Molecular architecture of the luminal ring of the Xenopus laevis nuclear pore complex

Nuclear pore complex (NPC) mediates the flow of substances between the nucleus and cytoplasm in eukaryotic cells. Here we report the cryo-electron tomography (cryo-ET) structure of the luminal ring (LR) of the NPC from Xenopus laevis oocyte. The observed key structural features of the LR are independently confirmed by single-particle cryo-electron microscopy (cryo-EM) analysis. The LR comprises eight butterfly-shaped subunits, each containing two symmetric wings. Each wing consists of four elongated, tubular protomers. Within the LR subunit, the eight protomers form a Finger domain, which directly contacts the fusion between the inner and outer nuclear membranes, and a Grid domain, which serves as a rigid base for the Finger domain. Two neighbouring LR subunits interact with each other through the lateral edges of their wings to constitute a Bumper domain, which displays two major conformations and appears to cushion neighbouring NPCs. Our study reveals previously unknown features of the LR and potentially explains the elastic property of the NPC.

[1]  Friedrich Förster,et al.  Snapshots of nuclear pore complexes in action captured by cryo-electron tomography , 2007, Nature.

[2]  Thomas Strecker,et al.  Acidic pH-Induced Conformations and LAMP1 Binding of the Lassa Virus Glycoprotein Spike , 2016, PLoS pathogens.

[3]  K. Wilson,et al.  Nuclear pore protein gp210 is essential for viability in HeLa cells and Caenorhabditis elegans. , 2003, Molecular biology of the cell.

[4]  D. Agard,et al.  MotionCor2: anisotropic correction of beam-induced motion for improved cryo-electron microscopy , 2017, Nature Methods.

[5]  Daniel Castaño-Díez,et al.  Dynamo Catalogue: Geometrical tools and data management for particle picking in subtomogram averaging of cryo-electron tomograms. , 2017, Journal of structural biology.

[6]  M. Beck,et al.  The nuclear pore complex: understanding its function through structural insight , 2016, Nature Reviews Molecular Cell Biology.

[7]  Qinghua Tao,et al.  Structure of the Cytoplasmic Ring of the Xenopus laevis Nuclear Pore Complex , 2020, bioRxiv.

[8]  Andrea Rothballer,et al.  Poring over pores: nuclear pore complex insertion into the nuclear envelope. , 2013, Trends in biochemical sciences.

[9]  O. Medalia,et al.  Structure and gating of the nuclear pore complex , 2015, Nature Communications.

[10]  Orestis Faklaris,et al.  Nuclear pore complex plasticity during developmental process as revealed by super-resolution microscopy , 2017, Scientific Reports.

[11]  David N Mastronarde,et al.  Automated electron microscope tomography using robust prediction of specimen movements. , 2005, Journal of structural biology.

[12]  G. Blobel,et al.  An integral membrane protein of the pore membrane domain of the nuclear envelope contains a nucleoporin-like region , 1993, The Journal of cell biology.

[13]  G. Blobel,et al.  Electron microscopy of Chaetomium pom152 shows the assembly of ten-bead string , 2018, Cell Discovery.

[14]  M. L. Watson.  Further Observations on the Nuclear Envelope of the Animal Cell , 1959, The Journal of biophysical and biochemical cytology.

[15]  B. Chait,et al.  The molecular architecture of the nuclear pore complex , 2007, Nature.

[16]  T. Schwartz,et al.  The Nuclear Pore Complex as a Flexible and Dynamic Gate , 2016, Cell.

[17]  M. Rout,et al.  POM152 is an integral protein of the pore membrane domain of the yeast nuclear envelope , 1994, The Journal of cell biology.

[18]  U. Aebi,et al.  Toward a more complete 3-D structure of the nuclear pore complex. , 1991, Journal of structural biology.

[19]  K. Wilson,et al.  Interference with the cytoplasmic tail of gp210 disrupts “close apposition” of nuclear membranes and blocks nuclear pore dilation , 2002, The Journal of cell biology.

[20]  M. Radermacher,et al.  Architecture of the Xenopus nuclear pore complex revealed by three- dimensional cryo-electron microscopy , 1993, The Journal of cell biology.

[21]  E. Hartmann,et al.  NDC1: a crucial membrane-integral nucleoporin of metazoan nuclear pore complexes , 2006, The Journal of cell biology.

[22]  W. Baumeister,et al.  In situ architecture of the algal nuclear pore complex , 2018, Nature Communications.

[23]  J. Briggs,et al.  Structure and assembly of the Ebola virus nucleocapsid , 2017, Nature.

