In Vitro FRAP Identifies the Minimal Requirements for Mad2 Kinetochore Dynamics

Vink, Martin; Simonetta, Marco; Transidico, Pietro; Ferrari, Karin Johanna; Mapelli, Marina; A, De Antoni; Massimiliano, Lucia; Ciliberto, Andrea; Faretta, Mario; Salmon, Edward D.; Musacchio, Andrea

doi:10.1016/j.cub.2006.03.057

Cited by 96 publications

(122 citation statements)

References 31 publications

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“…Like Aurora B, FRAP-experiments demonstrated that all MCC-components are dynamic and cycle on and off kinetochores, suggesting that the MCC is assembled at or near kinetochores. [21][22][23][24][25][26] Consistently, subunits of APC were found to be localized at the kinetochores and along chromosome arms 60 and the spindle checkpoint machinery is required to recruit the APC/ C to kinetochores. 61 Because inactivation of the Cul3/KLHL9/KLHL13 E3-ligase leads to ectopic accumulation of active Aurora B along chromosome arms, 44 we propose a model in which Cul3/KLHL9/ KLHL13 E3-ligase maintains SAC activity by directly regulating Aurora B-mediated association of the SAC components at kinetochores (Fig.…”

Section: How Does Cul3/klhl9/klhl13 E3-ligase Maintain Sac Signaling mentioning

confidence: 82%

“…14 In Xenopus egg extracts, the Aurora B-INCENP complex induces kinetochore localization of MPS1 (multipolar spindle 1), BUB1, BUB3 and CENP-E. 15 In mammalian cells, CENP-E and BUBR1 are part of a complex involved in force sensing, 16 and BUBR1, MAD2 and CENP-E are recruited to kinetochores by a mechanism that requires the kinase activity of Aurora B. [17][18][19][20] Kinetochore localization of these SAC components is dynamic, [21][22][23][24][25] similarly to the transient association of Aurora B with centromeric regions of mitotic chromosomes. 26 These results imply that the generation of the SAC-inhibitory signal might be achieved by regulated cycles of assembly and disassembly of kinetochore proteins.…”

Section: Sac Signaling At the Kinetochoresmentioning

confidence: 99%

“…35,36 Alternatively, SAC-inactivation may involve the MAD2-binding protein p31 comet , which functions as a negative regulator of the SAC. 25,38,39 Two recent studies 40,41 propose that checkpoint inactivation is an energy-consuming process, in which APC-dependent multi-ubiquitination leads to the dissociation of MAD2 and BUBR1 from CDC20. Interestingly, this process is reversed by the de-ubiquitinating enzyme USP44 (ubiquitin-specific protease 44) or Protectin, which deubiquitinates CDC20 and thereby directly counteracts the APC-driven disassembly of MCC complexes.…”

Section: Active Sac Inhibits the Apc E3-ligasementioning

confidence: 99%

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A Cul3-Based E3 Ligase Regulates Mitosis and is Required to Maintain the Spindle Assembly Checkpoint in Human Cells

Sumara

Peter

2007

Cell Cycle

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Section: How Does Cul3/klhl9/klhl13 E3-ligase Maintain Sac Signaling mentioning

confidence: 82%

Section: Sac Signaling At the Kinetochoresmentioning

confidence: 99%

Section: Active Sac Inhibits the Apc E3-ligasementioning

confidence: 99%

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A Cul3-Based E3 Ligase Regulates Mitosis and is Required to Maintain the Spindle Assembly Checkpoint in Human Cells

Sumara

Peter

2007

Cell Cycle

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“…Potentially, p31 comet could be up-regulated by using a two-step reaction process; however, such a scheme would have to be conceptually similar to that discussed in Model with an amplification step. We emphasize that p31 comet also could play other important roles, for example, in switching off kinetochore signaling after microtubule attachment (33). Model with an amplification step.…”

Section: Mechanisms With Diffusivementioning

confidence: 99%

Modeling dual pathways for the metazoan spindle assembly checkpoint

Sear

Howard

2006

Proc. Natl. Acad. Sci. U.S.A.

