26 results on '"*NUCLEAR pore complex"'
Search Results
2. Comprehensive structure and functional adaptations of the yeast nuclear pore complex.
- Author
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Akey, Christopher W, Singh, Digvijay, Ouch, Christna, Echeverria, Ignacia, Nudelman, Ilona, Varberg, Joseph M, Yu, Zulin, Fang, Fei, Shi, Yi, Wang, Junjie, Salzberg, Daniel, Song, Kangkang, Xu, Chen, Gumbart, James C, Suslov, Sergey, Unruh, Jay, Jaspersen, Sue L, Chait, Brian T, Sali, Andrej, Fernandez-Martinez, Javier, Ludtke, Steven J, Villa, Elizabeth, and Rout, Michael P
- Subjects
Nuclear Envelope ,Nuclear Pore ,Saccharomyces cerevisiae ,Nuclear Pore Complex Proteins ,Saccharomyces cerevisiae Proteins ,Reproducibility of Results ,Adaptation ,Physiological ,Amino Acid Sequence ,Amino Acid Motifs ,Fluorescence ,Molecular Docking Simulation ,Protein Domains ,NPC evolution ,Nuclear pore complex ,cryo-electron microscopy ,cryo-electron tomography ,inner ring dilation ,nuclear basket ,nucleocytoplasmic transport ,nucleoporins ,structural isoforms ,1.1 Normal biological development and functioning ,Generic health relevance ,Biological Sciences ,Medical and Health Sciences ,Developmental Biology - Abstract
Nuclear pore complexes (NPCs) mediate the nucleocytoplasmic transport of macromolecules. Here we provide a structure of the isolated yeast NPC in which the inner ring is resolved by cryo-EM at sub-nanometer resolution to show how flexible connectors tie together different structural and functional layers. These connectors may be targets for phosphorylation and regulated disassembly in cells with an open mitosis. Moreover, some nucleoporin pairs and transport factors have similar interaction motifs, which suggests an evolutionary and mechanistic link between assembly and transport. We provide evidence for three major NPC variants that may foreshadow functional specializations at the nuclear periphery. Cryo-electron tomography extended these studies, providing a model of the in situ NPC with a radially expanded inner ring. Our comprehensive model reveals features of the nuclear basket and central transporter, suggests a role for the lumenal Pom152 ring in restricting dilation, and highlights structural plasticity that may be required for transport.
- Published
- 2022
3. The molecular architecture of the nuclear basket.
- Author
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Singh, Digvijay, Soni, Neelesh, Hutchings, Joshua, Echeverria, Ignacia, Shaikh, Farhaz, Duquette, Madeleine, Suslov, Sergey, Li, Zhixun, van Eeuwen, Trevor, Molloy, Kelly, Shi, Yi, Wang, Junjie, Guo, Qiang, Chait, Brian T., Fernandez-Martinez, Javier, Rout, Michael P., Sali, Andrej, and Villa, Elizabeth
- Subjects
- *
NUCLEAR pore complex , *NUCLEAR transport , *NUCLEOCYTOPLASMIC interactions , *PROTEIN domains , *NUCLEOPORINS - Abstract
The nuclear pore complex (NPC) is the sole mediator of nucleocytoplasmic transport. Despite great advances in understanding its conserved core architecture, the peripheral regions can exhibit considerable variation within and between species. One such structure is the cage-like nuclear basket. Despite its crucial roles in mRNA surveillance and chromatin organization, an architectural understanding has remained elusive. Using in-cell cryo-electron tomography and subtomogram analysis, we explored the NPC's structural variations and the nuclear basket across fungi (yeast; S. cerevisiae), mammals (mouse; M. musculus), and protozoa (T. gondii). Using integrative structural modeling, we computed a model of the basket in yeast and mammals that revealed how a hub of nucleoporins (Nups) in the nuclear ring binds to basket-forming Mlp/Tpr proteins: the coiled-coil domains of Mlp/Tpr form the struts of the basket, while their unstructured termini constitute the basket distal densities, which potentially serve as a docking site for mRNA preprocessing before nucleocytoplasmic transport. [Display omitted] • A stable basket is anchored by a hub of Nups into a double nuclear ring • Mlps/Tprs form struts; their N/C termini form a distal density that docks mRNA • A 20-nm exclusion zone around the basket suggests its role in chromatin organization • The stoichiometry of the outer rings is variable across and within species The elusive architecture of the nuclear basket reveals that a double ring is necessary to stabilize a hub of Nups from which the struts emanate. The basket's distal density contains the N/C termini of the strut-forming proteins that serve as a docking platform for cargo and nuclear periphery elements. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
4. Structure and Function of the Nuclear Pore Complex Cytoplasmic mRNA Export Platform
- Author
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Fernandez-Martinez, Javier, Kim, Seung Joong, Shi, Yi, Upla, Paula, Pellarin, Riccardo, Gagnon, Michael, Chemmama, Ilan E, Wang, Junjie, Nudelman, Ilona, Zhang, Wenzhu, Williams, Rosemary, Rice, William J, Stokes, David L, Zenklusen, Daniel, Chait, Brian T, Sali, Andrej, and Rout, Michael P
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,Genetics ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,Active Transport ,Cell Nucleus ,Fungal Proteins ,Nuclear Pore ,Nuclear Pore Complex Proteins ,RNA ,Messenger ,Saccharomyces cerevisiae ,Saccharomyces cerevisiae Proteins ,Yeasts ,Nup4 complex ,Nup82 complex ,computational structural biology ,cross-linking and mass spectrometry ,electron microscopy ,integrative structure determination ,mRNA export ,mRNP remodeling ,nuclear pore complex ,small-angle X-ray scattering ,Medical and Health Sciences ,Developmental Biology ,Biological sciences ,Biomedical and clinical sciences - Abstract
The last steps in mRNA export and remodeling are performed by the Nup82 complex, a large conserved assembly at the cytoplasmic face of the nuclear pore complex (NPC). By integrating diverse structural data, we have determined the molecular architecture of the native Nup82 complex at subnanometer precision. The complex consists of two compositionally identical multiprotein subunits that adopt different configurations. The Nup82 complex fits into the NPC through the outer ring Nup84 complex. Our map shows that this entire 14-MDa Nup82-Nup84 complex assembly positions the cytoplasmic mRNA export factor docking sites and messenger ribonucleoprotein (mRNP) remodeling machinery right over the NPC's central channel rather than on distal cytoplasmic filaments, as previously supposed. We suggest that this configuration efficiently captures and remodels exporting mRNP particles immediately upon reaching the cytoplasmic side of the NPC.
