SEARCH · PubMed Health

Results for “Nuclear Export Signals”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 127 records · Page 7Linked to original sources

PubMed1999

Nuclear export of LIM-kinase 1, mediated by two leucine-rich nuclear-export signals within the PDZ domain.

LIM-kinase 1 (LIMK1) is a serine/threonine kinase that phosphorylates cofilin and regulates actin-filament dynamics. LIMK1, which contains two LIM domains and a single PDZ domain, localizes predominantly in the cytoplasm, but its mutant, deleted with the PDZ domain, localizes mainly in the nucleus, thereby indicating that the PDZ domain plays a role in the cytoplasmic localization of LIMK1. Here we provide evidence that the PDZ domain of LIMK1 contains two functional leucine-rich nuclear-export signals (NESs). The PDZ domain of LIMK1 fused with glutathione S-transferase (GST-PDZ), when injected into the nucleus, was rapidly excluded from the nucleus, but its mutant with replacements of conserved hydrophobic residues in two putative NESs by alanines remained in the nucleus. The nuclear export of GST-PDZ was sensitive to leptomycin B (LMB), a specific inhibitor of nuclear export mediated by leucine-rich NESs. Malfunctional mutation of two NESs or LMB treatment prevented the nuclear export of full-length LIMK1 and induced its nuclear accumulation. These results suggest that the predominant localization of LIMK1 in the cytoplasm is supported by two NESs within the PDZ domain and that LIMK1 normally shuttles between the cytoplasm and the nucleus. We also provide evidence that a short basic cluster sequence within the protein-kinase domain is involved in the nuclear import of LIMK1.

Amino Acid Sequence↗

PubMed1999

A leucine-rich nuclear export signal in the p53 tetramerization domain: regulation of subcellular localization and p53 activity by NES masking.

Appropriate subcellular localization is crucial for regulating p53 function. We show that p53 export is mediated by a highly conserved leucine-rich nuclear export signal (NES) located in its tetramerization domain. Mutation of NES residues prevented p53 export and hampered tetramer formation. Although the p53-binding protein MDM2 has an NES and has been proposed to mediate p53 export, we show that the intrinsic p53 NES is both necessary and sufficient for export. This report also demonstrates that the cytoplasmic localization of p53 in neuroblastoma cells is due to its hyperactive nuclear export: p53 in these cells can be trapped in the nucleus by the export-inhibiting drug leptomycin B or by binding a p53-tetramerization domain peptide that masks the NES. We propose a model in which regulated p53 tetramerization occludes its NES, thereby ensuring nuclear retention of the DNA-binding form. We suggest that attenuation of p53 function involves the conversion of tetramers into monomers or dimers, in which the NES is exposed to the proteins which mediate their export to the cytoplasm.

3T3 Cells↗

PubMed2004

Alternative splicing of Staufen2 creates the nuclear export signal for CRM1 (Exportin 1).

Mammalian Staufen2 (Stau2), a brain-specific double-stranded RNA-binding protein, is involved in the localization of mRNA in neurons. To gain insights into the function of Stau2, the subcellular localization of Stau2 isoforms fused to the green fluorescence protein was examined. Fluorescence microscopic analysis showed that Stau2 functions as a nucleocytoplasmic shuttle protein. The nuclear export of the 62-kDa isoform of Stau2 (Stau2(62)) is mediated by the double-stranded RNA-binding domain 3 (RBD3) because a mutation to RBD3 led to nuclear accumulation. On the other hand, the shorter isoform of Stau2, Stau2(59), is exported from the nucleus by two distinct pathways, one of which is RBD3-mediated and the other of which is CRM1 (exportin 1)-dependent. The nuclear export signal recognized by CRM1 was found to be located in the N-terminal region of Stau2(59). These results suggest that Stau2 may carry a variety of RNAs out of the nucleus, using the two export pathways. The present study addresses the issue of why plural Stau2 isoforms are expressed in neurons.

