Showing papers on "Importin published in 2009"
TL;DR: Six different NLS classes that specifically bind to distinct binding pockets of importin α are reported, finding that they fundamentally require the regions outside the core basic residues for their activity and have specific residues or patterns that confer the activities differently between yeast, plants, and mammals.
549 citations
TL;DR: A novel family of NPC proteins, the FG-nucleoporins (FG-Nups), coordinates and potentially regulates NPC translocation and are essential components of the nuclear permeability barrier.
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.
237 citations
TL;DR: This work shows that AY-WB phy toplasma produces a protein that targets the nuclei of plant host cells; this protein is a potential phytoplasma effector that may alter plant cell physiology.
Abstract: The fully sequenced genome of aster yellows phytoplasma strain witches' broom (AY-WB; Candidatus Phytoplasma asteris) was mined for the presence of genes encoding secreted proteins based on the presence of N-terminal signal peptides (SP). We identified 56 secreted AY-WB proteins (SAP). These SAP are candidate effector proteins potentially involved in interaction with plant and insect cell components. One of these SAP, SAP11, contains an N-terminal SP sequence and a eukaryotic bipartite nuclear localization signal (NLS). Transcripts for SAP11 were detected in AY-WB-infected plants. Yellow fluorescence protein (YFP)-tagged SAP11 accumulated in Nicotiana benthamiana cell nuclei, whereas the nuclear targeting of YFP-tagged SAP11 mutants with disrupted NLS was inhibited. The nuclear transport of YFP-SAP11 was also inhibited in N. benthamiana plants in which the expression of importin α was knocked down using virus-induced gene silencing (VIGS). Furthermore, SAP11 was detected by immunocytology in nuclei of you...
194 citations
TL;DR: This review examines several of these novel pathways, including facilitation of Imp‐dependent transport by microtubule motors, and Imp‐independent pathways involving either other transport molecules such as the calcium‐binding protein calmodulin or through direct binding to the components of the NPC.
Abstract: The movement of proteins between the cytoplasm and the nucleus conventionally involves the recognition of nuclear targeting signals by members of the importin (Imp) superfamily of nuclear transporters, followed by translocation through the nuclear envelope-embedded nuclear pore complexes (NPCs). It is becoming increasingly apparent, however, that distinct alternative pathways for nuclear transport exist and are relatively abundant. This review examines several of these novel pathways, including facilitation of Imp-dependent transport by microtubule motors, and Imp-independent pathways involving either other transport molecules such as the calcium-binding protein calmodulin or through direct binding to the components of the NPC. The existence of these pathways and the fact that many proteins appear to possess separate Imp-dependent and -independent nuclear import mechanisms ensure that the cell can function under conditions in which Imp-dependent transport is inhibited and/or modulate the efficiency of Imp-dependent transport itself, according to the need.
173 citations
TL;DR: GR cross-linked to the hsp90 heterocomplex was able to translocate to the nucleus in digitonin-permeabilized cells treated with steroid, suggesting that GR could pass through the pore in its untransformed state.
Abstract: Glucocorticoid receptor (GR) is cytoplasmic in the absence of ligand and localizes to the nucleus after steroid binding. Previous evidence demonstrated that the hsp90-based heterocomplex bound to GR is required for the efficient retrotransport of the receptor to the nuclear compartment. We examined the putative association of GR and its associated chaperone heterocomplex with structures of the nuclear pore. We found that importin β and the integral nuclear pore glycoprotein Nup62 interact with hsp90, hsp70, p23, and the TPR domain proteins FKBP52 and PP5. Nup62 and GR were able to interact in a more efficient manner when chaperoned by the hsp90-based heterocomplex. Interestingly, the binding of hsp70 and p23 to Nup62 does not require the presence of hsp90, whereas the association of FKBP52 and PP5 is hsp90 dependent, as indicated by the results of experiments where the hsp90 function was disrupted with radicicol. The ability of both FKBP52 and PP5 to interact with Nup62 was abrogated in cells overexpressing the TPR peptide. Importantly, GR cross-linked to the hsp90 heterocomplex was able to translocate to the nucleus in digitonin-permeabilized cells treated with steroid, suggesting that GR could pass through the pore in its untransformed state.