- Published
- 2016
5. Comprehensive maturity of nuclear pore complexes regulates zygotic genome activation.
- Author
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Shen, Weimin, Gong, Bo, Xing, Cencan, Zhang, Lin, Sun, Jiawei, Chen, Yuling, Yang, Changmei, Yan, Lu, Chen, Luxi, Yao, Likun, Li, Guangyuan, Deng, Haiteng, Wu, Xiaotong, and Meng, Anming
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NUCLEOCYTOPLASMIC interactions , *CARRIER proteins , *NUCLEAR transport , *GENOMES , *TRANSCRIPTION factors , *EMBRYOLOGY - Abstract
Nuclear pore complexes (NPCs) are channels for nucleocytoplasmic transport of proteins and RNAs. However, it remains unclear whether composition, structure, and permeability of NPCs dynamically change during the cleavage period of vertebrate embryos and affect embryonic development. Here, we report that the comprehensive NPC maturity (CNM) controls the onset of zygotic genome activation (ZGA) during zebrafish early embryogenesis. We show that more nucleoporin proteins are recruited to and assembled into NPCs with development, resulting in progressive increase of NPCs in size and complexity. Maternal transcription factors (TFs) transport into nuclei more efficiently with increasing CNM. Deficiency or dysfunction of Nup133 or Ahctf1/Elys impairs NPC assembly, maternal TFs nuclear transport, and ZGA onset, while nup133 overexpression promotes these processes. Therefore, CNM may act as a molecular timer for ZGA by controlling nuclear transport of maternal TFs that reach nuclear concentration thresholds at a given time to initiate ZGA. [Display omitted] • Maternal TFs cannot transport into nuclei efficiently in embryos before ZGA • Comprehensive NPC maturity gradually increases in embryos at pre-MBT stages • Comprehensive NPC maturity correlates with TF nuclear transport and ZGA • NPC assembly impairment downregulates TF nuclear transport and delays ZGA Experiments in zebrafish indicate that nuclear pore complex (NPC) maturation serves as a molecular timer of zygote genome activation, a key part of the oocyte-to-embryo transition during vertebrate development. Increased size and complexity of NPCs leads to increased nuclear transport of maternal transcription factors, allowing transcription factors to reach thresholds that initiate zygotic genome activation. [ABSTRACT FROM AUTHOR]
- Published
- 2022
- Full Text
- View/download PDF
6. Pre-assembled Nuclear Pores Insert into the Nuclear Envelope during Early Development.
- Author
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Hampoelz, Bernhard, Mackmull, Marie-Therese, Machado, Pedro, Ronchi, Paolo, Bui, Khanh Huy, Schieber, Nicole, Santarella-Mellwig, Rachel, Necakov, Aleksandar, Andrés-Pons, Amparo, Philippe, Jean Marc, Lecuit, Thomas, Schwab, Yannick, and Beck, Martin
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NUCLEAR pore complex , *NUCLEAR membranes , *NUCLEOCYTOPLASMIC interactions , *ENDOPLASMIC reticulum , *BLASTODERM , *EMBRYOS , *TISSUE scaffolds - Abstract
Summary Nuclear pore complexes (NPCs) span the nuclear envelope (NE) and mediate nucleocytoplasmic transport. In metazoan oocytes and early embryos, NPCs reside not only within the NE, but also at some endoplasmic reticulum (ER) membrane sheets, termed annulate lamellae (AL). Although a role for AL as NPC storage pools has been discussed, it remains controversial whether and how they contribute to the NPC density at the NE. Here, we show that AL insert into the NE as the ER feeds rapid nuclear expansion in Drosophila blastoderm embryos. We demonstrate that NPCs within AL resemble pore scaffolds that mature only upon insertion into the NE. We delineate a topological model in which NE openings are critical for AL uptake that nevertheless occurs without compromising the permeability barrier of the NE. We finally show that this unanticipated mode of pore insertion is developmentally regulated and operates prior to gastrulation. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
7. The Nuclear Pore Complex as a Flexible and Dynamic Gate.
- Author
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Knockenhauer, Kevin E. and Schwartz, Thomas U.
