RMD-1, a novel microtubule-associated protein, functions in chromosome segregation in Caenorhabditis elegans

For proper chromosome segregation, the sister kinetochores must attach to microtubules extending from the opposite spindle poles. Any errors in microtubule attachment can induce aneuploidy. In this study, we identify a novel conserved Caenorhabditis elegans microtubule-associated protein, regulator of microtubule dynamics 1 (RMD-1), that localizes to spindle microtubules and spindle poles. Depletion of RMD-1 induces severe defects in chromosome segregation, probably through merotelic attachments between microtubules and chromosomes. Although rmd-1 embryos also have a mild defect in microtubule growth, we find that mutants of the microtubule growth regulator XMAP215/ZYG-9 show much weaker segregation defects. This suggests that the microtubule growth defect in rmd-1 embryos does not cause abnormal chromosome segregation. We also see that RMD-1 interacts with aurora B in vitro. Our results suggest that RMD-1 functions in chromosome segregation in C. elegans embryos, possibly through the aurora B–mediated pathway. Human homologues of RMD-1 could also bind microtubules, which would suggest a function for these proteins in chromosome segregation during mitosis in other organisms as well.

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CSC-1

The Journal of Cell Biology ◽

10.1083/jcb.200207117 ◽

2003 ◽

Vol 161 (2) ◽

pp. 229-236 ◽

Cited By ~ 67

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.

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The aurora kinase AIR-2 functions in the release of chromosome cohesion in Caenorhabditis elegans meiosis

The Journal of Cell Biology ◽

10.1083/jcb.200110045 ◽

2002 ◽

Vol 157 (2) ◽

pp. 219-229 ◽

Cited By ~ 137

Author(s):

Eric Rogers ◽

John D. Bishop ◽

James A. Waddle ◽

Jill M. Schumacher ◽

Rueyling Lin

Keyword(s):

Caenorhabditis Elegans ◽

Chromosome Segregation ◽

Aurora Kinase ◽

Aurora B ◽

Localization Pattern ◽

Meiotic Chromosomes ◽

Sister Chromatids ◽

Chromosome Separation ◽

Major Amino Acid

Accurate chromosome segregation during cell division requires not only the establishment, but also the precise, regulated release of chromosome cohesion. Chromosome dynamics during meiosis are more complicated, because homologues separate at anaphase I whereas sister chromatids remain attached until anaphase II. How the selective release of chromosome cohesion is regulated during meiosis remains unclear. We show that the aurora-B kinase AIR-2 regulates the selective release of chromosome cohesion during Caenorhabditis elegans meiosis. AIR-2 localizes to subchromosomal regions corresponding to last points of contact between homologues in metaphase I and between sister chromatids in metaphase II. Depletion of AIR-2 by RNA interference (RNAi) prevents chromosome separation at both anaphases, with concomitant prevention of meiotic cohesin REC-8 release from meiotic chromosomes. We show that AIR-2 phosphorylates REC-8 at a major amino acid in vitro. Interestingly, depletion of two PP1 phosphatases, CeGLC-7α and CeGLC-7β, abolishes the restricted localization pattern of AIR-2. In Ceglc-7α/β(RNAi) embryos, AIR-2 is detected on the entire bivalent. Concurrently, chromosomal REC-8 is dramatically reduced and sister chromatids are separated precociously at anaphase I in Ceglc-7α/β(RNAi) embryos. We propose that AIR-2 promotes the release of chromosome cohesion via phosphorylation of REC-8 at specific chromosomal locations and that CeGLC-7α/β, directly or indirectly, antagonize AIR-2 activity.