138 citations
TL;DR: The key mechanisms, pathways, and models underlying the transport of proteins across nuclear pores are analysed and it is shown that nuclear import of proteins is an essential step in regulating gene expression and the replication cycle of several viruses.
Abstract: In eukaryotes, the nuclear membrane provides a physical barrier to the passive diffusion of macromolecules from and into the cytoplasm. Nucleocytoplasmic traffic occurs through highly specialized structures known as nuclear pores, and involves the participation of a special class of transport proteins. Active transport across the nuclear pores is an energy-dependent process that relies on the activity of Ran-GTPases both in the nuclear and cytoplasmic compartments. Nuclear import of proteins is an essential step in regulating gene expression and the replication cycle of several viruses. In this review, the key mechanisms, pathways, and models underlying the transport of proteins across nuclear pores are ana- lysed.
131 citations
TL;DR: It was found that RNA interference silencing of CUL4B led to an inhibition of cell proliferation and a prolonged S phase, due to the overaccumulation of cyclin E, a substrate targeted by Cul4B for ubiquitination.
105 citations
TL;DR: The results suggest that imp7 facilitates nuclear trafficking of DNA and that HIV-1 exploits imp7 to maximize nuclear import of its DNA genome.
Abstract: Background
Nuclear import of the HIV-1 reverse transcription complex (RTC) is critical for infection of non dividing cells, and importin 7 (imp7) has been implicated in this process. To further characterize the function of imp7 in HIV-1 replication we generated cell lines stably depleted for imp7 and used them in conjunction with infection, cellular fractionation and pull-down assays.
98 citations
TL;DR: It is suggested that Crm1-dependent nuclear export of M is central to RSV infection, representing the first report of such a mechanism for a paramyxovirus M protein and with important implications for relatedParamyxoviruses.
Abstract: The respiratory syncytial virus (RSV) matrix (M) protein is localized in the nucleus of infected cells early in infection but is mostly cytoplasmic late in infection. We have previously shown that M localizes in the nucleus through the action of the importin β1 nuclear import receptor. Here, we establish for the first time that M's ability to shuttle to the cytoplasm is due to the action of the nuclear export receptor Crm1, as shown in infected cells, and in cells transfected to express green fluorescent protein (GFP)-M fusion proteins. Specific inhibition of Crm1-mediated nuclear export by leptomycin B increased M nuclear accumulation. Analysis of truncated and point-mutated M derivatives indicated that Crm1-dependent nuclear export of M is attributable to a nuclear export signal (NES) within residues 194 to 206. Importantly, inhibition of M nuclear export resulted in reduced virus production, and a recombinant RSV carrying a mutated NES could not be rescued by reverse genetics. That this is likely to be due to the inability of a nuclear export deficient M to localize to regions of virus assembly is indicated by the fact that a nuclear-export-deficient GFP-M fails to localize to regions of virus assembly when expressed in cells infected with wild-type RSV. Together, our data suggest that Crm1-dependent nuclear export of M is central to RSV infection, representing the first report of such a mechanism for a paramyxovirus M protein and with important implications for related paramyxoviruses.
98 citations
TL;DR: Analysis of expressed sequence tags from a bovine oocyte cDNA library identified a new member of the importin alpha family, KPNA7, which has a strong binding affinity for the nuclear protein nucleoplasmin 2 relative to that of other importin alphas.
Abstract: Nuclear proteins such as transcription and chromatin remodeling factors are required for initiation of transcription in early embryos before embryonic genome activation. The nuclear transport of these proteins is mediated by transport factors such as importins. Through analysis of expressed sequence tags from a bovine oocyte cDNA library, we identified a new member of the importin alpha family (named importin alpha8). The cloned cDNA for bovine importin alpha8 (KPNA7) is 1817 base pair in length, encoding a protein of 522 amino acids that contains a conserved importin beta-binding domain and seven armadillo motifs. The RT-PCR analysis revealed that KPNA7 mRNA is specifically expressed in ovaries and mature oocytes. Real-time PCR demonstrated that KPNA7 expression in germinal vesicle (GV) oocytes is 33 to 2396 times higher than that of other importin alpha genes and that KPNA7 mRNA is abundant in GV and metaphase II oocytes, as well as in early-stage embryos collected before embryonic genome activation, but is barely detectable in morula- and blastocyst-stage embryos. Similarly, expression of KPNA7 protein is very high in oocytes and early embryos but is low in blastocysts. A glutathione S-transferase pull-down assay revealed that KPNA7 has a strong binding affinity for the nuclear protein nucleoplasmin 2 relative to that of other importin alphas. RNA interference experiments demonstrated that knockdown of KPNA7 in early embryos results in a decreased proportion of embryos developing to the blastocyst stage. These results suggest that KPNA7 may have an important role in the transport of essential nuclear proteins required for early embryogenesis.
