Letteratura scientifica selezionata sul tema "RanBP2 (Nup358)"

ABSTRACT Nuclear pore complexes (NPCs) traverse the nuclear envelope (NE), providing a channel through which nucleocytoplasmic transport occurs. Nup358/RanBP2, Nup214/CAN, and Nup88 are components of the cytoplasmic face of the NPC. Here we show that Nup88 localizes midway between Nup358 and Nup214 and physically interacts with them. RNA interference of either Nup88 or Nup214 in human cells caused a strong reduction of Nup358 at the NE. Nup88 and Nup214 showed an interdependence at the NPC and were not affected by the absence of Nup358. These data indicate that Nup88 and Nup214 mediate the attachment of Nup358 to the NPC. We show that localization of the export receptor CRM1 at the cytoplasmic face of the NE is Nup358 dependent and represents its empty state. Also, removal of Nup358 causes a distinct reduction in nuclear export signal-dependent nuclear export. We propose that Nup358 provides both a platform for rapid disassembly of CRM1 export complexes and a binding site for empty CRM1 recycling into the nucleus.

Mutations in RanBP2 (also known as Nup358), one of the main components of the cytoplasmic filaments of the nuclear pore complex, contribute to the overproduction of acute necrotizing encephalopathy (ANE1)-associated cytokines. Here we report that RanBP2 represses the translation of the interleukin 6 (IL6) mRNA, which encodes a cytokine that is aberrantly up-regulated in ANE1. Our data indicates that soon after its production, the IL6 messenger ribonucleoprotein (mRNP) recruits Argonautes bound to let-7 microRNA. After this mRNP is exported to the cytosol, RanBP2 sumoylates mRNP-associated Argonautes, thereby stabilizing them and enforcing mRNA silencing. Collectively, these results support a model whereby RanBP2 promotes an mRNP remodelling event that is critical for the miRNA-mediated suppression of clinically relevant mRNAs, such as IL6.

The RanGTPase activating protein RanGAP1 has essential functions in both nucleocytoplasmic transport and mitosis. In interphase, a significant fraction of vertebrate SUMO1-modified RanGAP1 forms a stable complex with the nucleoporin RanBP2/Nup358 at nuclear pore complexes. RanBP2 not only acts in the RanGTPase cycle but also is a SUMO1 E3 ligase. Here, we show that RanGAP1 is phosphorylated on residues T409, S428, and S442. Phosphorylation occurs before nuclear envelope breakdown and is maintained throughout mitosis. Nocodazole arrest leads to quantitative phosphorylation. The M-phase kinase cyclin B/Cdk1 phosphorylates RanGAP1 efficiently in vitro, and T409 phosphorylation correlates with nuclear accumulation of cyclin B1 in vivo. We find that phosphorylated RanGAP1 remains associated with RanBP2/Nup358 and the SUMO E2–conjugating enzyme Ubc9 in mitosis, hence mitotic phosphorylation may have functional consequences for the RanGTPase cycle and/or for RanBP2-dependent sumoylation.

In vertebrate cells, the nucleoporin Nup358/RanBP2 is a major component of the filaments that emanate from the nuclear pore complex into the cytoplasm. Nup358 forms a complex with SUMOylated RanGAP1, the GTPase activating protein for Ran. RanGAP1 plays a pivotal role in the establishment of a RanGTP gradient across the nuclear envelope and, hence, in the majority of nucleocytoplasmic transport pathways. Here, we investigate the roles of the Nup358-RanGAP1 complex and of soluble RanGAP1 in nuclear protein transport, combining in vivo and in vitro approaches. Depletion of Nup358 by RNA interference led to a clear reduction of importin α/β-dependent nuclear import of various reporter proteins. In vitro, transport could be partially restored by the addition of importin β, RanBP1, and/or RanGAP1 to the transport reaction. In intact Nup358-depleted cells, overexpression of importin β strongly stimulated nuclear import, demonstrating that the transport receptor is the most rate-limiting factor at reduced Nup358-concentrations. As an alternative approach, we used antibody-inhibition experiments. Antibodies against RanGAP1 inhibited the enzymatic activity of soluble and nuclear pore–associated RanGAP1, as well as nuclear import and export. Although export could be fully restored by soluble RanGAP, import was only partially rescued. Together, these data suggest a dual function of the Nup358-RanGAP1 complex as a coordinator of importin β recycling and reformation of novel import complexes.