- Subjects
- *
NUCLEAR pore complex , *NUCLEAR membranes , *CELL compartmentation , *HETEROGENEITY , *CYTOPLASM - Abstract
Nuclear pore complexes (NPCs) perforate the nuclear envelope and serve as the primary transport gates for molecular exchange between nucleus and cytoplasm. Stripping the megadalton complex down to its most essential organizational elements, one can divide the NPC into scaffold components and the disordered elements attached to them that generate a selective barrier between compartments. These structural elements exhibit flexibility, which may hold a clue in understanding NPC assembly and function. Here we review the current status of NPC research with a focus on the functional implications of its structural and compositional heterogeneity. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
8. The Nuclear Pore-Associated TREX-2 Complex Employs Mediator to Regulate Gene Expression.
- Author
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Schneider, Maren, Hellerschmied, Doris, Schubert, Tobias, Amlacher, Stefan, Vinayachandran, Vinesh, Reja, Rohit, Pugh, B. Franklin, Clausen, Tim, and Köhler, Alwin
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NUCLEAR pore complex , *GENETIC regulation , *MESSENGER RNA , *EXONUCLEASES , *GENETIC transcription - Abstract
Summary Nuclear pore complexes (NPCs) influence gene expression besides their established function in nuclear transport. The TREX-2 complex localizes to the NPC basket and affects gene-NPC interactions, transcription, and mRNA export. How TREX-2 regulates the gene expression machinery is unknown. Here, we show that TREX-2 interacts with the Mediator complex, an essential regulator of RNA Polymerase (Pol) II. Structural and biochemical studies identify a conserved region on TREX-2, which directly binds the Mediator Med31/Med7N submodule. TREX-2 regulates assembly of Mediator with the Cdk8 kinase and is required for recruitment and site-specific phosphorylation of Pol II. Transcriptome and phenotypic profiling confirm that TREX-2 and Med31 are functionally interdependent at specific genes. TREX-2 additionally uses its Mediator-interacting surface to regulate mRNA export suggesting a mechanism for coupling transcription initiation and early steps of mRNA processing. Our data provide mechanistic insight into how an NPC-associated adaptor complex accesses the core transcription machinery. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
9. Surveillance of Nuclear Pore Complex Assembly by ESCRT-III/Vps4.
- Author
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Webster, Brant M., Colombi, Paolo, Jäger, Jens, and Lusk, C. Patrick
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NUCLEAR pore complex , *CELL compartmentation , *NUCLEAR membranes , *CELLULAR aging , *CELL physiology , *MEMBRANE proteins - Abstract
Summary The maintenance of nuclear compartmentalization by the nuclear envelope and nuclear pore complexes (NPCs) is essential for cell function; loss of compartmentalization is associated with cancers, laminopathies, and aging. We uncovered a pathway that surveils NPC assembly intermediates to promote the formation of functional NPCs. Surveillance is mediated by Heh2, a member of the LEM (Lap2-emerin-MAN1) family of integral inner nuclear membrane proteins, which binds to an early NPC assembly intermediate, but not to mature NPCs. Heh2 recruits the endosomal sorting complex required for transport (ESCRT)—III subunit Snf7 and the AAA-ATPase Vps4 to destabilize and clear defective NPC assembly intermediates. When surveillance or clearance is compromised, malformed NPCs accumulate in a storage of improperly assembled nuclear pore complexes compartment, or SINC. The SINC is retained in old mothers to prevent loss of daughter lifespan, highlighting a continuum of mechanisms to ensure nuclear compartmentalization. [ABSTRACT FROM AUTHOR]
- Published
- 2014
- Full Text
- View/download PDF
10. Parental genome unification is highly error-prone in mammalian embryos
- Author
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Patrick Aldag, Martyn Blayney, Melina Schuh, Andrea Lucas-Hahn, Jonas Bucevičius, Tommaso Cavazza, Clara Baker, M. Aushev, Antonio Z. Politi, Yuko Takeda, Meenakshi Choudhary, Heiner Niemann, Mary Herbert, Kay Elder, and Gražvydas Lukinavičius
- Subjects
Male ,Zygote ,chromosome segregation ,Embryonic Development ,Mitosis ,Aneuploidy ,Biology ,Microtubules ,Genome ,Article ,Chromosomes ,General Biochemistry, Genetics and Molecular Biology ,Chromosome segregation ,03 medical and health sciences ,0302 clinical medicine ,nuclear pore complex ,medicine ,Animals ,Humans ,genome organization ,aneuploidy ,centrosomes ,030304 developmental biology ,Cell Nucleus ,Centrosome ,Genetics ,0303 health sciences ,dynein ,Pronucleus ,spindle ,Embryo, Mammalian ,medicine.disease ,human embryo ,Spermatozoa ,fertilization ,embryonic structures ,Oocytes ,Cattle ,Ploidy ,Cell Nucleolus ,030217 neurology & neurosurgery - Abstract
Summary Most human embryos are aneuploid. Aneuploidy frequently arises during the early mitotic divisions of the embryo, but its origin remains elusive. Human zygotes that cluster their nucleoli at the pronuclear interface are thought to be more likely to develop into healthy euploid embryos. Here, we show that the parental genomes cluster with nucleoli in each pronucleus within human and bovine zygotes, and clustering is required for the reliable unification of the parental genomes after fertilization. During migration of intact pronuclei, the parental genomes polarize toward each other in a process driven by centrosomes, dynein, microtubules, and nuclear pore complexes. The maternal and paternal chromosomes eventually cluster at the pronuclear interface, in direct proximity to each other, yet separated. Parental genome clustering ensures the rapid unification of the parental genomes on nuclear envelope breakdown. However, clustering often fails, leading to chromosome segregation errors and micronuclei, incompatible with healthy embryo development., Graphical abstract, Highlights • The parental genomes cluster at the pronuclear interface in human and bovine zygotes • Clustering is driven by centrosomes, which often reside at the pronuclear interface • Dynein orients chromosomes toward centrosomes via nuclear pore complexes as adaptors • Clustering defects lead to aneuploidy and micronuclei, impairing embryo development, In human and bovine zygotes, parental genomes cluster and polarize toward each other in a highly error-prone process driven by centrosomes, dynein, microtubules, and nuclear pore complexes. Failure to cluster the parental genomes leads to chromosome segregation errors and micronuclei, which are incompatible with healthy embryo development.
