5-Flurouracil disrupts nuclear export and nuclear pore permeability in a calcium dependent manner

APOPTOSIS ◽  
2016 ◽  
Vol 22 (3) ◽  
pp. 393-405 ◽  
Author(s):  
Kelly J. Higby ◽  
Melissa M. Bischak ◽  
Christina A. Campbell ◽  
Rebecca G. Anderson ◽  
Sarah A. Broskin ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Pore ◽  
Dependent Manner ◽  
Calcium Dependent ◽  
Pore Permeability

scholarly journals Steady-State Nuclear Localization of Exportin-t Involves RanGTP Binding and Two Distinct Nuclear Pore Complex Interaction Domains

2002 ◽  
Vol 22 (16) ◽  
pp. 5708-5720 ◽  
Author(s):  
Scott Kuersten ◽  
Gert-Jan Arts ◽  
Tobias C. Walther ◽  
Ludwig Englmeier ◽  
Iain W. Mattaj
Keyword(s):  
Steady State ◽  
Nuclear Export ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Dependent Manner ◽  
C Terminus ◽  
Transport Cycle ◽  
Interaction Domains ◽  
Pore Complexes

ABSTRACT Vertebrate tRNA export receptor exportin-t (Xpo-t) binds to RanGTP and mature tRNAs cooperatively to form a nuclear export complex. Xpo-t shuttles bidirectionally through nuclear pore complexes (NPCs) but is mainly nuclear at steady state. The steady-state distribution of Xpo-t is shown to depend on its interaction with RanGTP. Two distinct Xpo-t NPC interaction domains that bind differentially to peripherally localized nucleoporins in vitro are identified. The N terminus binds to both Nup153 and RanBP2/Nup358 in a RanGTP-dependent manner, while the C terminus binds to CAN/Nup214 independently of Ran. We propose that these interactions increase the concentration of tRNA export complexes and of empty Xpo-t in the vicinity of NPCs and thus increase the efficiency of the Xpo-t transport cycle.

scholarly journals The hsp90-FKBP52 Complex Links the Mineralocorticoid Receptor to Motor Proteins and Persists Bound to the Receptor in Early Nuclear Events

2009 ◽  
Vol 30 (5) ◽  
pp. 1285-1298 ◽  
Author(s):  
Mario D. Galigniana ◽  
Alejandra G. Erlejman ◽  
Martín Monte ◽  
Celso Gomez-Sanchez ◽  
Graciela Piwien-Pilipuk
Keyword(s):  
Nuclear Export ◽  
Mineralocorticoid Receptor ◽  
Soluble Fraction ◽  
Molecular System ◽  
Nuclear Pore ◽  
Dependent Manner ◽  
Cytoplasmic Localization ◽  
Nucleocytoplasmic Trafficking ◽  
Cytoplasmic Transport ◽  
Nuclear Events

ABSTRACT In this study, we demonstrate that the subcellular localization of the mineralocorticoid receptor (MR) is regulated by tetratricopeptide domain (TPR) proteins. The high-molecular-weight immunophilin (IMM) FKBP52 links the MR-hsp90 complex to dynein/dynactin motors favoring the cytoplasmic transport of MR to the nucleus. Replacement of this hsp90-binding IMM by FKBP51 or the TPR peptide favored the cytoplasmic localization of MR. The complete movement machinery, including dynein and tubulin, could be recovered from paclitaxel/GTP-stabilized cytosol and was fully reassembled on stripped MR immune pellets. The whole MR-hsp90-based heterocomplex was transiently recovered in the soluble fraction of the nucleus after 10 min of incubation with aldosterone. Moreover, cross-linked MR-hsp90 heterocomplexes accumulated in the nucleus in a hormone-dependent manner, demonstrating that the heterocomplex can pass undissociated through the nuclear pore. On the other hand, a peptide that comprises the DNA-binding domain of MR impaired the nuclear export of MR, suggesting the involvement of this domain in the process. This study represents the first report describing the entire molecular system that commands MR nucleocytoplasmic trafficking and proposes that the MR-hsp90-TPR protein heterocomplex is dissociated in the nucleus rather than in the cytoplasm.

