Exchange Factor RCC1 Research Articles

The nucleoporin Nup50 activates the Ran guanine nucleotide exchange factor RCC1 to promote NPC assembly at the end of mitosis

During mitotic exit, thousands of nuclear pore complexes (NPCs) assemble concomitant with the nuclear envelope to build a transport‐competent nucleus. Here, we show that Nup50 plays a crucial role in NPC assembly independent of its well‐established function in nuclear transport. RNAi‐mediated downregulation in cells or immunodepletion of Nup50 protein in Xenopus egg extracts interferes with NPC assembly. We define a conserved central region of 46 residues in Nup50 that is crucial for Nup153 and MEL28/ELYS binding, and for NPC interaction. Surprisingly, neither NPC interaction nor binding of Nup50 to importin α/β, the GTPase Ran, or chromatin is crucial for its function in the assembly process. Instead, an N‐terminal fragment of Nup50 can stimulate the Ran GTPase guanine nucleotide exchange factor RCC1 and NPC assembly, indicating that Nup50 acts via the Ran system in NPC reformation at the end of mitosis. In support of this conclusion, Nup50 mutants defective in RCC1 binding and stimulation cannot replace the wild‐type protein in in vitro NPC assembly assays, whereas excess RCC1 can compensate the loss of Nup50.

Modulation of innate immune signaling by a Coxiella burnetii eukaryotic-like effector protein

The Q fever agent Coxiella burnetii uses a defect in organelle trafficking/intracellular multiplication (Dot/Icm) type 4b secretion system (T4SS) to silence the host innate immune response during infection. By investigating C. burnetii effector proteins containing eukaryotic-like domains, here we identify NopA (nucleolar protein A), which displays four regulator of chromosome condensation (RCC) repeats, homologous to those found in the eukaryotic Ras-related nuclear protein (Ran) guanine nucleotide exchange factor (GEF) RCC1. Accordingly, NopA is found associated with the chromatin nuclear fraction of cells and uses the RCC-like domain to interact with Ran. Interestingly, NopA triggers an accumulation of Ran-GTP, which accumulates at nucleoli of transfected or infected cells, thus perturbing the nuclear import of transcription factors of the innate immune signaling pathway. Accordingly, qRT-PCR analysis on a panel of cytokines shows that cells exposed to the C. burnetii nopA::Tn or a Dot/Icm-defective dotA::Tn mutant strain present a functional innate immune response, as opposed to cells exposed to wild-type C. burnetii or the corresponding nopA complemented strain. Thus, NopA is an important regulator of the innate immune response allowing Coxiella to behave as a stealth pathogen.

Three-dimensional context rather than NLS amino acid sequence determines importin \u03b1 subtype specificity for RCC1

Active nuclear import of Ran exchange factor RCC1 is mediated by importin α3. This pathway is essential to generate a gradient of RanGTP on chromatin that directs nucleocytoplasmic transport, mitotic spindle assembly and nuclear envelope formation. Here we identify the mechanisms of importin α3 selectivity for RCC1. We find this isoform binds RCC1 with one order of magnitude higher affinity than the generic importin α1, although the two isoforms share an identical NLS-binding groove. Importin α3 uses its greater conformational flexibility to wedge the RCC1 β-propeller flanking the NLS against its lateral surface, preventing steric clashes with its Armadillo-core. Removing the β-propeller, or inserting a linker between NLS and β-propeller, disrupts specificity for importin α3, demonstrating the structural context rather than NLS sequence determines selectivity for isoform 3. We propose importin α3 evolved to recognize topologically complex NLSs that lie next to bulky domains or are masked by quaternary structures.

Crystal structure of importin-α3 bound to the nuclear localization signal of Ran-binding protein 3

