scholarly journals Individual pericentromeres display coordinated motion and stretching in the yeast spindle

2013 ◽  
Vol 203 (3) ◽  
pp. 407-416 ◽  
Author(s):  
Andrew D. Stephens ◽  
Chloe E. Snider ◽  
Julian Haase ◽  
Rachel A. Haggerty ◽  
Paula A. Vasquez ◽  
...  
Keyword(s):  
Stochastic Dynamics ◽  
Attachment Site ◽  
Coordinated Motion ◽  
Daughter Cells ◽  
Microtubule Attachment ◽  
Sister Chromatids ◽  
Attachment Sites ◽  
Physical Linkage ◽  
Pulling Force ◽  
Structural Maintenance Of Chromosomes

The mitotic segregation apparatus composed of microtubules and chromatin functions to faithfully partition a duplicated genome into two daughter cells. Microtubules exert extensional pulling force on sister chromatids toward opposite poles, whereas pericentric chromatin resists with contractile springlike properties. Tension generated from these opposing forces silences the spindle checkpoint to ensure accurate chromosome segregation. It is unknown how the cell senses tension across multiple microtubule attachment sites, considering the stochastic dynamics of microtubule growth and shortening. In budding yeast, there is one microtubule attachment site per chromosome. By labeling several chromosomes, we find that pericentromeres display coordinated motion and stretching in metaphase. The pericentromeres of different chromosomes exhibit physical linkage dependent on centromere function and structural maintenance of chromosomes complexes. Coordinated motion is dependent on condensin and the kinesin motor Cin8, whereas coordinated stretching is dependent on pericentric cohesin and Cin8. Linking of pericentric chromatin through cohesin, condensin, and kinetochore microtubules functions to coordinate dynamics across multiple attachment sites.

scholarly journals Hec1 and Nuf2 Are Core Components of the Kinetochore Outer Plate Essential for Organizing Microtubule Attachment Sites

2005 ◽  
Vol 16 (2) ◽  
pp. 519-531 ◽  
Author(s):  
Jennifer G. DeLuca ◽  
Yimin Dong ◽  
Polla Hergert ◽  
Joshua Strauss ◽  
Jennifer M. Hickey ◽  
...  
Keyword(s):  
Spindle Checkpoint ◽  
Attachment Site ◽  
Outer Plate ◽  
Microtubule Attachment ◽  
Attachment Sites ◽  
Outer Domain ◽  
Core Components ◽  
Fluorescence Light ◽  
Microtubule Formation ◽  
Stable Core

A major goal in the study of vertebrate mitosis is to identify proteins that create the kinetochore-microtubule attachment site. Attachment sites within the kinetochore outer plate generate microtubule dependent forces for chromosome movement and regulate spindle checkpoint protein assembly at the kinetochore. The Ndc80 complex, comprised of Ndc80 (Hec1), Nuf2, Spc24, and Spc25, is essential for metaphase chromosome alignment and anaphase chromosome segregation. It has also been suggested to have roles in kinetochore microtubule formation, production of kinetochore tension, and the spindle checkpoint. Here we show that Nuf2 and Hec1 localize throughout the outer plate, and not the corona, of the vertebrate kinetochore. They are part of a stable “core” region whose assembly dynamics are distinct from other outer domain spindle checkpoint and motor proteins. Furthermore, Nuf2 and Hec1 are required for formation and/or maintenance of the outer plate structure itself. Fluorescence light microscopy, live cell imaging, and electron microscopy provide quantitative data demonstrating that Nuf2 and Hec1 are essential for normal kinetochore microtubule attachment. Our results indicate that Nuf2 and Hec1 are required for organization of stable microtubule plus-end binding sites in the outer plate that are needed for the sustained poleward forces required for biorientation at kinetochores.

scholarly journals An unbiased, quantitative and versatile method for determining misaligned and lagging chromosome during mitosis

2020 ◽  
Author(s):  
Luciano Gama Braga ◽  
Diogjena Katerina Prifti ◽  
Chantal Garand ◽  
Pawan Kumar Saini ◽  
Sabine Elowe
Keyword(s):  
Chromosome Segregation ◽  
Hallmarks Of Cancer ◽  
Analytical Geometry ◽  
Daughter Cells ◽  
Chromosome Missegregation ◽  
Microtubule Attachment ◽  
Sister Chromatids ◽  
Chromosome Congression ◽  
Chromosome Alignment ◽  
Lagging Chromosome

