scholarly journals 14-3-3 transits to the nucleus and participates in dynamic nucleocytoplasmic transport

2002 ◽  
Vol 156 (5) ◽  
pp. 817-828 ◽  
Author(s):  
Anne Brunet ◽  
Fumihiko Kanai ◽  
Justine Stehn ◽  
Jian Xu ◽  
Dilara Sarbassova ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Ligand Binding ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Rich Region ◽  
Present Evidence ◽  
Α Helix ◽  
Export Signal

14-3-3 proteins regulate the cell cycle and prevent apoptosis by controlling the nuclear and cytoplasmic distribution of signaling molecules with which they interact. Although the majority of 14-3-3 molecules are present in the cytoplasm, we show here that in the absence of bound ligands 14-3-3 homes to the nucleus. We demonstrate that phosphorylation of one important 14-3-3 binding molecule, the transcription factor FKHRL1, at the 14-3-3 binding site occurs within the nucleus immediately before FKHRL1 relocalization to the cytoplasm. We show that the leucine-rich region within the COOH-terminal α-helix of 14-3-3, which had been proposed to function as a nuclear export signal (NES), instead functions globally in ligand binding and does not directly mediate nuclear transport. Efficient nuclear export of FKHRL1 requires both intrinsic NES sequences within FKHRL1 and phosphorylation/14-3-3 binding. Finally, we present evidence that phosphorylation/14-3-3 binding may also prevent FKHRL1 nuclear reimport. These results indicate that 14-3-3 can mediate the relocalization of nuclear ligands by several mechanisms that ensure complete sequestration of the bound 14-3-3 complex in the cytoplasm.

scholarly journals Exportin Crm1 is important for Swi6 nucleocytoplasmic shuttling and MBF transcription activation in Saccharomyces cerevisiae

2020 ◽  
Author(s):  
Kenneth D. Belanger ◽  
William T. Yewdell ◽  
Matthew F. Barber ◽  
Amy N. Russo ◽  
Mark A. Pettit ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Transcription Factor ◽  
Nuclear Export ◽  
Intracellular Localization ◽  
Nuclear Export Signal ◽  
Transcription Activation ◽  
Cell Phenotype ◽  
Export Signal

AbstractThe Swi6 protein acts as a transcription factor in budding yeast, functioning in two different heterodimeric complexes, SBF and MBF, that activate the expression of distinct but overlapping sets of genes. Swi6 undergoes regulated changes in nucleocytoplasmic localization throughout the cell cycle that correlate with changes in gene expression. While the process of Swi6 nuclear import is well understood, mechanisms underlying its nuclear export remain unclear. Here we investigate Swi6 nuclear export and its impact on Swi6 function. We show that the exportin Crm1, in addition to three other karyopherins previously shown to affect Swi6 localization, is important for Swi6 nuclear export and activity. A truncation of Swi6 that removes a putative Crm1 nuclear export signal results in the loss of changes in nucleocytoplasmic Swi6 localization that normally occur during progression through the cell cycle. Mutagenesis of the NES-like sequence or removal of Crm1 activity using leptomycin B results in a similar decrease in nuclear export as cells enter S-phase. Using two-hybrid analysis, we also show that Swi6 associates with Crm1 in vivo. Alteration of the Crm1 NES in Swi6 results in a decrease in MBF-mediated gene expression, but does not affect expression of an SBF reporter, suggesting that export of Swi6 by Crm1 regulates a subset of Swi6 transcription activation activity. Finally, alteration of the Crm1 NES in Swi6 results in cells that are larger than wild type, but not to the extent of those with a complete Swi6 deletion. Expressing a Swi6 NES mutant in combination with a deletion of Msn5, an exportin involved in Swi6 nuclear export and specifically affecting SBF activation, further increases the large cell phenotype, but still not to the extent observed in a Swi6 deletion mutant. These data suggest that Swi6 has at least two different exportins, Crm1 and Msn5, each of which interacts with a distinct nuclear export signal and influences expression of a different subset of Swi6-controlled genes.Summary StatementPrecise intracellular localization is important for the proper activity of proteins. Here we provide evidence that the Swi6 transcription factor important for cell cycle progression shuttles between the cell nucleus and cytoplasm, its nuclear export is important for its activity, and that it contains a nuclear export signal (NES) recognized by the Crm1 nuclear transport factor.

