scholarly journals The UL69 Transactivator Protein of Human Cytomegalovirus Interacts with DEXD/H-Box RNA Helicase UAP56 To Promote Cytoplasmic Accumulation of Unspliced RNA

2006 ◽  
Vol 26 (5) ◽  
pp. 1631-1643 ◽  
Author(s):  
Peter Lischka ◽  
Zsolt Toth ◽  
Marco Thomas ◽  
Regina Mueller ◽  
Thomas Stamminger
Keyword(s):  
Human Cytomegalovirus ◽  
Nuclear Export ◽  
Mrna Export ◽  
Regulatory Protein ◽  
Nuclear Export Signal ◽  
Cellular Transport ◽  
Nucleocytoplasmic Shuttling ◽  
Rna Helicases ◽  
Cytoplasmic Accumulation ◽  
Acid Domain

ABSTRACT The UL69 gene product of human cytomegalovirus belongs to a family of regulatory proteins conserved among all herpesviruses that have in part been characterized as posttranscriptional transactivators participating in the nuclear export of RNA. Recent experiments suggested that pUL69 also acts as a posttranscriptional activator since it was demonstrated that nucleocytoplasmic shuttling via a CRM1-independent nuclear export signal is a prerequisite for its stimulatory effect on gene expression. Based on these findings we initiated studies to investigate the role of pUL69 in mRNA export and demonstrate that pUL69 efficiently promotes the cytoplasmic accumulation of unspliced RNA. Furthermore, we show that this pUL69 activity is linked to the cellular mRNA export machinery by direct protein interaction with the highly related DEXD/H-box RNA helicases UAP56 and URH49. Particularly, we identified a 12-amino-acid domain within the N terminus of pUL69 which is required for binding to UAP56 and URH49, and we could demonstrate that UAP56 interaction and nucleocytoplasmic shuttling are both prerequisites for pUL69-mediated mRNA export. Thus, we identified a novel cellular target which provides a herpesviral regulatory protein with access to a conserved cellular transport system in order to promote nuclear export of unspliced RNA.

scholarly journals Human Cytomegalovirus UL84 Protein Contains Two Nuclear Export Signals and Shuttles between the Nucleus and the Cytoplasm

2006 ◽  
Vol 80 (20) ◽  
pp. 10274-10280 ◽  
Author(s):  
Peter Lischka ◽  
Claudia Rauh ◽  
Regina Mueller ◽  
Thomas Stamminger
Keyword(s):  
Human Cytomegalovirus ◽  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Nuclear Export ◽  
Signal Sequence ◽  
Regulatory Protein ◽  
Nucleocytoplasmic Shuttling ◽  
Viral Dna ◽  
Nuclear Localization Signal Sequence ◽  
Localization Signal

ABSTRACT Previous studies defined pUL84 of human cytomegalovirus as an essential regulatory protein with nuclear localization that was proposed to act during initiation of viral-DNA synthesis. Recently, we demonstrated that a complex domain of 282 amino acids within pUL84 functions as a nonconventional nuclear localization signal. Sequence inspection of this domain revealed the presence of motifs with homology to leucine-rich nuclear export signals. Here, we report the identification of two functional, autonomous nuclear export signals and show that pUL84 acts as a CRM-1-dependent nucleocytoplasmic shuttling protein. This suggests an unexpected cytoplasmic role for this essential viral regulatory protein.

scholarly journals RNA-Regulated Interaction of Transportin-1 and Exportin-5 with the Double-Stranded RNA-Binding Domain Regulates Nucleocytoplasmic Shuttling of ADAR1

2009 ◽  
Vol 29 (6) ◽  
pp. 1487-1497 ◽  
Author(s):  
Jutta Fritz ◽  
Alexander Strehblow ◽  
Andreas Taschner ◽  
Sandy Schopoff ◽  
Pawel Pasierbek ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Rna Binding ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Nucleocytoplasmic Shuttling ◽  
Double Stranded Rna ◽  
Binding Domains ◽  
Dsrna Binding ◽  
Localization Signal ◽  
Editing Enzyme

