Showing papers on "Importin published in 2012"
TL;DR: It is established for the first time that ivermectin has potent antiviral activity towards both HIV-1 and dengue virus, both of which are strongly reliant on importin α/β nuclear import, with respect to the HIV- 1 integrase and NS5 (non-structural protein 5) polymerase proteins respectively.
Abstract: The movement of proteins between the cytoplasm and nucleus mediated by the importin superfamily of proteins is essential to many cellular processes, including differentiation and development, and is critical to disease states such as viral disease and oncogenesis We recently developed a high-throughput screen to identify specific and general inhibitors of protein nuclear import, from which ivermectin was identified as a potential inhibitor of importin α/β-mediated transport In the present study, we characterized in detail the nuclear transport inhibitory properties of ivermectin, demonstrating that it is a broad-spectrum inhibitor of importin α/β nuclear import, with no effect on a range of other nuclear import pathways, including that mediated by importin β1 alone Importantly, we establish for the first time that ivermectin has potent antiviral activity towards both HIV-1 and dengue virus, both of which are strongly reliant on importin α/β nuclear import, with respect to the HIV-1 integrase and NS5 (non-structural protein 5) polymerase proteins respectively Ivermectin would appear to be an invaluable tool for the study of protein nuclear import, as well as the basis for future development of antiviral agents
543 citations
TL;DR: It is demonstrated that cytoplasmic and nuclear actin pools are dynamically connected and the nuclear import and export mechanisms of actin are identified, which suggest an active transport mechanism in both directions.
Abstract: Besides its essential and well established role as a component of the cytoskeleton, actin is also present in the cell nucleus, where it has been linked to many processes that control gene expression. For example, nuclear actin regulates the activity of specific transcription factors, associates with all three RNA polymerases, and is a component of many chromatin remodelling complexes. Despite the fact that two export receptors, Crm1 and exportin 6, have been linked to nuclear export of actin, the mechanism by which actin enters the nucleus to elicit these essential functions has not been determined. It is also unclear whether actin is actively exchanged between the nucleus and the cytoplasm, and whether this connection has any functional significance for the cell. By applying a variety of live-cell imaging techniques we revealed that actin constantly shuttles in and out of the nucleus. The fast transport rates, which depend on the availability of actin monomers, suggest an active transport mechanism in both directions. Importantly, we identified importin 9 as the nuclear import factor for actin. Furthermore, our RNAi experiments showed that the active maintenance of nuclear actin levels by importin 9 is required for maximal transcriptional activity. Measurements of nuclear export rates and depletion studies also clarified that nuclear export of actin is mediated by exportin 6, and not by Crm1. These results demonstrate that cytoplasmic and nuclear actin pools are dynamically connected and identify the nuclear import and export mechanisms of actin.
217 citations
TL;DR: In this article, a long 3' untranslated region (3' UTR) was found to direct axonal localization of Importin β1, which is required for efficient retrograde signaling in injured axons.
169 citations
TL;DR: In living cells, depletion of Hikeshi inhibits heat shock-induced nuclear import of Hsp70s, reduces cell viability after heat shock stress, and significantly delays the attenuation and reversion of multiple heat shock -induced nuclear phenotypes.
135 citations
TL;DR: A novel role for PAK4 is uncovered in modulating intracellular translocation and signaling of β-catenin, which is involved in cytoskeletal reorganization, cell proliferation, gene transcription and oncogenic transformation.
113 citations
TL;DR: It is proposed that c-Myc and p53 counter each other in the regulation of elements within the nuclear transport machinery, thereby exerting opposing effects on the rate of ribosome biogenesis.
112 citations
TL;DR: The study defines an extended set of binding cavities on the importin‐α surface, and expands on recent observations that longer linker sequences are allowed, and that long‐range electrostatic complementarity can contribute to cNLS‐binding affinity.