A Cytoplasmic FANCA-FANCC Complex Interacts and Stabilizes the Leukemic NPMc Protein.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3098-3098
Author(s):  
Wei Du ◽  
Suzette Maynard ◽  
Jared Sipple ◽  
Jie Li ◽  
Qishen Pang
Keyword(s):  
Bone Marrow ◽  
Nuclear Export ◽  
Biological Function ◽  
Bone Marrow Cells ◽  
Conflicts Of Interest ◽  
Nuclear Export Signal ◽  
Normal Karyotype ◽  
Present Evidence ◽  
Normal Human ◽  
Export Signal

Abstract Abstract 3098 Poster Board III-35 In a subset of AML with a normal karyotype, a frame-shift mutation in the extreme C-terminal of the nucleophosmin (NPM) gene results in the creation of a nuclear export signal, generating a mutant NPM protein (NPMc) that is permanently dislocated in the cytoplasm. In the present study, we have analyzed the interaction between NPMc and a cytoplasmic subcomplex of Fanconi anemia (FA) proteins. Sequence analysis of bone marrow samples from 46 FA patients shows that NPMc mutations were excluded from FA genome. NPMc was degraded more rapidly in AML bone marrow cells from FA patients (t1/2 < 30 min) than in AML cell line HL60 (t1/2 > 90 min). Further analysis revealed that inducible knockdown of FANCA or FANCC in leukemic OCI/AML3 cells carrying the NPMc mutation induced degradation of the cytoplasmic NPMc protein. Forced localization of FANCC to the nucleus also caused rapid NPMc degradation. We also show that NPMc degradation was mediated by the proteasome and that correction of mutant lymphoblasts from FA-A or FA-C patients with a functional FANCA or FANCC protein prevented NPMc ubiquitination and consequently degradation. Moreover, we demonstrate that the cytoplasmic FANCA and FANCC interacted with NPMc in the cytosolic fractions of normal human lymphoblaststic cells and that the acidic domains of NPM were required for the cytoplasmic FA-NPMc complex formation. Finally, using patient-derived FANCC mutant, a nuclearized FANCC and a NOG/SGM3 xenotansplant model, we present evidence that the cytoplasmic FANCA-FANCC complex was essential for NPMc stability and biological function. Thus, these findings reveal the potential molecular mechanism involved in the cytoplasmic retention of the leukemic NPMc. Disclosures No relevant conflicts of interest to declare.

scholarly journals Targeting levels or oligomerization of nucleophosmin 1 induces differentiation and loss of survival of human AML cells with mutant NPM1

Blood ◽  
2011 ◽  
Vol 118 (11) ◽  
pp. 3096-3106 ◽  
Author(s):  
Ramesh Balusu ◽  
Warren Fiskus ◽  
Rekha Rao ◽  
Daniel G. Chong ◽  
Srilatha Nalluri ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Molecular Chaperone ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
S Phase ◽  
Wild Type ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Induced Apoptosis ◽  
Export Signal