93 citations
TL;DR: This study identified Exp4 (exportin 4) as an interaction partner of Sox2 in mouse embryonic stem cells and neural progenitors and shows that, besides its established function in nuclear export, Exp4 acts as a bona fide nuclear import receptor for Sox2 and SRY.
Abstract: SRY and other Sox-type transcription factors are important developmental regulators with various implications in human disease. In this study, we identified Exp4 (exportin 4) as an interaction partner of Sox2 in mouse embryonic stem cells and neural progenitors. We show that, besides its established function in nuclear export, Exp4 acts as a bona fide nuclear import receptor for Sox2 and SRY. Thus, Exp4 is an example of a nuclear transport receptor carrying distinct cargoes into different directions. In contrast to a published study, we observed that the import activity of Imp-α (importin-a) isoforms toward Sox2 is negligible. Instead, we found that Imp9 and the Imp-β/7 heterodimer mediate nuclear import of Sox2 in parallel to Exp4. Import signals for the three pathways overlap and include conserved residues in the Sox2 high-mobility group (HMG) box domain that are also critical for DNA binding. This suggests that nuclear import of Sox proteins is facilitated by several parallel import pathways.
TL;DR: It is shown that an MgcRacGAP mutant lacking its nuclear localizing signal (NLS) blocks nuclear translocation of p-STATs both in vitro and in vivo, and functions as both a critical mediator of STAT's tyrosine phosphorylation and an NLS-containing nuclear chaperone of p -STATs.
Abstract: In addition to their pleiotropic functions under physiological conditions, transcription factors STAT3 and STAT5 also have oncogenic activities, but how activated STATs are transported to the nucleus has not been fully understood. Here we show that an MgcRacGAP mutant lacking its nuclear localizing signal (NLS) blocks nuclear translocation of p-STATs both in vitro and in vivo. Unlike wild-type MgcRacGAP, this mutant did not promote complex formation of phosphorylated STATs (p-STATs) with importin α in the presence of GTP-bound Rac1, suggesting that MgcRacGAP functions as an NLS-containing nuclear chaperone. We also demonstrate that mutants of STATs lacking the MgcRacGAP binding site (the strand βb) are hardly tyrosine phosphorylated after cytokine stimulation. Intriguingly, mutants harboring small deletions in the C′-adjacent region (βb-βc loop region) of the strand βb became constitutively active with the enhanced binding to MgcRacGAP. The molecular basis of this phenomenon will be discussed, based on the computer-assisted tertiary structure models of STAT3. Thus, MgcRacGAP functions as both a critical mediator of STAT's tyrosine phosphorylation and an NLS-containing nuclear chaperone of p-STATs.
TL;DR: It is shown that the initial step of chromatin seeding is negatively regulated by importin beta, and suggested that initiation sites for NPC assembly contain single copies of Chromatin-bound ELYS/Nup107-160 and that the lateral oligomerization of these subunits depends on the recruitment of membrane components.
Abstract: The nuclear envelope of higher eukaryotic cells reforms at the exit from mitosis, in concert with the assembly of nuclear pore complexes (NPCs). The first step in postmitotic NPC assembly involves the “seeding” of chromatin with ELYS and the Nup107-160 complex. Subsequent steps in the assembly process are poorly understood and different mechanistic models have been proposed to explain the formation of the full supramolecular structure. Here, we show that the initial step of chromatin seeding is negatively regulated by importin β. Direct imaging of the chromatin attachment sites reveals single sites situated predominantly on the highest substructures of chromatin surface and lacking any sign of annular structures or oligomerized pre-NPCs. Surprisingly, the inhibition by importin β is only partially reversed by RanGTP. Importin β forms a high-molecular-weight complex with both ELYS and the Nup107-160 complex in cytosol. We suggest that initiation sites for NPC assembly contain single copies of chromatin-bound ELYS/Nup107-160 and that the lateral oligomerization of these subunits depends on the recruitment of membrane components. We predict that additional regulators, besides importin β and Ran, may be involved in coordinating the initial seeding of chromatin with subsequent steps in the NPC assembly pathway.
