scholarly journals PDZ domain from Dishevelled -- a specificity study.

2011 ◽  
Vol 58 (2) ◽  
Author(s):  
Katarzyna Śmietana ◽  
Agnieszka Mateja ◽  
Artur Krężel ◽  
Jacek Otlewski
Keyword(s):  
Wnt Signaling ◽  
Nuclear Export ◽  
Signal Sequence ◽  
Consensus Sequence ◽  
Pdz Domain ◽  
Nuclear Export Signal ◽  
Signaling Cascades ◽  
Binding Motif ◽  
Wnt Proteins ◽  
Export Signal

Intracellular signaling cascades induced by Wnt proteins play a key role in developmental processes and are implicated in cancerogenesis. It is still unclear how the cell determines which of the three possible Wnt response mechanisms should be activated, but the decision process is most likely dependent on Dishevelled proteins. Dishevelled family members interact with many diverse targets, however, molecular mechanisms underlying these binding events have not been comprehensively described so far. Here, we investigated the specificity of the PDZ domain from human Dishevelled-2 using C-terminal phage display, which led us to identification of a leucine-rich binding motif strongly resembling the consensus sequence of a nuclear export signal. PDZ interactions with several peptide and protein motifs (including the nuclear export signal sequence from Dishevelled-2 protein) were investigated in detail using fluorescence spectroscopy, mutational analysis and immunoenzymatic assays. The experiments showed that the PDZ domain can bind the nuclear export signal sequence of the Dishevelled-2 protein. Since the intracellular localization of Dishevelled is governed by nuclear localization and nuclear export signal sequences, it is possible that the intramolecular interaction between PDZ domain and the export signal could modulate the balance between nuclear and cytoplasmic pool of the Dishevelled protein. Such a regulatory mechanism would be of utmost importance for the differential activation of Wnt signaling cascades, leading to selective promotion of the nucleus-dependent Wnt β-catenin pathway at the expense of non-canonical Wnt signaling.

scholarly journals Nuclear Export of L-Periaxin, Mediated by Its Nuclear Export Signal in the PDZ Domain

PLoS ONE ◽  
2014 ◽  
Vol 9 (3) ◽  
pp. e91953 ◽  
Author(s):  
Yawei Shi ◽  
Lei Zhang ◽  
Ting Yang
Keyword(s):  
Nuclear Export ◽  
Pdz Domain ◽  
Nuclear Export Signal ◽  
Export Signal

scholarly journals Nuclear Import of Hepatic Glucokinase Depends upon Glucokinase Regulatory Protein, whereas Export Is Due to a Nuclear Export Signal Sequence in Glucokinase

1999 ◽  
Vol 274 (52) ◽  
pp. 37125-37130 ◽  
Author(s):  
Chiyo Shiota ◽  
Jack Coffey ◽  
Joseph Grimsby ◽  
Joseph F. Grippo ◽  
Mark A. Magnuson
Keyword(s):  
Nuclear Import ◽  
Nuclear Export ◽  
Signal Sequence ◽  
Regulatory Protein ◽  
Nuclear Export Signal ◽  
Glucokinase Regulatory Protein ◽  
Export Signal

scholarly journals Phosphorylation in the accessory domain of yeast histone chaperone protein 1 exposes the nuclear export signal sequence

2021 ◽  
Author(s):  
Sho Ashida ◽  
Rikuri Morita ◽  
Yasuteru Shigeta ◽  
Ryuhei Harada
Keyword(s):  
Cell Nucleus ◽  
Nuclear Export ◽  
Signal Sequence ◽  
Nuclear Export Signal ◽  
Md Simulations ◽  
Histone Chaperone ◽  
Chaperone Protein ◽  
Accessory Domain ◽  
Export Signal

