scholarly journals Cryo-EM structure of the Inner Ring from Xenopus laevis Nuclear Pore Complex

2021 ◽  
Author(s):  
Gaoxingyu Huang ◽  
Xiechao Zhan ◽  
Chao Zeng ◽  
Ke Liang ◽  
Xuechen Zhu ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Xenopus Laevis ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Single Particle ◽  
Nuclear Pore ◽  
Nucleocytoplasmic Shuttling ◽  
Cryo Electron Microscopy

Nuclear pore complex (NPC) mediates nucleocytoplasmic shuttling. Here we present single-particle cryo-electron microscopy (cryo-EM) structure of the inner ring (IR) subunit from Xenopus laevis NPC at an average resolution of 4.4 Å. The symmetric IR subunit comprises a cytoplasmic half and a nuclear half. A homo-dimer of Nup205 resides at the center of the IR subunit, flanked by two molecules of Nup188. Four molecules of Nup93 each places an extended helix into the axial groove of Nup205 or Nup188, together constituting the central scaffold. The channel nucleoporin heterotrimer (CNT) of Nup54/58/62 is anchored on the central scaffold. Six Nup155 molecules interact with the central scaffold and together with the NDC1-ALADIN hetero-dimers anchor the IR subunit to the nuclear envelope and to outer rings. The scarce inter-subunit contacts may allow sufficient latitude in conformation and diameter of the IR. Our structure of vertebrate IR reveals key insights that are functionally important.

scholarly journals Structure of the Cytoplasmic Ring of the Xenopus laevis Nuclear Pore Complex

2020 ◽  
Author(s):  
Gaoxingyu Huang ◽  
Yanqing Zhang ◽  
Xuechen Zhu ◽  
Chao Zeng ◽  
Qifan Wang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Xenopus Laevis ◽  
Nuclear Pore Complex ◽  
Single Particle ◽  
Coiled Coil ◽  
Structural Features ◽  
Nuclear Pore ◽  
Structural Elements ◽  
Cryo Electron Microscopy ◽  
Secondary Structural Elements

Nuclear pore complex (NPC) exhibits structural plasticity and has only been characterized at local resolutions of up to 15 Å for the cytoplasmic ring (CR). Here we present a single-particle cryo-electron microscopy (cryo-EM) structure of the CR from Xenopus laevis NPC at average resolutions of 5.5-7.9 Å, with local resolutions reaching 4.5 Å. Improved resolutions allow identification and placement of secondary structural elements in the majority of the CR components. The two Y complexes in each CR subunit interact with each other and associate with those from flanking subunits, forming a circular scaffold. Within each CR subunit, the Nup358-containing region wraps around the stems of both Y complexes, likely stabilizing the scaffold. Nup205 connects the short arms of the two Y complexes and associates with the stem of a neighbouring Y complex. The Nup214-containing region uses an extended coiled-coil to link Nup85 of the two Y complexes and protrudes into the axial pore of the NPC. These previously uncharacterized structural features reveal insights into NPC assembly.

scholarly journals Architecture of the Xenopus nuclear pore complex revealed by three-dimensional cryo-electron microscopy

1993 ◽  
Vol 122 (1) ◽  
pp. 1-19 ◽  
Author(s):  
CW Akey ◽  
M Radermacher
Keyword(s):  
Electron Microscopy ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Three Dimensional ◽  
Nucleocytoplasmic Transport ◽  
Domain Model ◽  
Nuclear Pore ◽  
Transport Channel ◽  
Macromolecular Transport ◽  
Cryo Electron Microscopy

The nuclear pore complex spans the nuclear envelope and functions as a macromolecular transporter in the ATP-dependent process of nucleocytoplasmic transport. In this report, we present three dimensional (3D) structures for both membrane-associated and detergent-extracted Xenopus NPCs, imaged in frozen buffers by cryo-electron microscopy. A comparison of the differing configurations present in the 3D maps suggests that the spokes may possess an intrinsic conformational flexibility. When combined with recent data from a 3D map of negatively stained NPCs (Hinshaw, J. E., B. O. Carragher, and R. A. Milligan. 1992. Cell. 69:1133-1141), these observations suggest a minimal domain model for the spoke-ring complex which may account for the observed plasticity of this assembly. Moreover, lumenal domains in adjacent spokes are interconnected by radial arm dimers, forming a lumenal ring that may be responsible for anchoring the NPC within the nuclear envelope pore. Importantly, the NPC transporter is visualized as a centrally tapered cylinder that spans the entire width of the NPC, in a direction normal to the nuclear envelope. The central positioning, tripartite structure, and hollow nature of the transporter suggests that it may form a macromolecular transport channel, with a globular gating domain at each end. Finally, the packing of the transporter within the spokes creates a set of eight internal channels that may be responsible, in part, for the diffusion of ions and small molecules across the nuclear envelope.