[24]  A. C. Meinema,et al.  Long Unfolded Linkers Facilitate Membrane Protein Import Through the Nuclear Pore Complex , 2011, Science.

[25]  U. Kutay,et al.  Transport between the cell nucleus and the cytoplasm. , 1999, Annual review of cell and developmental biology.

[26]  Sjors H.W. Scheres,et al.  RELION: Implementation of a Bayesian approach to cryo-EM structure determination , 2012, Journal of structural biology.

[27]  T. Schwartz.  The Structure Inventory of the Nuclear Pore Complex. , 2016, Journal of molecular biology.

[28]  Daniel Castaño-Díez,et al.  Dynamo: a flexible, user-friendly development tool for subtomogram averaging of cryo-EM data in high-performance computing environments. , 2012, Journal of structural biology.

[29]  D. Spector,et al.  Macromolecular domains within the cell nucleus. , 1993, Annual review of cell biology.

[30]  F. Förster,et al.  Structural basis for coupling protein transport and N-glycosylation at the mammalian endoplasmic reticulum , 2018, Science.

[31]  Wolfram Antonin,et al.  The conserved transmembrane nucleoporin NDC1 is required for nuclear pore complex assembly in vertebrate cells. , 2006, Molecular cell.

[32]  M. Beck,et al.  Structure and Assembly of the Nuclear Pore Complex. , 2019, Annual review of biophysics.

[33]  A. Hyman,et al.  Visualizing the molecular sociology at the HeLa cell nuclear periphery , 2016, Science.

[34]  V. Doye,et al.  Pom33, a novel transmembrane nucleoporin required for proper nuclear pore complex distribution , 2010, The Journal of cell biology.

[35]  Bernd Rieger,et al.  Super-resolution imaging visualizes the eightfold symmetry of gp210 proteins around the nuclear pore complex and resolves the central channel with nanometer resolution , 2012, Journal of Cell Science.

[36]  J R Kremer,et al.  Computer visualization of three-dimensional image data using IMOD. , 1996, Journal of structural biology.

[37]  Ohad Medalia,et al.  Functional architecture of the nuclear pore complex. , 2012, Annual review of biophysics.

[38]  G. Blobel,et al.  Primary structure analysis of an integral membrane glycoprotein of the nuclear pore , 1989, The Journal of cell biology.

[39]  F. Förster,et al.  Structure of the native Sec61 protein-conducting channel , 2015, Nature Communications.

[40]  C. Akey.  Structural plasticity of the nuclear pore complex. , 1995, Journal of molecular biology.

[41]  J. Aitchison,et al.  The multifunctional nuclear pore complex: a platform for controlling gene expression. , 2014, Current opinion in cell biology.

[42]  J. Briggs,et al.  Implementation of a cryo-electron tomography tilt-scheme optimized for high resolution subtomogram averaging , 2017, Journal of structural biology.

[43]  O. Medalia,et al.  The human nuclear pore complex as revealed by cryo-electron tomography. , 2012, Structure.

[44]  P. Bork,et al.  In situ structural analysis of the human nuclear pore complex , 2015, Nature.

[45]  L. Gerace,et al.  Identification of a major polypeptide of the nuclear pore complex , 1982, The Journal of cell biology.

[46]  C. Akey,et al.  Interactions and structure of the nuclear pore complex revealed by cryo- electron microscopy , 1989, The Journal of cell biology.

[47]  André Hoelz,et al.  The structure of the nuclear pore complex. , 2011, Annual review of biochemistry.

[48]  A. Hoelz,et al.  The Structure of the Nuclear Pore Complex (An Update). , 2019, Annual review of biochemistry.

[49]  U. Greber,et al.  A major glycoprotein of the nuclear pore complex is a membrane‐spanning polypeptide with a large lumenal domain and a small cytoplasmic tail. , 1990, The EMBO journal.

[50]  Beata Turoňová,et al.  Efficient 3D-CTF correction for cryo-electron tomography using NovaCTF improves subtomogram averaging resolution to 3.4 Å , 2017, Journal of structural biology.

[51]  Seung Joong Kim,et al.  Molecular Architecture of the Major Membrane Ring Component of the Nuclear Pore Complex. , 2017, Structure.

[52]  Ueli Aebi,et al.  Cryo-electron tomography provides novel insights into nuclear pore architecture: implications for nucleocytoplasmic transport. , 2003, Journal of molecular biology.

[53]  N. Grigorieff,et al.  CTFFIND4: Fast and accurate defocus estimation from electron micrographs , 2015, bioRxiv.

[54]  O. Medalia,et al.  Structural analysis of a metazoan nuclear pore complex reveals a fused concentric ring architecture. , 2010, Journal of molecular biology.