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chTOG is a conserved mitotic error correction factor

10.1101/2020.08.03.235325 ◽

2020 ◽

Author(s):

Jacob A. Herman ◽

Matthew P. Miller ◽

Sue Biggins

Keyword(s):

Protein Kinase ◽

Error Correction ◽

Chromosome Segregation ◽

Correction Factor ◽

Budding Yeast ◽

Aurora B ◽

Spindle Poles ◽

Low Tension ◽

Intrinsic Error

AbstractAccurate chromosome segregation requires kinetochores on duplicated chromatids to biorient by attaching to dynamic microtubules from opposite spindle poles, which exerts forces to bring kinetochores under tension. However, kinetochores initially bind to MTs indiscriminately, resulting in errors that must be corrected. While the Aurora B protein kinase destabilizes low-tension attachments by phosphorylating kinetochores, low-tension attachments are intrinsically less stable than those under higher tension in vitro independent of Aurora activity. Intrinsic tensionsensitive behavior requires the microtubule regulator Stu2 (budding yeast Dis1/XMAP215 ortholog), which we demonstrate here is likely a conserved function for the TOG protein family. The human TOG protein, chTOG, localizes to kinetochores independent of microtubules by interacting with Hec1. We identify a chTOG mutant that regulates microtubule dynamics but accumulates erroneous kinetochore-microtubule attachments that Aurora B fails to destabilize. Thus, TOG proteins confer a unique, intrinsic error correction activity to kinetochores that ensures accurate chromosome segregation.

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

10.1101/455873 ◽

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.

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Tension on kinetochore substrates is insufficient to prevent Aurora-triggered detachment

10.1101/415992 ◽

2018 ◽

Cited By ~ 3

Author(s):

Anna K. de Regt ◽

Charles L. Asbury ◽

Sue Biggins

Keyword(s):

Chromosome Segregation ◽

Direct Evidence ◽

Underlying Mechanism ◽

Aurora B ◽

Trial And Error ◽

Spindle Poles ◽

Pulling Forces ◽

Low Tension

Introduction / AbstractChromosome segregation requires large macromolecular structures called kinetochores to attach dynamic microtubules from opposite spindle poles1,2. Attachments are made iteratively, through a trial-and-error process, and proper attachments come under tension from the pulling forces of microtubules3,4. However, if sister kinetochores bind microtubules from the same pole1,2, these defective attachments lack tension and must be destabilized to give another chance for proper attachments to form. This vital error correction process requires Aurora B kinase, which phosphorylates kinetochores lacking tension to reduce their affinity for microtubules5-11. An unresolved question is how Aurora B distinguishes the level of tension on kinetochores. There are conflicting reports on the underlying mechanism12-16, owing in part to the difficulties of manipulating kinetochore tension in vivo and distinguishing kinase from opposing phosphatase activity. To address these issues, we have reconstituted Aurora B-triggered kinetochore detachment in an in vitro optical trapping-based flow assay. Here, we test an outstanding model by determining whether kinetochore tension is sufficient to prevent kinase-triggered detachments. Strikingly, Aurora B detaches kinetochores from microtubules under both high and low tension, providing direct evidence that the kinase does not distinguish correct versus incorrect attachments by recognizing tension-dependent changes in the conformation of its kinetochore substrates.

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chTOG is a conserved mitotic error correction factor

eLife ◽

10.7554/elife.61773 ◽

2020 ◽

Vol 9 ◽

Author(s):

Jacob A Herman ◽

Matthew P Miller ◽

Sue Biggins

Keyword(s):

Protein Kinase ◽

Error Correction ◽

Chromosome Segregation ◽

Correction Factor ◽

Aurora B ◽

Spindle Poles ◽

Low Tension ◽

Intrinsic Error ◽

Sensitive Behavior

Accurate chromosome segregation requires kinetochores on duplicated chromatids to biorient by attaching to dynamic microtubules from opposite spindle poles, which exerts forces to bring kinetochores under tension. However, kinetochores initially bind to microtubules indiscriminately, resulting in errors that must be corrected. While the Aurora B protein kinase destabilizes low-tension attachments by phosphorylating kinetochores, low-tension attachments are intrinsically less stable than those under higher tension in vitro independent of Aurora activity. Intrinsic tension-sensitive behavior requires the microtubule regulator Stu2 (budding yeast Dis1/XMAP215 ortholog), which we demonstrate here is likely a conserved function for the TOG protein family. The human TOG protein, chTOG, localizes to kinetochores independent of microtubules by interacting with Hec1. We identify a chTOG mutant that regulates microtubule dynamics but accumulates erroneous kinetochore-microtubule attachments that are not destabilized by Aurora B. Thus, TOG proteins confer a unique, intrinsic error correction activity to kinetochores that ensures accurate chromosome segregation.

