scholarly journals Structural view of the yeast Dam1 complex, a ring-shaped molecular coupler for the dynamic microtubule end

2020 ◽  
Vol 64 (2) ◽  
pp. 359-370
Author(s):  
Shaowen Wu ◽  
Ekaterina L. Grishchuk
Keyword(s):  
Self Assembly ◽  
Eukaryotic Cell ◽  
Chromosome Loss ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ndc80 Complex ◽  
Unified View ◽  
Dam1 Complex ◽  
Ring Architecture ◽  
Chromosome Motion ◽  
Dynamic Microtubule

Abstract In a dividing eukaryotic cell, proper chromosome segregation requires the dynamic yet persistent attachment of kinetochores to spindle microtubules. In the budding yeast Saccharomyces cerevisiae, this function is especially crucial because each kinetochore is attached to a single microtubule; consequently, loss of attachment could lead to unrecoverable chromosome loss. The highly specialized heterodecameric Dam1 protein complex achieves this coupling by assembling into a microtubule-encircling ring that glides near the end of the dynamic microtubule to mediate chromosome motion. In recent years, we have learned a great deal about the structural properties of the Dam1 heterodecamer, its mechanism of self-assembly into rings, and its tethering to the kinetochore by the elongated Ndc80 complex. The most remarkable progress has resulted from defining the fine structures of helical bundles within Dam1 heterodecamer. In this review, we critically analyze structural observations collected by diverse approaches with the goal of obtaining a unified view of Dam1 ring architecture. A considerable consistency between different studies supports a coherent model of the circular core of the Dam1 ring. However, there are persistent uncertainties about the composition of ring protrusions and flexible extensions, as well as their roles in mediating ring core assembly and interactions with the Ndc80 complex and microtubule.

Fission yeast Pds5 is required for accurate chromosome segregation and for survival after DNA damage or metaphase arrest

2002 ◽  
Vol 115 (3) ◽  
pp. 587-598 ◽  
Author(s):  
Shao-Win Wang ◽  
Rebecca L. Read ◽  
Chris J. Norbury
Keyword(s):  
Dna Damage ◽  
Fission Yeast ◽  
Sister Chromatid ◽  
Budding Yeast ◽  
Eukaryotic Cell ◽  
Sister Chromatid Cohesion ◽  
Chromosome Loss ◽  
Dependent Manner ◽  
Yeast Saccharomyces Cerevisiae ◽  
Chromatid Cohesion

Sister chromatid cohesion, which is established during the S phase of the eukaryotic cell cycle and persists until the onset of anaphase, is essential for the maintenance of genomic integrity. Cohesion requires the multi-protein complex cohesin, as well as a number of accessory proteins including Pds5/BIMD/Spo76. In the budding yeast Saccharomyces cerevisiae Pds5 is an essential protein that localises to chromosomes in a cohesin-dependent manner. Here we describe the characterisation in the fission yeast Schizosaccharomyces pombe of pds5+, a novel,non-essential orthologue of S. cerevisiae PDS5. The S. pombePds5 protein was localised to punctate nuclear foci in a manner that was dependent on the Rad21 cohesin component. This, together with additional genetic evidence, points towards an involvement of S. pombe Pds5 in sister chromatid cohesion. S. pombe pds5 mutants were hypersensitive to DNA damage and to mitotic metaphase delay, but this sensitivity was apparently not due to precocious loss of sister chromatid cohesion. These cells also suffered increased spontaneous chromosome loss and meiotic defects and their viability was dependent on the spindle checkpoint protein Bub1. Thus, while S. pombe Pds5 has an important cohesin-related role, this differs significantly from that of the equivalent budding yeast protein.

scholarly journals The Dam1 complex confers microtubule plus end–tracking activity to the Ndc80 kinetochore complex

2010 ◽  
Vol 189 (4) ◽  
pp. 641-649 ◽  
Author(s):  
Fabienne Lampert ◽  
Peter Hornung ◽  
Stefan Westermann
Keyword(s):  
Fluorescence Microscopy ◽  
Chromosome Segregation ◽  
Molecular Basis ◽  
Total Internal Reflection ◽  
Functional Relationship ◽  
Budding Yeast ◽  
Ndc80 Complex ◽  
Dam1 Complex ◽  
Necessary And Sufficient ◽  
Dynamic Microtubule

Kinetochores must remain associated with microtubule ends, as they undergo rapid transitions between growth and shrinkage. The molecular basis for this essential activity that ensures correct chromosome segregation is unclear. In this study, we have used reconstitution of dynamic microtubules and total internal reflection fluorescence microscopy to define the functional relationship between two important budding yeast kinetochore complexes. We find that the Dam1 complex is an autonomous plus end–tracking complex. The Ndc80 complex, despite being structurally related to the general tip tracker EB1, fails to recognize growing ends efficiently. Dam1 oligomers are necessary and sufficient to recruit Ndc80 to dynamic microtubule ends, where both complexes remain continuously associated. The interaction occurs specifically in the presence of microtubules and is subject to regulation by Ipl1 phosphorylation. These findings can explain how the force harvested by Dam1 is transmitted to the rest of the kinetochore via the Ndc80 complex.

