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A quantitative map of nuclear pore assembly reveals two distinct mechanisms

View ORCID ProfileShotaro Otsuka, View ORCID ProfileJeremy O. B. Tempkin, View ORCID ProfileWanlu Zhang, View ORCID ProfileAntonio Z. Politi, View ORCID ProfileArina Rybina, View ORCID ProfileM. Julius Hossain, View ORCID ProfileMoritz Kueblbeck, Andrea Callegari, View ORCID ProfileBirgit Koch, View ORCID ProfileAndrej Sali, View ORCID ProfileJan Ellenberg
doi: https://doi.org/10.1101/2021.05.17.444137
Shotaro Otsuka
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
3Max Perutz Labs, a joint venture of the University of Vienna and the Medical University of Vienna, Vienna Biocenter (VBC), Dr. Bohr Gasse 9, 1030 Vienna, Austria
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  • For correspondence: shotaro.otsuka@univie.ac.at jan.ellenberg@embl.de
Jeremy O. B. Tempkin
2Department of Bioengineering and Therapeutic Sciences, Department of Pharmaceutical Chemistry, Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, CA 94143, USA
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Wanlu Zhang
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
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Antonio Z. Politi
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
4Max Planck Institute for Multidisciplinary Sciences, Göttingen, Germany
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Arina Rybina
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
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M. Julius Hossain
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
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Moritz Kueblbeck
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
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Andrea Callegari
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
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Birgit Koch
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
5Max Planck Institute for Medical Research, Heidelberg, Germany
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Andrej Sali
2Department of Bioengineering and Therapeutic Sciences, Department of Pharmaceutical Chemistry, Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, CA 94143, USA
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Jan Ellenberg
1Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany
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  • ORCID record for Jan Ellenberg
  • For correspondence: shotaro.otsuka@univie.ac.at jan.ellenberg@embl.de
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Summary

Understanding how the nuclear pore complex (NPC) assembles is of fundamental importance to grasp the mechanisms behind its essential function and understand its role during evolution of eukaryotes1–4. While we know that at least two NPC assembly pathways exist, one during exit from mitosis and one during nuclear growth in interphase, we currently lack a quantitative map of their molecular events. Here, we use fluorescence correlation spectroscopy (FCS) calibrated live imaging of endogenously fluorescently-tagged nucleoporins to map the changes in composition and stoichiometry of seven major modules of the human NPC during its assembly in single dividing cells. This systematic quantitative map reveals that the two assembly pathways employ strikingly different molecular mechanisms, inverting the order of addition of two large structural components, the central ring complex and nuclear filaments. Our dynamic stoichiometry data allows us to perform the first computational simulation that predicts the structure of postmitotic NPC assembly intermediates.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

  • Results strengthened by new experiments with 3D-STED (new Fig. 4) and live imaging of a newly-generated Nup cell lines (Extended Figs. 7, 8). Variance of integrative models highlighted more clearly (Extended Fig. 6). Clarified points in the discussion. Author list updated.

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 May 11, 2022.
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A quantitative map of nuclear pore assembly reveals two distinct mechanisms
Shotaro Otsuka, Jeremy O. B. Tempkin, Wanlu Zhang, Antonio Z. Politi, Arina Rybina, M. Julius Hossain, Moritz Kueblbeck, Andrea Callegari, Birgit Koch, Andrej Sali, Jan Ellenberg
bioRxiv 2021.05.17.444137; doi: https://doi.org/10.1101/2021.05.17.444137
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A quantitative map of nuclear pore assembly reveals two distinct mechanisms
Shotaro Otsuka, Jeremy O. B. Tempkin, Wanlu Zhang, Antonio Z. Politi, Arina Rybina, M. Julius Hossain, Moritz Kueblbeck, Andrea Callegari, Birgit Koch, Andrej Sali, Jan Ellenberg
bioRxiv 2021.05.17.444137; doi: https://doi.org/10.1101/2021.05.17.444137

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