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3D super-resolution fluorescence microscopy maps the variable molecular architecture of the Nuclear Pore Complex

View ORCID ProfileVilma Jimenez Sabinina, View ORCID ProfileM. Julius Hossain, View ORCID ProfileJean-Karim Hériché, View ORCID ProfilePhilipp Hoess, View ORCID ProfileBianca Nijmeijer, View ORCID ProfileShyamal Mosalaganti, View ORCID ProfileMoritz Kueblbeck, Andrea Callegari, View ORCID ProfileAnna Szymborska, View ORCID ProfileMartin Beck, View ORCID ProfileJonas Ries, View ORCID ProfileJan Ellenberg
doi: https://doi.org/10.1101/2020.11.27.386599
Vilma Jimenez Sabinina
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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M. Julius Hossain
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Jean-Karim Hériché
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Philipp Hoess
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
2Collaboration for joint PhD degree between EMBL and Heidelberg University, Faculty of Biosciences
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Bianca Nijmeijer
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Shyamal Mosalaganti
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Moritz Kueblbeck
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Andrea Callegari
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Anna Szymborska
3Max Delbrück Center for Molecular Medicine, Berlin, Germany
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Martin Beck
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
4Max Planck Institute of Biophysics, Frankfurt am Main, Germany
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Jonas Ries
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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Jan Ellenberg
1Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany
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  • For correspondence: jan.ellenberg@embl.de
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Abstract

Nuclear pore complexes (NPCs) are large macromolecular machines that mediate the traffic between the nucleus and the cytoplasm. In vertebrates, each NPC consists of ~1000 proteins, termed nucleoporins, and has a mass of over 100 MDa. While a pseudo-atomic static model of the central scaffold of the NPC has recently been assembled by integrating data from isolated proteins and complexes, many structural components still remain elusive due to the enormous size and flexibility of the NPC. Here, we explored the power of 3D super-resolution microscopy combined with computational classification and averaging to explore the 3D structure of the NPC in single human cells. We show that this approach can build the first integrated 3D structural map containing both central as well as peripheral NPC subunits with molecular specificity and nanoscale resolution. Our unbiased classification of over ten thousand individual NPCs indicates that the nuclear ring and the nuclear basket can adopt different conformations. Our approach opens up the exciting possibility to relate different structural states of the NPC to function in situ.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

  • ↵# Department of Obstetrics and Gynaecology, Faculty of Medical and Health Sciences, The University of Auckland, Auckland, New Zealand

  • ↵§ Life Science Institute, University of Michigan, Ann Arbor, MI, USA

  • In summary, we have included new figures and added more detail to the manuscript in order to assist the reader in navigating the data and make the clustering approach easier to understand. For example, we have added a new schematic figure to illustrate the position of all analyzed Nups and the particle averaging workflow (new Figure 1A) and have provided a confusion matrix to better support the performance of our clustering approach (new Figure 4B).

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted April 16, 2021.
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3D super-resolution fluorescence microscopy maps the variable molecular architecture of the Nuclear Pore Complex
Vilma Jimenez Sabinina, M. Julius Hossain, Jean-Karim Hériché, Philipp Hoess, Bianca Nijmeijer, Shyamal Mosalaganti, Moritz Kueblbeck, Andrea Callegari, Anna Szymborska, Martin Beck, Jonas Ries, Jan Ellenberg
bioRxiv 2020.11.27.386599; doi: https://doi.org/10.1101/2020.11.27.386599
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3D super-resolution fluorescence microscopy maps the variable molecular architecture of the Nuclear Pore Complex
Vilma Jimenez Sabinina, M. Julius Hossain, Jean-Karim Hériché, Philipp Hoess, Bianca Nijmeijer, Shyamal Mosalaganti, Moritz Kueblbeck, Andrea Callegari, Anna Szymborska, Martin Beck, Jonas Ries, Jan Ellenberg
bioRxiv 2020.11.27.386599; doi: https://doi.org/10.1101/2020.11.27.386599

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