Visualizing nuclear pore complex assembly in situ in human cells at nanometer resolution by correlating live imaging with electron microscopy
Visualizing nuclear pore complex assembly in situ in human cells at nanometer resolution by correlating live imaging with electron microscopy
In eukaryotic cells that undergo open mitosis, nuclear pore complex assembly proceeds via two distinct pathways: postmitotic and interphase assembly. Studying both assembly processes is challenging because postmitotic assembly is fast, interphase assembly is rare and sporadic, and assembly intermediates in both pathways are very small with a diameter below 100 nm. Here, we present a protocol for studying nuclear pore complex biogenesis in situ in cultured human cells in a spatiotemporally resolved and quantitative manner by combining live imaging with three-dimensional electron microscopy. The method described here can also be applied for studying other cell cycle–associated events with high spatiotemporal resolution.
Cell cycle, Correlative light-electron microscopy, Electron tomography, High-pressure freezing, Live-cell imaging, Mitosis, Nuclear envelope
493-512
Bragulat-Teixidor, Helena
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Hossain, M. Julius
bba1b875-7604-462b-a55b-ba0b54f728e8
Otsuka, Shotaro
f379d77f-b076-43cf-9359-b177f5a2015a
13 April 2022
Bragulat-Teixidor, Helena
a2043846-d66e-4b37-8734-67defa7ccf59
Hossain, M. Julius
bba1b875-7604-462b-a55b-ba0b54f728e8
Otsuka, Shotaro
f379d77f-b076-43cf-9359-b177f5a2015a
Bragulat-Teixidor, Helena, Hossain, M. Julius and Otsuka, Shotaro
(2022)
Visualizing nuclear pore complex assembly in situ in human cells at nanometer resolution by correlating live imaging with electron microscopy.
In,
Methods in Molecular Biology.
(Methods in Molecular Biology, 2502)
New York, USA.
Springer, .
(doi:10.1007/978-1-0716-2337-4_31).
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Book Section
Abstract
In eukaryotic cells that undergo open mitosis, nuclear pore complex assembly proceeds via two distinct pathways: postmitotic and interphase assembly. Studying both assembly processes is challenging because postmitotic assembly is fast, interphase assembly is rare and sporadic, and assembly intermediates in both pathways are very small with a diameter below 100 nm. Here, we present a protocol for studying nuclear pore complex biogenesis in situ in cultured human cells in a spatiotemporally resolved and quantitative manner by combining live imaging with three-dimensional electron microscopy. The method described here can also be applied for studying other cell cycle–associated events with high spatiotemporal resolution.
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Published date: 13 April 2022
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Funding Information:
We thank the Electron Microscopy Core Facility in the European Molecular Biology Laboratory (especially Rachel Santarella-Mellwig and Martin Schorb) for technical and computational support, and the members of the Jan Ellenberg and Martin Beck groups, Anna M Steyer, and Yannick Schwab for advice and discussion.
Publisher Copyright:
© 2022, Springer Science+Business Media, LLC, part of Springer Nature.
Keywords:
Cell cycle, Correlative light-electron microscopy, Electron tomography, High-pressure freezing, Live-cell imaging, Mitosis, Nuclear envelope
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Local EPrints ID: 458133
URI: http://eprints.soton.ac.uk/id/eprint/458133
ISSN: 1064-3745
PURE UUID: f4da6025-171c-4a63-9fa6-b07a2dbfbd2b
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Date deposited: 29 Jun 2022 16:35
Last modified: 18 Mar 2024 04:05
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Author:
Helena Bragulat-Teixidor
Author:
M. Julius Hossain
Author:
Shotaro Otsuka
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