scholarly journals JLP-centrosome is essential for the microtubule-mediated nucleocytoplasmic transport induced by extracellular stimuli

2019 ◽  
Vol 5 (8) ◽  
pp. eaav0318 ◽  
Author(s):  
Clement M. Lee ◽  
Ken Aizawa ◽  
Joshua Jiang ◽  
Sam K. P. Kung ◽  
Rinku Jain
Keyword(s):  
Intracellular Transport ◽  
Nuclear Translocation ◽  
Nucleocytoplasmic Transport ◽  
Scaffolding Proteins ◽  
Nuclear Pores ◽  
Focus Formation ◽  
Microtubule Array ◽  
Localization Domain ◽  
Nucleus Shape ◽  
Extracellular Stimuli

JLP belongs to the JIP family whose members serve as scaffolding proteins that link motor proteins and their cargo for intracellular transport. Although JLP is mainly cytoplasmic, it accumulates as a focus in the perinuclear region when stimulated by extracellular stimuli. Focus formation, which changes the nucleus shape and concentrates the nuclear pores, depends on p38MAPK activation and the dynein retrograde motor protein complex. Extracellular stimuli trigger the tethering of PLK1 to the centrosome by JLP, leading to centrosome maturation and microtubule array formation. The centrosome localization domain of JLP is important for the binding of the centrosome and the formation of the JLP focus and the microtubule array. Furthermore, the formation of the JLP focus and the microtubule array is interdependent and important for the transport of NF-κB p65 to the nucleus and its unloading therein. In conclusion, JLP exhibits multiple functions in the nuclear translocation of NF-κB p65.

scholarly journals Nuclear ERK: Mechanism of Translocation, Substrates, and Role in Cancer

2019 ◽  
Vol 20 (5) ◽  
pp. 1194 ◽  
Author(s):  
Galia Maik-Rachline ◽  
Avital Hacohen-Lev-Ran ◽  
Rony Seger
Keyword(s):  
Nuclear Translocation ◽  
Resting Cells ◽  
Nuclear Pores ◽  
Cellular Processes ◽  
Extracellular Signal ◽  
Anti Cancer ◽  
Proliferation And Differentiation ◽  
The One ◽  
Signaling Components ◽  
Proof Of Principle

The extracellular signal-regulated kinases 1/2 (ERK) are central signaling components that regulate stimulated cellular processes such as proliferation and differentiation. When dysregulated, these kinases participate in the induction and maintenance of various pathologies, primarily cancer. While ERK is localized in the cytoplasm of resting cells, many of its substrates are nuclear, and indeed, extracellular stimulation induces a rapid and robust nuclear translocation of ERK. Similarly to other signaling components that shuttle to the nucleus upon stimulation, ERK does not use the canonical importinmechanism of nuclear translocation. Rather, it has its own unique nuclear translocation signal (NTS) that interacts with importin7 to allow stimulated shuttling via the nuclear pores. Prevention of the nuclear translocation inhibits proliferation of B-Raf- and N/K-Ras-transformed cancers. This effect is distinct from the one achieved by catalytic Raf and MEK inhibitors used clinically, as cells treated with the translocation inhibitors develop resistance much more slowly. In this review, we describe the mechanism of ERK translocation, present all its nuclear substrates, discuss its role in cancer and compare its translocation to the translocation of other signaling components. We also present proof of principle data for the use of nuclear ERK translocation as an anti-cancer target. It is likely that the prevention of nuclear ERK translocation will eventually serve as a way to combat Ras and Raf transformed cancers with less side-effects than the currently used drugs.

scholarly journals Nuclear import of isoforms of the cytomegalovirus kinase pUL97 is mediated by differential activity of NLS1 and NLS2 both acting through classical importin-α binding

2012 ◽  
Vol 93 (8) ◽  
pp. 1756-1768 ◽  
Author(s):  
Rike Webel ◽  
Sara M. Ø. Solbak ◽  
Christian Held ◽  
Jens Milbradt ◽  
Andrea Groß ◽  
...  
Keyword(s):  
Virus Replication ◽  
Nuclear Localization ◽  
Evaluation System ◽  
Nuclear Translocation ◽  
Helical Structure ◽  
Nucleocytoplasmic Transport ◽  
Resonance Spectroscopy ◽  
Translational Initiation ◽  
Importin Α ◽  
Arginine Residues