Active Transport, Cell Nucleus↗

PubMed2000

A nuclear export signal in the N-terminal regulatory domain of IkappaBalpha controls cytoplasmic localization of inactive NF-kappaB/IkappaBalpha complexes.

Appropriate subcellular localization is crucial for regulation of NF-kappaB function. Herein, we show that latent NF-kappaB complexes can enter and exit the nucleus in preinduction states. The nuclear export inhibitor leptomycin B (LMB) sequestered NF-kappaB/IkappaBalpha complexes in the nucleus. Using deletion and site-directed mutagenesis, we identified a previously uncharacterized nuclear export sequence in residues 45-54 of IkappaBalpha that was required for cytoplasmic localization of inactive complexes. This nuclear export sequence also caused nuclear exclusion of heterologous proteins in a LMB-sensitive manner. Importantly, a LMB-insensitive CRM1 mutant (Crm1-K1) abolished LMB-induced nuclear accumulation of the inactive complexes. Moreover, a cell-permeable p50 NF-kappaB nuclear localization signal peptide also blocked these LMB effects. These results suggest that NF-kappaB/IkappaBalpha complexes shuttle between the cytoplasm and nucleus by a nuclear localization signal-dependent nuclear import and a CRM1-dependent nuclear export. The LMB-induced nuclear complexes could not bind DNA and were inaccessible to signaling events, because LMB inhibited NF-kappaB activation without affecting the subcellular localization of upstream kinases IKKbeta and NIK. Our findings indicate that the dominant nuclear export over nuclear import contributes to the largely cytoplasmic localization of the inactive complexes to achieve efficient NF-kappaB activation by extracellular signals.

Amino Acid Motifs↗

PubMed2002

Mapping of functional regions in the amino-terminal portion of the herpes simplex virus ICP27 regulatory protein: importance of the leucine-rich nuclear export signal and RGG Box RNA-binding domain.

Infected-cell protein 27 (ICP27) is an essential herpes simplex virus type 1 (HSV-1) regulatory protein that activates a subset of viral delayed-early and late genes, at least in part through posttranscriptional mechanisms. Previous studies have shown that the amino (N)-terminal half of the protein contains important functional regions, including a leucine-rich nuclear export signal (NES). However, to date, the phenotype of an HSV-1 ICP27 NES mutant has not been reported. In this study, we engineered and characterized dLeu, an HSV-1 deletion mutant that specifically lacks ICP27's NES (amino acids 6 to 19). The phenotype of dLeu was analyzed alongside those of eight other ICP27 N-terminal deletion mutants. We found that in Vero cells, dLeu displays modest defects in viral gene expression and an approximately 100-fold reduction in the production of viral progeny. Unlike wild-type (WT) ICP27, which exhibits a cytoplasmic distribution in addition to its predominant nuclear localization, dLeu ICP27 is highly restricted to the cell nucleus. This strongly suggests that the N-terminal leucine-rich sequence functions as an NES during viral infection. Our analysis of dLeu and the other mutants has enabled us to genetically define the regions in the N-terminal 200 residues of ICP27 which are required for efficient viral growth in Vero cells. Only two regions appear to be important: (i) the leucine-rich NES and (ii) the RGG box RNA-binding domain, encoded by residues 139 to 153. A virus lacking the RGG box-encoding sequence, d4-5, has a phenotype similar to that of dLeu in that it displays modest defects in viral gene expression and grows poorly. Interestingly, deletion of both the NES and RGG box, as well as the sequences in between, is lethal. The resulting virus, d1-5, displays severe defects in viral gene expression and DNA synthesis and is unable to produce significant amounts of infectious progeny. Therefore, the N-terminal portion of ICP27 contains at least two functional domains which collectively are absolutely essential for viral infection.

Amino Acid Sequence↗

PubMed2004

Identification of the nuclear export signal and STAT-binding domains of the Nipah virus V protein reveals mechanisms underlying interferon evasion.