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Using computational modeling, we investigate mechanisms of signal transduction. We focus on the spindle assembly checkpoint, where a single unattached kinetochore is able to signal to prevent cell cycle progression. The inhibitory signal switches off rapidly once spindle microtubules have attached to all kinetochores. This requirement tightly constrains the possible mechanisms. Here we investigate two possible mechanisms for spindle checkpoint operation in metazoan cells, both supported by recent experiments. The first involves the free diffusion and sequestration of cell cycle regulators. This mechanism is severely constrained both by experimental fluorescence recovery data and by the large volumes involved in open mitosis in metazoan cells. By using a simple mathematical analysis and computer simulation, we find that this mechanism can generate the inhibition found in experiment but likely requires a two-stage signal amplification cascade. The second mechanism involves spatial gradients of a short-lived inhibitory signal that propagates first by diffusion but then primarily by active transport along spindle microtubules. We propose that both mechanisms may be operative in the metazoan spindle assembly checkpoint, with either able to trigger anaphase onset even without support from the other pathway.kinetochore ͉ mathematical modeling ͉ signal transduction ͉ concentration gradients T he question of how a signal emanating from a small, compact structure in a cell can be amplified and propagated to an entire cell is fundamental to cell biology (1). An excellent example is provided by the spindle assembly checkpoint (SAC) (2), which regulates cell cycle progression from metaphase to anaphase during mitosis. The segregation of sister chromatids that occurs during anaphase is permitted only after all of the kinetochores are attached by microtubules to the mitotic spindle. Even a single unattached kinetochore can signal to the rest of the cell and prevent cell cycle progression (3, 4). A fundamental issue is how a relatively small structure, such as a kinetochore, can generate sufficient signal to robustly communicate with distant subcellular locations (1). Moreover, this signal must switch off rapidly, within a period of minutes, after complete kinetochore attachment to spindle microtubules (4). These requirements strongly constrain the possible signal transduction mechanisms. In this paper, we focus particularly on the SAC in cases where the nuclear envelope breaks down before SAC activity (open mitosis), as in metazoan cells. In this context, we examine two distinct models: a diffusive sequestration model and a model involving active signal transport along spindle microtubules. We believe that both of these pathways may be in simultaneous operation in the metazoan SAC.The metaphase͞anaphase transition is triggered by an intricate sequence of events centered around the proteins securin, cyclin B, and separase. The first step is the ubiquitination of securin and cyclin B by the anaphase-promoting complex͞cyclosome (...

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“…Central to the SACnetwork is a kinetochore-bound template complex made up from Mad1 and C-Mad2. This template complex recruits O-Mad2 and stabilizes an intermediate conformation (O-Mad2*), which can bind Cdc20 efficiently and switches to closed conformation upon Cdc20-binding, tightening the connection between both with the 'safety-belt' (De Antoni et al, 2005;Luo et al, 2004;Vink et al, 2006).The CMad2:Cdc20 complex formed by this mechanism has been given the name ''template-model'' (De Antoni et al, 2005). Mad2-activation at the kinetochores is commonly seen as the central element of the SAC.…”

Section: Template Model Of Mad2-activationmentioning

confidence: 99%

Systems Biology Modeling of Five Pathways for Regulation and Potent Inhibition of the Anaphase-Promoting Complex (APC/C): Pivotal Roles for MCC and BubR1

Ibrahim

2015

OMICS: A Journal of Integrative Biology

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Correct DNA segregation is a fundamental process that ensures the precise and reliable inheritance of genomic information for the propagation of cell life. Eukaryotic cells have evolved a conserved surveillance control mechanism for DNA segregation named the Spindle Assembly Checkpoint (SAC).The SAC ensures that the sister chromatids of the duplicated genome are not separated and distributed to the spindle poles before all chromosomes have been properly linked to the microtubules of the mitotic spindle. Biochemically, the SAC delays cell cycle progression by preventing activation of the anaphase-promoting complex (APC/C) or cyclosome whose activation by Cdc20 is required for sister-chromatid separation; this marks the transition into anaphase. In response to activation of the checkpoint, various species control the activity of both APC/C and Cdc20. However, the underlying regulatory pathways remain largely elusive. In this study, five possible model variants of APC/C regulation were constructed, namely BubR1, Mad2, MCC, MCF2, and an all-pathways model variant. These models were validated with experimental data from the literature. A wide range of parameter values has been tested to find the critical values of the APC/C binding rate. The results show that all variants are able to capture the wild-type behavior of the APC/C. However, only one model variant, which included both MCC as well as BubR1 as potent inhibitors of the APC/C, was able to reproduce both wild-type and mutant type behavior of APC/C regulation. In conclusion, the presented work informs the regulation of fundamental processes such as SAC and APC/C in cell biology and has successfully distinguished between five competing dynamical models using a systems biology approach. The results attest that systems-level approaches are vital for molecular and cell biology.

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In Vitro FRAP Identifies the Minimal Requirements for Mad2 Kinetochore Dynamics

Cited by 96 publications

References 31 publications

A Cul3-Based E3 Ligase Regulates Mitosis and is Required to Maintain the Spindle Assembly Checkpoint in Human Cells

A Cul3-Based E3 Ligase Regulates Mitosis and is Required to Maintain the Spindle Assembly Checkpoint in Human Cells

Modeling dual pathways for the metazoan spindle assembly checkpoint

Systems Biology Modeling of Five Pathways for Regulation and Potent Inhibition of the Anaphase-Promoting Complex (APC/C): Pivotal Roles for MCC and BubR1

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