scholarly journals ELYS coordinates NF-κB pathway dynamics during development in Drosophila

2018 ◽  
Author(s):  
Saurabh Jayesh Kumar Mehta ◽  
Vimlesh Kumar ◽  
Ram Kumar Mishra
Keyword(s):  
Nuclear Lamina ◽  
Nucleocytoplasmic Transport ◽  
Structural Features ◽  
Cellular Level ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Nuclear Pores ◽  
Summary Statement ◽  
Quality Check ◽  
Pore Complexes

Summary StatementELYS, a nucleoporin spatiotemporally regulates NF-κB pathway dynamics during development in Drosophila and its misregulation in post-embryonic stages leads to apoptosis mediated abnormalities.AbstractNuclear pores are the exclusive conduit to facilitate the nucleocytoplasmic transport in a precisely regulated manner. ELYS, a constituent protein of nuclear pores, initiates assembly of nuclear pore complexes (NPCs) into functional nuclear pores towards the end of mitosis. Using cellular, molecular and genetic tools, here, we report that ELYS orthologue (dElys) plays critical roles during Drosophila development. Through in silico analyses, we find all conserved structural features in dElys except for the presence of non-canonical AT-hook motif strongly binding with DNA. dElys localized to nuclear rim in interphase cells, but during mitosis, it was present on chromatin. RNAi mediated depletion of dElys leads to aberrant development and defects in the nuclear lamina and NPCs assembly at the cellular level. Furthermore, we demonstrate that in dElys depletion NF-κB is activated and accumulates inside the nucleus which results in illimed expression of critical molecules. dElys depletion sustains NF-κB into the nucleus in post-embryonic stages. Prolonged NF-κB inside nucleus induces apoptosis in response to hitherto unknown quality check mechanism and highlights on the under-appreciated apoptotic paradigm of NF-κB pathway.

scholarly journals Motor-driven motility of fungal nuclear pores organizes chromosomes and fosters nucleocytoplasmic transport

2012 ◽  
Vol 198 (3) ◽  
pp. 343-355 ◽  
Author(s):  
Gero Steinberg ◽  
Martin Schuster ◽  
Ulrike Theisen ◽  
Sreedhar Kilaru ◽  
Andrew Forge ◽  
...  
Keyword(s):  
Nuclear Lamina ◽  
Ustilago Maydis ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Nuclear Pores ◽  
Gfp Reporter ◽  
Import And Export ◽  
Reporter Proteins ◽  
Pore Complexes

Exchange between the nucleus and the cytoplasm is controlled by nuclear pore complexes (NPCs). In animals, NPCs are anchored by the nuclear lamina, which ensures their even distribution and proper organization of chromosomes. Fungi do not possess a lamina and how they arrange their chromosomes and NPCs is unknown. Here, we show that motor-driven motility of NPCs organizes the fungal nucleus. In Ustilago maydis, Aspergillus nidulans, and Saccharomyces cerevisiae fluorescently labeled NPCs showed ATP-dependent movements at ∼1.0 µm/s. In S. cerevisiae and U. maydis, NPC motility prevented NPCs from clustering. In budding yeast, NPC motility required F-actin, whereas in U. maydis, microtubules, kinesin-1, and dynein drove pore movements. In the latter, pore clustering resulted in chromatin organization defects and led to a significant reduction in both import and export of GFP reporter proteins. This suggests that fungi constantly rearrange their NPCs and corresponding chromosomes to ensure efficient nuclear transport and thereby overcome the need for a structural lamina.

scholarly journals Drosophila ELYS regulates Dorsal dynamics during development

2020 ◽  
Vol 295 (8) ◽  
pp. 2421-2437 ◽  
Author(s):  
Saurabh Jayesh Kumar Mehta ◽  
Vimlesh Kumar ◽  
Ram Kumar Mishra
Keyword(s):  
Target Genes ◽  
Developmental Stages ◽  
Nuclear Lamina ◽  
Structural Features ◽  
Cellular Level ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Nuclear Pores ◽  
Developmental Abnormalities ◽  
Larval Stages