Nuclear export of signal recognition particle RNA is a facilitated process that involves the Alu sequence domain

1994 ◽  
Vol 107 (4) ◽  
pp. 903-912 ◽  
Author(s):  
X.P. He ◽  
N. Bataille ◽  
H.M. Fried
Keyword(s):  
Nuclear Export ◽  
Signal Recognition Particle ◽  
Nuclear Pore ◽  
Dependent Manner ◽  
Signal Recognition ◽  
Cytoplasmic Rna ◽  
Alu Sequence ◽  
Alu Element ◽  
Sequence Domain ◽  
Srp Rna

The signal recognition particle is a cytoplasmic RNA-protein complex that mediates translocation of secretory polypeptides into the endoplasmic reticulum. We have used a Xenopus oocyte microinjection assay to determine how signal recognition particle (SRP) RNA is exported from the nucleus. Following nuclear injection, SRP RNA accumulated in the cytoplasm while cytoplasmically injected SRP RNA did not enter the nucleus. Cytoplasmic accumulation of SRP RNA was an apparently facilitated process dependent on limiting trans-acting factors, since nuclear export exhibited saturation kinetics and was completely blocked either at low temperature or by wheat germ agglutinin, a known inhibitor of nuclear pore-mediated transport. At least one target for trans-acting factors that promote nuclear export of SRP RNA appears to be the Alu element of the molecule, since a transcript consisting of only the Alu sequence was exported from the nucleus in a temperature-dependent manner and the Alu transcript competed in the nucleus for transport with intact SRP RNA. Although the identities of trans-acting factors responsible for SRP RNA transport are at present unknown, we suggest that proteins contained within the cytoplasmic form of SRP are candidates. Consistent with this idea were the effects of a mutation in SRP RNA that prevented binding of two known SRP proteins to the Alu sequence.

Preclinical evaluation of selective inhibitors of nuclear export (SINE) in basal-like breast cancer (BLBC).

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. 1055-1055 ◽  
Author(s):  
Dilara McCauley ◽  
Yosef Landesman ◽  
William Senapedis ◽  
Trinayan Kashyap ◽  
Jean-Richard Saint-Martin ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Tumor Growth ◽  
Cell Lines ◽  
Nuclear Export ◽  
The Cancer Genome Atlas ◽  
Nuclear Accumulation ◽  
Nuclear Pore ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Multiple Tumor

1055 Background: Basal-like breast cancers (BLBC) compose up to 15% of breast cancer (BC) and are usually triple negative characterized by lack of ER, PR, and HER-2 amplification. In addition, most BRCA1-associated BCs are BLBC and TNBC, expressing basal cytokeratins and EGFR. BLBC is characterized by an aggressive phenotype, high histological grade, and poor clinical outcomes: high recurrence and metastasis rates. CRM1 (XPO1) is the exclusive nuclear exporter of multiple Tumor Suppressor Proteins (TSP) including p53, p21, BRCA1&2, pRB, FOXO. CRM1 inhibition forces nuclear accumulation of TSPs, inducing apoptosis in cancer cells. KPT-SINE are novel, small molecule, irreversible inhibitors of CRM1 with potent anti cancer activity. Methods: The Cancer Genome Atlas (TCGA) and BC cell line databases were used for mRNA analyses. MTT assay was used to determine the cytotoxic effect of KPT-SINE (KPT-185 and KPT-330) on 44 breast cell lines including luminal A, luminal B, HER2 positive, BLBC and TNBC cells. The effect of KPT-330 treatment on tumor growth was tested in vivo in the TNBC model MDA-MB-468 xenograft. Results: Analyses of nuclear pore complex (NPC)-related mRNA levels (including CRM1) showed clear separation of BCs into high and low NPC expression. BLBC subtype was enriched with high NPC transcripts while luminal BC was enriched in low NPC levels (p<1.53e-20). High NPC levels had higher mutation levels in BRCA2 (cor=0.33, p=1.83e-8) and ABL1. NPC expression was inversely correlated with ER mRNA expression (cor=-0.58, p=1.37e-7). KPT-SINE showed potent cytotoxicity on >75% of the cell lines (IC50 values <1 μM). Only three of 24 TNBC cell lines displayed IC50 values >1.5 μM upon KPT-SINE treatment. Genomic analyses on all BC lines indicated that p53, PI3K/AKT and BRCA1 or 2 status did not affect cytotoxicity. In MDA-MB-468 xenograft, KPT-330 displayed efficacy in a dose-dependent manner inhibiting nearly 100% of tumor growth compared with vehicle treated animals, and was well tolerated. Conclusions: These data show that NPC/CRM1 mRNAs are overexpressed in BLBC/TNBC and that CRM-1 mediated nuclear export inhibition by SINE represents a potentially novel and well tolerated therapy for BLBC / TNBC.

scholarly journals Mex67p of Schizosaccharomyces pombeInteracts with Rae1p in Mediating mRNA Export