ABSTRACT Double-stranded RNA (dsRNA)-binding proteins interact with substrate RNAs via dsRNA-binding domains (dsRBDs). Several proteins harboring these domains exhibit nucleocytoplasmic shuttling and possibly remain associated with their substrate RNAs bound in the nucleus during nuclear export. In the human RNA-editing enzyme ADAR1-c, the nuclear localization signal overlaps the third dsRBD, while the corresponding import factor is unknown. The protein also lacks a clear nuclear export signal but shuttles between the nucleus and the cytoplasm. Here we identify transportin-1 as the import receptor for ADAR1. Interestingly, dsRNA binding interferes with transportin-1 binding. At the same time, each of the dsRBDs in ADAR1 interacts with the export factor exportin-5. RNA binding stimulates this interaction but is not a prerequisite. Thus, our data demonstrate a role for some dsRBDs as RNA-sensitive nucleocytoplasmic transport signals. dsRBD3 in ADAR1 can mediate nuclear import, while interaction of all dsRBDs might control nuclear export. This finding may have implications for other proteins containing dsRBDs and suggests a selective nuclear export mechanism for substrates interacting with these proteins.

scholarly journals Involvement of Nucleocytoplasmic Shuttling of Yeast Nap1 in Mitotic Progression

2003 ◽  
Vol 23 (18) ◽  
pp. 6672-6684 ◽  
Author(s):  
Mary Miyaji-Yamaguchi ◽  
Kohsuke Kato ◽  
Ryosuke Nakano ◽  
Tomohiro Akashi ◽  
Akihiko Kikuchi ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Temperature Sensitive ◽  
Nucleosome Assembly ◽  
Mitotic Progression ◽  
Nucleocytoplasmic Shuttling ◽  
Nucleosome Formation ◽  
Mitotic Cyclin ◽  
Carboxy Terminal

ABSTRACT Nucleosome assembly protein 1 (Nap1) is widely conserved from yeasts to humans and facilitates nucleosome formation in vitro as a histone chaperone. Nap1 is generally localized in the cytoplasm, except that subcellular localization of Drosophila melanogaster Nap1 is dynamically regulated between the cytoplasm and nucleus during early development. The cytoplasmic localization of Nap1 is seemingly incompatible with the proposed role of Nap1 in nucleosome formation, which should occur in the nucleus. Here, we have examined the roles of a putative nuclear export signal (NES) sequence in yeast Nap1 (yNap1). yNap1 mutants lacking the NES-like sequence were localized predominantly in the nucleus. Deletion of NAP1 in cells harboring a single mitotic cyclin gene is known to cause mitotic delay and temperature-sensitive growth. A wild-type NAP1 complemented these phenotypes while nap1 mutant genes lacking the NES-like sequence or carboxy-terminal region did not. These and other results suggest that yNap1 is a nucleocytoplasmic shuttling protein and that its shuttling is important for yNap1 function during mitotic progression. This study also provides a possible explanation for Nap1's involvement in nucleosome assembly and/or remodeling in the nucleus.

scholarly journals A Novel Nuclear Export Signal and a REF Interaction Domain Both Promote mRNA Export by the Epstein-Barr Virus EB2 Protein

2002 ◽  
Vol 278 (1) ◽  
pp. 335-342 ◽  
Author(s):  
Edwige Hiriart ◽  
Géraldine Farjot ◽  
Henri Gruffat ◽  
Minh Vu Chuong Nguyen ◽  
Alain Sergeant ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Epstein Barr Virus ◽  
Mrna Export ◽  
Nuclear Export Signal ◽  
Interaction Domain ◽  
Barr Virus ◽  
Epstein Barr ◽  
Export Signal

scholarly journals Mice develop normally in the absence of Smad4 nucleocytoplasmic shuttling

