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Augmin is a Ran-regulated spindle assembly factor

Mitotic spindles are composed of microtubules (MTs) that must nucleate at the right place and time. Ran regulates this process by directly controlling the release of spindle assembly factors (SAFs) from nucleocytoplasmic shuttle proteins importin-αβ and subsequently forms a biochemical gradient of S...

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Autores principales: Kraus, Jodi, Travis, Sophie M., King, Matthew R., Petry, Sabine
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Biochemistry and Molecular Biology 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10318467/
https://www.ncbi.nlm.nih.gov/pubmed/37086784
http://dx.doi.org/10.1016/j.jbc.2023.104736
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author Kraus, Jodi
Travis, Sophie M.
King, Matthew R.
Petry, Sabine
author_facet Kraus, Jodi
Travis, Sophie M.
King, Matthew R.
Petry, Sabine
author_sort Kraus, Jodi
collection PubMed
description Mitotic spindles are composed of microtubules (MTs) that must nucleate at the right place and time. Ran regulates this process by directly controlling the release of spindle assembly factors (SAFs) from nucleocytoplasmic shuttle proteins importin-αβ and subsequently forms a biochemical gradient of SAFs localized around chromosomes. The majority of spindle MTs are generated by branching MT nucleation, which has been shown to require an eight-subunit protein complex known as augmin. In Xenopus laevis, Ran can control branching through a canonical SAF, TPX2, which is nonessential in Drosophila melanogaster embryos and HeLa cells. Thus, how Ran regulates branching MT nucleation when TPX2 is not required remains unknown. Here, we use in vitro pulldowns and total internal reflection fluorescence microscopy to show that augmin is a Ran-regulated SAF. We demonstrate that augmin directly interacts with both importin-α and importin-β through two nuclear localization sequences on the Haus8 subunit, which overlap with the MT-binding site. Moreover, we show that Ran controls localization of augmin to MTs in both Xenopus egg extract and in vitro. Our results demonstrate that RanGTP directly regulates augmin, which establishes a new way by which Ran controls branching MT nucleation and spindle assembly both in the absence and presence of TPX2.
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spelling pubmed-103184672023-07-05 Augmin is a Ran-regulated spindle assembly factor Kraus, Jodi Travis, Sophie M. King, Matthew R. Petry, Sabine J Biol Chem Research Article Mitotic spindles are composed of microtubules (MTs) that must nucleate at the right place and time. Ran regulates this process by directly controlling the release of spindle assembly factors (SAFs) from nucleocytoplasmic shuttle proteins importin-αβ and subsequently forms a biochemical gradient of SAFs localized around chromosomes. The majority of spindle MTs are generated by branching MT nucleation, which has been shown to require an eight-subunit protein complex known as augmin. In Xenopus laevis, Ran can control branching through a canonical SAF, TPX2, which is nonessential in Drosophila melanogaster embryos and HeLa cells. Thus, how Ran regulates branching MT nucleation when TPX2 is not required remains unknown. Here, we use in vitro pulldowns and total internal reflection fluorescence microscopy to show that augmin is a Ran-regulated SAF. We demonstrate that augmin directly interacts with both importin-α and importin-β through two nuclear localization sequences on the Haus8 subunit, which overlap with the MT-binding site. Moreover, we show that Ran controls localization of augmin to MTs in both Xenopus egg extract and in vitro. Our results demonstrate that RanGTP directly regulates augmin, which establishes a new way by which Ran controls branching MT nucleation and spindle assembly both in the absence and presence of TPX2. American Society for Biochemistry and Molecular Biology 2023-04-21 /pmc/articles/PMC10318467/ /pubmed/37086784 http://dx.doi.org/10.1016/j.jbc.2023.104736 Text en © 2023 The Authors https://creativecommons.org/licenses/by/4.0/This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Research Article
Kraus, Jodi
Travis, Sophie M.
King, Matthew R.
Petry, Sabine
Augmin is a Ran-regulated spindle assembly factor
title Augmin is a Ran-regulated spindle assembly factor
title_full Augmin is a Ran-regulated spindle assembly factor
title_fullStr Augmin is a Ran-regulated spindle assembly factor
title_full_unstemmed Augmin is a Ran-regulated spindle assembly factor
title_short Augmin is a Ran-regulated spindle assembly factor
title_sort augmin is a ran-regulated spindle assembly factor
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10318467/
https://www.ncbi.nlm.nih.gov/pubmed/37086784
http://dx.doi.org/10.1016/j.jbc.2023.104736
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