In vivo analysis of the Saccharomyces cerevisiae centromere CDEIII sequence: requirements for mitotic chromosome segregation

1991 ◽  
Vol 11 (10) ◽  
pp. 5212-5221
Author(s):  
B Jehn ◽  
R Niedenthal ◽  
J H Hegemann
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Artificial Chromosome ◽  
Complete Information ◽  
Saturation Mutagenesis ◽  
Chromosome Loss ◽  
Single Mutation ◽  
Yeast Saccharomyces Cerevisiae ◽  
Double Base

In the yeast Saccharomyces cerevisiae, the complete information needed in cis to specify a fully functional mitotic and meiotic centromere is contained within 120 bp arranged in the three conserved centromeric (CEN) DNA elements CDEI, -II, and -III. The 25-bp CDEIII is most important for faithful chromosome segregation. We have constructed single- and double-base substitutions in all highly conserved residues and one nonconserved residue of this element and analyzed the mitotic in vivo function of the mutated CEN DNAs, using an artificial chromosome. The effects of the mutations on chromosome segregation vary between wild-type-like activity (chromosome loss rate of 4.8 x 10(-4)) and a complete loss of CEN function. Data obtained by saturation mutagenesis of the palindromic core sequence suggest asymmetric involvement of the palindromic half-sites in mitotic CEN function. The poor CEN activity of certain single mutations could be improved by introducing an additional single mutation. These second-site suppressors can be found at conserved and nonconserved positions in CDEIII. Our suppression data are discussed in the context of natural CDEIII sequence variations found in the CEN sequences of different yeast chromosomes.

scholarly journals Casein kinase I-like protein kinases encoded by YCK1 and YCK2 are required for yeast morphogenesis.

1993 ◽  
Vol 13 (5) ◽  
pp. 2870-2881 ◽  
Author(s):  
L C Robinson ◽  
M M Menold ◽  
S Garrett ◽  
M R Culbertson
Keyword(s):  
Protein Kinase ◽  
Eukaryotic Cell ◽  
Casein Kinase ◽  
Temperature Sensitive ◽  
Protein Kinase Activity ◽  
Restrictive Temperature ◽  
Loss Of Function ◽  
Casein Kinase I ◽  
Yeast Saccharomyces Cerevisiae

Casein kinase I is an acidotropic protein kinase class that is widely distributed among eukaryotic cell types. In the yeast Saccharomyces cerevisiae, the casein kinase I isoform encoded by the gene pair YCK1 and YCK2 is a 60- to 62-kDa membrane-associated form. The Yck proteins perform functions essential for growth and division; either alone supports growth, but loss of function of both is lethal. We report here that casein kinase I-like activity is associated with a soluble Yck2-beta-galactosidase fusion protein in vitro and that thermolabile protein kinase activity is exhibited by a protein encoded by fusion of a temperature-sensitive yck2 allele with lacZ. Cells carrying the yck2-2ts allele arrest at restrictive temperature with multiple, elongated buds containing multiple nuclei. This phenotype suggests that the essential functions of the Yck proteins include roles in bud morphogenesis, possibly in control of cell growth polarity, and in cytokinesis or cell separation. Further, a genetic relationship between the yck2ts allele and deletion of CDC55 indicates that the function of Yck phosphorylation may be related to that of protein phosphatase 2A activity.

Hybrid microhandling: a unified view of robotic handling and self-assembly

2008 ◽  
Vol 4 (1-2) ◽  
pp. 5-16 ◽  
Author(s):  
Veikko Sariola ◽  
Quan Zhou ◽  
Heikki N. Koivo
Keyword(s):  
Self Assembly ◽  
Unified View

scholarly journals Reduced Mad2 expression keeps relaxed kinetochores from arresting budding yeast in mitosis

2011 ◽  
Vol 22 (14) ◽  
pp. 2448-2457 ◽  
Author(s):  
Erin L. Barnhart ◽  
Russell K. Dorer ◽  
Andrew W. Murray ◽  
Scott C. Schuyler
Keyword(s):  
Chromosome Segregation ◽  
Budding Yeast ◽  
Spindle Checkpoint ◽  
Chromosome Loss ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Sister Chromatids ◽  
Chromatid Cohesion ◽  
Diploid Cells ◽  
Antimicrotubule Drugs

Chromosome segregation depends on the spindle checkpoint, which delays anaphase until all chromosomes have bound microtubules and have been placed under tension. The Mad1–Mad2 complex is an essential component of the checkpoint. We studied the consequences of removing one copy of MAD2 in diploid cells of the budding yeast, Saccharomyces cerevisiae. Compared to MAD2/MAD2 cells, MAD2/mad2Δ heterozygotes show increased chromosome loss and have different responses to two insults that activate the spindle checkpoint: MAD2/mad2Δ cells respond normally to antimicrotubule drugs but cannot respond to chromosomes that lack tension between sister chromatids. In MAD2/mad2Δ cells with normal sister chromatid cohesion, removing one copy of MAD1 restores the checkpoint and returns chromosome loss to wild-type levels. We conclude that cells need the normal Mad2:Mad1 ratio to respond to chromosomes that are not under tension.

scholarly journals The JNM1 gene in the yeast Saccharomyces cerevisiae is required for nuclear migration and spindle orientation during the mitotic cell cycle.