The V proteins of Nipah virus and Hendra virus have been demonstrated to bind to cellular STAT1 and STAT2 proteins to form high-molecular-weight complexes that inhibit interferon (IFN)-induced antiviral transcription by preventing STAT nuclear accumulation. Analysis of the Nipah virus V protein has revealed a region between amino acids 174 and 192 that functions as a CRM1-dependent nuclear export signal (NES). This peptide is sufficient to complement an export-defective human immunodeficiency virus Rev protein, and deletion and substitution mutagenesis revealed that this peptide is necessary for both V protein shuttling and cytoplasmic retention of STAT1 and STAT2 proteins. However, the NES is not required for V-dependent IFN signaling inhibition. IFN signaling is blocked primarily by interaction between Nipah virus V residues 100 to 160 and STAT1 residues 509 to 712. Interaction with STAT2 requires a larger Nipah virus V segment between amino acids 100 and 300, but deletion of residues 230 to 237 greatly reduced STAT2 coprecipitation. Further, V protein interactions with cellular STAT1 is a prerequisite for STAT2 binding, and sequential immunoprecipitations demonstrate that V, STAT1, and STAT2 can form a tripartite complex. These findings characterize essential regions for Henipavirus V proteins that represent potential targets for therapeutic intervention.

Active Transport, Cell Nucleus↗

PubMed1996

Protein sequence requirements for function of the human T-cell leukemia virus type 1 Rex nuclear export signal delineated by a novel in vivo randomization-selection assay.

The Rex protein of human T-cell leukemia virus type 1, like the functionally equivalent Rev protein of human immunodeficiency virus type 1, contains a leucine-rich activation domain that specifically interacts with the human nucleoporin-like Rab/hRIP cofactor. Here, this Rex sequence is shown to function also as a protein nuclear export signal (NES). Rex sequence libraries containing randomized forms of the activation domain/NES were screened for retention of the ability to bind Rab/hRIP by using the yeast two-hybrid assay. While the selected sequences differed widely in primary sequence, all were functional as Rex activation domains. In contrast, randomized sequences that failed to bind Rab/hRIP lacked Rex activity. The selected sequences included one with homology to the Rev activation domain/NES and a second that was similar to the NES found in the cellular protein kinase inhibitor alpha. A highly variant, yet fully active, activation domain sequence selected on the basis of Rab/hRIP binding retained full NES function even though this sequence preserved only a single leucine residue. In contrast, nonfunctional activation domain mutants that were unable to bind Rab/hRIP had also lost NES function. These data demonstrate that NES activity is a defining characteristic of the activation domains found in the Rev/Rex class of retroviral regulatory proteins and strongly support the hypothesis that the Rab/hRIP cofactor plays a critical role in mediating the biological activity of these NESs. In addition, these data suggest a consensus sequence for NESs of the Rev/Rex class.

Amino Acid Sequence↗

PubMed2003

Hic-5 communicates between focal adhesions and the nucleus through oxidant-sensitive nuclear export signal.

hic-5 was originally isolated as an H(2)O(2)-inducible cDNA clone whose product was normally found at focal adhesions. In this study, we found that Hic-5 accumulated in the nucleus in response to oxidants such as H(2)O(2). Other focal adhesion proteins including paxillin, the most homologous to Hic-5, remained in the cytoplasm. Mutation analyses revealed that the C- and N-terminal halves of Hic-5 contributed to its nuclear localization in a positive and negative manner, respectively. After the finding that leptomycin B (LMB), an inhibitor of nuclear export signal (NES), caused Hic-5 to be retained in the nucleus, Hic-5 was demonstrated to harbor NES in the N-terminal, which was sensitive to oxidants, thereby regulating the nuclear accumulation of Hic-5. NES consisted of a leucine-rich stretch and two cysteines with a limited similarity to Yap/Pap-type NES. In the nucleus, Hic-5 was suggested to participate in the gene expression of c-fos. Using dominant negative mutants, we found that Hic-5 was actually involved in endogenous c-fos gene expression upon H(2)O(2) treatment. Hic-5 was thus proposed as a focal adhesion protein with the novel aspect of shuttling between focal adhesions and the nucleus through an oxidant-sensitive NES, mediating the redox signaling directly to the nucleus.

