an important role for SAC malfunction in the development of AML with karyotypic abnormalities. Mitotic kinases, such as Plk1 and Aurora, are already promising targets for modern antineoplastic therapies. A deeper understanding of mitotic control in AML might contribute to even more sophisticated targeted therapeutic approaches.

scholarly journals Absence of the Spindle Assembly Checkpoint Restores Mitotic Fidelity upon Loss of Sister Chromatid Cohesion

2018 ◽  
Vol 28 (17) ◽  
pp. 2837-2844.e3 ◽  
Author(s):  
Rui D. Silva ◽  
Mihailo Mirkovic ◽  
Leonardo G. Guilgur ◽  
Om S. Rathore ◽  
Rui Gonçalo Martinho ◽  
...  
Keyword(s):  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Sister Chromatid Cohesion ◽  
Chromatid Cohesion

scholarly journals Distinct responses to reduplicated chromosomes require distinct Mad2 responses

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Benjamin M Stormo ◽  
Donald T Fox
Keyword(s):  
Cell Cycle ◽  
Cancer Progression ◽  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Polytene Chromosomes ◽  
Spindle Assembly ◽  
Tissue Development ◽  
Checkpoint Protein ◽  
Independent Mechanism ◽  
Spindle Assembly Checkpoint Protein

Duplicating chromosomes once each cell cycle produces sister chromatid pairs, which separate accurately at anaphase. In contrast, reduplicating chromosomes without separation frequently produces polytene chromosomes, a barrier to accurate mitosis. Chromosome reduplication occurs in many contexts, including: polytene tissue development, polytene tumors, and following treatment with mitosis-blocking chemotherapeutics. However, mechanisms responding to or resolving polyteny during mitosis are poorly understood. Here, using Drosophila, we uncover two distinct reduplicated chromosome responses. First, when reduplicated polytene chromosomes persist into metaphase, an anaphase delay prevents tissue malformation and apoptosis. Second, reduplicated polytene chromosomes can also separate prior to metaphase through a spindle-independent mechanism termed Separation-Into-Recent-Sisters (SIRS). Both reduplication responses require the spindle assembly checkpoint protein Mad2. While Mad2 delays anaphase separation of metaphase polytene chromosomes, Mad2’s control of overall mitotic timing ensures efficient SIRS. Our results pinpoint mechanisms enabling continued proliferation after genome reduplication, a finding with implications for cancer progression and prevention.

scholarly journals Securin and Separase Phosphorylation Act Redundantly to Maintain Sister Chromatid Cohesion in Mammalian Cells

2005 ◽  
Vol 16 (10) ◽  
pp. 4725-4732 ◽  
Author(s):  
Xingxu Huang ◽  
Rashieda Hatcher ◽  
J. Philippe York ◽  
Pumin Zhang
Keyword(s):  
Sister Chromatid ◽  
Mammalian Cells ◽  
Genome Stability ◽  
Spindle Assembly Checkpoint ◽  
Budding Yeast ◽  
Embryonic Stem ◽  
Spindle Assembly ◽  
Sister Chromatid Cohesion ◽  
Chromatid Cohesion ◽  
Spindle Microtubules

The spindle assembly checkpoint monitors the integrity of the spindle microtubules, which attach to sister chromatids at kinetochores and play a vital role in preserving genome stability by preventing missegregation. A key target of the spindle assembly checkpoint is securin, the separase inhibitor. In budding yeast, loss of securin results in precocious sister chromatid separation when the microtubule spindle is disrupted. However, in contrast to budding yeast, mammalian securin is not required for spindle checkpoint, suggesting that there are redundant mechanisms controlling the dissolution of sister chromatid cohesion in the absence of securin. One candidate mechanism is the inhibitory phosphorylation of separase. We generated a nonphosphorylable point mutant (S1121A) separase allele in securin-/- mouse embryonic stem cells. Securin-/-separase+/S1121A cells are viable but fail to maintain sister chromatid cohesion in response to the disruption of spindle microtubules, show enhanced sensitivity to nocodazole, and cannot recover from prometaphase arrest.

scholarly journals Accumulation of Mad2–Cdc20 complex during spindle checkpoint activation requires binding of open and closed conformers of Mad2 in Saccharomyces cerevisiae

2006 ◽  
Vol 174 (1) ◽  
pp. 39-51 ◽  
Author(s):  
Luigi Nezi ◽  
Giulia Rancati ◽  
Anna De Antoni ◽  
Sebastiano Pasqualato ◽  
Simonetta Piatti ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Sister Chromatid ◽  
Hela Cells ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Mitotic Progression ◽  
Checkpoint Activation ◽  
Mutant Proteins ◽  
Checkpoint Protein

The spindle assembly checkpoint (SAC) coordinates mitotic progression with sister chromatid alignment. In mitosis, the checkpoint machinery accumulates at kinetochores, which are scaffolds devoted to microtubule capture. The checkpoint protein Mad2 (mitotic arrest deficient 2) adopts two conformations: open (O-Mad2) and closed (C-Mad2). C-Mad2 forms when Mad2 binds its checkpoint target Cdc20 or its kinetochore receptor Mad1. When unbound to these ligands, Mad2 folds as O-Mad2. In HeLa cells, an essential interaction between C- and O-Mad2 conformers allows Mad1-bound C-Mad2 to recruit cytosolic O-Mad2 to kinetochores. In this study, we show that the interaction of the O and C conformers of Mad2 is conserved in Saccharomyces cerevisiae. MAD2 mutant alleles impaired in this interaction fail to restore the SAC in a mad2 deletion strain. The corresponding mutant proteins bind Mad1 normally, but their ability to bind Cdc20 is dramatically impaired in vivo. Our biochemical and genetic evidence shows that the interaction of O- and C-Mad2 is essential for the SAC and is conserved in evolution.

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