ABSTRACTAccurate chromosome alignment at metaphase facilitates the equal segregation of sister chromatids to each of the nascent daughter cells. Lack of proper metaphase alignment is an indicator of defective chromosome congression and aberrant kinetochore-microtubule attachments which in turn promotes chromosome missegregation and aneuploidy, hallmarks of cancer. Therefore, tools to sensitively and quantitatively measure chromosome alignment at metaphase will facilitate understanding of how changes in the composition and regulation of the microtubule attachment machinery impinge on this process. In this work, we have developed and validated a method based on analytical geometry to measure several indicators of chromosome misalignment. We generated semi-automated and flexible ImageJ2/Fiji pipelines to quantify kinetochore misalignment at metaphase plates as well as lagging chromosomes at anaphase. These tools will ultimately allow sensitive, unbiased, and systematic quantitation of these chromosome segregation defects in cells undergoing mitosis.

scholarly journals Spindle assembly checkpoint proteins are positioned close to core microtubule attachment sites at kinetochores

2013 ◽  
Vol 202 (5) ◽  
pp. 735-746 ◽  
Author(s):  
Dileep Varma ◽  
Xiaohu Wan ◽  
Dhanya Cheerambathur ◽  
Reto Gassmann ◽  
Aussie Suzuki ◽  
...  
Keyword(s):  
Spindle Assembly Checkpoint ◽  
Attachment Site ◽  
Spindle Assembly ◽  
Nanometer Scale ◽  
Outer Plate ◽  
Checkpoint Signaling ◽  
Checkpoint Proteins ◽  
Microtubule Attachment ◽  
C Terminus ◽  
Attachment Sites

Spindle assembly checkpoint proteins have been thought to reside in the peripheral corona region of the kinetochore, distal to microtubule attachment sites at the outer plate. However, recent biochemical evidence indicates that checkpoint proteins are closely linked to the core kinetochore microtubule attachment site comprised of the Knl1–Mis12–Ndc80 (KMN) complexes/KMN network. In this paper, we show that the Knl1–Zwint1 complex is required to recruit the Rod–Zwilch–Zw10 (RZZ) and Mad1–Mad2 complexes to the outer kinetochore. Consistent with this, nanometer-scale mapping indicates that RZZ, Mad1–Mad2, and the C terminus of the dynein recruitment factor Spindly are closely juxtaposed with the KMN network in metaphase cells when their dissociation is blocked and the checkpoint is active. In contrast, the N terminus of Spindly is ∼75 nm outside the calponin homology domain of the Ndc80 complex. These results reveal how checkpoint proteins are integrated within the substructure of the kinetochore and will aid in understanding the coordination of microtubule attachment and checkpoint signaling during chromosome segregation.

The mitosis-regulating and protein-protein interaction activities of Astrin are controlled by Aurora-A-induced phosphorylation

2014 ◽  
Vol 307 (5) ◽  
pp. C466-C478 ◽  
Author(s):  
Shao-Chih Chiu ◽  
Jo-Mei Maureen Chen ◽  
Tong-You Wade Wei ◽  
Tai-Shan Cheng ◽  
Ya-Hui Candice Wang ◽  
...  
Keyword(s):  
Regulatory Networks ◽  
Spindle Assembly Checkpoint ◽  
Morphological Changes ◽  
Spindle Assembly ◽  
Aurora A ◽  
Protein Protein Interaction ◽  
Daughter Cells ◽  
Sister Chromatids ◽  
Complicated Process ◽  
Spindle Stability

Cells display dramatic morphological changes in mitosis, where numerous factors form regulatory networks to orchestrate the complicated process, resulting in extreme fidelity of the segregation of duplicated chromosomes into two daughter cells. Astrin regulates several aspects of mitosis, such as maintaining the cohesion of sister chromatids by inactivating Separase and stabilizing spindle, aligning and segregating chromosomes, and silencing spindle assembly checkpoint by interacting with Src kinase-associated phosphoprotein (SKAP) and cytoplasmic linker-associated protein-1α (CLASP-1α). To understand how Astrin is regulated in mitosis, we report here that Astrin acts as a mitotic phosphoprotein, and Aurora-A phosphorylates Astrin at Ser115. The phosphorylation-deficient mutant Astrin S115A abnormally activates spindle assembly checkpoint and delays mitosis progression, decreases spindle stability, and induces chromosome misalignment. Mechanistic analyses reveal that Astrin phosphorylation mimicking mutant S115D, instead of S115A, binds and induces ubiquitination and degradation of securin, which sequentially activates Separase, an enzyme required for the separation of sister chromatids. Moreover, S115A fails to bind mitosis regulators, including SKAP and CLASP-1α, which results in the mitotic defects observed in Astrin S115A-transfected cells. In conclusion, Aurora-A phosphorylates Astrin and guides the binding of Astrin to its cellular partners, which ensures proper progression of mitosis.

scholarly journals The effect of attachment site mutations on strand exchange in bacteriophage lambda site-specific recombination.