Active Transport, Cell Nucleus↗

PubMed2000

The nuclear export signal within the E4orf6 protein of adenovirus type 5 supports virus replication and cytoplasmic accumulation of viral mRNA.

During the late phase of adenovirus infection, viral mRNA is efficiently transported from the nucleus to the cytoplasm while most cellular mRNA species are retained in the nucleus. Two viral proteins, E1B-55 kDa and E4orf6, are both necessary for these effects. The E4orf6 protein of adenovirus type 5 binds and relocalizes E1B-55 kDa, and the complex of the two proteins was previously shown to shuttle continuously between the nucleus and cytoplasm. Nucleocytoplasmic transport of the complex is achieved by a nuclear export signal (NES) within E4orf6. Mutation of this signal sequence severely reduces the ability of the E1B-55 kDa-E4orf6 complex to leave the nucleus. Here, we examined the role of functional domains within E4orf6 during virus infection. E4orf6 or mutants derived from it were transiently expressed, followed by infection with recombinant adenovirus lacking the E4 region and determination of virus yield. An arginine-rich putative alpha helix near the carboxy terminus of E4orf6 contributes to E1B-55 kDa binding and relocalization as well as to the synthesis of viral DNA, mRNA, and proteins. Further mutational analysis revealed that mutation of the NES within E4orf6 considerably reduces its ability to support virus production. The same effect was observed when nuclear export was blocked with a competitor. Further, a functional NES within E4orf6 contributed to the efficiency of late virus protein synthesis and viral DNA replication, as well as total and cytoplasmic accumulation of viral late mRNA. Our data support the view that NES-mediated nucleocytoplasmic shuttling strongly enhances most, if not all, intracellular activities of E4orf6 during the late phase of adenovirus infection.

Adenovirus E1A Proteins↗

PubMed2005

Nrf2 Possesses a redox-insensitive nuclear export signal overlapping with the leucine zipper motif.

Basic leucine zipper (bZIP) protein Nrf2 is a key transcription factor mediating the antioxidant response. Under homeostatic conditions Nrf2 is anchored to cysteine-rich Keap1 and sequestered in the cytoplasm. When challenged with oxidative stress, Keap1 functions as a redox-sensitive switch and releases Nrf2. Subsequently, Nrf2 translocates into the cell nucleus and binds to a cis-acting enhancer called the antioxidant response element located in the promoters of a battery of cytoprotective genes and initiates their transcription. In this study we identify a canonical nuclear export signal (NES) ((537)LKKQLSTLYL(546)) located in the leucine zipper (ZIP) domain of the Nrf2 protein. The enhanced green fluorescent protein-tagged ZIP domain of Nrf2 (amino acids 503-589) exhibited a CRM1-dependent cytosolic distribution that could be abrogated by site-directed mutations or treatment with the nuclear export inhibitor, leptomycin B. Ectotopic expression of the Nrf2-NES could also exclude the GAL4 DNA binding domain into the cytoplasm. This NES overlapped with the ZIP motif in Nrf2, suggesting that the formation of heterodimers between Nrf2 and other bZIP proteins may simultaneously mask the NES and attenuate Nrf2 nuclear export. The Nrf2-NES appeared to be redox-insensitive. Neither oxidants (sulforaphane and diethyl maleate) nor reducing compounds (N-acetyl-l-cysteine and reducing glutathione) could disrupt the cytosolic distribution of Nrf2zip. Because Nrf2 activation is generally redox-sensitive, the redox insensitivity of this Nrf2-NES indicates the importance of Keap1 retention as a key rate-limiting step in Nrf2 activation. The characterization of the Nrf2 NES may help decipher the mechanisms governing nuclear localization and subsequent transcriptional activation of Nrf2-mediated cytoprotective genes.