1994 ◽  
Vol 125 (1) ◽  
pp. 143-158 ◽  
Author(s):  
J N McMillan ◽  
K Tatchell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Low Temperature ◽  
Mother Cell ◽  
Significant Proportion ◽  
Nuclear Migration ◽  
Cold Sensitivity ◽  
Chain Gene ◽  
Chromosome Loss ◽  
Lacz Gene ◽  
Yeast Saccharomyces Cerevisiae

JNM1, a novel gene on chromosome XIII in the yeast Saccharomyces cerevisiae, is required for proper nuclear migration. jnm1 null mutants have a temperature-dependent defect in nuclear migration and an accompanying alteration in astral microtubules. At 30 degrees C, a significant proportion of the mitotic spindles is not properly located at the neck between the mother cell and the bud. This defect is more severe at low temperature. At 11 degrees C, 60% of the cells accumulate with large buds, most of which have two DAPI staining regions in the mother cell. Although mitosis is delayed and nuclear migration is defective in jnm1 mutant, we rarely observe more than two nuclei in a cell, nor do we frequently observe anuclear cells. No loss of viability is observed at 11 degrees C and cells continue to grow exponentially with increased doubling time. At low temperature the large budded cells of jnm1 mutants exhibit extremely long astral microtubules that often wind around the periphery of the cell. jnm1 mutants are not defective in chromosome segregation during mitosis, as assayed by the rate of chromosome loss, or nuclear migration during conjugation, as assayed by the rate of mating and cytoduction. The phenotype of a jnm1 mutant is strikingly similar to that for mutants in the dynein heavy chain gene (Eshel, D., L. A. Urrestarazu, S. Vissers, J.-C. Jauniaux, J. C. van Vliet-Reedijk, R. J. Plants, and I. R. Gibbons. 1993. Proc. Natl. Acad. Sci. USA. 90:11172-11176; Li, Y. Y., E. Yeh, T. Hays, and K. Bloom. 1993. Proc. Natl. Acad. Sci. USA. 90:10096-10100). The JNM1 gene product is predicted to encode a 44-kD protein containing three coiled coil domains. A JNM1:lacZ gene fusion is able to complement the cold sensitivity and microtubule phenotype of a jnm1 deletion strain. This hybrid protein localizes to a single spot in the cell, most often near the spindle pole body in unbudded cells and in the bud in large budded cells. Together these results point to a specific role for Jnm1p in spindle migration, possibly as a subunit or accessory protein for yeast dynein.

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 Author response: The Ndc80 complex bridges two Dam1 complex rings

2017 ◽  
Author(s):  
Jae ook Kim ◽  
Alex Zelter ◽  
Neil T Umbreit ◽  
Athena Bollozos ◽  
Michael Riffle ◽  
...  
Keyword(s):  
Author Response ◽  
Ndc80 Complex ◽  
Dam1 Complex

scholarly journals Astrin-SKAP complex reconstitution reveals its kinetochore interaction with microtubule-bound Ndc80

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
David M Kern ◽  
Julie K Monda ◽  
Kuan-Chung Su ◽  
Elizabeth M Wilson-Kubalek ◽  
Iain M Cheeseman
Keyword(s):  
Chromosome Segregation ◽  
Human Cells ◽  
Cross Linking ◽  
Microtubule Binding ◽  
Binding Domains ◽  
Anaphase Onset ◽  
Ndc80 Complex ◽  
Outstanding Question ◽  
Dynamic Microtubule

Chromosome segregation requires robust interactions between the macromolecular kinetochore structure and dynamic microtubule polymers. A key outstanding question is how kinetochore-microtubule attachments are modulated to ensure that bi-oriented attachments are selectively stabilized and maintained. The Astrin-SKAP complex localizes preferentially to properly bi-oriented sister kinetochores, representing the final outer kinetochore component recruited prior to anaphase onset. Here, we reconstitute the 4-subunit Astrin-SKAP complex, including a novel MYCBP subunit. Our work demonstrates that the Astrin-SKAP complex contains separable kinetochore localization and microtubule binding domains. In addition, through cross-linking analysis in human cells and biochemical reconstitution, we show that the Astrin-SKAP complex binds synergistically to microtubules with the Ndc80 complex to form an integrated interface. We propose a model in which the Astrin-SKAP complex acts together with the Ndc80 complex to stabilize correctly formed kinetochore-microtubule interactions.

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