Genetics ◽  
1989 ◽  
Vol 122 (4) ◽  
pp. 727-736
Author(s):  
C E Bauer ◽  
J F Gardner ◽  
R I Gumport ◽  
R A Weisberg
Keyword(s):  
Attachment Site ◽  
Strand Exchange ◽  
Segregation Pattern ◽  
Base Substitution ◽  
Base Pairs ◽  
Multiple Mutation ◽  
Attachment Sites ◽  
Dna Strands ◽  
Substitution Mutations ◽  
Overlap Region

Abstract Recombination of phage lambda attachment sites occurs by sequential exchange of the DNA strands at two specific locations. The first exchange produces a Holliday structure, and the second resolves it to recombinant products. Heterology for base substitution mutations in the region between the two strand exchange points (the overlap region) reduces recombination; some mutations inhibit the accumulation of Holliday structures, others inhibit their resolution to recombinant products. To see if heterology also alters the location of the strand exchange points, we determined the segregation pattern of three single and one multiple base pair substitution mutations of the overlap region in crosses with wild type sites. The mutations are known to differ in the severity of their recombination defect and in the stage of strand exchange they affect. The three single mutations behaved similarly: each segregated into both products of recombination, and the two products of a single crossover were frequently nonreciprocal in the overlap region. In contrast, the multiple mutation preferentially segregated into one of the two recombinant products, and the two products of a single crossover appeared to be fully reciprocal. The simplest explanation of the segregation pattern of the single mutations is that strand exchanges occur at the normal locations to produce recombinants with mismatched base pairs that are frequently repaired. The segregation pattern of the multiple mutation is consistent with the view that both strand exchanges usually occur to one side of the mutant site. We suggest that the segregation pattern of a particular mutation is determined by which stage of strand exchange it inhibits and by the severity of the inhibition.

scholarly journals A major role for Eco1 in regulating cohesin-mediated mitotic chromosome folding

2019 ◽  
Author(s):  
Lise Dauban ◽  
Rémi Montagne ◽  
Agnès Thierry ◽  
Luciana Lazar-Stefanita ◽  
Olivier Gadal ◽  
...  
Keyword(s):  
Mitotic Chromosome ◽  
Dna Looping ◽  
Loop Formation ◽  
Dna Loops ◽  
Sister Chromatids ◽  
Topologically Associating Domains ◽  
Dna Loop ◽  
Main Barrier ◽  
Structural Maintenance Of Chromosomes ◽  
Mitotic Chromatin

AbstractUnderstanding how chromatin organizes spatially into chromatid and how sister chromatids are maintained together during mitosis is of fundamental importance in chromosome biology. Cohesin, a member of the Structural Maintenance of Chromosomes (SMC) complex family, holds sister chromatids together 1–3 and promotes long-range intra-chromatid DNA looping 4,5. These cohesin-mediated DNA loops are important for both higher-order mitotic chromatin compaction6,7 and, in some organisms, compartmentalization of chromosomes during interphase into topologically associating domains (TADs) 8,9. Our understanding of the mechanism(s) by which cohesin generates large DNA loops remains incomplete. It involves a combination of molecular partners and active expansion/extrusion of DNA loops. Here we dissect the roles on loop formation of three partners of the cohesin complex: Pds5 10, Wpl1 11 and Eco1 acetylase 12, during yeast mitosis. We identify a new function for Eco1 in negatively regulating cohesin translocase activity, which powers loop extrusion. In the absence of negative regulation, the main barrier to DNA loop expansion appears to be the centromere. Those results provide new insights on the mechanisms regulating cohesin dependent DNA looping.

scholarly journals rough deal: a gene required for proper mitotic segregation in Drosophila.

