scholarly journals The Ndc80 complex bridges two Dam1 complex rings

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Jae ook Kim ◽  
Alex Zelter ◽  
Neil T Umbreit ◽  
Athena Bollozos ◽  
Michael Riffle ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Aurora B ◽  
Aurora B Kinase ◽  
Microtubule Attachment ◽  
Sister Chromatids ◽  
Ndc80 Complex ◽  
Interaction Regions ◽  
Dam1 Complex

Strong kinetochore-microtubule attachments are essential for faithful segregation of sister chromatids during mitosis. The Dam1 and Ndc80 complexes are the main microtubule binding components of the Saccharomyces cerevisiae kinetochore. Cooperation between these two complexes enhances kinetochore-microtubule coupling and is regulated by Aurora B kinase. We show that the Ndc80 complex can simultaneously bind and bridge across two Dam1 complex rings through a tripartite interaction, each component of which is regulated by Aurora B kinase. Mutations in any one of the Ndc80p interaction regions abrogates the Ndc80 complex’s ability to bind two Dam1 rings in vitro, and results in kinetochore biorientation and microtubule attachment defects in vivo. We also show that an extra-long Ndc80 complex, engineered to space the two Dam1 rings further apart, does not support growth. Taken together, our work suggests that each kinetochore in vivo contains two Dam1 rings and that proper spacing between the rings is vital.

scholarly journals Cooperation of the Dam1 and Ndc80 kinetochore complexes enhances microtubule coupling and is regulated by aurora B

2010 ◽  
Vol 189 (4) ◽  
pp. 713-723 ◽  
Author(s):  
Jerry F. Tien ◽  
Neil T. Umbreit ◽  
Daniel R. Gestaut ◽  
Andrew D. Franck ◽  
Jeremy Cooper ◽  
...  
Keyword(s):  
Error Correction ◽  
Cell Cycle Progression ◽  
Aurora B ◽  
Cycle Progression ◽  
Microtubule Attachment ◽  
Ndc80 Complex ◽  
Tensile Forces ◽  
Processivity Factor ◽  
Dam1 Complex

The coupling of kinetochores to dynamic spindle microtubules is crucial for chromosome positioning and segregation, error correction, and cell cycle progression. How these fundamental attachments are made and persist under tensile forces from the spindle remain important questions. As microtubule-binding elements, the budding yeast Ndc80 and Dam1 kinetochore complexes are essential and not redundant, but their distinct contributions are unknown. In this study, we show that the Dam1 complex is a processivity factor for the Ndc80 complex, enhancing the ability of the Ndc80 complex to form load-bearing attachments to and track with dynamic microtubule tips in vitro. Moreover, the interaction between the Ndc80 and Dam1 complexes is abolished when the Dam1 complex is phosphorylated by the yeast aurora B kinase Ipl1. This provides evidence for a mechanism by which aurora B resets aberrant kinetochore–microtubule attachments. We propose that the action of the Dam1 complex as a processivity factor in kinetochore–microtubule attachment is regulated by conserved signals for error correction.

scholarly journals CSC-1

2003 ◽  
Vol 161 (2) ◽  
pp. 229-236 ◽  
Author(s):  
Alper Romano ◽  
Annika Guse ◽  
Ivica Krascenicova ◽  
Heinke Schnabel ◽  
Ralf Schnabel ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Genetic Analysis ◽  
Chromosome Segregation ◽  
Kinase Activity ◽  
Aurora B ◽  
Aurora B Kinase ◽  
C Elegans ◽  
The Individual