Abstract: Classical nuclear localization signals (cNLSs), comprising one (monopartite cNLSs) or two clusters of basic residues connected by a 10–12 residue linker (bipartite cNLSs), are recognized by the nuclear import factor importin-α. The cNLSs bind along a concave groove on importin-α; however, specificity determinants of cNLSs remain poorly understood. We present a structural and interaction analysis study of importin-α binding to both designed and naturally occurring high-affinity cNLS-like sequences; the peptide inhibitors Bimax1 and Bimax2, and cNLS peptides of cap-binding protein 80. Our data suggest that cNLSs and cNLS-like sequences can achieve high affinity through maximizing interactions at the importin-α minor site, and by taking advantage of multiple linker region interactions. Our study defines an extended set of binding cavities on the importin-α surface, and also expands on recent observations that longer linker sequences are allowed, and that long-range electrostatic complementarity can contribute to cNLS-binding affinity. Altogether, our study explains the molecular and structural basis of the results of a number of recent studies, including systematic mutagenesis and peptide library approaches, and provides an improved level of understanding on the specificity determinants of a cNLS. Our results have implications for identifying cNLSs in novel proteins.
90 citations
TL;DR: It is shown that A3B nuclear import is an active process requiring at least one amino acid within an N-terminal motif analogous to the nuclear localization determinant of the antibody gene diversification enzyme AID (activation-induced cytosine deaminase), and highlighted the likelihood that specialized mechanisms exist to guide these enzymes to their respective physiological substrates and prevent gratuitous chromosomal DNA damage.
80 citations
TL;DR: Together, Nup358 functions as a cargo‐ and receptor‐specific assembly platform, increasing the efficiency of nuclear import of proteins through various mechanisms.
Abstract: In vertebrates, the nuclear pore complex (NPC), the gate for transport of macromolecules between the nucleus and the cytoplasm, consists of approximately 30 different nucleoporins (Nups). The Nup and SUMO E3-ligase Nup358/RanBP2 are the major components of the cytoplasmic filaments of the NPC. In this study, we perform a structure-function analysis of Nup358 and describe its role in nuclear import of specific proteins. In a screen for nuclear proteins that accumulate in the cytoplasm upon Nup358 depletion, we identified proteins that were able to interact with Nup358 in a receptor-independent manner. These included the importin α/β-cargo DBC-1 (deleted in breast cancer 1) and DMAP-1 (DNA methyltransferase 1 associated protein 1). Strikingly, a short N-terminal fragment of Nup358 was sufficient to promote import of DBC-1, whereas DMAP-1 required a larger portion of Nup358 for stimulated import. Neither the interaction of RanGAP with Nup358 nor its SUMO-E3 ligase activity was required for nuclear import of all tested cargos. Together, Nup358 functions as a cargo- and receptor-specific assembly platform, increasing the efficiency of nuclear import of proteins through various mechanisms.
75 citations
TL;DR: It is shown here that PPARα and PPARγ dynamically shuttle between nucleus and cytoplasm, although they constitutively and predominantly appear in nucleus, and that nuclear–cy toplasmic shuttling of PPARs is regulated by respective PPAR ligands and Ca2+ concentration.
Abstract: Peroxisome proliferator-activated receptors (PPARs) play important roles in diverse biological processes including metabolisms of sugars and lipids and differentiation of cells such as adipocytes. PPARs are transcription factors belonging to the ligand-dependent hormone receptor group. To function as transcription factors, PPARs translocate into nucleus where they associate with transcription apparatus. However, mechanisms underlying nuclear transport of PPARs remain enigmatic. We show here that PPARα and PPARγ dynamically shuttle between nucleus and cytoplasm, although they constitutively and predominantly appear in nucleus. With a series of truncation mutants, we identify that PPAR nuclear transport is mediated by at least two nuclear localization signals (NLSs) in DNA-binding domain (DBD)-hinge and activation function 1 (AF1) regions and their respective receptors including importinα/β, importin 7, and an unidentified receptor. PPARs also harbor two nuclear export signals in DBD and ligand-binding domain regions that are recognized by distinct export receptors, calreticulin and CRM1. Moreover, we show that nuclear-cytoplasmic shuttling of PPARs is regulated by respective PPAR ligands and Ca2+ concentration. Taken together, we suggest that the multiple pathways for the nuclear-cytoplasmic transport of PPARs regulate the biological functions of PPARs in response to external signals.