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Bod1, a novel kinetochore protein required for chromosome biorientation

The Journal of Cell Biology ◽

10.1083/jcb.200704098 ◽

2007 ◽

Vol 179 (2) ◽

pp. 187-197 ◽

Cited By ~ 41

Author(s):

Iain M. Porter ◽

Sarah E. McClelland ◽

Guennadi A. Khoudoli ◽

Christopher J. Hunter ◽

Jens S. Andersen ◽

...

Keyword(s):

Chromosome Segregation ◽

Hela Cells ◽

Aurora B ◽

Macromolecular Complex ◽

Mitotic Spindles ◽

Kinetochore Protein ◽

Spindle Poles ◽

Interfering Rna ◽

Rna Depletion

We have combined the proteomic analysis of Xenopus laevis in vitro–assembled chromosomes with RNA interference and live cell imaging in HeLa cells to identify novel factors required for proper chromosome segregation. The first of these is Bod1, a protein conserved throughout metazoans that associates with a large macromolecular complex and localizes with kinetochores and spindle poles during mitosis. Small interfering RNA depletion of Bod1 in HeLa cells produces elongated mitotic spindles with severe biorientation defects. Bod1-depleted cells form syntelic attachments that can oscillate and generate enough force to separate sister kinetochores, suggesting that microtubule–kinetochore interactions were intact. Releasing Bod1-depleted cells from a monastrol block increases the frequency of syntelic attachments and the number of cells displaying biorientation defects. Bod1 depletion does not affect the activity or localization of Aurora B but does cause mislocalization of the microtubule depolymerase mitotic centromere- associated kinesin and prevents its efficient phosphorylation by Aurora B. Therefore, Bod1 is a novel kinetochore protein that is required for the detection or resolution of syntelic attachments in mitotic spindles.

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Streblus asper Lour. exerts MAPK and SKN-1 mediated anti-aging, anti-photoaging activities and imparts neuroprotection by ameliorating Aβ in Caenorhabditis elegans

Nutrition and Healthy Aging ◽

10.3233/nha-210121 ◽

2021 ◽

pp. 1-17

Author(s):

Mani Iyer Prasanth ◽

James Michael Brimson ◽

Dicson Sheeja Malar ◽

Anchalee Prasansuklab ◽

Tewin Tencomnao

Keyword(s):

Oxidative Stress ◽

Caenorhabditis Elegans ◽

Stress Resistance ◽

Vital Role ◽

Transgenic Strain ◽

Wild Type ◽

Neuroprotective Effects ◽

C Elegans ◽

Streblus Asper

BACKGROUND: Streblus asper Lour., has been reported to have anti-aging and neuroprotective efficacies in vitro. OBJECTIVE: To analyze the anti-aging, anti-photoaging and neuroprotective efficacies of S. asper in Caenorhabditis elegans. METHODS: C. elegans (wild type and gene specific mutants) were treated with S. asper extract and analyzed for lifespan and other health benefits through physiological assays, fluorescence microscopy, qPCR and Western blot. RESULTS: The plant extract was found to increase the lifespan, reduce the accumulation of lipofuscin and modulate the expression of candidate genes. It could extend the lifespan of both daf-16 and daf-2 mutants whereas the pmk-1 mutant showed no effect. The activation of skn-1 was observed in skn-1::GFP transgenic strain and in qPCR expression. Further, the extract can extend the lifespan of UV-A exposed nematodes along with reducing ROS levels. Additionally, the extract also extends lifespan and reduces paralysis in Aβ transgenic strain, apart from reducing Aβ expression. CONCLUSIONS: S. asper was able to extend the lifespan and healthspan of C. elegans which was independent of DAF-16 pathway but dependent on SKN-1 and MAPK which could play a vital role in eliciting the anti-aging, anti-photoaging and neuroprotective effects, as the extract could impart oxidative stress resistance and neuroprotection.