The Aurora B kinase complex is a critical regulator of chromosome segregation and cytokinesis. In Caenorhabditis elegans, AIR-2 (Aurora B) function requires ICP-1 (Incenp) and BIR-1 (Survivin). In various systems, Aurora B binds to orthologues of these proteins. Through genetic analysis, we have identified a new subunit of the Aurora B kinase complex, CSC-1. C. elegans embryos depleted of CSC-1, AIR-2, ICP-1, or BIR-1 have identical phenotypes. CSC-1, BIR-1, and ICP-1 are interdependent for their localization, and all are required for AIR-2 localization. In vitro, CSC-1 binds directly to BIR-1. The CSC-1/BIR-1 complex, but not the individual subunits, associates with ICP-1. CSC-1 associates with ICP-1, BIR-1, and AIR-2 in vivo. ICP-1 dramatically stimulates AIR-2 kinase activity. This activity is not stimulated by CSC-1/BIR-1, suggesting that these two subunits function as targeting subunits for AIR-2 kinase.

scholarly journals The Topoisomerase I Poison CPT-11 Enhances the Effect of the Aurora B Kinase Inhibitor AZD1152 both In vitro and In vivo

2009 ◽  
Vol 15 (6) ◽  
pp. 2022-2030 ◽  
Author(s):  
Jayasree S. Nair ◽  
Elisa de Stanchina ◽  
Gary K. Schwartz
Keyword(s):  
Topoisomerase I ◽  
Kinase Inhibitor ◽  
Aurora B ◽  
Aurora B Kinase

scholarly journals Aurora B switches relative strength between kinetochore–microtubule attachment modes to promote error correction

2018 ◽  
Author(s):  
Harinath Doodhi ◽  
Taciana Kasciukovic ◽  
Lesley Clayton ◽  
Tomoyuki U. Tanaka
Keyword(s):  
Error Correction ◽  
Chromosome Segregation ◽  
Relative Strength ◽  
Lateral Side ◽  
Aurora B ◽  
Spindle Poles ◽  
Microtubule Attachment ◽  
Attachment Strength ◽  
Dam1 Complex

AbstractFor proper chromosome segregation, sister kinetochores must interact with microtubules from opposite spindle poles; this is called bi-orientation. To establish bi-orientation prior to chromosome segregation, any aberrant kinetochore–microtubule interaction must be resolved (error correction) by Aurora B kinase that phosphorylates outer kinetochore components. Aurora B differentially regulates kinetochore attachment to the microtubule plus end and its lateral side (end-on and lateral attachment, respectively). However, it is still not fully understood how kinetochore–microtubule interactions are exchanged during error correction. Here we reconstituted the kinetochore–microtubule interface of budding yeast in vitro by attaching the Ndc80 complexes (Ndc80C) to nanobeads. These Ndc80C–nanobeads recapitulated in vitro the lateral and end-on attachments of authentic kinetochores, on dynamic microtubules loaded with the Dam1 complex. This in vitro assay enabled the direct comparison of lateral and end-on attachment strength and showed that Dam1 phosphorylation by Aurora B makes the end-on attachment weaker than the lateral attachment. We suggest that the Dam1 phosphorylation weakens interaction with the Ndc80 complex, disrupts the end-on attachment and promotes the exchange to a new lateral attachment, leading to error correction. Our study reveals a fundamental mechanism of error correction for establishment of bi-orientation.

scholarly journals Nucleation of microtubules in vitro by isolated spindle pole bodies of the yeast Saccharomyces cerevisiae.

1978 ◽  
Vol 78 (2) ◽  
pp. 401-414 ◽  
Author(s):  
J S Hyams ◽  
G G Borisy
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Spindle Pole ◽  
Yeast Saccharomyces Cerevisiae ◽  
Microtubule Attachment ◽  
Spindle Pole Bodies ◽  
Attachment Sites ◽  
Air Water Interface ◽  
Microtubule Growth