2000 ◽  
Vol 20 (23) ◽  
pp. 8767-8782 ◽  
Author(s):  
Jin Ho Yoon ◽  
Dona C. Love ◽  
Anjan Guhathakurta ◽  
John A. Hanover ◽  
Ravi Dhar
Keyword(s):  
Nuclear Export ◽  
Rna Binding ◽  
Fluorescent Protein ◽  
Synthetic Lethality ◽  
Sequence Similarity ◽  
Mrna Export ◽  
Nuclear Periphery ◽  
Nuclear Pore ◽  
Multicopy Suppressor ◽  
Accessory Factor

ABSTRACT We identified the Schizosaccharomyces pombe mex67 gene (spmex67) as a multicopy suppressor of rae1-167 nup184-1 synthetic lethality and the rae1-167 tsmutation. spMex67p, a 596-amino-acid-long protein, has considerable sequence similarity to the Saccharomyces cerevisiae Mex67p (scMex67p) and human Tap. In contrast toscMEX67, spmex67 is essential for neither growth nor nuclear export of mRNA. However, an spmex67 null mutation (Δmex67) is synthetically lethal with therae1-167 mutation and accumulates poly(A)+ RNA in the nucleus. We identified a central region (149 to 505 amino acids) within spMex67p that associates with a complex containing Rae1p that complements growth and mRNA export defects of therae1-167 Δmex67 synthetic lethality. This region is devoid of RNA-binding, N-terminal nuclear localization, and the C-terminal nuclear pore complex-targeting regions. The (149–505)-green fluorescent protein (GFP) fusion is found diffused throughout the cell. Overexpression of spMex67p inhibits growth and mRNA export and results in the redistribution of the diffused localization of the (149–505)-GFP fusion to the nucleus and the nuclear periphery. These results suggest that spMex67p competes for essential mRNA export factor(s). Finally, we propose that the 149–505 region of spMex67p could act as an accessory factor in Rae1p-dependent transport and that spMex67p participates at various common steps with Rae1p export complexes in promoting the export of mRNA.

scholarly journals Ca2+-Dependent Protein Kinase 6 Enhances KAT2 Shaker Channel Activity in Arabidopsis thaliana

2021 ◽  
Vol 22 (4) ◽  
pp. 1596
Author(s):  
Elsa Ronzier ◽  
Claire Corratgé-Faillie ◽  
Frédéric Sanchez ◽  
Christian Brière ◽  
Tou Cheu Xiong
Keyword(s):  
Arabidopsis Thaliana ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Physical Interaction ◽  
Fluorescence Lifetime Imaging ◽  
Dependent Manner ◽  
In Planta ◽  
Knock Out ◽  
Calcium Dependent ◽  
Dependent Protein

Post-translational regulations of Shaker-like voltage-gated K+ channels were reported to be essential for rapid responses to environmental stresses in plants. In particular, it has been shown that calcium-dependent protein kinases (CPKs) regulate Shaker channels in plants. Here, the focus was on KAT2, a Shaker channel cloned in the model plant Arabidopsis thaliana, where is it expressed namely in the vascular tissues of leaves. After co-expression of KAT2 with AtCPK6 in Xenopuslaevis oocytes, voltage-clamp recordings demonstrated that AtCPK6 stimulates the activity of KAT2 in a calcium-dependent manner. A physical interaction between these two proteins has also been shown by Förster resonance energy transfer by fluorescence lifetime imaging (FRET-FLIM). Peptide array assays support that AtCPK6 phosphorylates KAT2 at several positions, also in a calcium-dependent manner. Finally, K+ fluorescence imaging in planta suggests that K+ distribution is impaired in kat2 knock-out mutant leaves. We propose that the AtCPK6/KAT2 couple plays a role in the homeostasis of K+ distribution in leaves.

scholarly journals The ESCRT-III protein VPS4, but not CHMP4B or CHMP2B, is pathologically increased in familial and sporadic ALS neuronal nuclei

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Alyssa N. Coyne ◽  
Jeffrey D. Rothstein
Keyword(s):  
Nuclear Pore Complex ◽  
Mammalian Cells ◽  
Nuclear Accumulation ◽  
Nuclear Pore ◽  
Dependent Manner ◽  
Structured Illumination Microscopy ◽  
Neuronal Nuclei ◽  
Complex Injury ◽  
Sporadic Als ◽  
Human Neurons