When higher eukaryotic cells transition into mitosis, the nuclear envelope, nuclear pore complexes, and nuclear lamina are coordinately disassembled. The COPI coatomer complex, which plays a major role in membrane remodeling at the Golgi, has been implicated in the process of nuclear envelope breakdown and requires interactions at the nuclear pore complex for recruitment to this new site of action at mitosis. Nup153, a resident of the nuclear pore basket, was found to be involved in COPI recruitment, but the molecular nature of the interface between COPI and the nuclear pore has not been fully elucidated. To better understand what occurs at the nuclear pore at this juncture, we have probed the role of the nucleoporin Nup358/RanBP2. Nup358 contains a repetitive zinc finger domain with overall organization similar to a region within Nup153 that is critical to COPI association, yet inspection of these two zinc finger domains reveals features that also clearly distinguish them. Here, we found that the Nup358 zinc finger domain, but not a zinc finger domain from an unrelated protein, binds to COPI and dominantly inhibits progression of nuclear envelope breakdown in an assay that robustly recapitulates this process in vitro. Moreover, the Nup358 zinc finger domain interferes with COPI recruitment to the nuclear rim. Consistent with a role for this pore protein in coordinating nuclear envelope breakdown, Nup358-specific antibodies impair nuclear disassembly. Significantly, targeting either Nup153 or Nup358 for inhibition perturbs nuclear envelope breakdown, supporting a model in which these nucleoporins play nonredundant roles, perhaps contributing to COPI recruitment platforms on both the nuclear and cytoplasmic faces of the pore. We found that an individual zinc finger is the minimal interface for COPI association, although tandem zinc fingers are optimal. These results provide new information about the critical components of nuclear membrane remodeling and lay the foundation for a better understanding of how this process is regulated.

ABSTRACT Nucleoporins mediate transport of macromolecules across the nuclear pore complex, yet the function of many individual nucleoporins is largely unresolved. To address this question, we depleted cells of the cytoplasmic nucleoporins Nup214/CAN and Nup358/RanBP2 by RNA interference. Depletion of Nup214 resulted in codepletion of its binding partner, Nup88. Nuclear pore complexes assembled in the absence of Nup214/Nup88 or Nup358 were fully functional in nuclear protein import, whereas nuclear mRNA export was slightly impaired. Depletion of Nup358 had only a minor effect on nuclear protein export. In contrast, depletion of Nup214/Nup88 led to strongly reduced CRM1-mediated export of the shuttling transcription factor NFAT as well as a human immunodeficiency virus-Rev derivative. A specific role of Nup214 in protein export is furthered by the biochemical properties of a high-affinity complex containing Nup214, CRM1, RanGTP, and an export cargo. Our results show that the Nup214/Nup88 complex is required for efficient CRM1-mediated transport, supporting a model involving a high-affinity binding site for CRM1 at Nup214 in the terminal steps of export.

Ran Binding Protein 2 (RanBP2 or Nucleoporin358) is one of the main components of the cytoplasmic filaments of the nuclear pore complex. Mutations in the RANBP2 gene are associated with acute necrotizing encephalopathy type 1 (ANE1), a rare condition where patients experience a sharp rise in cytokine production in response to viral infection and undergo hyperinflammation, seizures, coma, and a high rate of mortality. Despite this, it remains unclear howRanBP2 and its ANE1-associated mutations contribute to pathology. Mounting evidence has shown that RanBP2 interacts with distinct viruses to regulate viral infection. In addition, RanBP2 may regulate innate immune response pathways. This review summarizes recent advances in our understanding of how mutations in RANBP2 contribute to ANE1 and discusses how RanBP2 interacts with distinct viruses and affects viral infection. Recent findings indicate that RanBP2 might be an important therapeutic target, not only in the suppression of ANE1-driven cytokine storms, but also to combat hyperinflammation in response to viral infections.