Embryonic large molecule derived from yolk sac (ELYS) is a constituent protein of nuclear pores. It initiates assembly of nuclear pore complexes into functional nuclear pores toward the end of mitosis. Using cellular, molecular, and genetic tools, including fluorescence and Electron microscopy, quantitative PCR, and RNAi-mediated depletion, we report here that the ELYS ortholog (dElys) plays critical roles during Drosophila development. dElys localized to the nuclear rim in interphase cells, but during mitosis it was absent from kinetochores and enveloped chromatin. We observed that RNAi-mediated dElys depletion leads to aberrant development and, at the cellular level, to defects in the nuclear pore and nuclear lamina assembly. Further genetic analyses indicated that dElys depletion re-activates the Dorsal (NF-κB) pathway during late larval stages. Re-activated Dorsal caused untimely expression of the Dorsal target genes in the post-embryonic stages. We also demonstrate that activated Dorsal triggers apoptosis during later developmental stages by up-regulating the pro-apoptotic genes reaper and hid. The apoptosis induced by Reaper and Hid was probably the underlying cause for developmental abnormalities observed upon dElys depletion. Moreover, we noted that dElys has conserved structural features, but contains a noncanonical AT-hook–like motif through which it strongly binds to DNA. Together, our results uncover a novel epistatic interaction that regulates Dorsal dynamics by dElys during development.

scholarly journals Flexible Gates: Dynamic Topologies and Functions for FG Nucleoporins in Nucleocytoplasmic Transport

2009 ◽  
Vol 8 (12) ◽  
pp. 1814-1827 ◽  
Author(s):  
Laura J. Terry ◽  
Susan R. Wente
Keyword(s):  
Nuclear Envelope ◽  
Regulation Of Gene Expression ◽  
Nucleocytoplasmic Transport ◽  
Structural Features ◽  
Nuclear Pore ◽  
Permeability Barrier ◽  
Nuclear Pore Complexes ◽  
Cellular Functions ◽  
Essential Components ◽  
Pore Complexes

ABSTRACT The nuclear envelope is a physical barrier between the nucleus and cytoplasm and, as such, separates the mechanisms of transcription from translation. This compartmentalization of eukaryotic cells allows spatial regulation of gene expression; however, it also necessitates a mechanism for transport between the nucleus and cytoplasm. Macromolecular trafficking of protein and RNA occurs exclusively through nuclear pore complexes (NPCs), specialized channels spanning the nuclear envelope. A novel family of NPC proteins, the FG-nucleoporins (FG-Nups), coordinates and potentially regulates NPC translocation. The extensive repeats of phenylalanine-glycine (FG) in each FG-Nup directly bind to shuttling transport receptors moving through the NPC. In addition, FG-Nups are essential components of the nuclear permeability barrier. In this review, we discuss the structural features, cellular functions, and evolutionary conservation of the FG-Nups.

scholarly journals In Vivo Dynamics of Nuclear Pore Complexes in Yeast

1997 ◽  
Vol 136 (6) ◽  
pp. 1185-1199 ◽  
Author(s):  
Mirella Bucci ◽  
Susan R. Wente
Keyword(s):  
Nuclear Envelope ◽  
Recipient Cell ◽  
Nucleocytoplasmic Transport ◽  
Yeast Cells ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Central Location ◽  
Nuclear Membranes ◽  
Mutant Strains ◽  
Pore Complexes