TL;DR: Ph phylogenetics and gene structure analysis suggest a parsimonious path for the evolution of the mammalian importin α gene family from an ancestral α1-like progenitor and establish a rational basis for a unified nomenclature of the importinα gene family.
Abstract: Importin αs are import receptors for nuclear localization signal-containing proteins Most animal importin αs assort into α1, α2, and α3 groups Studies in Drosophila melanogaster, Caenorhabditis elegans, and mouse suggest that the animal importin α gene family evolved from ancestral plant-like genes to serve paralog-specific roles in gametogenesis To explore this hypothesis we extended the phylogenetic analysis of the importin α gene family to nonbilateral animals and investigated whether animal-like genes occur in premetazoan taxa Maximum likelihood analysis suggests that animal-like importin α genes occur in the Choanoflaggelate Monosiga brevicollis and the amoebozoan Dictyostelium; however, both of these results are caused by long-branch attraction effects The absence of animal-like α genes in premetazoan taxa is consistent with the hypothesis that they duplicated and then specialized to function in animal gametogenesis The gene structures of the importin αs provide insight into how the animal importin α gene family may have evolved from the most likely ancestral gene Interestingly, animal α1s are more similar to plant and fungal α1-like sequences than they are to animal α2s or α3s We show that animal α1 genes share most of their introns with plant α1-like genes, and α2s and α3s share many more intron positions with each other than with the α1s Together, phylogenetics and gene structure analysis suggests a parsimonious path for the evolution of the mammalian importin α gene family from an ancestral α1-like progenitor Finally, these results establish a rational basis for a unified nomenclature of the importin α gene family
TL;DR: The eukaryotic cell nuclear transport system selectively mediates molecular trafficking to facilitate the regulation of cellular processes as discussed by the authors, where the components of this system include diverse transport factors such as importins and nuclear pore components.
TL;DR: The data show that shuttling of HIF-1alpha between cytoplasm and nucleus is a complex process involving several members of the nuclear transport receptor family.
TL;DR: It is shown that several importins mediate the nuclear import of the human Snail proteins and a unique nuclear localization signal (NLS) is identified, recognized by all the importins, that has been conserved during the evolution of the Snail family.
Abstract: Snail proteins are C2H2 class zinc finger transcription factors involved in different processes during embryonic development, as well as in several adult pathologies including cancer and organ fibrosis. The expression of Snail transcription factors is tightly regulated at the transcriptional level and their activity is modulated by their subcellular localization. Given the importance of this gene family in physiology and pathology, it is essential to understand the mechanisms by which Snail proteins are imported into or exported out of the nucleus. Here we show that several importins mediate the nuclear import of the human Snail proteins and we identify a unique nuclear localization signal (NLS), recognized by all the importins, that has been conserved during the evolution of the Snail family. This NLS is characterized by the presence of basic residues at defined positions in at least three consecutive zinc fingers. Interestingly, the consensus residues for importin-binding are also involved in DNA binding, suggesting that importins could prevent non-specific binding of these transcription factors to cytoplasmic polyanions. Importantly, the identified basic residues are also conserved in other families of C2H2 transcription factors whose nuclear localization requires the zinc finger region.
TL;DR: Evidence is provided for a molecular mechanism that might control nuclear import of specific ADARs and, in turn, nuclear RNA editing through controlled subcellular localization ofADARs, which in turn is governed by the coordinated local expression of importin α proteins and ADAR protein variants.