Histone is a scaffold protein that constitutes nucleosomes with DNA in the cell nucleus. When forming histone, hetero octamer is assisted by histone chaperone proteins. As a histone chaperone protein, the crystal structure of yeast nucleosome assembly protein (yNap1) has been determined. For yNap1, a nuclear export signal/sequence (NES) has been identified as a part of the long -helix. Experimental evidence via mutagenesis on budding yeast suggests the NES is necessary for transport out from the cell nucleus. However, the NES is masked by a region defined as an accessory domain (AD). In addition, the role of the AD in nuclear transport has not been elucidated yet. To address the role of the AD, we focused on phosphorylation in the AD because proteome experiments have identified multiple phosphorylation sites of yNap1. To computationally treat phosphorylation, we performed all-atom molecular dynamics (MD) simulations for a set of non-phosphorylated and phosphorylated yNap1 (Nap1-nonP and Nap1-P). As an analysis, we addressed how the NES is exposed to the protein surface by measuring its solvent-access surface area (SASA). As a result, there was a difference in the SASA distributions between both systems. Quantitatively, the median of the SASA distribution of Nap1-P was greater than that of Nap1-nonP, meaning that phosphorylation in the AD exposed to the NES, resulting in increasing its accessibility. In conclusion, yNap1 might modulate the accessibility of the NES by dislocating the AD through phosphorylation.

scholarly journals Identification of a nuclear export signal sequence for bovine papillomavirus E1 protein

Virology ◽  
2008 ◽  
Vol 373 (1) ◽  
pp. 149-162 ◽  
Author(s):  
Germán Rosas-Acosta ◽  
Van G. Wilson
Keyword(s):  
Nuclear Export ◽  
Signal Sequence ◽  
Nuclear Export Signal ◽  
Bovine Papillomavirus ◽  
E1 Protein ◽  
Export Signal

scholarly journals Epstein-Barr Virus EB2 Protein Exports Unspliced RNA via a Crm-1-Independent Pathway

2000 ◽  
Vol 74 (13) ◽  
pp. 6068-6076 ◽  
Author(s):  
Géraldine Farjot ◽  
Monique Buisson ◽  
Madeleine Duc Dodon ◽  
Louis Gazzolo ◽  
Alain Sergeant ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Epstein Barr Virus ◽  
Consensus Sequence ◽  
Nuclear Export Signal ◽  
Site Directed Mutagenesis ◽  
Virus Protein ◽  
Nuclear Pore ◽  
Barr Virus ◽  
Epstein Barr ◽  
Export Signal

ABSTRACT Human herpesviruses encode posttranscriptional activators that are believed to up-regulate viral replication by facilitating early and late gene expression. We have reported previously that the Epstein-Barr virus protein EB2 (also called M or SM) promotes nuclear export of RNAs that are poor substrates for spliceosome assembly, an effect that closely resembles the human immunodeficiency virus type 1 Rev-dependent nuclear export of unspliced viral RNA. Here we present experimental data showing that EB2 efficiently promotes the nuclear export of unspliced RNA expressed from a Rev reporter construct. Site-directed mutagenesis as well as domain swapping experiments indicate that a leucine-rich region found in the EB2 protein, which matches the consensus sequence for the leucine-rich nuclear export signal, is not a nuclear export signal per se. Accordingly, leptomycin B (LMB), a specific Crm-1 inhibitor, impairs Rev- but not EB2-dependent nuclear export of unspliced RNA. Moreover, EB2 nucleocytoplasmic shuttling visualized by a heterokaryon assay is, unlike Rev shuttling, not affected by LMB. We also show that overexpression of an N-terminal deletion mutant of Nup214/can, a major nucleoporin of the nuclear pore complex involved in several aspects of nuclear transport, blocks both Rev- and EB2-dependent nuclear export of RNA. These results strongly suggest that EB2 nuclear export of unspliced RNA is mediated by a Crm-1-independent pathway.

scholarly journals Keap1 Regulates the Oxidation-Sensitive Shuttling of Nrf2 into and out of the Nucleus via a Crm1-Dependent Nuclear Export Mechanism

2005 ◽  
Vol 25 (11) ◽  
pp. 4501-4513 ◽  
Author(s):  
Michaella Velichkova ◽  
Tama Hasson
Keyword(s):  
Nuclear Export ◽  
Consensus Sequence ◽  
Nuclear Export Signal ◽  
Antioxidant Response ◽  
Negative Regulator ◽  
Nuclear Accumulation ◽  
Leptomycin B ◽  
Hydrophobic Amino Acids ◽  
Export Signal ◽  
Full Activation