AbstractNuclear pore complex injury has recently emerged as an early and significant contributor to familial and sporadic ALS disease pathogenesis. However, the molecular events leading to this pathological phenomenon characterized by the reduction of specific nucleoporins from neuronal nuclear pore complexes remain largely unknown. This is due in part to a lack of knowledge regarding the biological pathways and proteins underlying nuclear pore complex homeostasis specifically in human neurons. We have recently uncovered that aberrant nuclear accumulation of the ESCRT-III protein CHMP7 initiates nuclear pore complex in familial and sporadic ALS neurons. In yeast and non-neuronal mammalian cells, nuclear relocalization of CHMP7 has been shown to recruit the ESCRT-III proteins CHMP4B, CHMP2B, and VPS4 to facilitate nuclear pore complex and nuclear envelope repair and homeostasis. Here, using super resolution structured illumination microscopy, we find that neither CHMP4B nor CHMP2B are increased in ALS neuronal nuclei. In contrast, VPS4 expression is significantly increased in ALS neuronal nuclei prior to the emergence of nuclear pore injury in a CHMP7 dependent manner. However, unlike our prior CHMP7 knockdown studies, impaired VPS4 function does not mitigate alterations to the NPC and the integral transmembrane nucleoporin POM121. Collectively our data suggest that while alterations in VPS4 subcellular localization appear to be coincident with nuclear pore complex injury, therapeutic efforts to mitigate this pathogenic cascade should be targeted towards upstream events such as the nuclear accumulation of CHMP7 as we have previously described.

scholarly journals Nuclear Export of Smads by RanBP3L Regulates Bone Morphogenetic Protein Signaling and Mesenchymal Stem Cell Differentiation

2015 ◽  
Vol 35 (10) ◽  
pp. 1700-1711 ◽  
Author(s):  
Fenfang Chen ◽  
Xia Lin ◽  
Pinglong Xu ◽  
Zhengmao Zhang ◽  
Yanzhen Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Mesenchymal Stem Cell ◽  
Nuclear Export ◽  
Stem Cell Differentiation ◽  
Bmp Signaling ◽  
Dependent Manner ◽  
Mesenchymal Stem Cell Differentiation

Bone morphogenetic proteins (BMPs) play vital roles in regulating stem cell maintenance and differentiation. BMPs can induce osteogenesis and inhibit myogenesis of mesenchymal stem cells. Canonical BMP signaling is stringently controlled through reversible phosphorylation and nucleocytoplasmic shuttling of Smad1, Smad5, and Smad8 (Smad1/5/8). However, how the nuclear export of Smad1/5/8 is regulated remains unclear. Here we report that the Ran-binding protein RanBP3L acts as a nuclear export factor for Smad1/5/8. RanBP3L directly recognizes dephosphorylated Smad1/5/8 and mediates their nuclear export in a Ran-dependent manner. Increased expression of RanBP3L blocks BMP-induced osteogenesis of mouse bone marrow-derived mesenchymal stem cells and promotes myogenic induction of C2C12 mouse myoblasts, whereas depletion of RanBP3L expression enhances BMP-dependent stem cell differentiation activity and transcriptional responses. In conclusion, our results demonstrate that RanBP3L, as a nuclear exporter for BMP-specific Smads, plays a critical role in terminating BMP signaling and regulating mesenchymal stem cell differentiation.

scholarly journals SUMO-1 Modification and Its Role in Targeting the Ran GTPase-activating Protein, RanGAP1, to the Nuclear Pore Complex

1998 ◽  
Vol 140 (3) ◽  
pp. 499-509 ◽  
Author(s):  
Michael J. Matunis ◽  
Jian Wu ◽  
Günter Blobel
Keyword(s):  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Gtpase Activating Protein ◽  
Terminal Domain ◽  
Localization Signal

RanGAP1 is the GTPase-activating protein for Ran, a small ras-like GTPase involved in regulating nucleocytoplasmic transport. In vertebrates, RanGAP1 is present in two forms: one that is cytoplasmic, and another that is concentrated at the cytoplasmic fibers of nuclear pore complexes (NPCs). The NPC-associated form of RanGAP1 is covalently modified by the small ubiquitin-like protein, SUMO-1, and we have recently proposed that SUMO-1 modification functions to target RanGAP1 to the NPC. Here, we identify the domain of RanGAP1 that specifies SUMO-1 modification and demonstrate that mutations in this domain that inhibit modification also inhibit targeting to the NPC. Targeting of a heterologous protein to the NPC depended on determinants specifying SUMO-1 modification and also on additional determinants in the COOH-terminal domain of RanGAP1. SUMO-1 modification and these additional determinants were found to specify interaction between the COOH-terminal domain of RanGAP1 and a region of the nucleoporin, Nup358, between Ran-binding domains three and four. Together, these findings indicate that SUMO-1 modification targets RanGAP1 to the NPC by exposing, or creating, a Nup358 binding site in the COOH-terminal domain of RanGAP1. Surprisingly, the COOH-terminal domain of RanGAP1 was also found to harbor a nuclear localization signal. This nuclear localization signal, and the presence of nine leucine-rich nuclear export signal motifs, suggests that RanGAP1 may shuttle between the nucleus and the cytoplasm.

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