The multifunctional protein kinase pUL97 of human cytomegalovirus (HCMV) strongly determines the efficiency of virus replication. Previously, the existence of two pUL97 isoforms that arise from alternative translational initiation and show a predominant nuclear localization was described. Two bipartite nuclear localization sequences, NLS1 and NLS2, were identified in the N terminus of the large isoform, whilst the small isoform exclusively contained NLS2. The current study found the following: (i) pUL97 nuclear localization in HCMV-infected primary fibroblasts showed accumulations in virus replication centres and other nuclear sections; (ii) in a quantitative evaluation system for NLS activity, the large isoform showed higher efficiency of nuclear translocation than the small isoform; (iii) NLS1 was mapped to aa 6–35 and NLS2 to aa 190–213; (iv) using surface plasmon resonance spectroscopy, the binding of both NLS1 and NLS2 to human importin-α was demonstrated, stressing the importance of individual arginine residues in the bipartite consensus motifs; (v) nuclear magnetic resonance spectroscopy of pUL97 peptides confirmed an earlier statement about the functional requirement of NLS1 embedding into an intact α-helical structure; and (vi) a bioinformatics investigation of the solvent-accessible surface suggested a high accessibility of NLS1 and an isoform-specific, variable accessibility of NLS2 for interaction with importin-α. Thus, the nucleocytoplasmic transport mechanism of the isoforms appeared to be differentially regulated, and this may have consequences for isoform-dependent functions of pUL97 during virus replication.

scholarly journals Nuclear Localization Sequences in Cytomegalovirus Capsid Assembly Proteins (UL80 Proteins) Are Required for Virus Production: Inactivating NLS1, NLS2, or Both Affects Replication to Strikingly Different Extents

2008 ◽  
Vol 82 (11) ◽  
pp. 5381-5389 ◽  
Author(s):  
Nang L. Nguyen ◽  
Amy N. Loveland ◽  
Wade Gibson
Keyword(s):  
Nuclear Localization ◽  
Infectious Virus ◽  
Nuclear Translocation ◽  
Virus Production ◽  
Wild Type Virus ◽  
Capsid Assembly ◽  
Scaffolding Proteins ◽  
Hcmv Replication ◽  
Protease Precursor ◽  
Nuclear Localization Sequences

ABSTRACT Scaffolding proteins of spherical prokaryotic and eukaryotic viruses have critical roles in capsid assembly. The primary scaffolding components of cytomegalovirus, called the assembly protein precursor (pAP, pUL80.5) and the maturational protease precursor (pPR, pUL80a), contain two nuclear localization sequences (NLS1 and NLS2), at least one of which is required in coexpression experiments to translocate the major capsid protein (MCP, pUL85) into the nucleus. In the work reported here, we have mutated NLS1 and NLS2, individually or together, in human cytomegalovirus (HCMV, strain AD169) bacmid-derived viruses to test their effects on virus replication. Consistent with results from earlier transfection/coexpression experiments, both single-mutant bacmids gave rise to infectious virus but the double mutant did not. In comparisons with the wild-type virus, both mutants showed slower cell-to-cell spread; decreased yields of infectious virus (3-fold lower for NLS1− and 140-fold lower for NLS2−); reduced efficiency of pAP, pPR, and MCP nuclear translocation (sixfold lower for NLS1− and eightfold lower for NLS2−); increased amounts of a 120-kDa MCP fragment; and reduced numbers of intranuclear capsids. All effects were more severe for the NLS2− mutant than the NLS1− mutant, and a distinguishing feature of cells infected with the NLS2− mutant was the accumulation of large, UL80 protein-containing structures within the nucleus. We conclude that these NLS assist in the nuclear translocation of MCP during HCMV replication and that NLS2, which is unique to the betaherpesvirus UL80 homologs, may have additional involvements during replication.

scholarly journals The Nsp1p Carboxy-Terminal Domain Is Organized into Functionally Distinct Coiled-Coil Regions Required for Assembly of Nucleoporin Subcomplexes and Nucleocytoplasmic Transport

2001 ◽  
Vol 21 (23) ◽  
pp. 7944-7955 ◽  
Author(s):  
Susanne M. Bailer ◽  
Carolin Balduf ◽  
Ed Hurt
Keyword(s):  
Protein Import ◽  
Coiled Coil ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Temperature Sensitive ◽  
Nuclear Pores ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Carboxy Terminal

ABSTRACT Nucleoporin Nsp1p, which has four predicted coiled-coil regions (coils 1 to 4) in the essential carboxy-terminal domain, is unique in that it is part of two distinct nuclear pore complex (NPC) subcomplexes, Nsp1p-Nup57p-Nup49p-Nic96p and Nsp1p-Nup82p-Nup159p. As shown by in vitro reconstitution, coiled-coil region 2 (residues 673 to 738) is sufficient to form heterotrimeric core complexes and can bind either Nup57p or Nup82p. Accordingly, interaction of Nup82p with Nsp1p coil 2 is competed by excess Nup57p. Strikingly, coil 3 and 4 mutants are still assembled into the core Nsp1p-Nup57p-Nup49p complex but no longer associate with Nic96p. Consistently, the Nsp1p-Nup57p-Nup49p core complex dissociates from the nuclear pores in nsp1coil 3 and 4 mutant cells, and as a consequence, defects in nuclear protein import are observed. Finally, the nsp1-L640Stemperature-sensitive mutation, which maps in coil 1, leads to a strong nuclear mRNA export defect. Thus, distinct coiled-coil regions within Nsp1p-C have separate functions that are related to the assembly of different NPC subcomplexes, nucleocytoplasmic transport, and incorporation into the nuclear pores.