Ran-binding protein 3 (RanBP3) is a primarily nuclear Ran-binding protein that functions as an accessory factor in the Ran GTPase system. RanBP3 associates with Ran-specific nucleotide exchange factor RCC1 and enhances its catalytic activity towards Ran. RanBP3 also promotes CRM1-mediated nuclear export as well as CRM1-independent nuclear export of β-catenin, Smad2, and Smad3. Nuclear import of RanBP3 is dependent on the nuclear import adaptor protein importin-α and, RanBP3 is imported more efficiently by importin-α3 than by other members of the importin-α family. Protein kinase signaling pathways control nucleocytoplasmic transport through phosphorylation of RanBP3 at Ser58, immediately C-terminal to the nuclear localization signal (NLS) in the N-terminal region of RanBP3. Here we report the crystal structure of human importin-α3 bound to an N-terminal fragment of human RanBP3 containing the NLS sequence that is necessary and sufficient for nuclear import. The structure reveals that RanBP3 binds to importin-α3 residues that are strictly conserved in all seven isoforms of human importin-α at the major NLS-binding site, indicating that the region of importin-α outside the NLS-binding site, possibly the autoinhibotory importin-β1-binding domain, may be the key determinant for the preferential binding of RanBP3 to importin-α3. Computational docking simulation indicates that phosphorylation of RanBP3 at Ser58 could potentially stabilize the association of RanBP3 with importin-α through interactions between the phosphate moiety of phospho-Ser58 of RanBP3 and a cluster of basic residues (Arg96 and Lys97 in importin-α3) on armadillo repeat 1 of importin-α.

MYCBP2 Is a Guanosine Exchange Factor for Ran Protein and Determines Its Localization in Neurons of Dorsal Root Ganglia

The small GTPase Ran coordinates retrograde axonal transport in neurons, spindle assembly during mitosis, and the nucleo-cytoplasmic transport of mRNA. Its localization is tightly regulated by the GTPase-activating protein RanGAP1 and the nuclear guanosine exchange factor (GEF) RCC1. We show that loss of the neuronal E3 ubiquitin ligase MYCBP2 caused the up-regulation of Ran and RanGAP1 in dorsal root ganglia (DRG) under basal conditions and during inflammatory hyperalgesia. SUMOylated RanGAP1 physically interacted with MYCBP2 and inhibited its E3 ubiquitin ligase activity. Stimulation of neurons induced a RanGAP1-dependent translocation of MYCBP2 to the nucleus. In the nucleus of DRG neurons MYCBP2 co-localized with Ran and facilitated through its RCC1-like domain the GDP/GTP exchange of Ran. In accordance with the necessity of a GEF to promote GTP-binding and nuclear export of Ran, the nuclear localization of Ran was strongly increased in MYCBP2-deficient DRGs. The finding that other GEFs for Ran besides RCC1 exist gives new insights in the complexity of the regulation of the Ran signaling pathway.

TD-60 links RalA GTPase function to the CPC in mitosis.

TD-60 (also known as RCC2) is a highly conserved protein that structurally resembles the Ran guanine exchange factor (GEF) RCC1, but has not previously been shown to have GEF activity. TD-60 has a typical chromosomal passenger complex (CPC) distribution in mitotic cells, but associates with integrin complexes and is involved in cell motility during interphase. Here we show that TD-60 exhibits GEF activity, in vitro and in cells, for the small GTPase RalA. TD-60 or RalA depletion causes spindle abnormalities in prometaphase associated with abnormal centromeric accumulation of CPC components. TD-60 and RalA apparently work together to contribute to the regulation of kinetochore–microtubule interactions in early mitosis. Importantly, several mitotic phenotypes caused by TD-60 depletion are reverted by the expression of a GTP-locked mutant, RalA (Q72L). The demonstration that a small GTPase participates in the regulation of the CPC reveals a level of mitotic regulation not suspected in previous studies.

Disruption of the ran system by cysteine oxidation of the nucleotide exchange factor RCC1.

Transport regulation by the Ran GTPase requires its nuclear localization and GTP loading by the chromatin-associated exchange factor RCC1. These reactions generate Ran protein and Ran nucleotide gradients between the nucleus and the cytoplasm. Cellular stress disrupts the Ran gradients, but the specific mechanisms underlying this disruption have not been elucidated. We used biochemical approaches to determine how oxidative stress disrupts the Ran system. RCC1 exchange activity was reduced by diamide-induced oxidative stress and restored with dithiothreitol. Using mass spectrometry, we found that multiple solvent-exposed cysteines in RCC1 are oxidized in cells treated with diamide. The cysteines oxidized in RCC1 included Cys93, which is solvent exposed and unique because it becomes buried upon contact with Ran. A Cys93Ser substitution dramatically reduced exchange activity through an effect on RCC1 binding to RanGDP. Diamide treatment reduced the size of the mobile fraction of RCC1-green fluorescent protein in cells and inhibited nuclear import in digitonin-permeabilized cell assays. The Ran protein gradient was also disrupted by UV-induced stress but without affecting RCC1 exchange activity. Our data suggest that stress can disrupt the Ran gradients through RCC1-dependent and RCC1-independent mechanisms, possibly dependent on the particular stress condition.