scholarly journals The role of the integral membrane nucleoporins Ndc1p and Pom152p in nuclear pore complex assembly and function

2006 ◽  
Vol 173 (3) ◽  
pp. 361-371 ◽  
Author(s):  
Alexis S. Madrid ◽  
Joel Mancuso ◽  
W. Zacheus Cande ◽  
Karsten Weis
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Nuclear Envelope ◽  
Lipid Bilayers ◽  
Nuclear Pore Complex ◽  
Transmembrane Proteins ◽  
Nuclear Pore ◽  
Large Channel ◽  
And Function

The nuclear pore complex (NPC) is a large channel that spans the two lipid bilayers of the nuclear envelope and mediates transport events between the cytoplasm and the nucleus. Only a few NPC components are transmembrane proteins, and the role of these proteins in NPC function and assembly remains poorly understood. We investigate the function of the three integral membrane nucleoporins, which are Ndc1p, Pom152p, and Pom34p, in NPC assembly and transport in Saccharomyces cerevisiae. We find that Ndc1p is important for the correct localization of nuclear transport cargoes and of components of the NPC. However, the role of Ndc1p in NPC assembly is partially redundant with Pom152p, as cells lacking both of these proteins show enhanced NPC disruption. Electron microscopy studies reveal that the absence of Ndc1p and Pom152p results in aberrant pores that have enlarged diameters and lack proteinaceous material, leading to an increased diffusion between the cytoplasm and the nucleus.

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

2020 ◽  
Author(s):  
Yanqing Zhang ◽  
Sai Li ◽  
Chao Zeng ◽  
Gaoxingyu Huang ◽  
Xuechen Zhu ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Nuclear Pore Complex ◽  
Electron Tomography ◽  
Structural Features ◽  
Rigid Base ◽  
Nuclear Pore ◽  
Molecular Architecture ◽  
Nuclear Membranes ◽  
Cryo Electron Microscopy ◽  
Cryo Electron Tomography

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.

scholarly journals Dissecting Torsin/cofactor function at the nuclear envelope: a genetic study

2016 ◽  
Vol 27 (25) ◽  
pp. 3964-3971 ◽  
Author(s):  
Ethan Laudermilch ◽  
Pei-Ling Tsai ◽  
Morven Graham ◽  
Elizabeth Turner ◽  
Chenguang Zhao ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Structural Analysis ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Compositional Analysis ◽  
Immunogold Labeling ◽  
Nuclear Pore ◽  
Functional Link ◽  
Experimental Platform ◽  
Bleb Formation

The human genome encodes four Torsin ATPases, the functions of which are poorly understood. In this study, we use CRISPR/Cas9 engineering to delete all four Torsin ATPases individually and in combination. Using nuclear envelope (NE) blebbing as a phenotypic measure, we establish a direct correlation between the number of inactivated Torsin alleles and the occurrence of omega-shaped herniations within the lumen of the NE. A similar, although not identical, redundancy is observed for LAP1 and LULL1, which serve as regulatory cofactors for a subset of Torsin ATPases. Unexpectedly, deletion of Tor2A in a TorA/B/3A-deficient background results in a stark increase of bleb formation, even though Tor2A does not respond to LAP1/LULL1 stimulation. The robustness of the observed phenotype in Torsin-deficient cells enables a structural analysis via electron microscopy tomography and a compositional analysis via immunogold labeling. Ubiquitin and nucleoporins were identified as distinctively localizing components of the omega-shaped bleb structure. These findings suggest a functional link between the Torsin/cofactor system and NE/nuclear pore complex biogenesis or homeostasis and establish a Torsin-deficient cell line as a valuable experimental platform with which to decipher Torsin function.

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