Bub1 and CENP-U redundantly recruit Plk1 to stabilize kinetochore-microtubule attachments and ensure accurate chromosome segregation
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA758021 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR15619176_1.fastq.gz | Fastqsanger.gz | |||
| SRR15619176_2.fastq.gz | Fastqsanger.gz | |||
| SRR15619177_1.fastq.gz | Fastqsanger.gz | |||
| SRR15619177_2.fastq.gz | Fastqsanger.gz | |||
| SRR15619178_1.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 18 |
Similar Datasets
Project description:Bub1 is required for the kinetochore/centromere localization of two essential mitotic kinases Plk1 and Aurora B. Surprisingly, stable depletion of Bub1 by ~95% in human cells marginally affects whole chromosome segregation fidelity. We show that CENP-U, which is recruited to kinetochores by the CENP-P and CENP-Q subunits of the CENP-O complex, is required to prevent chromosome mis-segregation in Bub1-depleted cells. Mechanistically, Bub1 and CENP-U redundantly recruit Plk1 to kinetochores to stabilize kinetochore-microtubule attachments, thereby ensuring accurate chromosome segregation. Furthermore, unlike its budding yeast homolog, the CENP-O complex does not regulate centromeric localization of Aurora B. Consistently, depletion of Bub1 or CENP-U sensitizes cells to the inhibition of Plk1 but not Aurora B kinase activity. Taken together, our findings provide mechanistic insight into the regulation of kinetochore function, which may have implications for targeted treatment of cancer cells with mutations perturbing kinetochore recruitment of Plk1 by Bub1 or the CENP-O complex.
2021-08-29 | GSE182827 | GEO
Project description:Kinetochore localized Mad1 is essential for generating a “wait anaphase” signal during mitosis hereby ensuring accurate chromosome segregation. Inconsistent models for the function and quantitative contribution of the two mammalian Mad1 kinetochore receptors: Bub1 and the Rod-Zwilch-ZW10 (RZZ) complex exist. By combining genome editing and RNAi we achieve penetrant removal of Bub1 and Rod in human cells, which reveals that efficient checkpoint signaling depends on the integrated activities of these proteins. Rod removal reduces the proximity of Bub1 and Mad1 and we can bypass the requirement for Rod by tethering Mad1 to kinetochores or increasing the strength of the Bub1-Mad1 interaction. We find that Bub1 has checkpoint functions independent of Mad1 localization that are supported by low levels of Bub1 suggesting a catalytic function. In conclusion, our results support an integrated model for the Mad1 receptors in which the primary role of RZZ is to localize Mad1 at kinetochores to generate the Mad1-Bub1 complex.
2019-02-19 | PXD012235 | Pride
Project description:The accurate segregation of chromosomes during mitosis relies on the attachment of sister chromatids to microtubules from opposite poles, called biorientation. Sister chromatid cohesion resists microtubule forces, generating tension which provides the signal that biorientation has occurred. How tension silences the surveillance pathways that prevent cell cycle progression and correct erroneous kinetochore-microtubule attachments remains unclear. Here we identify SUMOylation as a mechanism that promotes anaphase onset upon biorientation. SUMO ligases modify the tension-sensing pericentromere-localized chromatin protein, shugoshin, to stabilize bioriented sister kinetochore-microtubule attachments. In the absence of SUMOylation, Aurora B kinase removal from kinetochores is delayed. Shugoshin SUMOylation prevents its binding to protein phosphatase 2A (PP2A) and release of this interaction is important for stabilizing sister kinetochore biorientation. We propose that SUMOylation modulates the kinase-phosphatase network within pericentromeres to inactivate the error correction machinery, thereby allowing anaphase entry in response to biorientation.
2021-03-25 | PXD019287 | Pride
Project description:Kinetochore protein phosphorylation promotes the correction of erroneous microtubule attachments to ensure faithful chromosome segregation during cell division. Determining how phosphorylation executes error correction requires an understanding of whether kinetochore substrates are completely (i.e. all-or-none) or only fractionally phosphorylated. Using quantitative mass spectrometry (MS), we measured phospho-occupancy on the conserved kinetochore protein Hec1 (NDC80) that directly binds microtubules. None of the positions measured exceeded ~50% phospho-occupancy, and the cumulative phospho-occupancy changed by only ~20% in response to changes in microtubule attachment status. The narrow dynamic range of phospho-occupancy is maintained, in part, by ongoing phosphatase activity. Further, both Cdk1-Cyclin B1 and Aurora kinases phosphorylate Hec1 to enhance error correction in response to different types of microtubule attachment errors. The low inherent phospho-occupancy promotes microtubule attachment to kinetochores while the high sensitivity of kinetochore-microtubule attachments to small changes in phospho-occupancy drives error correction and ensures high mitotic fidelity.
2022-06-27 | MSV000089738 | MassIVE
Project description:Polo-like kinase 1 (PLK1) is a serine/threonine kinase required for mitosis and cytokinesis. As cancer cells are often hypersensitive to partial PLK1 inactivation, chemical inhibitors of PLK1 have been developed and tested in clinical trials. However, these molecules alone were not completely effective. PLK1 promotes numerous molecular and cellular events in the cell division cycle. To date, it is unclear which of these events most crucially depend on PLK1 activity. We used a CRISPR-based genome-wide screening strategy to identify genes whose inactivation enhances cell proliferation defects upon partial chemical inhibition of PLK1. Genes identified encode proteins that are functionally linked to PLK1 in multiple ways. Among them, factors that promote centromere and kinetochore function were clearly enriched. In particular, inactivation of the kinesin KIF18A or SKA1 in PLK1-compromised cells results in mitotic defects, activation of the spindle assembly checkpoint and nuclear reassembly defects. Our results suggest that functions of PLK1 at kinetochores are most sensitive to its inhibition, and point at KIF18A as a possible target for combinatorial therapies using existing PLK1 inhibitors.