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 17β-Estradiol Inhibits Inflammatory Gene Expression by Controlling NF-κB Intracellular Localization

2005 ◽  
Vol 25 (8) ◽  
pp. 2957-2968 ◽  
Author(s):  
Serena Ghisletti ◽  
Clara Meda ◽  
Adriana Maggi ◽  
Elisabetta Vegeto
Keyword(s):  
Inflammatory Response ◽  
Intracellular Transport ◽  
Time Course ◽  
Nuclear Translocation ◽  
De Novo ◽  
Intracellular Localization ◽  
Mitogen Activated Protein Kinase ◽  
Proinflammatory Mediators ◽  
17Β Estradiol ◽  
Anti Inflammatory

ABSTRACT Estrogen is an immunoregulatory agent, in that hormone deprivation increases while 17β-estradiol (E2) administration blocks the inflammatory response; however, the underlying mechanism is still unknown. The transcription factor p65/relA, a member of the nuclear factor κB (NF-κB) family, plays a major role in inflammation and drives the expression of proinflammatory mediators. Here we report a novel mechanism of action of E2 in inflammation. We observe that in macrophages E2 blocks lipopolysaccharide-induced DNA binding and transcriptional activity of p65 by preventing its nuclear translocation. This effect is selectively activated in macrophages to prevent p65 activation by inflammatory agents and extends to other members of the NF-κB family, including c-Rel and p50. We observe that E2 activates a rapid and persistent response that involves the activation of phosphatidylinositol 3-kinase, without requiring de novo protein synthesis or modifying Iκ-Bα degradation and mitogen-activated protein kinase activation. Using a time course experiment and the microtubule-disrupting agent nocodazole, we observe that the hormone inhibits p65 intracellular transport to the nucleus. This activity is selectively mediated by estrogen receptor alpha (ERα) and not ERβ and is not shared by conventional anti-inflammatory drugs. These results unravel a novel and unique mechanism for E2 anti-inflammatory activity, which may be useful for identifying more selective ligands for the prevention of the inflammatory response.

Nuclear pores and macromolecular assemblies involved in nucleocytoplasmic transport

1996 ◽  
Vol 6 (2) ◽  
pp. 162-165 ◽  
Author(s):  
Murray Stewart ◽  
W David Clarkson
Keyword(s):  
Nucleocytoplasmic Transport ◽  
Nuclear Pores ◽  
Macromolecular Assemblies

scholarly journals In vitro reconstitution of a heterotrimeric nucleoporin complex consisting of recombinant Nsp1p, Nup49p, and Nup57p.

1997 ◽  
Vol 8 (1) ◽  
pp. 33-46 ◽  
Author(s):  
N L Schlaich ◽  
M Häner ◽  
A Lustig ◽  
U Aebi ◽  
E C Hurt
Keyword(s):  
Nucleocytoplasmic Transport ◽  
Full Length ◽  
Yeast Cells ◽  
Heptad Repeat ◽  
Nuclear Pores ◽  
Terminal Domain ◽  
Carboxyl Terminal Domain ◽  
Carboxyl Terminal

The yeast nucleoporins Nsp1p, Nup49p, and Nup57p form a complex at the nuclear pores which is involved in nucleocytoplasmic transport. To investigate the molecular basis underlying complex formation, recombinant full-length Nup49p and Nup57p and the carboxyl-terminal domain of Nsp1p, which lacks the FXFG repeat domain, were expressed in Escherichia coli. When the three purified proteins were mixed together, they spontaneously associated to form a 150-kDa complex of 1:1:1 stoichiometry. In this trimeric complex, Nup57p fulfills the role of an organizing center, to which Nup49p and Nsp1p individually bind. For this interaction to occur, only two heptad repeat regions of the Nsp1p carboxyl-terminal domain are required, each region being about 50 amino acids in length. Finally, the reconstituted complex has the capability to bind to full-length Nic96p but not to mutant forms which also do not interact in vivo. When added to permeabilized yeast cells, the complex associates with the nuclear envelope and the nuclear pores. We conclude that Nsp1p, Nup49p, and Nup57p can reconstitute a complex in vitro which is competent for further assembly with other components of nuclear pores.

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