Architecture of the fungal nuclear pore inner ring complex
The nuclear pore complex (NPC) constitutes the sole gateway for bidirectional nucleocytoplasmic transport. We present the reconstitution and interdisciplinary analyses of the ~425-kDa inner ring complex (IRC), which forms the central transport channel and diffusion barrier of the NPC, revealing its interaction network and equimolar stoichiometry. The Nsp1•Nup49•Nup57 channel nucleoporin hetero-trimer (CNT) attaches to the IRC solely through the adaptor nucleoporin Nic96. The CNT•Nic96 structure reveals that Nic96 functions as an assembly sensor that recognizes the three dimensional architecture of the CNT, thereby mediating the incorporation of a defined CNT state into the NPC. We propose that the IRC adopts a relatively rigid scaffold that recruits the CNT to primarily form the diffusion barrier of the NPC, rather than enabling channel dilation.
© 2015 American Association for the Advancement of Science. 29 June 2015; accepted 12 August 2015 Published online 27 August 2015. We thank W. M. Clemons, A. Correia, G. Mobbs, A. Patke, D. C. Rees, S. O. Shan, and E. Stuwe for critical reading of the manuscript, M. Budd for help with yeast experiments, J. Herrmann, R. Kunze, and L. Zhang for technical support, and J. Kaiser and the scientific staff of the Stanford Synchrotron Radiation Laboratory (SSRL) Beamline 12-2, the National Institute of General Medical Sciences and National Cancer Institute Structural Biology Facility (GM/CA) at the Advanced Photon Source (APS), and the Advanced Light Source (ALS) beamline 8.2.1 for their support with x-ray diffraction measurements. We acknowledge the Gordon and Betty Moore Foundation, the Beckman Institute, and the Sanofi-Aventis Bioengineering Research Program for their support of the Molecular Observatory at the California Institute of Technology (Caltech). The operations at the SSRL, ALS, and APS are supported by the U.S. 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). T.S. was supported by a Postdoctoral Fellowship of the Deutsche Forschungsgemeinschaft. S.P. and D.H.L are Amgen Graduate Fellows, supported through the Caltech-Amgen Research Collaboration. F.M.H. was supported by a PhD student fellowship of the Boehringer Ingelheim Fonds. S.K. was supported by NIH Awards R01-GM090324 and U54-GM087519 and by the University of Chicago Comprehensive Cancer Center (P30-CA014599). A.A.K. was supported by NIH awards U01-GM094588 and U54-GM087519 and by Searle Funds at The Chicago Community Trust. A.H. was supported by Caltech startup funds, the Albert Wyrick V Scholar Award of he V Foundation for Cancer Research, the 54th Mallinckrodt Scholar Award of the Edward Mallinckrodt Jr. Foundation, a Kimmel Scholar Award of the Sidney Kimmel Foundation for Cancer Research, a Camille-Dreyfus Teacher Scholar Award of The Camille & Henry Dreyfus Foundation, and NIH grant R01-GM111461. The coordinates and structure factors have been deposited with the Protein Data Bank with accession codes 5CWV (Nup192 ^(TAIL)), 5CWU (Nup188^(TAIL)), 4JQ5 (hsNup49^(CCS2+3*)), 4JNV and 4JNU (hsNup57^(CCS3)*), 5CWT (Nup57^(CCS3*)), 4JO7 (hsNup49^(CCS2+3*)•hsNup57^(CCS3*), 2:2 stoichiometry), 4JO9 (hsNup49^(CCS2+3*)•hsNup57^(CCS3*), 1:2 stoichiometry), 5CWW (Nup82^(NTD)•Nup159^T•Nup145N^(APD)) and 5CWS (CNT•Nic96^(R1)•sAB-158). The authors declare no financial conflicts of interest. SK and AK are inventors on a patent application filed by the University of Chicago that covers a design of monobody libraries (US 13/813,409). Monobodies are available from SK under a material transfer agreement with the University of Chicago.
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Accepted Version - nihms773389.pdf
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