- Published
- 2021
11. Gating Immunity and Death at the Nuclear Pore Complex.
- Author
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Dasso, Mary and Fontoura, Beatriz M.A.
- Subjects
- *
IMMUNITY , *NUCLEAR pore complex , *CELL death , *CYCLIN-dependent kinase inhibitors , *IMMUNOLOGY - Abstract
The nuclear pore complex is the primary conduit for nuclear import and export of molecules. In this issue, Gu et al. uncover a novel mechanism in which immune signaling and programmed cell death require nuclear pore rearrangement and release of sequestered cyclin-dependent kinase inhibitors to elicit immunity and death. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
12. Nuclear Pores Assemble from Nucleoporin Condensates During Oogenesis
- Author
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Andre Schwarz, Bernhard Hampoelz, Helena Bragulat-Teixidor, Imre Gaspar, Anne Ephrussi, Christian Tischer, Yannick Schwab, Paolo Ronchi, and Martin Beck
- Subjects
NPC assembly ,Nup358 ,Active Transport, Cell Nucleus ,Biology ,Microtubules ,Article ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,local translation ,Oogenesis ,0302 clinical medicine ,nuclear pore complex ,otorhinolaryngologic diseases ,Animals ,Drosophila Proteins ,RNA, Messenger ,nucleoporin condensation ,Nuclear pore ,Mitosis ,030304 developmental biology ,0303 health sciences ,microtubule-dependent transport ,RNA localization ,Cell biology ,Nuclear Pore Complex Proteins ,stomatognathic diseases ,Ran ,Drosophila melanogaster ,ran GTP-Binding Protein ,Cytoplasm ,Nuclear Pore ,Female ,Interphase ,Nucleoporin ,Nuclear transport ,030217 neurology & neurosurgery ,Biogenesis ,Molecular Chaperones - Abstract
Summary The molecular events that direct nuclear pore complex (NPC) assembly toward nuclear envelopes have been conceptualized in two pathways that occur during mitosis or interphase, respectively. In gametes and embryonic cells, NPCs also occur within stacked cytoplasmic membrane sheets, termed annulate lamellae (AL), which serve as NPC storage for early development. The mechanism of NPC biogenesis at cytoplasmic membranes remains unknown. Here, we show that during Drosophila oogenesis, Nucleoporins condense into different precursor granules that interact and progress into NPCs. Nup358 is a key player that condenses into NPC assembly platforms while its mRNA localizes to their surface in a translation-dependent manner. In concert, Microtubule-dependent transport, the small GTPase Ran and nuclear transport receptors regulate NPC biogenesis in oocytes. We delineate a non-canonical NPC assembly mechanism that relies on Nucleoporin condensates and occurs away from the nucleus under conditions of cell cycle arrest., Graphical Abstract, Highlights • Nucleoporins condense into precursor granules that mature into nuclear pore complexes (NPCs) • Microtubule-dependent transport and RanGTP promote NPC biogenesis in vivo • nup358 mRNA localizes to sites of NPC biogenesis in a translation-dependent manner • Nup358 protein condensation is Ran regulated and controls NPC assembly beyond nuclei, Nuclear pores form through progressive condensation events during oogenesis
- Published
- 2019
- Full Text
- View/download PDF
13. Cone-shaped HIV-1 capsids are transported through intact nuclear pores
- Author
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Jona Rada, Barbara Müller, Marina Lusic, Thorsten G. Müller, Kathleen Börner, Simone Mattei, Martin Beck, Christian E. Zimmerli, Beata Turoňová, Vojtech Zila, Erica Margiotta, Matteo Allegretti, and Hans-Georg Kräusslich
- Subjects
viruses ,electron tomography ,Active Transport, Cell Nucleus ,Human immunodeficiency virus (HIV) ,HIV Infections ,Biology ,medicine.disease_cause ,Models, Biological ,Article ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,Capsid ,0302 clinical medicine ,Viral life cycle ,Correlative light and electron microscopy ,nuclear pore complex ,medicine ,Humans ,correlative light and electron microscopy ,Nuclear pore ,030304 developmental biology ,mRNA Cleavage and Polyadenylation Factors ,0303 health sciences ,human immunodeficiency virus ,Cryoelectron Microscopy ,Virion ,Reverse Transcription ,cryoelectron tomography ,Virus Internalization ,nuclear import ,Cell biology ,HEK293 Cells ,HIV-1 ,Nuclear Pore ,uncoating ,Nuclear transport ,030217 neurology & neurosurgery - Abstract
Summary Human immunodeficiency virus (HIV-1) remains a major health threat. Viral capsid uncoating and nuclear import of the viral genome are critical for productive infection. The size of the HIV-1 capsid is generally believed to exceed the diameter of the nuclear pore complex (NPC), indicating that capsid uncoating has to occur prior to nuclear import. Here, we combined correlative light and electron microscopy with subtomogram averaging to capture the structural status of reverse transcription-competent HIV-1 complexes in infected T cells. We demonstrated that the diameter of the NPC in cellulo is sufficient for the import of apparently intact, cone-shaped capsids. Subsequent to nuclear import, we detected disrupted and empty capsid fragments, indicating that uncoating of the replication complex occurs by breaking the capsid open, and not by disassembly into individual subunits. Our data directly visualize a key step in HIV-1 replication and enhance our mechanistic understanding of the viral life cycle., Graphical Abstract, Highlights • Nuclear translocation of HIV-1 capsids is captured by 3D CLEM/cryo-ET • Nuclear pore complexes in T cells are sufficiently dilated to accommodate HIV-1 capsids • Cone-shaped HIV-1 capsids translocate through nuclear pore complexes • Inside the nucleus HIV-1 capsids rupture and release their interior, Visualization of nuclear translocation of HIV-1 capsids by 3D correlative fluorescence light and electron microscope, combined with cryoelectron tomography, demonstrates that nuclear pore complexes in infected T cells are sufficiently dilated to allow cone-shaped HIV-1 capsids to pass through.