Spindle pole bodies (SPBs) were isolated from the yeast Saccharomyces cerevisiae by an adaptation of the Kleinschmidt monolayer technique. Spheroplasts prepared from the cells were lysed on an air-water interface. Spread preparations were picked up on grids, transferred to experimental test solutions, and prepared for whole-mount electron microscopy. Using purified exogenous tubulin from porcine brain tissue, the isolated SPBs were shown to nucleate the assembly of microtubules in vitro. Microtubule growth was directional and primarily onto the intranuclear face of the SPB. Neither the morphology nor the microtubule-initiating capacity of the SPB was affected by treatment with the enzymes DNase, RNase, or phospholipase although both properties were sensitive to trypsin. Analysis of SPBs at various stages of the cell cycle showed that newly replicated SPBs had the capacity to nucleate microtubules. SPBs isolated from exponentially growing cells initiated a subset of the yeast spindle microtubules equivalent to the number of pole-to-pole microtubules seen in vivo. However, SPBs isolated from cells in stationary phase and therefore arrested in G1 nucleated a number of microtubules equal to the total chromosomal and pole-to-pole tubules in the yeast spindle. This may mean that in G1-arrested cells, the SPB is associated with microtubule attachment sites of the yeast chromatin.

scholarly journals Targeting Aurora B kinase with Tanshinone IIA suppresses tumor growth and overcomes radioresistance

2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Ming Li ◽  
Haidan Liu ◽  
Qin Zhao ◽  
Shuangze Han ◽  
Li Zhou ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Kinase Inhibitor ◽  
Tanshinone Iia ◽  
Kinase Assay ◽  
Aurora B ◽  
Aurora B Kinase ◽  
Competitive Binding Assay ◽  
In Silico Docking

AbstractAurora B kinase is aberrantly overexpressed in various tumors and shown to be a promising target for anti-cancer therapy. In human oral squamous cell carcinoma (OSCC), the high protein level of Aurora B is required for maintaining of malignant phenotypes, including in vitro cell growth, colony formation, and in vivo tumor development. By molecular modeling screening of 74 commercially available natural products, we identified that Tanshinone IIA (Tan IIA), as a potential Aurora B kinase inhibitor. The in silico docking study indicates that Tan IIA docks into the ATP-binding pocket of Aurora B, which is further confirmed by in vitro kinase assay, ex vivo pull-down, and ATP competitive binding assay. Tan IIA exhibited a significant anti-tumor effect on OSCC cells both in vitro and in vivo, including reduction of Aurora B and histone H3 phosphorylation, induction of G2/M cell cycle arrest, increase the population of polyploid cells, and promotion of apoptosis. The in vivo mouse model revealed that Tan IIA delayed tumor growth of OSCC cells. Tan IIA alone or in combination with radiation overcame radioresistance in OSCC xenograft tumors. Taken together, our data indicate that Tan IIA is an Aurora B kinase inhibitor with therapeutic potentials for cancer treatment.

scholarly journals Kinetochore-associated Mps1 regulates the strength of kinetochore-microtubule attachments via Ndc80 phosphorylation

2021 ◽  
Author(s):  
Krishna K. Sarangapani ◽  
Lori B. Koch ◽  
Christian R. Nelson ◽  
Charles L. Asbury ◽  
Sue Biggins
Keyword(s):  
Error Correction ◽  
Genetic Interactions ◽  
Aurora B ◽  
Aurora B Kinase ◽  
Additional Mechanism ◽  
Microtubule Binding ◽  
Dividing Cells ◽  
Intrinsic Error

AbstractDividing cells detect and correct erroneous kinetochore-microtubule attachments during mitosis, thereby avoiding chromosome mis-segregation. Most studies of this process have focused on the Aurora B kinase, which phosphorylates microtubule-binding elements specifically at incorrectly attached kinetochores, promoting their release and providing another chance for proper attachments to form. However, growing evidence suggests additional mechanisms, potentially involving Mps1 kinase, that also underlie error correction. Because these mechanisms overlap in vivo, and because both Mps1 and Aurora B function in numerous other vital processes, their contributions to the correction of erroneous kinetochore attachments have been difficult to disentangle. Here we directly examine how Mps1 activity affects kinetochore-microtubule attachments using a reconstitution-based approach that allowed us to separate its effects from Aurora B activity. When endogenous Mps1 that co-purifies with isolated kinetochores is activated in vitro, it weakens their attachments to microtubules via phosphorylation of Ndc80, a major microtubule-binding element of the outer kinetochore. Mps1 phosphorylation of Ndc80 appears to contribute to error correction because phospho-deficient Ndc80 mutants exhibit genetic interactions and segregation defects when combined with mutants in an intrinsic error correction pathway. In addition, Mps1 phosphorylation of Ndc80 is stimulated on kinetochores lacking tension. These data suggest that Mps1 provides an additional mechanism for correcting erroneous kinetochore-microtubule attachments, complementing the well-known activity of Aurora B.