72 citations
TL;DR: It is found that nuclear‐retained importin α2 binds with DNase I‐sensitive nuclear component(s) and exhibits selective upregulation of mRNA encoding Serine/threonine kinase 35 (STK35) by microarray analysis and constitutive overexpression of STK35 proteins enhances caspase‐independent cell death under oxidative stress conditions.
Abstract: Various cellular stresses including oxidative stress induce a collapse of the Ran gradient, which causes accumulation of importin α in the nucleus and a subsequent block of nuclear protein import. However, it is unknown whether accumulated importin α performs roles in the nucleus after its migration in response to stress. In this study, we found that nuclear-retained importin α2 binds with DNase I-sensitive nuclear component(s) and exhibits selective upregulation of mRNA encoding Serine/threonine kinase 35 (STK35) by microarray analysis. Chromatin immunoprecipitation and promoter analysis demonstrated that importin α2 can access to the promoter region of STK35 and accelerate its transcription in response to hydrogen peroxide exposure. Furthermore, constitutive overexpression of STK35 proteins enhances caspase-independent cell death under oxidative stress conditions. These results collectively reveal that nuclear-localized importin α2 influences gene expression and contributes directly to cell fate outcomes including non-apoptotic cell death.
TL;DR: It is shown that knockdown of NUP153 prevents 53BP1, but not several other DDR factors, from entering the nuclei in the newly forming daughter cells, which translates into decreased IR-induced 53 BP1 focus formation, delayed DNA repair and impaired cell survival after IR.
Abstract: 53BP1 is a mediator of DNA damage response (DDR) and a tumor suppressor whose accumulation on damaged chromatin promotes DNA repair and enhances DDR signaling. Using foci formation of 53BP1 as a readout in two human cell lines, we performed an siRNA-based functional high-content microscopy screen for modulators of cellular response to ionizing radiation (IR). Here, we provide the complete results of this screen as an information resource, and validate and functionally characterize one of the identified 'hits': a nuclear pore component NUP153 as a novel factor specifically required for 53BP1 nuclear import. Using a range of cell and molecular biology approaches including live-cell imaging, we show that knockdown of NUP153 prevents 53BP1, but not several other DDR factors, from entering the nuclei in the newly forming daughter cells. This translates into decreased IR-induced 53BP1 focus formation, delayed DNA repair and impaired cell survival after IR. In addition, NUP153 depletion exacerbates DNA damage caused by replication stress. Finally, we show that the C-terminal part of NUP153 is required for effective 53BP1 nuclear import, and that 53BP1 is imported to the nucleus through the NUP153-importin-β interplay. Our data define the structure-function relationships within this emerging 53BP1-NUP153/importin-β pathway and implicate this mechanism in the maintenance of genome integrity.
TL;DR: Intrinsically disordered regions that undergo a stimulus-dependent disorder-to-order transition are common to a large number of signaling proteins and the N-terminal region of AtREM1.3 may constitute a regulatory domain, stabilizing these interactions.
TL;DR: This study demonstrates that nuclear translocation of JMY is controlled by the concentration of monomeric actin in the cytoplasm.
Abstract: Junction-mediating and regulatory protein (JMY) is a regulator of both transcription and actin filament assembly. In response to DNA damage, JMY accumulates in the nucleus and promotes p53-dependent apoptosis. JMY's actin-regulatory activity relies on a cluster of three actin-binding Wiskott-Aldrich syndrome protein homology 2 (WH2) domains that nucleate filaments directly and also promote nucleation activity of the Arp2/3 complex. In addition to these activities, we find that the WH2 cluster overlaps an atypical, bipartite nuclear localization sequence (NLS) and controls JMY's subcellular localization. Actin monomers bound to the WH2 domains block binding of importins to the NLS and prevent nuclear import of JMY. Mutations that impair actin binding, or cellular perturbations that induce actin filament assembly and decrease the concentration of monomeric actin in the cytoplasm, cause JMY to accumulate in the nucleus. DNA damage induces both cytoplasmic actin polymerization and nuclear import of JMY, and we find that damage-induced nuclear localization of JMY requires both the WH2/NLS region and importin β. On the basis of our results, we propose that actin assembly regulates nuclear import of JMY in response to DNA damage.