Abstract: The adenosine deaminases acting on RNA (ADARs) comprise a family of RNA editing enzymes that selectively modify single codons within RNA primary transcripts with often profound impact on protein function. Little is known about the mechanisms that regulate nuclear RNA editing activity. Editing levels show cell-type specific and developmental modulation that does not strictly coincide with observed expression levels of ADARs. Here, we provide evidence for a molecular mechanism that might control nuclear import of specific ADARs and, in turn, nuclear RNA editing. We identify an in vivo ADAR3 interaction partner, importin alpha 1 (KPNA2) that specifically recognizes an arginine-rich ADAR3 sequence motif and show that it acts as a functional nuclear localization sequence. Furthermore, whereas KPNA2, but not KPNA1 or KNPA3, recognizes the ADAR3 NLS, we observe the converse binding specificity with ADAR2. Interestingly, alternative splicing of ADAR2 pre-mRNA introduces an ADAR3-like NLS that alters the interaction profile with the importins. Thus, in vivo RNA editing might be regulated, in part, through controlled subcellular localization of ADARs, which in turn is governed by the coordinated local expression of importin alpha proteins and ADAR protein variants.
TL;DR: Results suggest that mDia2 continuously shuttles between the nucleus and the cytoplasm using specific nuclear transport machinery composing of importin-α/β and CRM1.
TL;DR: It is concluded that the interplay of the two negative regulators, transportin and importin beta, along with the positive regulator RanGTP, allows precise choreography of multiple cell cycle assembly events.
Abstract: Mitosis in higher eukaryotes is marked by the sequential assembly of two massive structures: the mitotic spindle and the nucleus. Nuclear assembly itself requires the precise formation of both nuclear membranes and nuclear pore complexes. Previously, importin alpha/beta and RanGTP were shown to act as dueling regulators to ensure that these assembly processes occur only in the vicinity of the mitotic chromosomes. We now find that the distantly related karyopherin, transportin, negatively regulates nuclear envelope fusion and nuclear pore assembly in Xenopus egg extracts. We show that transportin-and importin beta-initiate their regulation as early as the first known step of nuclear pore assembly: recruitment of the critical pore-targeting nucleoporin ELYS/MEL-28 to chromatin. Indeed, each karyopherin can interact directly with ELYS. We further define the nucleoporin subunit targets for transportin and importin beta and find them to be largely the same: ELYS, the Nup107/160 complex, Nup53, and the FG nucleoporins. Equally importantly, we find that transportin negatively regulates mitotic spindle assembly. These negative regulatory events are counteracted by RanGTP. We conclude that the interplay of the two negative regulators, transportin and importin beta, along with the positive regulator RanGTP, allows precise choreography of multiple cell cycle assembly events.
TL;DR: It is proposed that RCC1 reads the histone code created by caspase-activated Mst1 to initiate apoptosis by reducing the level of RanGTP in the nucleus.
Abstract: The guanine nucleotide exchange factor for Ran, RCC1, dynamically binds chromatin. During apoptosis, caspase-mediated activation of Mst1 induces histone H2B phosphorylation, which immobilizes RCC1 on the chromatin, leading to a reduction in nuclear RanGTP. A number of signalling pathways have been identified that regulate apoptosis, but the mechanism that initiates apoptosis remains incompletely understood. We have found that the nuclear RanGTP level is diminished during the early stages of apoptosis, which correlates with immobilization of RCC1 on the chromosomes. Furthermore, the expression of phosphomimetic histone H2B or caspase-activated Mst1 immobilizes RCC1 and causes reduction of nuclear RanGTP levels, which leads to inactivation of the nuclear transport machinery. As a consequence, nuclear localization signal (NLS)-containing proteins, including NF-κB–p65, remain bound to importins α and β in the cytoplasm. Knocking down Mst1 allows resumption of nuclear transport and the nuclear entry of NF-κB–p65, which have important roles in rescuing cells from apoptosis. Therefore, we propose that RCC1 reads the histone code created by caspase-activated Mst1 to initiate apoptosis by reducing the level of RanGTP in the nucleus.
TL;DR: The data represent the first study of a direct link between LaA mutant expression and reduced nuclear protein import, and both LaA mutants displayed similar effects on nuclear morphology and functions, despite their differences in disease severity.
TL;DR: It is concluded that the yeast nuclearExosome and the cytoplasmic exosome share a common stable core complex, but are decorated with quite a few differing peripheral proteins.
TL;DR: It is indicated that importin α is tethered at the postsynaptic density by binding to the NLS present in NR1-1a, which is activity dependent and released following NMDA receptor activation and phosphorylation rendering it available to bind soluble cargoes and transport them to the nucleus during transcription-dependent forms of neuronal plasticity.