Nucleosomal regulation of chromatin composition and nuclear assembly revealed by histone depletion.

Nucleosomes are the fundamental unit of chromatin, but analysis of transcription-independent nucleosome functions has been complicated by the gene-expression changes resulting from histone manipulation. Here we solve this dilemma by developing Xenopus laevis egg extracts deficient for nucleosome formation and by analyzing the proteomic landscape and behavior of nucleosomal chromatin and nucleosome-free DNA. We show that although nucleosome-free DNA can recruit nuclear-envelope membranes, nucleosomes are required for spindle assembly and for formation of the lamina and of nuclear pore complexes (NPCs). We show that, in addition to the Ran G-nucleotide exchange factor RCC1, ELYS, the initiator of NPC formation, fails to associate with naked DNA but directly binds histone H2A-H2B dimers and nucleosomes. Tethering ELYS and RCC1 to DNA bypasses the requirement for nucleosomes in NPC formation in a synergistic manner. Thus, the minimal essential function of nucleosomes in NPC formation is to recruit RCC1 and ELYS.

Cell Cycle-Dependent Binding Modes of the Ran Exchange Factor RCC1 to Chromatin

The formation of an activity gradient of the small G-protein Ran around chromatin depends on the differential partitioning of the opposing enzyme activities of the Ran guanine nucleotide exchange factor RCC1 that resides on chromatin, and the cytoplasmic Ran GTPase activating protein RanGAP. We studied the time-dependent interaction kinetics between RCC1 and chromatin and the mobility of the Ran-RCC1 complex in living cells by fluorescence correlation spectroscopy to investigate whether binding of RCC1 to chromatin regulates the exchange activity of RCC1, and whether the stability of the RCC1-chromatin interaction is regulated during the cell cycle. We found that RCC1 mobility is dominated by two states: a highly mobile state that is trapped within chromatin, and a transiently immobilized state that is stabilized during mitosis. We show that only the immobilized state of RCC1 interacts with Ran and conclude that its guanine nucleotide exchange activity is restricted to specific sites on chromatin.

Guanine-Nucleotide Exchange Factor RCC1 Facilitates a Tight Binding between the Encephalomyocarditis Virus Leader and Cellular Ran GTPase

The leader (L) protein of encephalomyocarditis virus (EMCV) shuts off host cell nucleocytoplasmic trafficking (NCT) by inducing hyperphosphorylation of nuclear pore proteins. This dramatic effect by a nonenzymatic protein of 6 kDa is not well understood but clearly involves L binding to cellular Ran GTPase, a critical factor of active NCT. Exogenous GDP and GTP are inhibitory to L-Ran binding, but the guanine-nucleotide exchange factor RCC1 can relieve this inhibition. In the presence of RCC1, L binds Ran with a KD (equilibrium dissociation constant) of ≈ 3 nM and reaches saturation within 20 min. The results of fluorescently tagged nucleotide experiments suggest that L-Ran interactions affect the nucleotide-binding pocket of Ran.

Spatiotemporal control of microtubule nucleation and assembly using magnetic nanoparticles

Decisions on the fate of cells and their functions are dictated by the spatiotemporal dynamics of molecular signalling networks. However, techniques to examine the dynamics of these intracellular processes remain limited. Here, we show that magnetic nanoparticles conjugated with key regulatory proteins can artificially control, in time and space, the Ran/RCC1 signalling pathway that regulates the cell cytoskeleton. In the presence of a magnetic field, RanGTP proteins conjugated to superparamagnetic nanoparticles can induce microtubule fibres to assemble into asymmetric arrays of polarized fibres in Xenopus laevis egg extracts. The orientation of the fibres is dictated by the direction of the magnetic force. When we locally concentrated nanoparticles conjugated with the upstream guanine nucleotide exchange factor RCC1, the assembly of microtubule fibres could be induced over a greater range of distances than RanGTP particles. The method shows how bioactive nanoparticles can be used to engineer signalling networks and spatial self-organization inside a cell environment.