While much is known about the role of nuclear pore complexes (NPCs) in nucleocytoplasmic transport, the mechanism of NPC assembly into pores formed through the double lipid bilayer of the nuclear envelope is not well defined. To investigate the dynamics of NPCs, we developed a live-cell assay in the yeast Saccharomyces cerevisiae. The nucleoporin Nup49p was fused to the green fluorescent protein (GFP) of Aequorea victoria and expressed in nup49 null haploid yeast cells. When the GFP–Nup49p donor cell was mated with a recipient cell harboring only unlabeled Nup49p, the nuclei fused as a consequence of the normal mating process. By monitoring the distribution of the GFP–Nup49p, we could assess whether NPCs were able to move from the donor section of the nuclear envelope to that of the recipient nucleus. We observed that fluorescent NPCs moved and encircled the entire nucleus within 25 min after fusion. When assays were done in mutant kar1-1 strains, where nuclear fusion does not occur, GFP–Nup49p appearance in the recipient nucleus occurred at a very slow rate, presumably due to new NPC biogenesis or to exchange of GFP– Nup49p into existing recipient NPCs. Interestingly, in a number of existing mutant strains, NPCs are clustered together at permissive growth temperatures. This has been explained with two different hypotheses: by movement of NPCs through the double nuclear membranes with subsequent clustering at a central location; or, alternatively, by assembly of all NPCs at a central location (such as the spindle pole body) with NPCs in mutant cells unable to move away from this point. Using the GFP–Nup49p system with a mutant in the NPCassociated factor Gle2p that exhibits formation of NPC clusters only at 37°C, it was possible to distinguish between these two models for NPC dynamics. GFP– Nup49p-labeled NPCs, assembled at 23°C, moved into clusters when the cells were shifted to growth at 37°C. These results indicate that NPCs can move through the double nuclear membranes and, moreover, can do so to form NPC clusters in mutant strains. Such clusters may result by releasing NPCs from a nuclear tether, or by disappearance of a protein that normally prevents pore aggregation. This system represents a novel approach for identifying regulators of NPC assembly and movement in the future.

scholarly journals O-GlcNAc modification of nuclear pore complexes accelerates bi-directional transport

2020 ◽  
Author(s):  
Tae Yeon Yoo ◽  
Timothy J Mitchison
Keyword(s):  
Nuclear Import ◽  
Nuclear Transport ◽  
Super Resolution ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Permeability Barrier ◽  
Nuclear Pore Complexes ◽  
Facilitated Diffusion ◽  
Import And Export ◽  
Pore Complexes

AbstractMacromolecular transport across the nuclear envelope depends on facilitated diffusion through nuclear pore complexes (NPCs). The interior of NPCs contains a permeability barrier made of phenylalanine-glycine (FG) repeat domains that selectively facilitates the permeation of cargoes bound to nuclear transport receptors (NTRs). FG repeats in NPC are a major site of O-linked N-acetylglucosamine (O-GlcNAc) modification, but the functional role of this modification in nucleocytoplasmic transport is unclear. We developed high-throughput assays based on optogenetic probes to quantify the kinetics of nuclear import and export in living human cells. We found that increasing O-GlcNAc modification of the NPC accelerated NTR-facilitated nucleocytoplasmic transport of proteins in both directions, and decreasing modification slowed transport. Super-resolution imaging revealed strong enrichment of O-GlcNAc at the FG-repeat barrier. O-GlcNAc modification also accelerated passive permeation of a small, inert protein through NPCs. We conclude that O-GlcNAc modification accelerates nucleocytoplasmic transport by enhancing the non-specific permeability the FG-repeat barrier, perhaps by steric inhibition of interactions between FG repeats.SummaryNuclear pore complexes mediate nuclear transport and are highly modified with O-linked N-acetylglucosamine (O-GlcNAc) on FG repeat domains. Using a new quantitative live-cell imaging assay, Yoo and Mitchison demonstrate acceleration of nuclear import and export by O-GlcNAc modification.

scholarly journals Concentration-dependent lamin assembly and its roles in the localization of other nuclear proteins

2014 ◽  
Vol 25 (8) ◽  
pp. 1287-1297 ◽  
Author(s):  
Yuxuan Guo ◽  
Youngjo Kim ◽  
Takeshi Shimi ◽  
Robert D. Goldman ◽  
Yixian Zheng
Keyword(s):  
Nuclear Lamina ◽  
Nuclear Pore ◽  
Lamin A ◽  
Nuclear Pore Complexes ◽  
Developmental Defects ◽  
Protein Levels ◽  
Lamin B1 ◽  
Mouse Cells ◽  
Pore Complexes ◽  
And Function