Abstract: Synaptic plasticity, the capacity of neurons to change the strength of their connections with experience, provides a mechanism for learning and memory in the brain. Long-term plasticity requires new transcription, indicating that synaptically generated signals must be transported to the nucleus. Previous studies have described a role for importin nuclear transport adaptors in mediating the retrograde transport of signals from synapse to nucleus during plasticity. Here, we investigated the possibility that stimulus-induced translocation of importins from synapse to nucleus involves activity-dependent anchoring of importins at the synapse. We show that importin α binds to a nuclear localization signal (NLS) present in the cytoplasmic tail of NR1-1a. This interaction is disrupted by activation of NMDA receptors in cultured neurons and by stimuli that trigger late-phase, but not early-phase, long-term potentiation of CA3–CA1 synapses in acute hippocampal slices. In vitro PKC phosphorylation of GST-NR1-1a abolishes its ability to bind importin α in brain lysates, and the interaction of importin α and NR1 in neurons is modulated by PKC activity. Together, our results indicate that importin α is tethered at the postsynaptic density by binding to the NLS present in NR1-1a. This interaction is activity dependent, with importin α being released following NMDA receptor activation and phosphorylation rendering it available to bind soluble cargoes and transport them to the nucleus during transcription-dependent forms of neuronal plasticity.
TL;DR: It is reported here that p68 shuttles between the nucleus and the cytoplasm via a classical RanGTPase-dependent pathway.
Abstract: P68 RNA helicase is a prototypical DEAD box RNA helicase. The protein plays a very important role in early organ development and maturation. Consistent with the function of the protein in transcriptional regulation and pre-mRNA splicing, p68 was found to predominately localize in the cell nucleus. However, recent experiments demonstrate a transient cytoplasmic localization of the protein. We report here that p68 shuttles between the nucleus and the cytoplasm. The nucleocytoplasmic shuttling of p68 is mediated by two nuclear localization signal and two nuclear exporting signal sequence elements. Our experiments reveal that p68 shuttles via a classical RanGTPase-dependent pathway.
TL;DR: The subcellular distribution and the molecular mechanisms regulating the intracellular allocation of PHD1 and PHD2 are investigated and it is revealed that the exit ofPHD2 from the nucleus requires CRM1 and the N-terminal 100 amino acids of the protein.
TL;DR: The extended roles recently discovered for importins in the nervous system might also be relevant in non-neuronal cells, and the localized modes of importin regulation in neurons offer new avenues to interrogate their cytoplasmic functions.
TL;DR: In this article, the authors describe procedures for in vitro nuclear import in permeabilized mammalian cells, and for the preparation of recombinant transport factors (importin alpha, importin beta, Importin 7, transportin, Ran, NTF2) and other reagents commonly used in the assay.
Abstract: The trafficking of protein and RNA cargoes between the cytoplasm and the nucleus of eukaryotic cells, which is a major pathway involved in cell regulation, is mediated by nuclear transport sequences in the cargoes and by shuttling transport factors. The latter include receptors (karyopherins) that recognize the cargoes and carry them across the nuclear pore complex (NPC), and the small GTPase Ran, which modulates karyopherin-cargo binding. Nuclear import can be studied in vitro using digitonin-permeabilized cells, which are depleted of shuttling transport factors. Nuclear import can be reconstituted in the permeabilized cells with exogenous cytosol or with purified recombinant transport factors, and can be quantified by light microscopy of fluorescently labeled cargoes or by immunofluorescence staining. Here we describe procedures for in vitro nuclear import in permeabilized mammalian cells, and for the preparation of recombinant transport factors (importin alpha, importin beta, importin 7, transportin, Ran, NTF2) and other reagents commonly used in the assay. This assay provides means to characterize the molecular mechanisms of nuclear import and to study the import requirements of specific cargoes.
TL;DR: Fluorescence-based methods are presented to quantitatively address the physicochemical details of NLS recognition by the receptor protein importin α (Impα) in living cells and indicate that the subsaturation of Impα with the expressed NLS cargo regulates nuclear import rates in live cells.
TL;DR: The molecular mechanism by which the C protein of arthropod-borne flavivirus enters the nucleus is illustrated and the importance of importin-alpha/C protein interaction in the context of flaviv virus life-cycle is shown.