Faculty Opinions recommendation of Human Blinkin/AF15q14 is required for chromosome alignment and the mitotic checkpoint through direct interaction with Bub1 and BubR1.

2007 ◽  
Author(s):  
Kevin Hardwick
Keyword(s):  
Direct Interaction ◽  
Mitotic Checkpoint ◽  
Chromosome Alignment

scholarly journals The association of Plk1 with the astrin–kinastrin complex promotes formation and maintenance of a metaphase plate

2020 ◽  
Vol 134 (1) ◽  
pp. jcs251025
Author(s):  
Zoë Geraghty ◽  
Christina Barnard ◽  
Pelin Uluocak ◽  
Ulrike Gruneberg
Keyword(s):  
Molecular Mechanisms ◽  
Metaphase Plate ◽  
Direct Interaction ◽  
Mitotic Chromosomes ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Mitotic Kinase ◽  
Spindle Poles ◽  
Chromosome Congression ◽  
Chromosome Alignment

ABSTRACTErrors in mitotic chromosome segregation can lead to DNA damage and aneuploidy, both hallmarks of cancer. To achieve synchronous error-free segregation, mitotic chromosomes must align at the metaphase plate with stable amphitelic attachments to microtubules emanating from opposing spindle poles. The astrin–kinastrin (astrin is also known as SPAG5 and kinastrin as SKAP) complex, also containing DYNLL1 and MYCBP, is a spindle and kinetochore protein complex with important roles in bipolar spindle formation, chromosome alignment and microtubule–kinetochore attachment. However, the molecular mechanisms by which astrin–kinastrin fulfils these diverse roles are not fully understood. Here, we characterise a direct interaction between astrin and the mitotic kinase Plk1. We identify the Plk1-binding site on astrin as well as four Plk1 phosphorylation sites on astrin. Regulation of astrin by Plk1 is dispensable for bipolar spindle formation and bulk chromosome congression, but promotes stable microtubule–kinetochore attachments and metaphase plate maintenance. It is known that Plk1 activity is required for effective microtubule–kinetochore attachment formation, and we suggest that astrin phosphorylation by Plk1 contributes to this process.

scholarly journals Kinetochore recruitment of CENP-F illustrates how paralog divergence shapes kinetochore composition and function

2018 ◽  
Author(s):  
Giuseppe Ciossani ◽  
Katharina Overlack ◽  
Arsen Petrovic ◽  
Pim Huis in ‘t Veld ◽  
Carolin Körner ◽  
...  
Keyword(s):  
Coiled Coil ◽  
Kinase Domain ◽  
Mitotic Checkpoint ◽  
Checkpoint Control ◽  
Directed Motion ◽  
Coil Structure ◽  
Evolutionary Relatedness ◽  
Chromosome Alignment ◽  
And Function ◽  
Paralogous Proteins

The metazoan proteins CENP-E and CENP-F are components of a fibrous layer of mitotic kinetochores named the corona. Several features suggest that CENP-E and CENP-F are paralogs: they are very large (approximately 2700 and 3200 residues, respectively), rich in predicted coiled-coil structure, C-terminally prenylated, and endowed with microtubule-binding sites at their termini. In addition, CENP-E contains an ATP-hydrolyzing motor domain that promotes microtubule plus-end directed motion. Here, we show that CENP-E and CENP- F are recruited to mitotic kinetochores independently of the Rod-Zwilch-ZW10 (RZZ) complex, the main corona constituent. We identify selective interactions of CENP-E and CENP-F respectively with BubR1 and Bub1, paralogous proteins involved in mitotic checkpoint control and chromosome alignment. While BubR1 is dispensable for kinetochore localization of CENP-E, Bub1 is stringently required for CENP-F localization. Through biochemical reconstitution, we demonstrate that the CENP-E:BubR1 and CENP-F:Bub1 interactions are direct and require similar determinants, a dimeric coiled-coil in CENP-E or CENP-F and a kinase domain in BubR1 or Bub1. Our findings are consistent with the existence of ‘pseudo-symmetric’, paralogous Bub1:CENP-F and BubR1:CENP-E axes, supporting evolutionary relatedness of CENP-E and CENP-F.