- Published
- 2021
14. Parental genome unification is highly error-prone in mammalian embryos.
- Author
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Cavazza, Tommaso, Takeda, Yuko, Politi, Antonio Z., Aushev, Magomet, Aldag, Patrick, Baker, Clara, Choudhary, Meenakshi, Bucevičius, Jonas, Lukinavičius, Gražvydas, Elder, Kay, Blayney, Martyn, Lucas-Hahn, Andrea, Niemann, Heiner, Herbert, Mary, and Schuh, Melina
- Subjects
- *
MAMMALIAN embryos , *NUCLEAR membranes , *CHROMOSOME segregation , *HUMAN embryos , *MICROTUBULES , *ZYGOTES - Abstract
Most human embryos are aneuploid. Aneuploidy frequently arises during the early mitotic divisions of the embryo, but its origin remains elusive. Human zygotes that cluster their nucleoli at the pronuclear interface are thought to be more likely to develop into healthy euploid embryos. Here, we show that the parental genomes cluster with nucleoli in each pronucleus within human and bovine zygotes, and clustering is required for the reliable unification of the parental genomes after fertilization. During migration of intact pronuclei, the parental genomes polarize toward each other in a process driven by centrosomes, dynein, microtubules, and nuclear pore complexes. The maternal and paternal chromosomes eventually cluster at the pronuclear interface, in direct proximity to each other, yet separated. Parental genome clustering ensures the rapid unification of the parental genomes on nuclear envelope breakdown. However, clustering often fails, leading to chromosome segregation errors and micronuclei, incompatible with healthy embryo development. [Display omitted] • The parental genomes cluster at the pronuclear interface in human and bovine zygotes • Clustering is driven by centrosomes, which often reside at the pronuclear interface • Dynein orients chromosomes toward centrosomes via nuclear pore complexes as adaptors • Clustering defects lead to aneuploidy and micronuclei, impairing embryo development In human and bovine zygotes, parental genomes cluster and polarize toward each other in a highly error-prone process driven by centrosomes, dynein, microtubules, and nuclear pore complexes. Failure to cluster the parental genomes leads to chromosome segregation errors and micronuclei, which are incompatible with healthy embryo development. [ABSTRACT FROM AUTHOR]
- Published
- 2021
- Full Text
- View/download PDF
15. Cone-shaped HIV-1 capsids are transported through intact nuclear pores.
- Author
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Zila, Vojtech, Margiotta, Erica, Turoňová, Beata, Müller, Thorsten G., Zimmerli, Christian E., Mattei, Simone, Allegretti, Matteo, Börner, Kathleen, Rada, Jona, Müller, Barbara, Lusic, Marina, Kräusslich, Hans-Georg, and Beck, Martin
- Subjects
- *
CAPSIDS , *VIRAL genomes , *ELECTRON microscopes , *MICROSCOPY , *T cells , *HIV - Abstract
Human immunodeficiency virus (HIV-1) remains a major health threat. Viral capsid uncoating and nuclear import of the viral genome are critical for productive infection. The size of the HIV-1 capsid is generally believed to exceed the diameter of the nuclear pore complex (NPC), indicating that capsid uncoating has to occur prior to nuclear import. Here, we combined correlative light and electron microscopy with subtomogram averaging to capture the structural status of reverse transcription-competent HIV-1 complexes in infected T cells. We demonstrated that the diameter of the NPC in cellulo is sufficient for the import of apparently intact, cone-shaped capsids. Subsequent to nuclear import, we detected disrupted and empty capsid fragments, indicating that uncoating of the replication complex occurs by breaking the capsid open, and not by disassembly into individual subunits. Our data directly visualize a key step in HIV-1 replication and enhance our mechanistic understanding of the viral life cycle. • Nuclear translocation of HIV-1 capsids is captured by 3D CLEM/cryo-ET • Nuclear pore complexes in T cells are sufficiently dilated to accommodate HIV-1 capsids • Cone-shaped HIV-1 capsids translocate through nuclear pore complexes • Inside the nucleus HIV-1 capsids rupture and release their interior Visualization of nuclear translocation of HIV-1 capsids by 3D correlative fluorescence light and electron microscope, combined with cryoelectron tomography, demonstrates that nuclear pore complexes in infected T cells are sufficiently dilated to allow cone-shaped HIV-1 capsids to pass through. [ABSTRACT FROM AUTHOR]