Amino Acid Sequence↗

PubMed2004

Both nuclear-cytoplasmic shuttling of the dual specificity phosphatase MKP-3 and its ability to anchor MAP kinase in the cytoplasm are mediated by a conserved nuclear export signal.

MAP kinase phosphatase (MKP)-3 is a cytoplasmic dual specificity protein phosphatase that specifically binds to and inactivates the ERK1/2 MAP kinases in mammalian cells. However, the molecular basis of the cytoplasmic localization of MKP-3 or its physiological significance is unknown. We have used MKP-3-green fluorescent protein fusions in conjunction with leptomycin B to show that the cytoplasmic localization of MKP-3 is mediated by a chromosome region maintenance-1 (CRM1)-dependent nuclear export pathway. Furthermore, the nuclear translocation of MKP-3 seen in the presence of leptomycin B is mediated by an active process, indicating that MKP-3 shuttles between the nucleus and cytoplasm. The amino-terminal noncatalytic domain of MKP-3 is both necessary and sufficient for nuclear export of the phosphatase and contains a single functional leucine-rich nuclear export signal (NES). Even though this domain of the protein also mediates the binding of MKP-3 to MAP kinase, we show that mutations of the kinase interaction motif which abrogate ERK2 binding do not affect MKP-3 localization. Conversely, mutation of the NES does not affect either the binding or phosphatase activity of MKP-3 toward ERK2, indicating that the kinase interaction motif and NES function independently. Finally, we demonstrate that the ability of MKP-3 to cause the cytoplasmic retention of ERK2 requires both a functional kinase interaction motif and NES. We conclude that in addition to its established function in the regulated dephosphorylation and inactivation of MAP kinase, MKP-3 may also play a role in determining the subcellular localization of its substrate. Our results reinforce the idea that regulatory proteins such as MKP-3 may play a key role in the spatio-temporal regulation of MAP kinase activity.

3T3 Cells↗

PubMed2004

The yeast splicing factor Prp40p contains functional leucine-rich nuclear export signals that are essential for splicing.

To investigate the function of the essential U1 snRNP protein Prp40p, we performed a synthetic lethal screen in Saccharomyces cerevisiae. Using an allele of PRP40 that deletes 47 internal residues and causes only a slight growth defect, we identified aphenotypic mutations in three distinct complementation groups that conferred synthetic lethality. The synthetic phenotypes caused by these mutations were suppressed by wild-type copies of CRM1 (XPO1), YNL187w, and SME1, respectively. The strains whose synthetic phenotypes were suppressed by CRM1 contained no mutations in the CRM1 coding sequence or promoter. This indicates that overexpression of CRM1 confers dosage suppression of the synthetic lethality. Interestingly, PRP40 and YNL187w encode proteins with putative leucine-rich nuclear export signal (NES) sequences that fit the consensus sequence recognized by Crm1p. One of Prp40p's two NESs lies within the internal deletion. We demonstrate here that the NES sequences of Prp40p are functional for nuclear export in a leptomycin B-sensitive manner. Furthermore, mutation of these NES sequences confers temperature-sensitive growth and a pre-mRNA splicing defect. Although we do not expect that yeast snRNPs undergo compartmentalized biogenesis like their metazoan counterparts, our results suggest that Prp40p and Ynl187wp contain redundant NESs that aid in an important, Crm1p-mediated nuclear export event.

Active Transport, Cell Nucleus↗

PubMed1999

Isolation and cloning of rat poly(ADP-ribose) glycohydrolase: presence of a potential nuclear export signal conserved in mammalian orthologs.