scholarly journals CENP-E–dependent BubR1 autophosphorylation enhances chromosome alignment and the mitotic checkpoint

2012 ◽  
Vol 198 (2) ◽  
pp. 205-217 ◽  
Author(s):  
Yige Guo ◽  
Christine Kim ◽  
Sana Ahmad ◽  
Jiayin Zhang ◽  
Yinghui Mao
Keyword(s):  
Mitotic Checkpoint ◽  
The State ◽  
Chromosome Loss ◽  
Spindle Microtubule ◽  
Single Chromosome ◽  
Checkpoint Signaling ◽  
Mitotic Kinase ◽  
Chromosome Alignment ◽  
Kinetochore Function ◽  
Microtubule Capture

How the state of spindle microtubule capture at the kinetochore is translated into mitotic checkpoint signaling remains largely unknown. In this paper, we demonstrate that the kinetochore-associated mitotic kinase BubR1 phosphorylates itself in human cells and that this autophosphorylation is dependent on its binding partner, the kinetochore motor CENP-E. This CENP-E–dependent BubR1 autophosphorylation at unattached kinetochores is important for a full-strength mitotic checkpoint to prevent single chromosome loss. Replacing endogenous BubR1 with a nonphosphorylatable BubR1 mutant, as well as depletion of CENP-E, the BubR1 kinase activator, results in metaphase chromosome misalignment and a decrease of Aurora B–mediated Ndc80 phosphorylation at kinetochores. Furthermore, expressing a phosphomimetic BubR1 mutant substantially reduces the incidence of polar chromosomes in CENP-E–depleted cells. Thus, the state of CENP-E–dependent BubR1 autophosphorylation in response to spindle microtubule capture by CENP-E is important for kinetochore function in achieving accurate chromosome segregation.

scholarly journals Identification of an Overlapping Binding Domain on Cdc20 for Mad2 and Anaphase-Promoting Complex: Model for Spindle Checkpoint Regulation

2001 ◽  
Vol 21 (15) ◽  
pp. 5190-5199 ◽  
Author(s):  
Yongke Zhang ◽  
Emma Lees
Keyword(s):  
Mammalian Cells ◽  
Peptide Mapping ◽  
Spindle Checkpoint ◽  
Direct Interaction ◽  
Mitotic Checkpoint ◽  
Complex Model ◽  
Anaphase Promoting Complex ◽  
Structural Explanation ◽  
Multinucleated Cells ◽  
Truncation Mutant

ABSTRACT Activation of the anaphase-promoting complex (APC) is required for anaphase initiation and for exit from mitosis in mammalian cells. Cdc20, which specifically recognizes APC substrates involved in the metaphase-to-anaphase transition, plays a pivotal role in APC activation through direct interaction with the APC. The activation of the APC by Cdc20 is prevented by the interaction of Cdc20 with Mad2 when the spindle checkpoint is activated. Using deletion mutagenesis and peptide mapping, we have identified the sequences in Cdc20 that target it to Mad2 and the APC, respectively. These sequences are distinct but overlapping, providing a possible structural explanation for the internal modulation of the APC-Cdc20 complex by Mad2. In the course of these studies, a truncation mutant of Cdc20 (1–153) that constitutively binds Mad2 but fails to bind the APC was identified. Overexpression of this mutant induces the formation of multinucleated cells and increases their susceptibility to undergoing apoptosis when treated with microtubule-inhibiting drugs. Our experiments demonstrate that disruption of the Mad2-Cdc20 interaction perturbs the mitotic checkpoint, leading to premature activation of the APC, sensitizing the cells to the cytotoxic effects of microtubule-inhibiting drugs.

scholarly journals Oncogenic Adenomatous Polyposis Coli Mutants Impair the Mitotic Checkpoint through Direct Interaction with Mad2