Abstract Nucleophosmin 1 (NPM1) is an oligomeric, nucleolar phosphoprotein that functions as a molecular chaperone for both proteins and nucleic acids. NPM1 is mutated in approximately one-third of patients with AML. The mutant NPM1c+ contains a 4-base insert that results in extra C-terminal residues encoding a nuclear export signal, which causes NPM1c+ to be localized in the cytoplasm. Here, we determined the effects of targeting NPM1 in cultured and primary AML cells. Treatment with siRNA to NPM1 induced p53 and p21, decreased the percentage of cells in S-phase of the cell cycle, as well as induced differentiation of the AML OCI-AML3 cells that express both NPMc+ and unmutated NPM1. Notably, knockdown of NPM1 by shRNA abolished lethal AML phenotype induced by OCI-AML3 cells in NOD/SCID mice. Knockdown of NPM1 also sensitized OCI-AML3 to all-trans retinoic acid (ATRA) and cytarabine. Inhibition of NPM1 oligomerization by NSC348884 induced apoptosis and sensitized OCI-AML3 and primary AML cells expressing NPM1c+ to ATRA. This effect was significantly less in AML cells coexpressing FLT3-ITD, or in AML or normal CD34+ progenitor cells expressing wild-type NPM1. Thus, attenuating levels or oligomerization of NPM1 selectively induces apoptosis and sensitizes NPM1c+ expressing AML cells to treatment with ATRA and cytarabine.

scholarly journals The microtubule plus-end tracking protein Bik1 is required for chromosome congression

2021 ◽  
Author(s):  
Alexander Julner ◽  
Marjan Abbasi ◽  
Victoria Menendez Benito
Keyword(s):  
Cell Cycle ◽  
Nuclear Export ◽  
Cell Cycle Progression ◽  
Nuclear Export Signal ◽  
Microtubule Dynamics ◽  
Dependent Manner ◽  
Chromosome Congression ◽  
A Cell ◽  
Cycle Dependent ◽  
Export Signal

During mitosis, sister chromatids congress on either side of the spindle equator to facilitate the correct partitioning of the genomic material. Chromosome congression requires a finely tuned control of microtubule dynamics by the kinesin motor proteins. In Saccharomyces cerevisiae, the kinesin proteins Cin8, Kip1, and Kip3 have pivotal roles in chromosome congression. It has been hypothesized that additional proteins that modulate microtubule dynamics are also involved. Here, we show that the microtubule plus-end tracking protein Bik1 (the budding yeast ortholog of CLIP-170) is essential for chromosome congression. We find that nuclear Bik1 localizes to the kinetochores in a cell-cycle-dependent manner. Disrupting the nuclear pool of Bik1 with a nuclear export signal (Bik1-NES) leads to a slower cell cycle progression characterized by a delayed metaphase-anaphase transition. Bik1-NES cells have mispositioned kinetochores along the spindle in metaphase. Furthermore, using proximity-dependent methods, we identify Cin8 as an interaction partner of Bik1. Deleting CIN8 reduces the amount of Bik1 at the spindle. In contrast, Cin8 retains its typical bilobed distribution in Bik1-NES and does not localize to the unclustered kinetochores characteristic of Bik1-NES cells. Thus, we propose that Bik1 functions together with Cin8 to regulate kinetochore-microtubule dynamics for correct kinetochore positioning and chromosome congression.

scholarly journals Nuclear Export Is Evolutionarily Conserved in CVC Paired-Like Homeobox Proteins and Influences Protein Stability, Transcriptional Activation, and Extracellular Secretion

2005 ◽  
Vol 25 (7) ◽  
pp. 2573-2582 ◽  
Author(s):  
Shirley K. Knauer ◽  
Gert Carra ◽  
Roland H. Stauber
Keyword(s):  
Transcription Factors ◽  
Protein Stability ◽  
Cell Fate ◽  
Transcriptional Activation ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Cell Fate Decisions ◽  
Extracellular Secretion ◽  
Export Signal

ABSTRACT Homeodomain transcription factors control a variety of essential cell fate decisions during development. To understand the developmental regulation by these transcription factors, we describe here the molecular analysis of paired-like CVC homeodomain protein (PLC-HDP) trafficking. Complementary experimental approaches demonstrated that PLC-HDP family members are exported by the Crm1 pathway and contain an evolutionary conserved leucine-rich nuclear export signal. Importantly, inactivation of the nuclear export signal enhanced protein stability, resulting in increased transactivation of transfected reporters and decreased extracellular secretion. In addition, PLC-HDPs harbor a conserved active nuclear import signal that could also function as a protein transduction domain. In our study, we characterized PLC-HDPs as mobile nucleocytoplasmic shuttle proteins with the potential for unconventional secretion and intercellular transfer. Nucleocytoplasmic transport may thus represent a conserved control mechanism to fine-tune the transcriptional activity of PLC-HDPs prerequisite for regulating and maintaining the complex expression pattern during development.