Conjugation with the small ubiquitin-like modifier (SUMO) modulates protein interactions and localisation. The kinase Aurora B, a key regulator of mitosis, was previously identified as a SUMOylation target in vitro and in assays with overexpressed components. However, where and when this modification genuinely occurs in human cells was not ascertained. Here, we have developed intramolecular Proximity Ligation Assays (PLA) to visualise SUMO-conjugated Aurora B in human cells in situ. We visualised Aurora B-SUMO products at centromeres in prometaphase and metaphase, which declined from anaphase onwards and became virtually undetectable at cytokinesis. In the mitotic window in which Aurora B/SUMO products are abundant, Aurora B co-localised and interacted with NUP358/RANBP2, a nucleoporin with SUMO ligase and SUMO-stabilising activity. Indeed, in addition to the requirement for the previously identified PIAS3 SUMO ligase, we found that NUP358/RANBP2 is also implicated in Aurora B-SUMO PLA product formation and centromere localisation. In summary, SUMOylation marks a distinctive window of Aurora B functions at centromeres in prometaphase and metaphase while being dispensable for functions exerted in cytokinesis, and RANBP2 contributes to this control, adding a novel layer to modulation of Aurora B functions during mitosis.

Type I interferon (IFN-I)-induced signaling plays a critical role in host antiviral innate immune responses. Despite this, the mechanisms that regulate this signaling pathway have yet to be fully elucidated. The nucleoporin Ran Binding Protein 2 (RanBP2) (also known as Nucleoporin 358 KDa, Nup358) has been implicated in a number of cellular processes, including host innate immune signaling pathways, and is known to influence viral infection. In this study, we documented that RanBP2 mediates the sumoylation of signal transducers and activators of transcription 1 (STAT1) and inhibits IFN-α-induced signaling. Specifically, we found that RanBP2-mediated sumoylation inhibits the interaction of STAT1 and Janus kinase 1 (JAK1), as well as the phosphorylation and nuclear accumulation of STAT1 after IFN-α stimulation, thereby antagonizing the IFN-α-mediated antiviral innate immune signaling pathway and promoting viral infection. Our findings not only provide insights into a novel function of RanBP2 in antiviral innate immunity but may also contribute to the development of new antiviral therapeutic strategies.

Les adénovirus (AdV), comme d'autres virus à réplication nucléaire, ont besoin d’arriver jusqu’aunoyau cellulaire afin de libérer leur génome. Pour ce faire, les particules des AdV contenant l’ADNviral sont transportées jusqu’au complexe du pore nucléaire (NPC), via le centre d’organisation desmicrotubules, par un mécanisme encore mal compris qui implique l’exportine cellulaire CRM1. Lacapside des AdV dépasse la taille limite d’entrée dans le noyau, et par conséquent, elle doit êtredésassemblée au niveau du NPC. Le mécanisme d’import de molécules d’ADN n’est pas un processusphysiologique. Pour cela, les AdV doivent détourner la machinerie cellulaire afin d’importer leurgénome dans le noyau. Le NPC est un complexe de protéines appelées nucléoporines. LaNup358/RanBP2, principal composant des filaments cytoplasmiques, sert de plateforme de liaison àdes karyopherines (e.g Importin-β, CRM1) et à la protéine GTPase Ran. Les karyopherinesreconnaissent des signaux spécifiques présents dans les cargos et facilitent leur transport d’unemanière très régulée dépendante de RanGTP. Nous avons constaté que l’import du génome AdV estmoins efficace en l’absence de Nup358. Dans ces conditions, nous avons observé que certaineskaryopherines deviennent limitantes pour l’import du génome viral, et identifié la région minimale deNup358 requise pour compenser ce défaut. D’autre part, nous avons confirmé l’implication de CRM1dans l’arrivé des particules virales au noyau et identifié un nouveau rôle de CRM1 dans ledésassemblage de la capside des AdV. Ces travaux contribuent à mieux connaître le mécanismed’entrée du génome AdV dans le noyau et donnent une idée de la façon dont les virus peuventcontourner la machinerie de transport cellulaire pour leur propre bénéfice
Nuclear delivery of viral genomes is an essential step for nuclear replicating DNA viruses such asAdenovirus (AdV). AdV particles reach the nuclear pore complex (NPC) in the form of genomecontaining, partially disassembled capsids, through a poorly understood CRM1-dependent mechanism.These capsids exceed the NPC size limit and therefore, they must disassemble at the NPC to releasethe viral genome. Nuclear import of DNA cargos is not a physiological process. Consequently, AdVneed to divert the cellular transport machinery for nuclear genome delivery. The NPC is a multiproteincomplex consisting of nucleoporins (Nups). The Nup358/RanBP2 is the major component ofthe cytoplasmic filaments of the NPC and serves as binding platform for factors includingkaryopherins (i.e Importin-β, CRM1) and the small GTPase Ran. Selective transport of cargo throughthe NPC is mediated by karyopherins, which recognize specific signals within the cargos and facilitatetheir transport in a RanGTP-dependent regulated manner. We identified that Nup358-depleted cellsreduce nuclear import efficiency of the AdV genome. Indeed, we observed that karyopherins are ratelimitingfor AdV genome import under these conditions and we mapped the minimal region ofNup358 necessary to compensate the import defect. On the other hand, we could confirm therequirement of CRM1 in nuclear targeting of AdV capsids and identified and additional role inmediating AdV capsid disassembly. This work helps to understand the strategy used by AdV todeliver their genome and gives insight about how viruses hijack the cellular transport machinery fortheir own benefit