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Caenorhabditis elegans as an in vivo Model to Assess Amphetamine Tolerance

Brain Behavior and Evolution ◽

10.1159/000514858 ◽

2021 ◽

pp. 1-9

Author(s):

Dayana Torres Valladares ◽

Sirisha Kudumala ◽

Murad Hossain ◽

Lucia Carvelli

Keyword(s):

Caenorhabditis Elegans ◽

Molecular Mechanisms ◽

Drugs Of Abuse ◽

Genetic Model ◽

In Vivo Model ◽

C Elegans ◽

Hyperactivity Disorder ◽

In Vitro Data

Amphetamine is a potent psychostimulant also used to treat attention deficit/hyperactivity disorder and narcolepsy. In vivo and in vitro data have demonstrated that amphetamine increases the amount of extra synaptic dopamine by both inhibiting reuptake and promoting efflux of dopamine through the dopamine transporter. Previous studies have shown that chronic use of amphetamine causes tolerance to the drug. Thus, since the molecular mechanisms underlying tolerance to amphetamine are still unknown, an animal model to identify the neurochemical mechanisms associated with drug tolerance is greatly needed. Here we took advantage of a unique behavior caused by amphetamine in <i>Caenorhabditis elegans</i> to investigate whether this simple, but powerful, genetic model develops tolerance following repeated exposure to amphetamine. We found that at least 3 treatments with 0.5 mM amphetamine were necessary to see a reduction in the amphetamine-induced behavior and, thus, to promote tolerance. Moreover, we found that, after intervals of 60/90 minutes between treatments, animals were more likely to exhibit tolerance than animals that underwent 10-minute intervals between treatments. Taken together, our results show that <i>C. elegans</i> is a suitable system to study tolerance to drugs of abuse such as amphetamines.

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Microtubules and microtubule-associated proteins from the nematode Caenorhabditis elegans: periodic cross-links connect microtubules in vitro.

The Journal of Cell Biology ◽

10.1083/jcb.103.1.23 ◽

1986 ◽

Vol 103 (1) ◽

pp. 23-31 ◽

Cited By ~ 35

Author(s):

E J Aamodt ◽

J G Culotti

Keyword(s):

Caenorhabditis Elegans ◽

Apparent Molecular Weight ◽

Cross Link ◽

Nematode Caenorhabditis Elegans ◽

Microtubule Associated Proteins ◽

C Elegans ◽

Associated Proteins ◽

Cross Links ◽

The Cross

The nematode Caenorhabditis elegans should be an excellent model system in which to study the role of microtubules in mitosis, embryogenesis, morphogenesis, and nerve function. It may be studied by the use of biochemical, genetic, molecular biological, and cell biological approaches. We have purified microtubules and microtubule-associated proteins (MAPs) from C. elegans by the use of the anti-tumor drug taxol (Vallee, R. B., 1982, J. Cell Biol., 92:435-44). Approximately 0.2 mg of microtubules and 0.03 mg of MAPs were isolated from each gram of C. elegans. The C. elegans microtubules were smaller in diameter than bovine microtubules assembled in vitro in the same buffer. They contained primarily 9-11 protofilaments, while the bovine microtubules contained 13 protofilaments. The principal MAP had an apparent molecular weight of 32,000 and the minor MAPs were 30,000, 45,000, 47,000, 50,000, 57,000, and 100,000-110,000 mol wt as determined by SDS-gel electrophoresis. The microtubules were observed, by electron microscopy of negatively stained preparations, to be connected by stretches of highly periodic cross-links. The cross-links connected the adjacent protofilaments of aligned microtubules, and occurred at a frequency of one cross-link every 7.7 +/- 0.9 nm, or one cross-link per tubulin dimer along the protofilament. The cross-links were removed when the MAPs were extracted from the microtubules with 0.4 M NaCl. The cross-links then re-formed when the microtubules and the MAPs were recombined in a low salt buffer. These results strongly suggest that the cross-links are composed of MAPs.

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