2007 ◽  
Vol 404 (2) ◽  
pp. 235-245 ◽  
Author(s):  
Christine A. Biondi ◽  
Debipriya Das ◽  
Michael Howell ◽  
Ayesha Islam ◽  
Elizabeth K. Bikoff ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Nuclear Export ◽  
Transforming Growth Factor ◽  
Es Cells ◽  
Nuclear Export Signal ◽  
Embryonic Stem ◽  
Nucleocytoplasmic Shuttling ◽  
Mouse Development ◽  
Developmental Defects

Smad4 in partnership with R-Smads (receptor-regulated Smads) activates TGF-β (transforming growth factor-β)-dependent signalling pathways essential for early mouse development. Smad4 null embryos die shortly after implantation due to severe defects in cell proliferation and visceral endoderm differentiation. In the basal state, Smad4 undergoes continuous shuttling between the cytoplasm and the nucleus due to the combined activities of an N-terminal NLS (nuclear localization signal) and an NES (nuclear export signal) located in its linker region. Cell culture experiments suggest that Smad4 nucleocytoplasmic shuttling plays an important role in TGF-β signalling. In the present study we have investigated the role of Smad4 shuttling in vivo using gene targeting to engineer two independent mutations designed to eliminate Smad4 nuclear export. As predicted this results in increased levels of Smad4 in the nucleus of homozygous ES cells (embryonic stem cells) and primary keratinocytes, in the presence or absence of ligand. Neither mutation affects Smad4 expression levels nor its ability to mediate transcriptional activation in homozygous cell lines. Remarkably mouse mutants lacking the Smad4 NES develop normally. Smad4 NES mutants carrying one copy of a Smad4 null allele also fail to display developmental defects. The present study clearly demonstrates that Smad4 nucleocytoplasmic shuttling is not required for embryonic development or tissue homoeostasis in normal, healthy adult mice.

scholarly journals NOVEL NPM1 EXON 5 MUTATIONS AND GENE FUSIONS LEADING TO ABERRANT CYTOPLASMIC NUCLEOPHOSMIN IN AML

Blood ◽  
2021 ◽  
Author(s):  
Maria Paola Martelli ◽  
Roberta Rossi ◽  
Alessandra Venanzi ◽  
Manja Meggendorfer ◽  
Vincenzo Maria Perriello ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Myeloid Leukemia ◽  
Nuclear Export Signal ◽  
Fusion Partner ◽  
Functional Studies ◽  
Exon 9 ◽  
Cytoplasmic Accumulation ◽  
Exon 11 ◽  
Export Signal

Nucleophosmin (NPM1) mutations in acute myeloid leukemia (AML) affect exon 12, but sporadically also exon 9 and 11, all causing changes at protein C-terminal end (loss of tryptophans and creation of a nuclear export signal-NES motif) that lead to aberrant cytoplasmic NPM1 (NPM1c+), detectable by immunohistochemistry. Combining immunohistochemistry and molecular analyses in 929 AML patients, we found non-exon 12 NPM1 mutations in 5/387 (1.3%) NPM1c+ cases. Besides mutations in exon 9 (n=1) and exon 11 (n=1), novel mutations in exon 5 were discovered (n=3). One more exon 5 mutation was identified in additional 141 AML patients selected for wild-type NPM1 exon 12. Furthermore, 3 NPM1 rearrangements (i.e. NPM1/RPP30, NPM1/SETBP1, NPM1/CCDC28A) were detected and characterized among 13,979 AML samples screened by cytogenetic/FISH and RNA sequencing. Functional studies demonstrated that in AML cases the new NPM1 proteins harboured an efficient extra NES, either newly created or already present in the fusion partner, ensuring its cytoplasmic accumulation. Our findings support NPM1 cytoplasmic relocation as critical for leukemogenesis and reinforce the role of immunohistochemistry in predicting any AML-associated NPM1 genetic lesions. Also, this study highlights the need for developing new specific assays for molecular diagnosis and monitoring of NPM1-mutated AML.

scholarly journals Borna Disease Virus Nucleoprotein Requires both Nuclear Localization and Export Activities for Viral Nucleocytoplasmic Shuttling