Requirement for Lamin B Receptor and Its Regulation by Importin β and Phosphorylation in Nuclear Envelope Assembly during Mitotic Exit

Lamin B receptor (LBR), a chromatin and lamin B-binding protein in the inner nuclear membrane, has been proposed to target the membrane precursor vesicles to chromatin mediated by importin β during the nuclear envelope (NE) assembly. However, the mechanisms for the binding of LBR with importin β and the membrane targeting by LBR in NE assembly remain largely unknown. In this report, we show that the amino acids (aa) 69-90 of LBR sequences are required to bind with importin β at aa 45-462, and the binding is essential for the NE membrane precursor vesicle targeting to the chromatin during the NE assembly at the end of mitosis. We also show that this binding is cell cycle-regulated and dependent on the phosphorylation of LBR Ser-71 by p34(cdc2) kinase. RNAi knockdown of LBR causes the NE assembly failure and abnormal chromatin decondensation of the daughter cell nuclei, leading to the daughter cell death at early G(1) phase by apoptosis. Perturbation of the interaction of LBR with importin β by deleting the LBR N-terminal spanning region or aa 69-73 also induces the NE assembly failure, the abnormal chromatin decondensation, and the daughter cell death. The first transmembrane domain of LBR promotes the NE production and expansion, because overexpressing this domain is sufficient to induce membrane overproduction of the NE. Thus, these results demonstrate that LBR targets the membrane precursor vesicles to chromatin by interacting with importin β in a LBR phosphorylation-dependent manner during the NE assembly at the end of mitosis and that the first transmembrane domain of LBR promotes the LBR-bearing membrane production and the NE expansion in interphase.

Allosteric beta-propeller signalling in TolB and its manipulation by translocating colicins.

The Tol system is a five-protein assembly parasitized by colicins and bacteriophages that helps stabilize the Gram-negative outer membrane (OM). We show that allosteric signalling through the six-bladed beta-propeller protein TolB is central to Tol function in Escherichia coli and that this is subverted by colicins such as ColE9 to initiate their OM translocation. Protein-protein interactions with the TolB beta-propeller govern two conformational states that are adopted by the distal N-terminal 12 residues of TolB that bind TolA in the inner membrane. ColE9 promotes disorder of this 'TolA box' and recruitment of TolA. In contrast to ColE9, binding of the OM lipoprotein Pal to the same site induces conformational changes that sequester the TolA box to the TolB surface in which it exhibits little or no TolA binding. Our data suggest that Pal is an OFF switch for the Tol assembly, whereas colicins promote an ON state even though mimicking Pal. Comparison of the TolB mechanism to that of vertebrate guanine nucleotide exchange factor RCC1 suggests that allosteric signalling may be more prevalent in beta-propeller proteins than currently realized.

Regulation of chromatin binding by a conformational switch in the tail of the Ran exchange factor RCC1

RCC1 is the only known exchange factor for the Ran guanosine triphosphatase and performs essential roles in nuclear transport, spindle organization, and nuclear envelope formation. RCC1 binds to chromatin through a bimodal attachment to DNA and histones, and defects in binding cause chromosome missegregation. Chromatin binding is enhanced by apo-Ran. However, the mechanism underlying this regulation has been unclear. We now demonstrate that the N-terminal tail of RCC1 is essential for association with DNA but inhibits histone binding. Apo-Ran significantly promotes RCC1 binding to both DNA and histones, and these effects are tail mediated. Using a fluorescence resonance energy transfer biosensor, we detect conformational changes in the tail of RCC1 coupled to the two binding modes and in response to interactions with Ran and importin-α. The biosensor also reports changes accompanying mitosis in living cells. We propose that Ran induces an allosteric conformational switch in the tail that exposes the histone-binding surface on RCC1 and facilitates association of the positively charged tail with DNA.