TL;DR: The configuration of the Nup153-Nup50 partnership suggests that the N up153 scaffold provides not just a means of pore targeting for Nup50 but also serves to provide a local environment that facilitates bringing N up50 and importin α together, as well as other soluble factors involved in transport.
TL;DR: It is shown that Rho-actin signalling induced by serum stimulation promotes the nuclear accumulation ofPhactr1, but not other Phactr family members, and this data support a role for PhActr1 in actomyosin assembly and suggest that PhactR1 G-actIn sensing allows its coordination with F-act in availability.
Abstract: Summary The Phactr family of PP1-binding proteins is implicated in human diseases including Parkinson’s, cancer and myocardial infarction. Each Phactr protein contains four G-actin binding RPEL motifs, including an N-terminal motif, abutting a basic element, and a C-terminal triple RPEL repeat, which overlaps a conserved C-terminus required for interaction with PP1. RPEL motifs are also found in the regulatory domains of the MRTF transcriptional coactivators, where they control MRTF subcellular localisation and activity by sensing signal-induced changes in G-actin concentration. However, whether G-actin binding controls Phactr protein function – and its relation to signalling – has not been investigated. Here, we show that Rho-actin signalling induced by serum stimulation promotes the nuclear accumulation of Phactr1, but not other Phactr family members. Actin binding by the three Phactr1 C-terminal RPEL motifs is required for Phactr1 cytoplasmic localisation in resting cells. Phactr1 nuclear accumulation is importin α-β dependent. G-actin and importin α-β bind competitively to nuclear import signals associated with the N- and C-terminal RPEL motifs. All four motifs are required for the inhibition of serum-induced Phactr1 nuclear accumulation when G-actin is elevated. G-actin and PP1 bind competitively to the Phactr1 C-terminal region, and Phactr1 C-terminal RPEL mutants that cannot bind G-actin induce aberrant actomyosin structures dependent on their nuclear accumulation and on PP1 binding. In CHL-1 melanoma cells, Phactr1 exhibits actin-regulated subcellular localisation and is required for stress fibre assembly, motility and invasiveness. These data support a role for Phactr1 in actomyosin assembly and suggest that Phactr1 G-actin sensing allows its coordination with F-actin availability.
TL;DR: A new model by whichPHD2-mediated hydroxylation of HIF-1&agr; predominantly occurs in the cell nucleus and is dependent on very dynamic subcellular trafficking of PHD2 is presented.
Abstract: Hypoxia-inducible factors (HIFs) regulate more than 200 genes involved in cellular adaptation to reduced oxygen availability. HIFs are heterodimeric transcription factors that consist of one of three HIF-α subunits and a HIF-β subunit. Under normoxic conditions the HIF-α subunit is hydroxylated by members of a family of prolyl-4-hydroxylase domain (PHD) proteins, PHD1, PHD2 and PHD3, resulting in recognition by von-Hippel-Lindau protein, ubiquitylation and proteasomal degradation. It has been suggested that PHD2 is the key regulator of HIF-1α stability in vivo. Previous studies on the intracellular distribution of PHD2 have provided evidence for a predominant cytoplasmic localisation but also nuclear activity of PHD2. Here, we investigated functional nuclear transport signals in PHD2 and identified amino acids 196-205 as having a crucial role in nuclear import, whereas amino acids 6-20 are important for nuclear export. Fluorescence resonance energy transfer (FRET) showed that an interaction between PHD2 and HIF-1α occurs in both the nuclear and cytoplasmic compartments. However, a PHD2 mutant that is restricted to the cytoplasm does not interact with HIF-1α and shows less prolyl hydroxylase activity for its target HIF-1α than wild-type PHD2 located in the nucleus. Here, we present a new model by which PHD2-mediated hydroxylation of HIF-1α predominantly occurs in the cell nucleus and is dependent on very dynamic subcellular trafficking of PHD2.
TL;DR: Alterations in the levels of the 70-kDa fragment during development and in the adult after spinal cord injury or in a mouse model of Alzheimer disease suggest that this fragment is functionally implicated in development, regeneration, neurodegeneration, tumorigenesis, and possibly synaptic plasticity in the mature nervous system.