The nuclear lamina (NL) consists of lamin polymers and proteins that bind to the polymers. Disruption of NL proteins such as lamin and emerin leads to developmental defects and human diseases. However, the expression of multiple lamins, including lamin-A/C, lamin-B1, and lamin-B2, in mammals has made it difficult to study the assembly and function of the NL. Consequently, it has been unclear whether different lamins depend on one another for proper NL assembly and which NL functions are shared by all lamins or are specific to one lamin. Using mouse cells deleted of all or different combinations of lamins, we demonstrate that the assembly of each lamin into the NL depends primarily on the lamin concentration present in the nucleus. When expressed at sufficiently high levels, each lamin alone can assemble into an evenly organized NL, which is in turn sufficient to ensure the even distribution of the nuclear pore complexes. By contrast, only lamin-A can ensure the localization of emerin within the NL. Thus, when investigating the role of the NL in development and disease, it is critical to determine the protein levels of relevant lamins and the intricate shared or specific lamin functions in the tissue of interest.

The structure and interactions of components of nuclear envelopes from Xenopus oocyte germinal vesicles observed by heavy metal shadowing

1988 ◽  
Vol 90 (3) ◽  
pp. 409-423 ◽  
Author(s):  
MURRAY STEWART ◽  
SUE WHYTOCK
Keyword(s):  
Xenopus Oocyte ◽  
Freeze Drying ◽  
Nuclear Lamina ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Air Drying ◽  
Freeze Dried ◽  
Germinal Vesicles ◽  
Pore Complexes ◽  
Different Levels

We have examined the structure of the nuclear envelope of oocytes of Xenopus laevis by electronmicroscopy of metal-shadowed specimens. Material was prepared by either freeze-drying ora rapid protocol using air-drying after dehydration in ethanol followed by amyl acetate. These methods emphasized different aspects of the structure and enabled an integrated view of the arrangement of nuclear pore complexes, nuclear lamina and pore-connecting fibrils to be assembled. In specimens prepared by either air drying or freeze-drying, the lamina meshwork beneath the nuclear face of the envelope was well preserved, but the fine structure of the nuclearpores was superior in freeze-dried preparations. Both methods also showed pore-connecting fibrils that were clearly not components of the lamina. By using stereo pairs, we established criteria for recognizing the cytoplasmic and nucleoplasmic faces of shadowed nuclear envelopes. These views also enabled us to identify the levels atwhich different fibrous components were attached to the pores. In particular, we were able to visualize the nuclear lamina fibres and poreconnecting fibrils simultaneously and show that they attach to the pore complexes at different levels. We supplemented this work by using arange of treatments to disrupt the nuclear envelopes lightly and gained several insights into this structure as a result. Sometimes pore complexes and their connecting fibrils were stripped from the envelope. This enabled a clearer view of these connections to be obtained without the lamina present. Moreover, in some conditions, the nuclearpore complexes and fibrous lamina began to disintegrate, there by showing some of the morphological components from which they were assembled.

scholarly journals Nuclear envelope organization in Dictyostelium discoideum

2019 ◽  
Vol 63 (8-9-10) ◽  
pp. 509-519 ◽  
Author(s):  
Petros Batsios ◽  
Ralph Gräf ◽  
Michael P. Koonce ◽  
Denis A. Larochelle ◽  
Irene Meyer
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Nuclear Lamina ◽  
Major Constituent ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Linc Complex ◽  
Family Protein ◽  
Import And Export ◽  
Pore Complexes