ABSTRACT Keap1 is a negative regulator of Nrf2, a transcription factor essential for antioxidant response element (ARE)-mediated gene expression. We find that Keap1 sequesters Nrf2 in the cytoplasm, not by docking it to the actin cytoskeleton but instead through an active Crm1/exportin-dependent nuclear export mechanism. Deletion and mutagenesis studies identified a nuclear export signal (NES) in the intervening region of Keap1 comprised of hydrophobic leucine and isoleucine residues in agreement with a traditional NES consensus sequence. Mutation of the hydrophobic amino acids resulted in nuclear accumulation of both Keap1 and Nrf2, as did treatment with the drug leptomycin B, which inactivates Crm1/exportin. ARE genes were partially activated under these conditions, suggesting that additional oxidation-sensitive elements are required for full activation of the antioxidant response. Based on these data, we propose a new model for regulation of Nrf2 by Keap1. Under normal conditions, Keap1 and Nrf2 are complexed in the cytoplasm where they are targeted for degradation. Oxidative stress inactivates Keap1's NES, allowing entry of both Keap1 and Nrf2 into the nucleus and transcriptional transactivation of ARE genes.

scholarly journals Generation of Influenza A Virus NS2 (NEP) Mutants with an Altered Nuclear Export Signal Sequence

2004 ◽  
Vol 78 (18) ◽  
pp. 10149-10155 ◽  
Author(s):  
Kiyoko Iwatsuki-Horimoto ◽  
Taisuke Horimoto ◽  
Yutaka Fujii ◽  
Yoshihiro Kawaoka
Keyword(s):  
Influenza A Virus ◽  
Nuclear Export ◽  
Influenza A ◽  
Signal Sequence ◽  
Nuclear Export Signal ◽  
Wild Type Virus ◽  
Wild Type ◽  
Amino Terminal ◽  
Ribonucleoprotein Complexes ◽  
Export Signal

ABSTRACT The NS2 (NEP) protein of influenza A virus contains a highly conserved nuclear export signal (NES) motif in its amino-terminal region (12ILMRMSKMQL21, A/WSN/33), which is thought to be required for nuclear export of viral ribonucleoprotein complexes (vRNPs) mediated by a cellular export factor, CRM1. However, simultaneous replacement of three hydrophobic residues in the NES with alanine does not affect NS2 (NEP) binding to CRM1, although the virus with these mutations is not viable. To determine the extent of sequence conservation required by the NS2 (NEP) NES for its export function during viral replication, we randomly introduced mutations by degenerative mutagenesis into the region of NS cDNA encoding the NS2 (NEP) NES and then attempted to generate mutant viruses containing these alterations by reverse genetics. Sequence analysis of the recovered viruses showed that although some of the mutants possessed amino acids other than those conserved in the NES, hydrophobicity within this motif was maintained. Nuclear export of vRNPs representing all of the mutant viruses was completely inhibited in the presence of a CRM1 inhibitor, leptomycin B, as was the transport of wild-type virus, indicating that the CRM1-mediated pathway is responsible for the nuclear export of both wild-type and mutant vRNPs. The vRNPs of some of the mutant viruses were exported in a delayed manner, resulting in limited viral growth in cell culture and in mice. These results suggest that the NES motif may be an attractive target for the introduction of attenuating mutations in the production of live vaccine viruses.