Going green: plants' alternative way to position the Ran gradient

Ran is a multi-functional small GTPase of the Ras super-family involved in nucleocytoplasmic transport, mitotic spindle assembly, cell cycle control and nuclear envelope (NE) formation. Its roles are accomplished by the asymmetric distribution of its GTP- and GDP-bound forms, enabled by the specific localization of Ran accessory proteins, the Ran GTPase-activating protein RanGAP and the nucleotide exchange factor RCC1. Mammalian RanGAP1 is targeted to the NE during interphase and to the spindle and kinetochores during mitosis via a SUMOylated C-terminal domain and interaction with the nucleoporin Nup358/RanBP2. Arabidopsis RanGAP1 (AtRanGAP1) lacks the SUMOylated C-terminal domain of vertebrate RanGAP, but contains a plant-specific N-terminal domain (WPP domain), which is necessary and sufficient for its targeting to the NE in interphase. AtRanGAP1 has a mitotic trafficking pattern uniquely different from that of vertebrate RanGAP, which includes targeting to the outward-growing rim of the cell plate. The WPP domain is necessary and sufficient for this targeting. Now, a novel family of plant-specific, nuclear pore-associated proteins has been identified in Arabidopsis, which is essential for anchoring RanGAP to the Arabidopsis nuclear envelope at the root meristem. This suggests that RanGAP anchoring to the nuclear pore has been solved in two fundamentally different ways in animals and plants. These findings support a separate evolution of RanGAP targeting mechanisms in different kingdoms, possibly related to different functional geometries of the Ran gradient in animal and higher plant cell division.

Ran GTPase guanine nucleotide exchange factor RCC1 is phosphorylated on serine 11 by cdc2 kinase in vitro

RCC1, a guanine nucleotide exchange factor for Ran GTPase, plays essential roles in the growth and viability of mammalian cells. Here, we examined the phosphorylation of specific serine and threonine residues of RCC1 in vivo and showed that RCC1 is indeed phosphorylated. Analysis by two-dimensional (2D) gel electrophoresis suggested that serine 11 (S11) of hamster RCC1 is phosphorylated in vivo. A point mutation of S11 of hamster RCC1 resulted in a decrease in the number of 2D gel spots, indicating a lack of phosphorylation at the mutant residue. S11 phosphorylation in vitro depended on cyclin B-cdc2 kinase. An RCC1 mutant in which all N-terminal serine and threonine residues were substituted with glutamate residues to mimic phosphorylation at these residues showed decreased binding to the karyopherin, KPNA4, compared with wild type RCC1. We conclude that RCC1 undergoes post-translational phosphorylation.

The RanGTP gradient – a GPS for the mitotic spindle

The GTPase Ran has a key role in nuclear import and export, mitotic spindle assembly and nuclear envelope formation. The cycling of Ran between its GTP- and GDP-bound forms is catalyzed by the chromatin-bound guanine nucleotide exchange factor RCC1 and the cytoplasmic Ran GTPase-activating protein RanGAP. The result is an intracellular concentration gradient of RanGTP that equips eukaryotic cells with a ;genome-positioning system' (GPS). The binding of RanGTP to nuclear transport receptors (NTRs) of the importin beta superfamily mediates the effects of the gradient and generates further downstream gradients, which have been elucidated by fluorescence resonance energy transfer (FRET) imaging and computational modeling. The Ran-dependent GPS spatially directs many functions required for genome segregation by the mitotic spindle during mitosis. Through exportin 1, RanGTP recruits essential centrosome and kinetochore components, whereas the RanGTP-induced release of spindle assembly factors (SAFs) from importins activates SAFs to nucleate, bind and organize nascent spindle microtubules. Although a considerable fraction of cytoplasmic SAFs is active and RanGTP induces only partial further activation near chromatin, bipolar spindle assembly is robustly induced by cooperativity and positive-feedback mechanisms within the network of Ran-activated SAFs. The RanGTP gradient is conserved, although its roles vary among different cell types and species, and much remains to be learned regarding its functions.

RanBP10 Is a Cytoplasmic Guanine Nucleotide Exchange Factor That Modulates Noncentrosomal Microtubules

Microtubule spindle assembly in mitosis is stimulated by Ran.GTP, which is generated along condensed chromosomes by the guanine nucleotide exchange factor (GEF) RCC1. This relationship suggests that similar activities might modulate other microtubule structures. Interphase microtubules usually extend from the centrosome, although noncentrosomal microtubules function in some differentiated cells, including megakaryocytes. In these cells, platelet biogenesis requires massive mobilization of microtubules in the cell periphery, where they form proplatelets, the immediate precursors of platelets, in the apparent absence of centrioles. Here we identify a cytoplasmic Ran-binding protein, RanBP10, as a factor that binds beta-tubulin and associates with megakaryocyte microtubules. Unexpectedly, RanBP10 harbors GEF activity toward Ran. A point mutation in the candidate GEF domain abolishes exchange activity, and our results implicate RanBP10 as a localized cytoplasmic Ran-GEF. RNA interference-mediated loss of RanBP10 in cultured megakaryocytes disrupts microtubule organization. These results lead us to propose that spatiotemporally restricted generation of cytoplasmic Ran.GTP may influence organization of the specialized microtubules required in thrombopoiesis and that RanBP10 might serve as a molecular link between Ran and noncentrosomal microtubules.