- Published
- 2021
- Full Text
- View/download PDF
16. Maturation Kinetics of a Multiprotein Complex Revealed by Metabolic Labeling.
- Author
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Onischenko, Evgeny, Noor, Elad, Fischer, Jonas S., Gillet, Ludovic, Wojtynek, Matthias, Vallotton, Pascal, and Weis, Karsten
- Subjects
- *
PROTEOMICS , *ANALYTICAL mechanics , *CELL anatomy , *NUCLEOPORINS , *DRUG labeling , *LABELS - Abstract
All proteins interact with other cellular components to fulfill their function. While tremendous progress has been made in the identification of protein complexes, their assembly and dynamics remain difficult to characterize. Here, we present a high-throughput strategy to analyze the native assembly kinetics of protein complexes. We apply our approach to characterize the co-assembly for 320 pairs of nucleoporins (NUPs) constituting the ≈ 50 MDa nuclear pore complex (NPC) in yeast. Some NUPs co-assemble fast via rapid exchange whereas others require lengthy maturation steps. This reveals a hierarchical principle of NPC biogenesis where individual subcomplexes form on a minute timescale and then co-assemble from center to periphery in a ∼ 1 h-long maturation process. Intriguingly, the NUP Mlp1 stands out as joining very late and associating preferentially with aged NPCs. Our approach is readily applicable beyond the NPC, making it possible to analyze the intracellular dynamics of a variety of multiprotein assemblies. • KARMA, an approach to characterize maturation and dynamics of protein complexes • Metabolic labeling rates reveal complex maturation time and assembly order • Nuclear pore complex maturation follows a modular principle • Nucleoporins assemble from center to periphery on a minute to hour timescale A general approach to determining assembly kinetics for large protein architectures reveals the time scale and assembly sequence for the nuclear pore complex. [ABSTRACT FROM AUTHOR]
- Published
- 2020
- Full Text
- View/download PDF
17. A New Path through the Nuclear Pore.
- Author
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Gozalo, Alejandro and Capelson, Maya
- Subjects
- *
NUCLEAR pore complex , *NUCLEOCYTOPLASMIC interactions , *RNA export , *MESSENGER RNA , *CYTOLOGICAL research - Abstract
Knowing the configuration of the nuclear pore is essential for appreciating the underlying mechanisms of nucleo-cytoplasmic communication. Now, Fernandez-Martinez et al. present a high-resolution structure of the cytoplasmic nuclear pore-mRNA export holo-complex, challenging our textbook depiction of this massive membrane-embedded complex. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
18. Baby Nuclear Pores Grow Up Faster All the Time.
- Author
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Lusk, C. Patrick
- Subjects
- *
NUCLEAR pore complex , *ENDOPLASMIC reticulum , *BIOLOGICAL membranes , *INTERPHASE , *NUCLEAR membranes - Abstract
Annulate lamellae (AL) are stacked ER-derived membranes containing nuclear pore complex-like structures whose fate and function have remained a mystery. During the short interphase of early embryonic cells, AL are rapidly delivered into the nuclear envelope through fenestrations, highlighting the remarkable dynamics of the nuclear envelope. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
19. Nuclear Pores Assemble from Nucleoporin Condensates During Oogenesis.
- Author
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Hampoelz B, Schwarz A, Ronchi P, Bragulat-Teixidor H, Tischer C, Gaspar I, Ephrussi A, Schwab Y, and Beck M
- Subjects
- Active Transport, Cell Nucleus, Animals, Drosophila Proteins genetics, Drosophila melanogaster, Female, Microtubules metabolism, Molecular Chaperones genetics, Nuclear Pore Complex Proteins genetics, RNA, Messenger genetics, RNA, Messenger metabolism, ran GTP-Binding Protein genetics, ran GTP-Binding Protein metabolism, Drosophila Proteins metabolism, Molecular Chaperones metabolism, Nuclear Pore metabolism, Nuclear Pore Complex Proteins metabolism, Oogenesis
- Abstract
The molecular events that direct nuclear pore complex (NPC) assembly toward nuclear envelopes have been conceptualized in two pathways that occur during mitosis or interphase, respectively. In gametes and embryonic cells, NPCs also occur within stacked cytoplasmic membrane sheets, termed annulate lamellae (AL), which serve as NPC storage for early development. The mechanism of NPC biogenesis at cytoplasmic membranes remains unknown. Here, we show that during Drosophila oogenesis, Nucleoporins condense into different precursor granules that interact and progress into NPCs. Nup358 is a key player that condenses into NPC assembly platforms while its mRNA localizes to their surface in a translation-dependent manner. In concert, Microtubule-dependent transport, the small GTPase Ran and nuclear transport receptors regulate NPC biogenesis in oocytes. We delineate a non-canonical NPC assembly mechanism that relies on Nucleoporin condensates and occurs away from the nucleus under conditions of cell cycle arrest., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2019
- Full Text
- View/download PDF
20. Nuclear Pores Promote Lethal Prostate Cancer by Increasing POM121-Driven E2F1, MYC, and AR Nuclear Import.
- Author
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Rodriguez-Bravo, Veronica, Pippa, Raffaella, Song, Won-Min, Carceles-Cordon, Marc, Dominguez-Andres, Ana, Fujiwara, Naoto, Woo, Jungreem, Koh, Anna P., Ertel, Adam, Lokareddy, Ravi K., Cuesta-Dominguez, Alvaro, Kim, Rosa S., Rodriguez-Fernandez, Irene, Li, Peiyao, Gordon, Ronald, Hirschfield, Hadassa, Prats, Josep M., Reddy, E. Premkumar, Fatatis, Alessandro, and Petrylak, Daniel P.
