Molecular basis of outer kinetochore assembly on CENP-T

Stable kinetochore-microtubule attachment is essential for cell division. It requires recruitment of outer kinetochore microtubule binders by centromere proteins C and T (CENP-C and CENP-T). To study the molecular requirements of kinetochore formation, we reconstituted the binding of the MIS12 and N...

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Autores principales: Huis in 't Veld, Pim J, Jeganathan, Sadasivam, Petrovic, Arsen, Singh, Priyanka, John, Juliane, Krenn, Veronica, Weissmann, Florian, Bange, Tanja, Musacchio, Andrea
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2016
Materias:
Biochemistry
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5241120/
https://www.ncbi.nlm.nih.gov/pubmed/28012276
http://dx.doi.org/10.7554/eLife.21007
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author Huis in 't Veld, Pim J
Jeganathan, Sadasivam
Petrovic, Arsen
Singh, Priyanka
John, Juliane
Krenn, Veronica
Weissmann, Florian
Bange, Tanja
Musacchio, Andrea
author_facet Huis in 't Veld, Pim J
Jeganathan, Sadasivam
Petrovic, Arsen
Singh, Priyanka
John, Juliane
Krenn, Veronica
Weissmann, Florian
Bange, Tanja
Musacchio, Andrea
author_sort Huis in 't Veld, Pim J
collection PubMed
description Stable kinetochore-microtubule attachment is essential for cell division. It requires recruitment of outer kinetochore microtubule binders by centromere proteins C and T (CENP-C and CENP-T). To study the molecular requirements of kinetochore formation, we reconstituted the binding of the MIS12 and NDC80 outer kinetochore subcomplexes to CENP-C and CENP-T. Whereas CENP-C recruits a single MIS12:NDC80 complex, we show here that CENP-T binds one MIS12:NDC80 and two NDC80 complexes upon phosphorylation by the mitotic CDK1:Cyclin B complex at three distinct CENP-T sites. Visualization of reconstituted complexes by electron microscopy supports this model. Binding of CENP-C and CENP-T to MIS12 is competitive, and therefore CENP-C and CENP-T act in parallel to recruit two MIS12 and up to four NDC80 complexes. Our observations provide a molecular explanation for the stoichiometry of kinetochore components and its cell cycle regulation, and highlight how outer kinetochore modules bridge distances of well over 100 nm. DOI: http://dx.doi.org/10.7554/eLife.21007.001
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spelling pubmed-52411202017-01-18 Molecular basis of outer kinetochore assembly on CENP-T Huis in 't Veld, Pim J Jeganathan, Sadasivam Petrovic, Arsen Singh, Priyanka John, Juliane Krenn, Veronica Weissmann, Florian Bange, Tanja Musacchio, Andrea eLife Biochemistry Stable kinetochore-microtubule attachment is essential for cell division. It requires recruitment of outer kinetochore microtubule binders by centromere proteins C and T (CENP-C and CENP-T). To study the molecular requirements of kinetochore formation, we reconstituted the binding of the MIS12 and NDC80 outer kinetochore subcomplexes to CENP-C and CENP-T. Whereas CENP-C recruits a single MIS12:NDC80 complex, we show here that CENP-T binds one MIS12:NDC80 and two NDC80 complexes upon phosphorylation by the mitotic CDK1:Cyclin B complex at three distinct CENP-T sites. Visualization of reconstituted complexes by electron microscopy supports this model. Binding of CENP-C and CENP-T to MIS12 is competitive, and therefore CENP-C and CENP-T act in parallel to recruit two MIS12 and up to four NDC80 complexes. Our observations provide a molecular explanation for the stoichiometry of kinetochore components and its cell cycle regulation, and highlight how outer kinetochore modules bridge distances of well over 100 nm. DOI: http://dx.doi.org/10.7554/eLife.21007.001 eLife Sciences Publications, Ltd 2016-12-24 /pmc/articles/PMC5241120/ /pubmed/28012276 http://dx.doi.org/10.7554/eLife.21007 Text en © 2016, Huis in 't Veld et al http://creativecommons.org/licenses/by/4.0/ This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Biochemistry
Huis in 't Veld, Pim J
Jeganathan, Sadasivam
Petrovic, Arsen
Singh, Priyanka
John, Juliane
Krenn, Veronica
Weissmann, Florian
Bange, Tanja
Musacchio, Andrea
Molecular basis of outer kinetochore assembly on CENP-T
title Molecular basis of outer kinetochore assembly on CENP-T
title_full Molecular basis of outer kinetochore assembly on CENP-T
title_fullStr Molecular basis of outer kinetochore assembly on CENP-T
title_full_unstemmed Molecular basis of outer kinetochore assembly on CENP-T
title_short Molecular basis of outer kinetochore assembly on CENP-T
title_sort molecular basis of outer kinetochore assembly on cenp-t
topic Biochemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5241120/
https://www.ncbi.nlm.nih.gov/pubmed/28012276
http://dx.doi.org/10.7554/eLife.21007
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