2023-10-02 | GSE228155 | GEO
Project description:Spindle assembly checkpoint (SAC) regulators such as the Mps1 kinase not only delay anaphase onset, but also correct improper chromosome-spindle linkages that would otherwise lead to missegregation and aneuploidy. However, the substrates and mechanisms involved in this pathway of error correction remain poorly understood. Using a chemically tuned kinetochore-targeting assay, we show that Mps1 destabilizes microtubule attachments (K-fibers) epistatically to Aurora B, the other major error-correcting kinase. Through chemical genetics and quantitative proteomics, we identify both known and novel sites of Mps1- regulated phosphorylation at the outer kinetochore. Modification of these substrates was sensitive to microtubule tension and counterbalanced by the PP2A-B56 phosphatase, a positive regulator of chromosome-spindle interactions. Consistently, Mps1 inhibition rescued K-fiber stability after depleting PP2A-B56. We also identify the hinge region of the W-shaped Ska complex as a key effector of Mps1 at the kinetochore-microtubule interface, as mutations that mimic constitutive phosphorylation strongly destabilized K-fibers in vivo and inhibited the Ska complex’s conversion from lattice diffusion to end-coupled microtubule binding in vitro. Together these results provide new insights into how Mps1 modulates the microtubule-binding properties of the kinetochore to promote the selective stabilization of bipolar attachments and error-free chromosome segregation.
2018-10-24 | PXD006135 | Pride
Project description:Plk1 is a key mitotic kinase that localizes to distinct subcellular structures to promote accurate mitotic progression. Plk1 recruitment depends on direct interaction between polo-box domain (PBD) on Plk1 and PBD binding motif (PBD BM) on the interactors. However, recent study showed that PBD BM alone is not enough for stable binding between CENP-U and Plk1 highlighting the complexity of the interaction which warrants further investigation. An important interactor for Plk1 during mitosis is the checkpoint protein BubR1. Plk1 bound to BubR1 via PBD interaction with pT620 phosphorylates BubR1 S676/T680 to promote BubR1-PP2A/B56 interaction. The BubR1-PP2A/B56 complex counteracts the destablizing effect on kinetochore-microtubule attachments by mitotic kinases to promote mitotic progression.
Here we show that Plk1 phosphorylates T600/T608 on BubR1 and the double phosphorylation is critical for BubR1-Plk1 interaction. A similar mechanism for Plk1-Bub1 interaction also exists indicating a general principle for Plk1 kinetochore recruitment through self-priming. Mechanistically preventing BubR1 T600/T608 phosphorylation impairs chromosome congression and checkpoint silencing by reducing Plk1 and PP2A/B56 binding to BubR1. Increasing the binding affinity towards Plk1 and PP2A/B56 in BubR1 through protein engineering bypasses the requirement of T600/T608 phosphorylation for mitotic progression. These results reveal a new layer of regulation for accurate mitotic progression.
2024-10-31 | MSV000096264 | MassIVE
Project description:Parthenolide is a natural compound that has shown highly promising anticancer activity. Even though its mode of action has been studied for decades, its antimitotic activity has been largely overlooked, limiting the understanding of its full anticancer potential. In this study, we combined click chemistry with quantitative mass spectrometry and cell biology to elucidate the mechanism of action of parthenolide in mitosis. We show that parthenolide does not act as a microtubule-targeting agent in cells. Instead, it binds to the kinetochore protein ZNF207/BUGZ, preventing the establishment of proper kinetochore-microtubule attachment. Our results show that parthenolide covalently binds to Cys54 of BUGZ via Michael addition to its α-methylene-γ-lactone moiety. Since Cys54 is located within the second zinc finger domain of BUGZ microtubule-targeting region, we propose that parthenolide interferes with the microtubule-binding ability of BUGZ, consequently preventing kinetochore-microtubule attachments required for accurate chromosome congression to the spindle equator.
2025-05-27 | PXD063178 | Pride
Project description: Not available
| S-EPMC3549975 | biostudies-literature
Project description:During metaphase, in response to improper kinetochore-microtubule attachments, the spindle assembly checkpoint (SAC) activates the mitotic checkpoint complex (MCC) to inhibit the anaphase-promoting complex/cyclosome (APC/C). The Mad1-Mad2 complex provides a catalytic platform for MCC assembly. Mad1-bound Mad2 recruits open-Mad2 through asymmetric dimerization and Mad1 phosphorylation by Mps1 promotes conversion of Mad2 from an open (O-Mad2) to closed (C-Mad2) state, which binds Cdc20 to form the MCC. How Mad1 phosphorylation catalytically activates MCC formation is poorly understood. This study characterises Mad1 phosphorylation by Mps1 and provides structural and biochemical insights into a phosphorylation-specific Mad1-Cdc20 interaction, which allows for a tripartite assembly of Bub1-Mad1-Cdc20 on the C-terminal domain of Mad1. We also identify a folded state of Mad1-Mad2 complex, suggesting a model by which the Cdc20-Mad1 interaction brings the Cdc20 MIM motif near Mad2. The Cdc20 MIM motif is then entrapped by the Mad2 safety belt to form a stable complex, allowing spontaneous MCC assembly.
2023-03-11 | PXD031872 | Pride