- Subjects
- *
PROSTATE cancer , *NUCLEAR transport , *NUCLEAR pore complex , *ANDROGEN receptors , *NUCLEOPORINS - Abstract
Summary Nuclear pore complexes (NPCs) regulate nuclear-cytoplasmic transport, transcription, and genome integrity in eukaryotic cells. However, their functional roles in cancer remain poorly understood. We interrogated the evolutionary transcriptomic landscape of NPC components, nucleoporins (Nups), from primary to advanced metastatic human prostate cancer (PC). Focused loss-of-function genetic screen of top-upregulated Nups in aggressive PC models identified POM121 as a key contributor to PC aggressiveness. Mechanistically, POM121 promoted PC progression by enhancing importin-dependent nuclear transport of key oncogenic (E2F1, MYC) and PC-specific (AR-GATA2) transcription factors, uncovering a pharmacologically targetable axis that, when inhibited, decreased tumor growth, restored standard therapy efficacy, and improved survival in patient-derived pre-clinical models. Our studies molecularly establish a role of NPCs in PC progression and give a rationale for NPC-regulated nuclear import targeting as a therapeutic strategy for lethal PC. These findings may have implications for understanding how NPC deregulation contributes to the pathogenesis of other tumor types. [ABSTRACT FROM AUTHOR]
- Published
- 2018
- Full Text
- View/download PDF
21. Natively Unfolded FG Repeats Stabilize the Structure of the Nuclear Pore Complex.
- Author
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Onischenko, Evgeny, Tang, Jeffrey H., Andersen, Kasper R., Knockenhauer, Kevin E., Vallotton, Pascal, Derrer, Carina P., Kralt, Annemarie, Mugler, Christopher F., Chan, Leon Y., Schwartz, Thomas U., and Weis, Karsten
- Subjects
- *
NUCLEAR pore complex , *PHENYLALANINE , *NUCLEAR transport , *ORIGIN of life , *TISSUE scaffolds - Abstract
Summary Nuclear pore complexes (NPCs) are ∼100 MDa transport channels assembled from multiple copies of ∼30 nucleoporins (Nups). One-third of these Nups contain phenylalanine-glycine (FG)-rich repeats, forming a diffusion barrier, which is selectively permeable for nuclear transport receptors that interact with these repeats. Here, we identify an additional function of FG repeats in the structure and biogenesis of the yeast NPC. We demonstrate that GLFG-containing FG repeats directly bind to multiple scaffold Nups in vitro and act as NPC-targeting determinants in vivo . Furthermore, we show that the GLFG repeats of Nup116 function in a redundant manner with Nup188, a nonessential scaffold Nup, to stabilize critical interactions within the NPC scaffold needed for late steps of NPC assembly. Our results reveal a previously unanticipated structural role for natively unfolded GLFG repeats as Velcro to link NPC subcomplexes and thus add a new layer of connections to current models of the NPC architecture. [ABSTRACT FROM AUTHOR]
- Published
- 2017
- Full Text
- View/download PDF
22. Single-Molecule Real-Time 3D Imaging of the Transcription Cycle by Modulation Interferometry.
- Author
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Wang, Guanshi, Hauver, Jesse, Thomas, Zachary, Darst, Seth A., and Pertsinidis, Alexandros
- Subjects
- *
INTERFEROMETRY , *RNA polymerases , *SIGMA factor (Transcription factor) , *ESCHERICHIA coli , *GENETIC transcription , *CHEMICAL kinetics - Abstract
Summary Many essential cellular processes, such as gene control, employ elaborate mechanisms involving the coordination of large, multi-component molecular assemblies. Few structural biology tools presently have the combined spatial-temporal resolution and molecular specificity required to capture the movement, conformational changes, and subunit association-dissociation kinetics, three fundamental elements of how such intricate molecular machines work. Here, we report a 3D single-molecule super-resolution imaging study using modulation interferometry and phase-sensitive detection that achieves <2 nm axial localization precision, well below the few-nanometer-sized individual protein components. To illustrate the capability of this technique in probing the dynamics of complex macromolecular machines, we visualize the movement of individual multi-subunit E. coli RNA polymerases through the complete transcription cycle, dissect the kinetics of the initiation-elongation transition, and determine the fate of σ 70 initiation factors during promoter escape. Modulation interferometry sets the stage for single-molecule studies of several hitherto difficult-to-investigate multi-molecular transactions that underlie genome regulation. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
23. An Ancient, Unified Mechanism for Metformin Growth Inhibition in C. elegans and Cancer.
- Author
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Wu, Lianfeng, Zhou, Ben, Oshiro-Rapley, Noriko, Li, Man, Paulo, Joao A., Webster, Christopher M., Mou, Fan, Kacergis, Michael C., Talkowski, Michael E., Carr, Christopher E., Gygi, Steven P., Zheng, Bin, and Soukas, Alexander A.