2009 ◽  
Vol 20 (9) ◽  
pp. 2381-2388 ◽  
Author(s):  
Jiayin Zhang ◽  
Roberto Neisa ◽  
Yinghui Mao
Keyword(s):  
Adenomatous Polyposis Coli ◽  
Mammalian Cells ◽  
Chromosome Instability ◽  
Direct Interaction ◽  
Suppressor Gene ◽  
Mitotic Checkpoint ◽  
Adenomatous Polyposis ◽  
Polyposis Coli ◽  
Egg Extracts

The majority of colorectal tumors are aneuploid because of the underlying chromosome instability (CIN) phenotype, in which a defective mitotic checkpoint is implicated. Adenomatous polyposis coli (APC), a tumor suppressor gene that is commonly mutated in colon cancers, has been suggested in causing CIN; however, the molecular mechanism remains unresolved. In this study, we report an interaction of tumor-associated N-terminal APC fragments (N-APC) with Mad2, an essential mitotic checkpoint protein, providing a direct molecular support for linking APC mutations to the generation of CIN. N-APC interacts with Mad2 in Xenopus egg extracts, colon cancer cells, and in vitro with purified components. The interaction between N-APC and Mad2 decreases the soluble pool of Mad2, which is essential for Mad2 cycling and releasing from unattached kinetochores to produce a diffusible |P`wait anaphase|P' signal. Addition of such an N-APC mutant of egg extracts inactivates the mitotic checkpoint. Expressing a tumor-associated N-APC mutant in mammalian cells with an intact mitotic checkpoint produces premature anaphase onset with missegregated chromosomes.

scholarly journals Kif18A promotes Hec1 dephosphorylation to coordinate chromosome alignment with kinetochore microtubule attachment

2018 ◽  
Author(s):  
Haein Kim ◽  
Jason Stumpff
Keyword(s):  
Molecular Mechanisms ◽  
Spindle Assembly Checkpoint ◽  
Primordial Germ Cells ◽  
Direct Interaction ◽  
Mitotic Arrest ◽  
Microtubule Dynamics ◽  
Mitotic Chromosomes ◽  
Microtubule Attachment ◽  
C Terminus ◽  
Chromosome Alignment

SUMMARYMitotic chromosomes are spatially confined at the spindle equator just prior to chromosome segregation through a process called chromosome alignment. Alignment requires temporal coordination of kinetochore microtubule attachment and dynamics. However, the molecular mechanisms that couple these activities are not understood. Kif18A (kinesin-8) suppresses the dynamics of kinetochore microtubules to promote chromosome alignment during metaphase. Loss of Kif18A function in HeLa and primordial germ cells leads to alignment defects accompanied by a spindle assembly checkpoint (SAC)-dependent mitotic arrest, suggesting the motor also plays a role in regulating kinetochore-microtubule attachments. We show here that Kif18A increases attachment by promoting dephosphorylation of the kinetochore protein Hec1, which provides the primary linkage between kinetochores and microtubules. This function requires a direct interaction between the Kif18A C-terminus and protein phosphatase-1 (PP1). However, the Kif18A-PP1 interaction is not required for chromosome alignment, indicating that regulation of kinetochore microtubule dynamics and attachments are separable Kif18A functions. Mitotic arrest in Kif18A-depleted cells is rescued by expression of a Hec1 variant that mimics a low-phosphorylation state, indicating that Kif18A-dependent Hec1 dephosphorylation is a key step for silencing the checkpoint and promoting mitotic progression. Our data support a model in which Kif18A provides positive feedback for kinetochore microtubule attachment by directly recruiting PP1 to dephosphorylate Hec1. We propose that this function works synergistically with Kif18A’s direct control of kinetochore microtubule dynamics to temporally coordinate chromosome alignment and attachment.

scholarly journals BubR1 and APC/EB1 cooperate to maintain metaphase chromosome alignment

2007 ◽  
Vol 178 (5) ◽  
pp. 773-784 ◽  
Author(s):  
Jiayin Zhang ◽  
Sana Ahmad ◽  
Yinghui Mao
Keyword(s):  
Metaphase Chromosome ◽  
Kinase Activity ◽  
Metaphase Plate ◽  
Mitotic Checkpoint ◽  
Microtubule Attachment ◽  
Chromosome Alignment ◽  
Egg Extracts ◽  
Associated Proteins ◽  
Potential Link ◽  
Segregation Of Chromosomes

