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).