A functional study of nucleocytoplasmic transport signals of the EhNCABP166 protein from Entamoeba histolytica

Parasitology ◽  
2012 ◽  
Vol 139 (13) ◽  
pp. 1697-1710 ◽  
Author(s):  
R. URIBE ◽  
J. ALMARAZ BARRERA MA DE ◽  
M. ROBLES-FLORES ◽  
G. MENDOZA HERNÁNDEZ ◽  
A. GONZÁLEZ-ROBLES ◽  
...  
Keyword(s):  
Entamoeba Histolytica ◽  
Nuclear Localization ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Cytoplasmic Domain ◽  
Nucleocytoplasmic Transport ◽  
Actin Binding ◽  
Functional Study ◽  
Nuclear Localization Signals ◽  
Export Signal

SUMMARYEhNCABP166 is an Entamoeba histolytica actin-binding protein that localizes to the nucleus and cytoplasm. Bioinformatic analysis of the EhNCABP166 amino acid sequence shows the presence of 3 bipartite nuclear localization signals (NLS) and a nuclear export signal (NES). The present study aimed to investigate the functionality of these signals in 3 ways. First, we fused each potential NLS to a cytoplasmic domain of ehFLN to determine whether the localization of this domain could be altered by the presence of the NLSs. Furthermore, the localization of each domain of EhNCABP166 was determined. Similarly, we generated mutations in the first block of bipartite signals from the domains that contained these signals. Additionally, we added an NES to 2 constructs that were then evaluated. We confirmed the intranuclear localization of EhNCABP166 using transmission electron microscopy. Fusion of each NLS resulted in shuttling of the cytoplasmic domain to the nucleus. With the exception of 2 domains, all of the evaluated domains localized within the nucleus. A mutation in the first block of bipartite signals affected the localization of the domains containing an NLS. The addition of an NES shifted the localization of these domains to the cytoplasm. The results presented here establish EhNCABP166 as a protein containing functional nuclear localization signals and a nuclear export signal.

Regulated nuclear export of the homeodomain transcription factor Prospero

Development ◽  
2001 ◽  
Vol 128 (8) ◽  
pp. 1359-1367
Author(s):  
Z. Demidenko ◽  
P. Badenhorst ◽  
T. Jones ◽  
X. Bi ◽  
M.A. Mortin
Keyword(s):  
Transcription Factor ◽  
Subcellular Distribution ◽  
Nuclear Export ◽  
Cultured Cells ◽  
System Development ◽  
Nuclear Export Signal ◽  
Nervous System Development ◽  
Embryonic Nervous System ◽  
Mother Cells ◽  
Export Signal

Subcellular distribution of the Prospero protein is dynamically regulated during Drosophila embryonic nervous system development. Prospero is first detected in neuroblasts where it becomes cortically localized and tethered by the adapter protein, Miranda. After division, Prospero enters the nucleus of daughter ganglion mother cells where it functions as a transcription factor. We have isolated a mutation that removes the C-terminal 30 amino acids from the highly conserved 100 amino acid Prospero domain. Molecular dissection of the homeo- and Prospero domains, and expression of chimeric Prospero proteins in mammalian and insect cultured cells indicates that Prospero contains a nuclear export signal that is masked by the Prospero domain. Nuclear export of Prospero, which is sensitive to the drug leptomycin B, is mediated by Exportin. Mutation of the nuclear export signal-mask in Drosophila embryos prevents Prospero nuclear localization in ganglion mother cells. We propose that a combination of cortical tethering and regulated nuclear export controls Prospero subcellular distribution and function in all higher eukaryotes.

Sign in / Sign up

Export Citation Format

Share Document