scholarly journals Aurora B switches relative strength of kinetochore–microtubule attachment modes for error correction

2021 ◽  
Vol 220 (6) ◽  
Author(s):  
Harinath Doodhi ◽  
Taciana Kasciukovic ◽  
Lesley Clayton ◽  
Tomoyuki U. Tanaka
Keyword(s):  
Error Correction ◽  
Relative Strength ◽  
In Vitro Assay ◽  
Lateral Side ◽  
Aurora B ◽  
Vitro Assay ◽  
Microtubule Attachment ◽  
Attachment Strength ◽  
Dam1 Complex

To establish chromosome biorientation, aberrant kinetochore–microtubule interaction must be resolved (error correction) by Aurora B kinase. Aurora B differentially regulates kinetochore attachment to the microtubule plus end and its lateral side (end-on and lateral attachment, respectively). However, it is still unclear how kinetochore–microtubule interactions are exchanged during error correction. Here, we reconstituted the budding yeast kinetochore–microtubule interface in vitro by attaching the Ndc80 complexes to nanobeads. These Ndc80C nanobeads recapitulated in vitro the lateral and end-on attachments of authentic kinetochores on dynamic microtubules loaded with the Dam1 complex. This in vitro assay enabled the direct comparison of lateral and end-on attachment strength and showed that Dam1 phosphorylation by Aurora B makes the end-on attachment weaker than the lateral attachment. Similar reconstitutions with purified kinetochore particles were used for comparison. We suggest the Dam1 phosphorylation weakens interaction with the Ndc80 complex, disrupts the end-on attachment, and promotes the exchange to a new lateral attachment, leading to error correction.

scholarly journals Targeting Aurora B kinase with Tanshinone IIA Suppresses Tumor Growth and Overcomes Radioresistance

2020 ◽  
Author(s):  
Ming Li ◽  
Feng Gao ◽  
haidan liu ◽  
Qin Zhao ◽  
Shuangze Han ◽  
...  
Keyword(s):  
Molecular Modeling ◽  
Tumor Growth ◽  
Kinase Inhibitor ◽  
Inhibitory Effect ◽  
Tanshinone Iia ◽  
Aurora B ◽  
Aurora B Kinase ◽  
M Cell

Abstract Background Aurora B kinase is aberrantly overexpressed in various tumors and shown to be a promising target for anti-cancer therapy. Methods A customized natural product library was used for natural compound screening through Molecular modeling. The expression of Aurora B in oral squamous cell carcinoma (OSCC) and the inhibitory effect of Tanshinone IIA (Tan IIA) on OSCC were examined by MTS and colony formation assays, immunoblot, immunofluorescence, immunohistochemical staining, and in vivo xenograft experiment. Results Aurora B is overexpressed in OSCC tumor tissues and cell lines. Knockdown of Aurora B inhibited the malignant phenotypes of OSCC cells in vitro and in vivo. With a molecular modeling screening of 74 commercially available natural products, we identified that Tan IIA, as a potential Aurora B kinase inhibitor. Tan IIA exhibited a significant anti-tumor effect on OSCC cells both in vitro and in vivo, including reduction of Aurora B and histone H3 phosphorylation, induction of G2/M cell cycle arrest, increase the population of polyploid cells, and promotion of apoptosis. The in vivo mouse model revealed that Tan IIA delayed tumor growth of OSCC cells. Tan IIA alone or in combination with radiation overcame radioresistance in OSCC xenograft tumors. Conclusion Our data indicates that targeting Aurora B kinase signaling is a promising anti-tumor strategy for OSCC treatment.

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