TL;DR: ICP27 competes with some host cell transport receptors for binding, resulting in inhibition of host pathways and supporting ICP27-mediated transport of HSV-1 mRNAs, and is proposed to provide additional binding sites at the nuclear pore for I CP27 shuttling.
TL;DR: It is shown that leucine‐rich repeat containing 59 (LRRC59) is strictly required for nuclear import of exogenous FGF1, and that the localization of LRRC59 to the NE depends on Kpnβ1.
Abstract: Fibroblast growth factor 1 (FGF1) taken up by cells into endocytic vesicles can be translocated across vesicular membranes into the cytosol and the nucleus where it has a growth regulatory activity. Previously, leucine-rich repeat containing 59 (LRRC59) was identified as an intracellular binding partner of FGF1, but its biological role remained unknown. Here, we show that LRRC59 is strictly required for nuclear import of exogenous FGF1. siRNA-mediated depletion of LRRC59 did not inhibit the translocation of FGF1 into cytosol, but blocked the nuclear import of FGF1. We also found that an nuclear localization sequence (NLS) in FGF1, Ran GTPase, karyopherin-α1 (Kpnα1), and Kpnβ1 were required for nuclear import of FGF1. Nuclear import of exogenous FGF2, which depends on CEP57/Translokin, was independent of LRRC59, but was dependent on Kpnα1 and Kpnβ1, while the nuclear import of FGF1 was independent of CEP57. LRRC59 is a membrane-anchored protein that localizes to the endoplasmic reticulum (ER) and the nuclear envelope (NE). We found that LRRC59 possesses NLS-like sequences in its cytosolic part that can mediate nuclear import of soluble LRRC59 variants, and that the localization of LRRC59 to the NE depends on Kpnβ1. We propose that LRRC59 facilitates transport of cytosolic FGF1 through nuclear pores by interaction with Kpns and movement of LRRC59 along the ER and NE membranes.
TL;DR: This work found Lys157 is critical to β-arrestin-1 nuclear localization and its regulation of NF-κB activation and identified a seven-residue candidate nuclear localization sequence (NLS) based on sequence analysis and mutational analysis.
Journal Article•
TL;DR: This review will elaborate the role of individual viral factors and some of current insights on their molecular mechanism(s) associated with HIV-1 nuclear import, and discuss the importance of nuclear import for subsequent step of viral DNA integration.
Abstract: The human immunodeficiency virus 1 (HIV-1) synthesizes its genomic DNA in cytoplasm as soon as it enters the cell. The newly synthesized DNA remains associated with viral/cellular proteins as a high molecular weight pre-integration complex (PIC), which precludes passive diffusion across intact nuclear membrane. However, HIV-1 successfully overcomes nuclear membrane barrier by actively delivering its DNA into nucleus with the help of host nuclear import machinery. Such ability allows HIV-1 to productively infect non-dividing cells as well as dividing cells at interphase. Further, HIV-1 nuclear import is also found important for the proper integration of viral DNA. Thus, nuclear import plays a crucial role in establishment of infection and disease progression. While several viral components, including matrix, viral protein R, integrase, capsid, and central DNA flap are implicated in HIV-1 nuclear import, their molecular mechanism remains poorly understood. In this review, we will elaborate the role of individual viral factors and some of current insights on their molecular mechanism(s) associated with HIV-1 nuclear import. In addition, we will discuss the importance of nuclear import for subsequent step of viral DNA integration. Hereby we aim to further our understanding on molecular mechanism of HIV-1 nuclear import and its potential usefulness for anti-HIV-1 strategies.
TL;DR: In this article, the authors used quantitative live cell imaging of 3 isogenic nontransformed/transformed cell pairs to show that nuclear accumulation of nuclear localization signal (NLS)-containing proteins, but not their NLS-mutated derivatives, is increased up to 7-fold in MCF10CA1h human epithelial breast carcinoma cells and in simian virus 40 (SV40)-transformed fibroblasts of human and monkey origin.