1989 ◽  
Vol 109 (6) ◽  
pp. 2951-2961 ◽  
Author(s):  
R E Karess ◽  
D M Glover
Keyword(s):  
Cell Death ◽  
Genetic Locus ◽  
Motile Sperm ◽  
High Frequencies ◽  
Morphological Process ◽  
Daughter Cells ◽  
Chromosome Transmission ◽  
Sister Chromatids ◽  
Mitotic Abnormalities ◽  
Homozygous Mutant

We describe a genetic locus rough deal (rod) in Drosophila melanogaster, identified by mutations that interfere with the faithful transmission of chromosomes to daughter cells during mitosis. Five mutant alleles were isolated, each associated with a similar set of mitotic abnormalities in the dividing neuroblasts of homozygous mutant larvae: high frequencies of aneuploid cells and abnormal anaphase figures, in which chromatids may lag, form bridges, or completely fail to separate. Surviving homozygous adults are sterile, and show cuticular defects associated with cell death, i.e., roughened eyes, sparse abdominal bristles, and notched wing margins. The morphological process of spermatogenesis is largely unaffected and motile sperm are produced, but meiocyte aneuploidy is common. The nature of the observed abnormalities in mitotic cells suggests that the reduced fidelity of chromosome transmission to the daughter cells is due to a failure in a mechanism involved in assuring the proper release of sister chromatids.

AN ANATOMICAL STUDY OF THE LIGAMENTS OF THE ULNAR COMPARTMENT OF THE WRIST

Hand Surgery ◽  
2003 ◽  
Vol 08 (02) ◽  
pp. 219-226 ◽  
Author(s):  
Saburo Sasao ◽  
Moroe Beppu ◽  
Hitoshi Kihara ◽  
Kazuaki Hirata ◽  
Masayuki Takagi
Keyword(s):  
Three Dimensional ◽  
Attachment Site ◽  
Ulnar Collateral Ligament ◽  
Anatomical Study ◽  
Styloid Process ◽  
Dimensional Structure ◽  
Collateral Ligament ◽  
Ulnar Styloid ◽  
Attachment Sites ◽  
Triangular Fibrocartilage

The ligamentous structures of the triangular fibrocartilage complex (TFCC) and their attachments were examined anatomically and histologically using fresh and embalmed cadavers. The TFCC was observed to have a three-dimensional structure consisting of three palmar ligaments — the short radiolunate (SRL), ulnolunate (UL), and ulnotriquetral (UT) ligaments. In addition, the attachment site of the ulnocarpal ligament (UC), which had been previously unknown, was identified. The dorsal components of the TFCC have been previously reported to consist solely of the extensor carpi ulnaris (ECU) subsheath; however, the ligamentous components running from the ulnar styloid process to the triquetrum were found at a layer deeper than the floor of the ECU subsheath. The UC has been reported previously as a two-dimensional structure, but there has been some disagreement as to its attachment sites.2–6,14,15 It is suggested that the dorsal UT ligament should be considered as a separate ligament, based on its different direction and distal attachment site as compared with those of the ulnar collateral ligament (UCL) and ECU subsheath.

Comparison Between Wrist-Worn and Waist-Worn Accelerometry

2017 ◽  
Vol 14 (7) ◽  
pp. 539-545 ◽  
Author(s):  
Paul D. Loprinzi ◽  
Brandee Smith
Keyword(s):  
Pearson Correlation ◽  
Attachment Site ◽  
Free Living ◽  
Extreme Caution ◽  
Attachment Sites ◽  
Midaxillary Line

Objective:To use the most recent ActiGraph model (GT9X) to compare counts per minute (CPM) estimates between wrist-worn and waist-worn attachment sites.Methods:Participants completed 2 conditions (laboratory [N = 13] and free-living conditions [N = 9]), in which during both of these conditions they wore 2 ActiGraph GT9X accelerometers on their nondominant wrist (side-by-side) and 2 ActiGraph GT9X accelerometers on their right hip in line with the midaxillary line (side-by-side). During the laboratory visit, participants completed 5 treadmill-based trials all lasting 5 min: walk at 3 mph, 3.5 mph, 4 mph, and a jog at 6 mph and 6.5 mph. During the free-living setting, participants wore the monitors for 8 hours. Paired t test, Pearson correlation and Bland-Altman analyses were employed to evaluate agreement of CPM between the attachment sites.Results:Across all intensity levels and setting (laboratory and free-living), CPM were statistically significantly and substantively different between waist- and wrist-mounted accelerometry.Conclusion:Attachment site drastically influences CPM. As such, extreme caution should be exercised when comparing CPM estimates among studies employing different attachment site methodologies, particularly waist versus wrist.

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