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Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments

During mitosis, individual microtubules make attachments to chromosomes via a specialized protein complex called the kinetochore to faithfully segregate the chromosomes to daughter cells. Translocation of kinetochores on the lateral surface of the microtubule has been proposed to contribute to high...

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Autores principales: Torvi, Julia R, Wong, Jonathan, Serwas, Daniel, Moayed, Amir, Drubin, David G, Barnes, Georjana
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9259035/
https://www.ncbi.nlm.nih.gov/pubmed/35791811
http://dx.doi.org/10.7554/eLife.78450
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author Torvi, Julia R
Wong, Jonathan
Serwas, Daniel
Moayed, Amir
Drubin, David G
Barnes, Georjana
author_facet Torvi, Julia R
Wong, Jonathan
Serwas, Daniel
Moayed, Amir
Drubin, David G
Barnes, Georjana
author_sort Torvi, Julia R
collection PubMed
description During mitosis, individual microtubules make attachments to chromosomes via a specialized protein complex called the kinetochore to faithfully segregate the chromosomes to daughter cells. Translocation of kinetochores on the lateral surface of the microtubule has been proposed to contribute to high fidelity chromosome capture and alignment at the mitotic midzone, but has been difficult to observe in vivo because of spatial and temporal constraints. To overcome these barriers, we used total internal reflection fluorescence (TIRF) microscopy to track the interactions between microtubules, kinetochore proteins, and other microtubule-associated proteins in lysates from metaphase-arrested Saccharomyces cerevisiae. TIRF microscopy and cryo-correlative light microscopy and electron tomography indicated that we successfully reconstituted interactions between intact kinetochores and microtubules. These kinetochores translocate on the lateral microtubule surface toward the microtubule plus end and transition to end-on attachment, whereupon microtubule depolymerization commences. The directional kinetochore movement is dependent on the highly processive kinesin-8, Kip3. We propose that Kip3 facilitates stable kinetochore attachment to microtubule plus ends through its abilities to move the kinetochore laterally on the surface of the microtubule and to regulate microtubule plus end dynamics.
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spelling pubmed-92590352022-07-07 Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments Torvi, Julia R Wong, Jonathan Serwas, Daniel Moayed, Amir Drubin, David G Barnes, Georjana eLife Cell Biology During mitosis, individual microtubules make attachments to chromosomes via a specialized protein complex called the kinetochore to faithfully segregate the chromosomes to daughter cells. Translocation of kinetochores on the lateral surface of the microtubule has been proposed to contribute to high fidelity chromosome capture and alignment at the mitotic midzone, but has been difficult to observe in vivo because of spatial and temporal constraints. To overcome these barriers, we used total internal reflection fluorescence (TIRF) microscopy to track the interactions between microtubules, kinetochore proteins, and other microtubule-associated proteins in lysates from metaphase-arrested Saccharomyces cerevisiae. TIRF microscopy and cryo-correlative light microscopy and electron tomography indicated that we successfully reconstituted interactions between intact kinetochores and microtubules. These kinetochores translocate on the lateral microtubule surface toward the microtubule plus end and transition to end-on attachment, whereupon microtubule depolymerization commences. The directional kinetochore movement is dependent on the highly processive kinesin-8, Kip3. We propose that Kip3 facilitates stable kinetochore attachment to microtubule plus ends through its abilities to move the kinetochore laterally on the surface of the microtubule and to regulate microtubule plus end dynamics. eLife Sciences Publications, Ltd 2022-07-05 /pmc/articles/PMC9259035/ /pubmed/35791811 http://dx.doi.org/10.7554/eLife.78450 Text en © 2022, Torvi, Wong et al https://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Cell Biology
Torvi, Julia R
Wong, Jonathan
Serwas, Daniel
Moayed, Amir
Drubin, David G
Barnes, Georjana
Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
title Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
title_full Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
title_fullStr Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
title_full_unstemmed Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
title_short Reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
title_sort reconstitution of kinetochore motility and microtubule dynamics reveals a role for a kinesin-8 in establishing end-on attachments
topic Cell Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9259035/
https://www.ncbi.nlm.nih.gov/pubmed/35791811
http://dx.doi.org/10.7554/eLife.78450
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