Anchorage of Plant RanGAP to the Nuclear Envelope Involves Novel Nuclear-Pore-Associated Proteins

The Ran GTPase controls multiple cellular processes including nucleocytoplasmic transport, spindle assembly, and nuclear envelope (NE) formation [1-4]. Its roles are accomplished by the asymmetric distribution of RanGTP and RanGDP enabled by the specific locations of the Ran GTPase-activating protein RanGAP and the nucleotide exchange factor RCC1 [5-8]. Mammalian RanGAP1 targeting to the NE and kinetochores requires interaction of its sumoylated C-terminal domain with the nucleoporin Nup358/RanBP2 [9-14]. In contrast, Arabidopsis RanGAP1 is associated with the NE and cell plate, mediated by an N-terminal, plant-specific WPP domain [15-18]. In the absence of RanBP2 in plants, the mechanism for spatially sequestering plant RanGAP is unknown. Here, Arabidopsis WPP-domain interacting proteins (WIPs) that interact with RanGAP1 in vivo and colocalize with RanGAP1 at the NE and cell plate were identified. Immunogold labeling indicates that WIP1 is associated with the outer NE. In a wip1-1/wip2-1/wip3-1 triple mutant, RanGAP1 is dislocated from the NE in undifferentiated root-tip cells, whereas NE targeting in differentiated root cells and targeting to the cell plate remain intact. We propose that WIPs are novel plant nucleoporins involved in RanGAP1 NE anchoring in specific cell types. Our data support a separate evolution of RanGAP targeting mechanisms in different kingdoms.

Calcium-Dependent Regulation of NEMO Nuclear Export in Response to Genotoxic Stimuli

The mechanisms involved in activation of the transcription factor NF-kappaB by genotoxic agents are not well understood. Previously, we provided evidence that a regulatory subunit of the IkappaB kinase (IKK) complex, NF-kappaB essential modulator (NEMO)/IKKgamma, is a component of a nuclear signal that is generated after DNA damage to mediate NF-kappaB activation. Here, we found that etoposide (VP16) and camptothecin induced increases in intracellular free calcium levels at 60 min after stimulation of CEM T leukemic cells. Inhibition of calcium increases by calcium chelators, BAPTA-AM and EGTA-AM, abrogated NF-kappaB activation by these agents in several cell types examined. Conversely, thapsigargin and ionomycin attenuated the BAPTA-AM effects and promoted NF-kappaB activation by the genotoxic stimuli. Analyses of nuclear NEMO levels in VP16-treated cells suggested that calcium was required for nuclear export of NEMO. Inhibition of the nuclear exporter CRM1 by leptomycin B did not interfere with NEMO nuclear export. Similarly, deficiency of a plausible calcium-dependent nuclear export receptor, calreticulin, failed to prevent NF-kappaB activation by VP16. However, temperature inactivation of the Ran guanine nucleotide exchange factor RCC1 in the tsBN2 cell line harboring a temperature-sensitive mutant of RCC1 blocked NF-kappaB activation induced by genotoxic stimuli. Overexpression of Ran in this cell model showed that DNA damage stimuli induced formation of a complex between Ran and NEMO, suggesting that RCC1 regulated NF-kappaB activation through the modulation of RanGTP. Indeed, evidence for VP16-inducible interaction between Ran-GTP and NEMO could be obtained by means of glutathione S-transferase (GST) pull-down assays using GST fused to the Ran binding domain of RanBP2, which specifically interacts with the GTP-bound form of Ran. BAPTA-AM did not alter these interactions, suggesting that calcium is a necessary step beyond the formation of a Ran-GTP-NEMO complex in the nucleus. These results suggest that calcium has a unique role in genotoxic stress-induced NF-kappaB signaling by regulating nuclear export of NEMO subsequent to the formation of a nuclear export complex composed of Ran-GTP, NEMO, and presumably, an undefined nuclear export receptor.