- Subjects
- *
METFORMIN , *CANCER treatment , *GENETIC testing , *LIFE spans , *MTOR protein - Abstract
Summary Metformin has utility in cancer prevention and treatment, though the mechanisms for these effects remain elusive. Through genetic screening in C. elegans , we uncover two metformin response elements: the nuclear pore complex (NPC) and acyl-CoA dehydrogenase family member-10 (ACAD10). We demonstrate that biguanides inhibit growth by inhibiting mitochondrial respiratory capacity, which restrains transit of the RagA-RagC GTPase heterodimer through the NPC. Nuclear exclusion renders RagC incapable of gaining the GDP-bound state necessary to stimulate mTORC1. Biguanide-induced inactivation of mTORC1 subsequently inhibits growth through transcriptional induction of ACAD10. This ancient metformin response pathway is conserved from worms to humans. Both restricted nuclear pore transit and upregulation of ACAD10 are required for biguanides to reduce viability in melanoma and pancreatic cancer cells, and to extend C. elegans lifespan. This pathway provides a unified mechanism by which metformin kills cancer cells and extends lifespan, and illuminates potential cancer targets. PaperClip [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
24. ESCRTs Take on a Job in Surveillance.
- Author
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Odorizzi, Greg
- Subjects
- *
NUCLEAR pore complex , *MEMBRANE proteins , *ENDOSOMES , *CELL membranes , *CELL physiology - Abstract
Nuclear pore assembly can go awry, but how the cell handles defective intermediates has been an ongoing question. In this issue, Lusk and colleagues describe a surveillance pathway during nuclear pore assembly and, in doing so, identify a new role for proteins that function at the endosome and plasma membrane. [ABSTRACT FROM AUTHOR]
- Published
- 2014
- Full Text
- View/download PDF
25. Surface Properties Determining Passage Rates of Proteins through Nuclear Pores.
- Author
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Frey S, Rees R, Schünemann J, Ng SC, Fünfgeld K, Huyton T, and Görlich D
- Subjects
- Amino Acid Motifs, Binding Sites, Green Fluorescent Proteins genetics, Green Fluorescent Proteins metabolism, HeLa Cells, Humans, Hydrophobic and Hydrophilic Interactions, Microscopy, Confocal, Mutagenesis, Site-Directed, Nuclear Pore Complex Proteins chemistry, Nuclear Pore Complex Proteins genetics, Protein Domains, Protein Structure, Quaternary, Recombinant Proteins biosynthesis, Recombinant Proteins chemistry, Recombinant Proteins isolation & purification, Surface Properties, Active Transport, Cell Nucleus physiology, Nuclear Pore metabolism, Nuclear Pore Complex Proteins metabolism
- Abstract
Nuclear pore complexes (NPCs) conduct nucleocytoplasmic transport through an FG domain-controlled barrier. We now explore how surface-features of a mobile species determine its NPC passage rate. Negative charges and lysines impede passage. Hydrophobic residues, certain polar residues (Cys, His), and, surprisingly, charged arginines have striking translocation-promoting effects. Favorable cation-π interactions between arginines and FG-phenylalanines may explain this apparent paradox. Application of these principles to redesign the surface of GFP resulted in variants that show a wide span of transit rates, ranging from 35-fold slower than wild-type to ∼500 times faster, with the latter outpacing even naturally occurring nuclear transport receptors (NTRs). The structure of a fast and particularly FG-specific GFP
NTR variant illustrates how NTRs can expose multiple regions for binding hydrophobic FG motifs while evading non-specific aggregation. Finally, we document that even for NTR-mediated transport, the surface-properties of the "passively carried" cargo can strikingly affect the translocation rate., (Copyright © 2018 Elsevier Inc. All rights reserved.)- Published
- 2018
- Full Text
- View/download PDF
26. Nuclear Pore Permeabilization Is a Convergent Signaling Event in Effector-Triggered Immunity.
- Author
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Gu, Yangnan, Zebell, Sophia G., Liang, Zizhen, Wang, Shui, Kang, Byung-Ho, and Dong, Xinnian
- Subjects
- *
NUCLEAR pore complex , *ANTIGEN receptors , *IMMUNITY , *CELLULAR signal transduction , *IMMUNOREGULATION - Abstract
Summary Nuclear transport of immune receptors, signal transducers, and transcription factors is an essential regulatory mechanism for immune activation. Whether and how this process is regulated at the level of the nuclear pore complex (NPC) remains unclear. Here, we report that CPR5, which plays a key inhibitory role in effector-triggered immunity (ETI) and programmed cell death (PCD) in plants, is a novel transmembrane nucleoporin. CPR5 associates with anchors of the NPC selective barrier to constrain nuclear access of signaling cargos and sequesters cyclin-dependent kinase inhibitors (CKIs) involved in ETI signal transduction. Upon activation by immunoreceptors, CPR5 undergoes an oligomer to monomer conformational switch, which coordinates CKI release for ETI signaling and reconfigures the selective barrier to allow significant influx of nuclear signaling cargos through the NPC. Consequently, these coordinated NPC actions result in simultaneous activation of diverse stress-related signaling pathways and constitute an essential regulatory mechanism specific for ETI/PCD induction. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
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