The accurate segregation of chromosomes in mitosis requires the stable attachment of microtubules to kinetochores. The details of this complex and dynamic process are poorly understood. In this study, we report the interaction of a kinetochore-associated mitotic checkpoint kinase, BubR1, with two microtubule plus end–associated proteins, adenomatous polyposis coli (APC) and EB1, providing a potential link in stable kinetochore microtubule attachment. Using immunodepletion from and antibody addition to Xenopus laevis egg extracts, we show that BubR1 and its kinase activity are essential for positioning chromosomes at the metaphase plate. BubR1 associates with APC and EB1 in egg extracts, and the complex formation is necessary for metaphase chromosome alignment. Using purified components, BubR1 directly phosphorylates APC and forms a ternary complex with APC and microtubules. These findings support a model in which BubR1 kinase may directly regulate APC function involved in stable kinetochore microtubule attachment.

scholarly journals The association of Plk1 with the Astrin-Kinastrin complex promotes formation and maintenance of a metaphase plate

2020 ◽  
Author(s):  
Zoë Geraghty ◽  
Christina Barnard ◽  
Pelin Uluocak ◽  
Ulrike Gruneberg
Keyword(s):  
Molecular Mechanisms ◽  
Metaphase Plate ◽  
Direct Interaction ◽  
Mitotic Chromosomes ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Mitotic Kinase ◽  
Spindle Poles ◽  
Chromosome Congression ◽  
Chromosome Alignment

AbstractErrors in mitotic chromosome segregation can lead to DNA damage and aneuploidy, both hallmarks of cancer. To achieve synchronous error-free segregation, mitotic chromosomes must align at the metaphase plate with stable amphitelic attachments to microtubules emanating from opposing spindle poles. The Astrin-Kinastrin/SKAP complex, also containing DYNLL1 and MYCBP, is a spindle and kinetochore protein complex with important roles in bipolar spindle formation, chromosome alignment and microtubule-kinetochore attachment. However, the molecular mechanisms by which Astrin-Kinastrin fulfils these diverse roles are not fully understood. Here we characterise a direct interaction between Astrin and the mitotic kinase Plk1. We identify the Plk1-binding site on Astrin as well as four Plk1 phosphorylation sites on Astrin. Regulation of Astrin-Kinastrin by Plk1 is dispensable for bipolar spindle formation and bulk chromosome congression but promotes stable microtubule-kinetochore attachments and metaphase plate maintenance. It is known that Plk1 activity is required for effective microtubule-kinetochore attachment formation, and we suggest that Astrin phosphorylation by Plk1 contributes to this process.SummaryWe demonstrate that Plk1 binds to and phosphorylates the N-terminus of Astrin. This interaction promotes recruitment of the Astrin-complex to kinetochores and stabilises microtubule-kinetochore-attachments in situations when mitosis is delayed.

Nutrient-regulated gene expression in eukaryotes

2006 ◽  
Vol 73 ◽  
pp. 85-96 ◽  
Author(s):  
Richard J. Reece ◽  
Laila Beynon ◽  
Stacey Holden ◽  
Amanda D. Hughes ◽  
Karine Rébora ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Control ◽  
Direct Interaction ◽  
Signalling Pathways ◽  
A Cell ◽  
One Step ◽  
Higher Eukaryotes ◽  
Regulated Gene Expression

The recognition of changes in environmental conditions, and the ability to adapt to these changes, is essential for the viability of cells. There are numerous well characterized systems by which the presence or absence of an individual metabolite may be recognized by a cell. However, the recognition of a metabolite is just one step in a process that often results in changes in the expression of whole sets of genes required to respond to that metabolite. In higher eukaryotes, the signalling pathway between metabolite recognition and transcriptional control can be complex. Recent evidence from the relatively simple eukaryote yeast suggests that complex signalling pathways may be circumvented through the direct interaction between individual metabolites and regulators of RNA polymerase II-mediated transcription. Biochemical and structural analyses are beginning to unravel these elegant genetic control elements.

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