Abstract: Fundamental to eukaryotic cell function, nucleocytoplasmic transport can be regulated at many levels, including through modulation of the importin/exportin (Imp/Exp) nuclear transport machinery itself. Although Imps/Exps are overexpressed in a number of transformed cell lines and patient tumor tissues, the efficiency of nucleocytoplasmic transport in transformed cell types compared with nontransformed cells has not been investigated. Here we use quantitative live cell imaging of 3 isogenic nontransformed/transformed cell pairs to show that nuclear accumulation of nuclear localization signal (NLS)-containing proteins, but not their NLS-mutated derivatives, is increased up to 7-fold in MCF10CA1h human epithelial breast carcinoma cells and in simian virus 40 (SV40)-transformed fibroblasts of human and monkey origin, compared with their nontransformed counterparts. The basis for this appears to be a significantly faster rate of nuclear import in transformed cell types, as revealed by analysis using fluorescence recovery after photobleaching for the human MCF10A/MCF10CA1h cell pair. Nuclear accumulation of NLS/nuclear export signal-containing (shuttling) proteins was also enhanced in transformed cell types, experiments using the nuclear export inhibitor leptomycin B demonstrating that efficient Exp-1-mediated nuclear export was not impaired in transformed compared with nontransformed cells. Enhanced nuclear import and export efficiencies were found to correlate with 2- to 4-fold higher expression of specific Imps/Exps in transformed cells, as indicated by quantitative Western blot analysis, with ectopic expression of Imps able to enhance NLS nuclear accumulation levels up to 5-fold in nontransformed MCF10A cells. The findings indicate that transformed cells possess altered nuclear transport properties, most likely due to the overexpression of Imps/Exps. The findings have important implications for the development of tumor-specific drug nanocarriers in anticancer therapy.
TL;DR: Importin-β binds RANBP2 and RANGAP1 and antagonizes CRM1 activity during RangAP1 recruitment to kinetochores.
Abstract: Importin-β is the main vector for interphase nuclear protein import and plays roles after nuclear envelope breakdown. Here we show that importin-β regulates multiple aspects of mitosis via distinct domains that interact with different classes of proteins in human cells. The C-terminal region (which binds importin-α) inhibits mitotic spindle pole formation. The central region (harboring nucleoporin-binding sites) regulates microtubule dynamic functions and interaction with kinetochores. Importin-β interacts through this region with NUP358/RANBP2, which in turn binds SUMO-conjugated RANGAP1 in nuclear pores. We show that this interaction continues after nuclear pore disassembly. Overexpression of importin-β, or of the nucleoporin-binding region, inhibited RANGAP1 recruitment to mitotic kinetochores, an event that is known to require microtubule attachment and the exportin CRM1. Co-expressing either importin-β–interacting RANBP2 fragments, or CRM1, restored RANGAP1 to kinetochores and rescued importin-β–dependent mitotic dynamic defects. These results reveal previously unrecognized importin-β functions at kinetochores exerted via RANBP2 and opposed by CRM1.
TL;DR: It is shown that the novel specific NLS brings to the cell nucleus not only the “nuclear” isoform of myosin I (NM1 protein) but also its “cytoplasmic”isoform (Myo1c protein), which opens a new field for exploring functions of this molecular motor in nuclear processes, and for exploring the signals between cy toplasm and the nucleus.
Abstract: Background
Nuclear myosin I (NM1) was the first molecular motor identified in the cell nucleus. Together with nuclear actin, they participate in crucial nuclear events such as transcription, chromatin movements, and chromatin remodeling. NM1 is an isoform of myosin 1c (Myo1c) that was identified earlier and is known to act in the cytoplasm. NM1 differs from the “cytoplasmic” myosin 1c only by additional 16 amino acids at the N-terminus of the molecule. This amino acid stretch was therefore suggested to direct NM1 into the nucleus.
Methodology/Principal Findings
We investigated the mechanism of nuclear import of NM1 in detail. Using over-expressed GFP chimeras encoding for truncated NM1 mutants, we identified a specific sequence that is necessary for its import to the nucleus. This novel nuclear localization sequence is placed within calmodulin-binding motif of NM1, thus it is present also in the Myo1c. We confirmed the presence of both isoforms in the nucleus by transfection of tagged NM1 and Myo1c constructs into cultured cells, and also by showing the presence of the endogenous Myo1c in purified nuclei of cells derived from knock-out mice lacking NM1. Using pull-down and co-immunoprecipitation assays we identified importin beta, importin 5 and importin 7 as nuclear transport receptors that bind NM1. Since the NLS sequence of NM1 lies within the region that also binds calmodulin we tested the influence of calmodulin on the localization of NM1. The presence of elevated levels of calmodulin interfered with nuclear localization of tagged NM1.