The nuclear envelope consists of the outer and the inner nuclear membrane, the nuclear lamina and the nuclear pore complexes, which regulate nuclear import and export. The major constituent of the nuclear lamina of Dictyostelium is the lamin NE81. It can form filaments like B-type lamins and it interacts with Sun1, as well as with the LEM/HeH-family protein Src1. Sun1 and Src1 are nuclear envelope transmembrane proteins involved in the centrosome-nucleus connection and nuclear envelope stability at the nucleolar regions, respectively. In conjunction with a KASH-domain protein, Sun1 usually forms a so-called LINC complex. Two proteins with functions reminiscent of KASH-domain proteins at the outer nuclear membrane of Dictyostelium are known; interaptin which serves as an actin connector and the kinesin Kif9 which plays a role in the microtubule-centrosome connector. However, both of these lack the conserved KASH-domain. The link of the centrosome to the nuclear envelope is essential for the insertion of the centrosome into the nuclear envelope and the appropriate spindle formation. Moreover, centrosome insertion is involved in permeabilization of the mitotic nucleus, which ensures access of tubulin dimers and spindle assembly factors. Our recent progress in identifying key molecular players at the nuclear envelope of Dictyostelium promises further insights into the mechanisms of nuclear envelope dynamics.

scholarly journals Nuclear Pore Proteins in Regulation of Chromatin State

Cells ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 1414 ◽  
Author(s):  
Terra M. Kuhn ◽  
Maya Capelson
Keyword(s):  
Nucleosome Occupancy ◽  
Regulation Of Gene Expression ◽  
Nucleocytoplasmic Transport ◽  
Chromatin State ◽  
Nuclear Pore ◽  
Regulation Of Transcription ◽  
Nuclear Pore Complexes ◽  
Functional Connection ◽  
Histone Modifiers ◽  
Pore Complexes

Nuclear pore complexes (NPCs) are canonically known to regulate nucleocytoplasmic transport. However, research efforts over the last decade have demonstrated that NPCs and their constituent nucleoporins (Nups) also interact with the genome and perform important roles in regulation of gene expression. It has become increasingly clear that many Nups execute these roles specifically through regulation of chromatin state, whether through interactions with histone modifiers and downstream changes in post-translational histone modifications, or through relationships with chromatin-remodeling proteins that can result in physical changes in nucleosome occupancy and chromatin compaction. This review focuses on these findings, highlighting the functional connection between NPCs/Nups and regulation of chromatin structure, and how this connection can manifest in regulation of transcription.

scholarly journals Nuclear pore complexes form immobile networks and have a very low turnover in live mammalian cells

2001 ◽  
Vol 154 (1) ◽  
pp. 71-84 ◽  
Author(s):  
Nathalie Daigle ◽  
Joël Beaudouin ◽  
Lisa Hartnell ◽  
Gabriela Imreh ◽  
Einar Hallberg ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Fluorescent Protein ◽  
Nuclear Lamina ◽  
Nuclear Pore ◽  
Global Changes ◽  
Nuclear Pore Complexes ◽  
Annulate Lamellae ◽  
Lamin B1 ◽  
Green Fluorescent ◽  
Pore Complexes

The nuclear pore complex (NPC) and its relationship to the nuclear envelope (NE) was characterized in living cells using POM121–green fluorescent protein (GFP) and GFP-Nup153, and GFP–lamin B1. No independent movement of single pore complexes was found within the plane of the NE in interphase. Only large arrays of NPCs moved slowly and synchronously during global changes in nuclear shape, strongly suggesting mechanical connections which form an NPC network. The nuclear lamina exhibited identical movements. NPC turnover measured by fluorescence recovery after photobleaching of POM121 was less than once per cell cycle. Nup153 association with NPCs was dynamic and turnover of this nucleoporin was three orders of magnitude faster. Overexpression of both nucleoporins induced the formation of annulate lamellae (AL) in the endoplasmic reticulum (ER). Turnover of AL pore complexes was much higher than in the NE (once every 2.5 min). During mitosis, POM121 and Nup153 were completely dispersed and mobile in the ER (POM121) or cytosol (Nup153) in metaphase, and rapidly redistributed to an immobilized pool around chromatin in late anaphase. Assembly and immobilization of both nucleoporins occurred before detectable recruitment of lamin B1, which is thus unlikely to mediate initiation of NPC assembly at the end of mitosis.

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