Les complexes des pores nucléaires (CPN) sont d'énormes assemblages protéiques intégrés dans l'enveloppe nucléaire (EN). Ils servent de structures pour le transport bidirectionnel. Essentiels, ils permettent le maintien de l'équilibre entre le noyau et le cytoplasme. Au-delà de leur résidence dans l'EN, les CPN se trouvent également dans des feuillets du réticulum endoplasmique (RE) empilés connus sous le nom de lamelles annulaires (LA). Cependant, la fonction et les voies régissant la biogenèse des LA restent énigmatiques. Notre investigation dans les cellules de mammifères révèle un mécanisme où la formation des LA résulte de la fusion de CPN cytosoliques préassemblés. Le mouvement des CPN cytosoliques est intimement lié à la dynamique du RE, car ils migrent vers l'EN et s'y intègrent en début d'interphase en conditions de croissance normales, ce processus est médié par les microtubules. RanBP2 (Nup358), un constituant des filaments cytoplasmiques des CPN, apparaît comme nécessaire et suffisant pour la formation des LA dans le cytoplasme. Mécaniquement, les répétitions FG dans le N-terminus de RanBP2 jouent un rôle crucial en orchestrant l'état d'oligomérisation des unités de l'anneau extérieur des CPN, connues sous le nom de complexes Y. Notre étude élucide un processus d'assemblage crucial pour nourrir l'EN, assurant la fonctionnalité des pores nucléaires et soulignant l'importance des CPN cytosoliques dans l'homéostasie cellulaire des mammifères
Nuclear pore complexes (NPCs), huge protein assemblies built into the nuclear envelope (NE), serve as pivotal structures for bidirectional transport, maintaining the equilibrium between the nucleus and cytoplasm. Beyond their residence within the NE, NPCs are also found in stacked cytoplasmic membranes known as annulate lamellae (AL). However, the function and pathways governing the biogenesis of AL remain enigmatic. Our investigation in mammalian cells unveils a mechanism wherein AL formation arises through the fusion of pre-assembled cytosolic NPCs. The movement of cytosolic NPCs is intricately linked to the dynamics of the endoplasmic reticulum (ER), as they migrate towards and integrate into NE during early interphase under normal growth conditions, a process mediated by microtubules. RanBP2 (Nup358), a constituent of the NPC cytoplasmic filaments, emerges as necessary and sufficient for AL formation in the cytoplasm. Mechanistically, the FG repeats in the N-terminus of RanBP2 play a pivotal role by orchestrating the oligomerization state of the NPC outer ring units, known as Y-complexes. Our study elucidates an assembly process crucial for NE nourishment, ensuring the functionality of nuclear pores and underscoring the significance of cytosolic NPCs in mammalian cellular homeostasis