Conclusions/Significance
We have shown that the novel specific NLS brings to the cell nucleus not only the “nuclear” isoform of myosin I (NM1 protein) but also its “cytoplasmic” isoform (Myo1c protein). This opens a new field for exploring functions of this molecular motor in nuclear processes, and for exploring the signals between cytoplasm and the nucleus.
TL;DR: It is shown that huntingtin has a karyopherin β2-dependent proline-tyrosine (PY)-NLS in the amino terminus of the protein, and it is proposed that this huntingtin PY-NLS may comprise a new class of multiple import factor-dependent NLSs with an internal structural component that may regulate NLS activity.
TL;DR: This study confirms a cancer-specific nuclear translocation of TβRI and demonstrates its potential function in regulating nuclear RNA processing, as well as a novel gain-of-function mechanism of TGF-β signaling in cancer.
Abstract: Signaling of transforming growth factor β (TGF-β) is redirected in cancer to promote malignancy, but how TGF-β function is altered in a transformed cell is not fully understood. We investigated TGF-β signaling by profiling proteins that differentially bound to type I TGF-β receptor (TβRI) in nontransformed, HER2-transformed, and HER2-negative breast cancer cells using immunoprecipitation followed by protein identification. Interestingly, several nuclear proteins implicated in posttranscriptional RNA processing were uniquely identified in the TβRI coprecipitates from HER2-transformed cells. Ligand-inducible nuclear translocation of TβRI was observed only in transformed cells, and the translocation required importin β1, nucleolin, and Smad2/3. This trafficking was dependent on the high Ran GTPase activity resulting from oncogenic transformation. In the nucleus, TβRI associated with purine-rich RNA sequences in a synergistic manner with the RNA-binding factor hnRNP A1. We further found that nuclear translocation of TβRI specifically induced epidermal growth factor receptor (EGFR) transcript isoform c, which encodes a soluble EGFR protein, through alternative splicing or 3'-end processing. Our study confirms a cancer-specific nuclear translocation of TβRI and demonstrates its potential function in regulating nuclear RNA processing, as well as a novel gain-of-function mechanism of TGF-β signaling in cancer.
TL;DR: The present analysis provides evidence of the necessity for and specificity of each nuclear transport factor and for nucleoporins during germ cell differentiation.
TL;DR: It is highlighted that miRNAs are also transported from the cytoplasm to the nucleus, where they function in a noncanonical manner to regulate noncoding RNAs.
01 Jan 2012
TL;DR: In this paper, the authors provided insight into the mechanism and regulation of hunting-tin nuclear entry and proposed a new class of multiple import factor-dependent NLSs with an internal struc-tural component that may regulate NLS activity.
Abstract: This work provides insight into the mechanism and regulation of huntingtin nuclear entry.Among the known pathways of protein nuclear import, thekaryopherin 2/transportinpathwayisonlythesecondtohaveadefined nuclear localization signal (NLS) consensus. Hunting-tin,a350-kDaprotein,hasdefinedrolesinthenucleus,aswellasa CRM1/exportin-dependent nuclear export signal; however,the NLS and exact pathway of import have remained elusive.Here, using a live cell assay and affinity chromatography, weshow that huntingtin has a karyopherin 2-dependent proline-tyrosine (PY)-NLS in the amino terminus of the protein. ThisNLS comprises three consensus components: a basic chargedsequence,adownstreamconservedarginine,andaPYsequence.UnliketheclassicPY-NLS,whichhasanunstructuredinterven-ingsequencebetweentheconsensuscomponents,weshowthata A second NLS consensus has been defined for karyopherinsheetstructuredregionseparatingtheconsensuselementsiscritical for huntingtin NLS function. The huntingtin PY-NLS isalso capable of import through the importin/karyopherin 1pathway but was not functional in all cell types tested. We pro-pose that this huntingtin PY-NLS may comprise a new class ofmultiple import factor-dependent NLSs with an internal struc-tural component that may regulate NLS activity.