2001 ◽  
Vol 75 (7) ◽  
pp. 3404-3412 ◽  
Author(s):  
Takeshi Kobayashi ◽  
Wataru Kamitani ◽  
Guoqi Zhang ◽  
Makiko Watanabe ◽  
Keizo Tomonaga ◽  
...  
Keyword(s):  
Disease Virus ◽  
Nuclear Localization ◽  
Nuclear Export ◽  
Critical Role ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Shuttling ◽  
Nuclear Targeting ◽  
Borna Disease Virus ◽  
Export Activity ◽  
Borna Disease

ABSTRACT Nuclear transport of viral nucleic acids is crucial to the life cycle of many viruses. Borna disease virus (BDV) belongs to the orderMononegavirales and replicates its RNA genome in the nucleus. Previous studies have suggested that BDV nucleoprotein (N) and phosphoprotein (P) have important functions in the nuclear import of the viral ribonucleoprotein (RNP) complexes via their nuclear targeting activity. Here, we showed that BDV N has cytoplasmic localization activity, which is mediated by a nuclear export signal (NES) within the sequence. Our analysis using deletion and substitution mutants of N revealed that NES of BDV N consists of a canonical leucine-rich motif and that the nuclear export activity of the protein is mediated through the chromosome region maintenance protein-dependent pathway. Interspecies heterokaryon assay indicated that BDV N shuttles between the nucleus and cytoplasm as a nucleocytoplasmic shuttling protein. Furthermore, interestingly, the NES region overlaps a binding site to the BDV P protein, and nuclear export of a 38-kDa form of BDV N is prevented by coexpression of P. These results suggested that BDV N has two contrary activities, nuclear localization and export activity, and plays a critical role in the nucleocytoplasmic transport of BDV RNP by interaction with other viral proteins.

scholarly journals Nucleocytoplasmic Shuttling Modulates Activity and Ubiquitination-Dependent Turnover of SUMO-Specific Protease 2

2006 ◽  
Vol 26 (12) ◽  
pp. 4675-4689 ◽  
Author(s):  
Yoko Itahana ◽  
Edward T. H. Yeh ◽  
Yanping Zhang
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Shuttling ◽  
Leptomycin B ◽  
Sumo Protease ◽  
Specific Protease ◽  
Localization Signal ◽  
Modified Proteins ◽  
Export Signal

ABSTRACT Small ubiquitin-related modifier (SUMO) proteins are conjugated to numerous polypeptides in cells, and attachment of SUMO plays important roles in regulating the activity, stability, and subcellular localization of modified proteins. SUMO modification of proteins is a dynamic and reversible process. A family of SUMO-specific proteases catalyzes the deconjugation of SUMO-modified proteins. Members of the Sentrin (also known as SUMO)-specific protease (SENP) family have been characterized with unique subcellular localizations. However, little is known about the functional significance of or the regulatory mechanism derived from the specific localizations of the SENPs. Here we identify a bipartite nuclear localization signal (NLS) and a CRM1-dependent nuclear export signal (NES) in the SUMO protease SENP2. Both the NLS and the NES are located in the nonhomologous domains of SENP2 and are not conserved among other members of the SENP family. Using a series of SENP2 mutants and a heterokaryon assay, we demonstrate that SENP2 shuttles between the nucleus and the cytoplasm and that the shuttling is blocked by mutations in the NES or by treating cells with leptomycin B. We show that SENP2 can be polyubiquitinated in vivo and degraded through proteolysis. Restricting SENP2 in the nucleus by mutations in the NES impairs its polyubiquitination, whereas a cytoplasm-localized SENP2 made by introducing mutations in the NLS can be efficiently polyubiquitinated, suggesting that SENP2 is ubiquitinated in the cytoplasm. Finally, treating cells with MG132 leads to accumulation of polyubiquitinated SENP2, indicating that SENP2 is degraded through the 26S proteolysis pathway. Thus, the function of SENP2 is regulated by both nucleocytoplasmic shuttling and polyubiquitin-mediated degradation.

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