Poly(ADP-ribose) glycohydrolase (Parg) is the main enzyme of poly(ADP-ribose) degradation. To understand its structure-and-function relationship, we purified Parg from rat testis 9,740-fold using an improved affinity column; the purified product was a 60 kDa protein. Based on the determined sequences of three peptide fragments, degenerated primers were synthesized and a Parg cDNA comprising 3,974 nucleotides, encoding a 109 kDa protein, was isolated. The 60 kDa Parg purified from rat testes corresponded to the C-terminal half of the 109 kDa deduced peptide. When recombinant rat Parg was expressed as a glutathione S-transferase fusion protein in Escherichia coli, Parg activity was observed for the full-length and C-terminal half proteins but not in for the N-terminal half protein. Taken together, these data indicate that the catalytic domain of Parg is located in the C-terminal half. Further, we newly identified the presence of a potential nuclear export signal in the N-terminal half in addition to the previously reported nuclear localization signals in rat and other mammalian Pargs. Northern blot analysis showed the ubiquitous expression of a single 4.0 kb Parg mRNA in various rat tissues. The findings suggest that the 60 kDa Parg is produced by post-transcriptional processing.

Amino Acid Sequence↗

PubMed1997

Anti-idiotype RNA selected with an anti-nuclear export signal antibody is actively transported in oocytes and inhibits Rev- and cap-dependent RNA export.

The anti-idiotype approach is based on the assumption that an antibody specific for a receptor-binding domain of a ligand could be structurally related to the receptor. Therefore, a structural mimic of a receptor-binding domain, selected with an anti-ligand antibody, might be a functional substrate for the receptor. This hypothesis was addressed here by generating antibodies recognizing the Rev-nuclear export signal (NES). A functional NES is required for active export, presumably by interacting directly or indirectly with the nuclear pore complex. Anti-NES antibodies were used to isolate RNA mimics of the NES peptide from combinatorial RNA libraries. The RNA-mimics are exported actively, block Rev-dependent export of a reporter RNA, and inhibit cap-dependent U1 snRNA export in Xenopus oocytes, properties previously reported for NES-peptide conjugates.

Animals↗

PubMed2000

Identification of a nuclear export signal in MKK6, an activator of the carp p38 mitogen-activated protein kinases.

Carp homologues of p38 mitogen-activated protein kinase (MAPK) and its activator MAPK kinase 6 (MAPKK6, referred to as MKK6) were identified. There exist at least two distinct carp p38s, cp38a and cp38b, both of which consist of 361 amino acids. The transcript of c38a was exclusively expressed in the ovary, whereas that of cp38b was ubiquitously expressed. Western blot analysis with anti-(phosphorylated MAPK) Ig specific to the active p38 or JNK has shown that p38 was activated in response to hypertonic stress (1 M sorbitol) in epithelioma papilosum cyprini carp epithelial cells (EPC) and that the activation of p38 proceeded faster to the maximal level than that of JNK. Carp homologue (cMKK6) of p38 activator MKK6 consists of 404 amino acids. It was expressed ubiquitously but was most abundant in the ovary. An in vitro kinase assay demonstrated that cMKK6 is an upstream activator of cp38 and cp38b in carp because it specifically phosphorylated and activated cp38a and cp38b. Interestingly, we found that cMKK6 has a nuclear export signal (NES) sequence in its N-terminal region although upstream activators of stress-activated MAPKs, p38 and JNK, do not in other animals. The NES sequence facilitated nuclear export of cMKK6 and ovalbumin. Leucine residues in the sequence were crucial for the NES activity, as the activity was lost on replacement of the leucines to alanines. The existence of an NES in cMKK6 implies the requisite of strict regulation of the p38 MAPK pathway in carp. The abundance of these components for the stress-activated pathway in the ovary might be related to ectogenetic early development.

Amino Acid Sequence↗

PubMed2005

Expression of fragile X mental retardation-1 gene with nuclear export signal mutation changes the expression profiling of mouse cerebella immortal neuronal cell.

