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ACKNOWLEDGMENTS
We thank the members of the Hoelz laboratory, M. Guttman, and
S. Shan for insightful discussions; A. Cohen, J. Chou, R. Gulati,
Y. Jeon, H. Margolis, E. Stuwe, T. Stuwe, and J. Thai for experimental
support; M. Beck for sharing the ~12-Å cryo-ET reconstruction
of the intact human NPC prior to publication; and V. Doye, U. Kutay,
E. Hurt, and I. Mattaj for providing material. We acknowledge
J. Kaiser and the scientific staff of the Stanford Synchrotron Radiation
Laboratory (SSRL) Beamline 12-2 and the National Institute of
General Medical Sciences and National Cancer Institute Structural
Biology Facility (GM/CA) at the Advanced Photon Source (APS)
for their support with x-ray diffraction measurements.Funding:
The Molecular Observatory at the California Institute of Technology
(Caltech) is supported by Donald and Judith Voet, the Gordon
and Betty Moore Foundation, and the Beckman Institute. The
operations at the SSRL and APS are supported by the US
Department of Energy and the National Institutes of Health (NIH).
GM/CA has been funded in whole or in part with federal funds from
the National Cancer Institute (ACB-12002) and the National
Institute of General Medical Sciences (AGM-12006). S.P. and
F.M.H. were supported by a PhD fellowship of the Boehringer
Ingelheim Fonds. D.H.L. was supported by an NIH Research Service
Award (5 T32 GM07616) and Amgen Graduate Fellowship
through the Caltech-Amgen Research Collaboration. S.C., S.G.R.,
and M.D. were supported by National Institute of Child Health andHuman Development Division of Intramural Research funding
ZIAHD001902 and ZIAHD008954. A.A.K. was supported by NIH
grants R01-GM117372 and P50-GM082545. A.H. was supported by
a Camille-Dreyfus Teacher Scholar Award (TC-15-082) and NIH
grants R01-GM117360 and R01-GM111461 and is an Investigator of
the Heritage Medical Research Institute (HMRI-15-09-01) and a
Faculty Scholar of the Howard Hughes Medical Institute (55108534).
Author contributions:A.H. conceived and coordinated the
study. C.J.B., S.N., G.W.M., S.P., A.T.G., X.L., S.M., S.H., F.M.H.,
D.H.L., B.B., A.W.T., E.J.R., A.R.C., S.C., S.G.R., T.A.S., C.A.J., M.D.,
A.P., A.F.P., A.A.K., and A.H. designed the research. C.J.B., S.N.,
G.W.M., S.P., A.T.G., X.L., S.M., S.H., F.M.H., D.H.L., B.B., A.W.T.,
E.J.R., A.R.C., S.C., S.G.R., T.A.S., C.A.J., and A.H. performed
the research. C.J.B., S.N., G.W.M., S.P., A.T.G., X.L., S.M., S.H.,
F.M.H., D.H.L., B.B., A.W.T., E.J.R., A.R.C., S.C., S.G.R., T.A.S.,
C.A.J., M.D., A.P., A.F.P., A.A.K., and A.H. analyzed the data.
S.M. and A.A.K. (synthetic antibodies) and S.C., S.G.R., and M.D.
(auxin-degron cell lines) provided new reagents. A.P. and A.F.P.
provided reagents and experimental guidance. C.J.B., S.N., G.W.M.,
S.P., A.T.G., X.L., A.P., and A.H. integrated and conceptualized
the results. C.J.B., S.N., G.W.M., S.P., A.P., and A.H. wrote and
revised the manuscript, with contributions from all authors.
Competing interests:The authors declare no conflicts of interest.
Data and materials availability:Materials generated in this
study are available on request from the corresponding author. The
auxin-inducible AID::NUP358 HCT116, AID::NUP358 DLD1, and
NUP160::NG AID DLD1 degron cell lines are subject to a materials
transfer agreement, which is available upon request. The
coordinates and structure factors have been deposited in the
Protein Data Bank (PDB) with accession codes 7MNJ (NUP358145-673),
7MNK (NUP358OE), 7MNI (NUP88NTD•NUP98APD), 7MNL
(NUP358NTD•sAB-14), 7MNM (NUP358NTDT585M•sAB-14), 7MNN
(NUP358NTDT653I•sAB-14), 7MNO (NUP358NTDI656V•sAB-14),
7MNP (NUP358ZnF2•Ran(GDP)), 7MNQ (NUP358ZnF2•Ran(GDP)),
7MNR (NUP358ZnF3•Ran(GDP)), 7MNS (NUP358ZnF4•Ran(GDP)),
7MNT (NUP358ZnF5/6•Ran(GDP)), 7MNU (NUP358ZnF7•Ran(GDP)),
7MNV (NUP358ZnF8•Ran(GDP)), 7MNW (NUP358RanBD-I•Ran
(GMPPNP)), 7MNX (Nup358RanBD-II•Ran(GMPPNP)), 7MNY
(NUP358RanBD-III•Ran(GMPPNP)), 7MNZ (NUP358RanBD-IV•
Ran(GMPPNP)), 7MO0 (NUP50RanBD•Ran(GMPPNP)), 7MO1
(NUP153ZnF1•Ran(GDP)), 7MO2 (NUP153ZnF2•Ran(GDP)), 7MO3
(NUP153ZnF3•Ran(GDP), 2.05 Å), 7MO4 (NUP153ZnF3•Ran(GDP),
2.4 Å), and 7MO5 (NUP153ZnF4•Ran(GDP)). PyMol and Chimera
sessions containing the composite structures of the constricted
and dilated human NPCs can be obtained from our website (http://
ahweb.caltech.edu), and coordinates are deposited in the PDB
with accession numbers 7TBL and 7TBM, respectively.
Quantitative docking data, workflow code, and PyMol and
Chimera sessions have been deposited in the CaltechDATA
repository ( 101 ). License information:Copyright © 2022 the
authors, some rights reserved; exclusive licensee American
Association for the Advancement of Science. No claim to original
US government works.https://www.science.org/about/science-
licenses-journal-article-reuseSUPPLEMENTARY MATERIALS
science.org/doi/10.1126/science.abm9129
Materials and Methods
Supplementary Text
Figs. S1 to S89
Tables S1 to S18
References ( 102 – 153 )
MDAR Reproducibility ChecklistView/request a protocol for this paper fromBio-protocol.Submitted 22 October 2021; accepted 15 April 2022
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RESEARCH | STRUCTURE OF THE NUCLEAR PORE