A factor required for nonsense-mediated mRNA decay in yeast is exported from the nucleus to the cytoplasm by a nuclear export signal sequence

1998 ◽  
Vol 111 (21) ◽  
pp. 3129-3143 ◽  
Author(s):  
R.L. Shirley ◽  
M.J. Lelivelt ◽  
L.R. Schenkman ◽  
J.N. Dahlseid ◽  
M.R. Culbertson
Keyword(s):  
Protein Function ◽  
Nuclear Export ◽  
Mrna Decay ◽  
Signal Sequence ◽  
Nuclear Export Signal ◽  
Reporter Protein ◽  
Nuclear Localization Signals ◽  
Sequence Elements ◽  
Nonsense Mediated Mrna Decay ◽  
Export Signal

In Saccharomyces cerevisiae, Upf3p is required for nonsense-mediated mRNA decay (NMD). Although localized primarily in the cytoplasm, Upf3p contains three sequence elements that resemble nuclear localization signals (NLSs) and two sequence elements that resemble nuclear export signals (NESs). We found that a cytoplasmic reporter protein localized to the nucleus when fused to any one of the three NLS-like sequences of Upf3p. A nuclear reporter protein localized to the cytoplasm when fused to one of the NES-like sequences (NES-A). We present evidence that NES-A functions to signal the export of Upf3p from the nucleus. Combined alanine substitutions in the NES-A element caused a re-distribution of Upf3p to a subnuclear location identified as the nucleolus and conferred an Nmd- phenotype. Single mutations in NES-A failed to affect the distribution of Upf3p and were Nmd+. When an NES element from HIV-1 Rev was inserted near the C terminus of a mutant Upf3p containing multiple mutations in NES-A, the cytoplasmic distribution typical of wild-type Upf3p was restored but the cells remained phenotypically Nmd-. These results suggest that NES-A is a functional nuclear export signal. Combined mutations in NES-A may cause multiple defects in protein function leading to an Nmd- phenotype even when export is restored.

Control of the nuclear-cytoplasmic partitioning of annexin II by a nuclear export signal and by p11 binding

2001 ◽  
Vol 114 (17) ◽  
pp. 3155-3166 ◽  
Author(s):  
David A. Eberhard ◽  
Larry R. Karns ◽  
Scott R. VandenBerg ◽  
Carl E. Creutz
Keyword(s):  
Nuclear Export ◽  
Consensus Sequence ◽  
Nuclear Export Signal ◽  
Nuclear Accumulation ◽  
Annexin Ii ◽  
Nuclear Entry ◽  
Nuclear Exclusion ◽  
Endogenous Proteins ◽  
Necessary And Sufficient ◽  
Export Signal

This study investigated mechanisms controlling the nuclear-cytoplasmic partitioning of annexin II (AnxII). AnxII and its ligand, p11, were localized by immunofluorescence to the cytoplasmic compartment of U1242MG cells, with minimal AnxII or p11 detected within nuclei. Similarly, GFP-AnxII and GFP-p11 chimeras localized to the endogenous proteins. Likewise, GFP-AnxII(1-22) was excluded from nuclei, whereas GFP-AnxII(23-338) and GFP alone were distributed throughout the cells. Immunoprecipitation and biochemical studies showed that GFP-AnxII did not form heteromeric complexes with endogenous p11 and AnxII. Thus, the AnxII N-tail is necessary and sufficient to cause nuclear exclusion of the GFP fusion protein but this does not involve p11 binding. A nuclear export signal consensus sequence was found in the AnxII 3-12 region. The consensus mutant GFP-AnxII(L10A/L12A) confirmed that these residues are necessary for nuclear exclusion. The nuclear exclusion of GFP-AnxII(1-22) was temperature-dependent and reversible, and the nuclear export inhibitor leptomycin B (LmB) caused GFP-AnxII or overexpressed AnxII monomer to accumulate in nuclei. Therefore, AnxII monomer can enter the nucleus and is actively exported. However, LmB had little effect on the localization of AnxII/p11 complex in U1242MG cells, indicating that the complex is sequestered in the cytoplasm. By contrast, LmB treatment of v-src-transformed fibroblasts caused endogenous AnxII to accumulate in nuclei. The LmB-induced nuclear accumulation of AnxII was accelerated by pervanadate and inhibited by genistein, suggesting that phosphorylation promotes nuclear entry of AnxII. Thus, nuclear exclusion of AnxII results from nuclear export of the monomer and sequestration of AnxII/p11 complex, and may be modulated by phosphorylation.

Sign in / Sign up

Export Citation Format

Share Document