Fragile X syndrome (FXS) is the most frequent cause of inherited mental retardation and is largely caused by a loss of expression of fragile X mental retardation protein (FMRP), encoded by fragile X retardation gene-1 (Fmr1). FMRP is a multifunction protein, with intrinsic RNA-binding properties, which is a component of ribonucleoprotein complex associated with polyribosomes. The properties of FMRP indicate that it might participate in post-transcriptional processes in the regulation of some mRNA species, including localization, stability and translational control. However, the function of FMRP related to the pathologenesis in FXS is largely unknown. Many efforts were undertaken to identify the putative specific RNA targets as well as the FMRP-related proteins and to identify the effect of FMRP absence on the corresponding proteins. Here we present our efforts using proteomics approach to explore the differential expression profiling of mouse cerebella immortal cell, in which we changed the expression of FMRP by expressing Fmr1 gene with nuclear export signal (NES) mutation. This mutation makes FMRP unable to shuttle from nucleus to cytoplasm and leads to nuclear instead of cytoplasmic location as usual, which was hypothesized to affect the pathways of groups of RNAs or proteins related with FMRP. In present study, 56 proteins were found to be differentially expressed in transfected R2 neuronal cells, including 16 decreased expressions and 40 increased expressions. The differentially expressed proteins play roles in diverse physiological processes, such as neuronal plasticity, spermatogenesis and craniofacial and limb development etc. In addition, the expressions of three mRNA identified as FMRP targets in fragile X cell were tested in present model cells. All these results provide new insights to the role of FMRP in the disease.

Active Transport, Cell Nucleus↗

PubMed2000

Identification of nuclear export signal in UL37 protein of herpes simplex virus type 2.

The UL37 gene of herpes simplex virus (HSV) encodes a 120-kDa phosphoprotein associated with the virion. In this study, we have generated a rabbit polyclonal antiserum against HSV-2 UL37 protein, and examined its intracellular localization by immunofluorescence study. In infected cells, specific fluorescence was detectable in the perinuclear region. In transfected cells, UL37 protein was observed mainly in the cytoplasm. Transfection assays of deletion mutants of UL37 protein suggested that the leucine rich region (LRR) containing amino acids 263-273 may be important for cytoplasmic localization. Deletion of the LRR or substitution of the leucine residues resulted in nuclear remaining of UL37 protein. Moreover, the LRR could export green fluorescent protein (GFP) to the cytoplasm as a fusion protein and this export was blocked by leptomycin B treatment, indicating that the LRR acted as a nuclear export signal. These results suggest that UL37 protein fulfills a role as a shuttle between the nucleus and the cytoplasm through the LRR.

Active Transport, Cell Nucleus↗

PubMed2003

Identification of nuclear export signals in antizyme-1.

Antizyme-1 (AZ1) is a protein that negatively regulates polyamine synthesis by inhibiting the key synthetic enzyme ornithine decarboxylase and targeting it for degradation by the 26 S proteasome. Recent work shows that antizyme protein translocates to the nucleus during mouse development (Gritli-Linde, A., Nilssom, J., Bohlooly, Y. M., Heby, O., and Linde, A. (2001) Dev. Dyn. 220, 259-275). However, the significance and mechanism of this phenomenon remain unclear. In this study, we expressed AZ1 fused with enhanced green fluorescent protein (EGFP) to study its localization in a living cell. We found that EGFP-AZ1 was predominantly localized in the cytoplasm and that treatment with leptomycin B, a specific inhibitor of chromosomal region maintenance 1 (CRM1) induced nuclear accumulation of EGFP-AZ1 in Chinese hamster ovary and NIH3T3 cells. Two independent nuclear export signal (NES) sequences, each containing essential hydrophobic residues, were identified in the 50 N-terminal amino acid residues and in the central part of AZ1. The activity of the second NES was inhibited by an N-terminal adjacent region and was only revealed in N-terminal truncated constructs. Both NESs were active when fused to an artificial nuclear protein SV40-NLS-EGFP-EGFP. The ability of AZ1 to shuttle between the nucleus and the cytoplasm suggests that it has a novel function in the nucleus.

Active Transport, Cell Nucleus↗