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 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 A Curious Novel Combination of Nucleophosmin (NPM1) Gene Mutations Leading to Aberrant Cytoplasmic Dislocation of NPM1 in Acute Myeloid Leukemia (AML)

Genes ◽  
2021 ◽  
Vol 12 (9) ◽  
pp. 1426
Author(s):  
Alessandra Venanzi ◽  
Roberta Rossi ◽  
Giovanni Martino ◽  
Ombretta Annibali ◽  
Giuseppe Avvisati ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Nuclear Export ◽  
Myeloid Leukemia ◽  
Nuclear Export Signal ◽  
Gene Mutations ◽  
Molecular Alteration ◽  
C Terminus ◽  
Molecular Sequencing ◽  
Acute Myeloid

Nucleophosmin (NPM1) mutations occurring in acute myeloid leukemia (AML) (about 50 so far identified) cluster almost exclusively in exon 12 and lead to common changes at the NPM1 mutants C-terminus, i.e., loss of tryptophans 288 and 290 (or 290 alone) and creation of a new nuclear export signal (NES), at the bases of exportin-1(XPO1)-mediated aberrant cytoplasmic NPM1. Immunohistochemistry (IHC) detects cytoplasmic NPM1 and is predictive of the molecular alteration. Besides IHC and molecular sequencing, Western blotting (WB) with anti-NPM1 mutant specific antibodies is another approach to identify NPM1-mutated AML. Here, we show that among 382 AML cases with NPM1 exon 12 mutations, one was not recognized by WB, and describe the discovery of a novel combination of two mutations involving exon 12. This appeared as a conventional mutation A with the known TCTG nucleotides insertion/duplication accompanied by a second event (i.e., an 8-nucleotide deletion occurring 15 nucleotides downstream of the TCTG insertion), resulting in a new C-terminal protein sequence. Strikingly, the sequence included a functional NES ensuring cytoplasmic relocation of the new mutant supporting the role of cytoplasmic NPM1 as critical in AML leukemogenesis.

scholarly journals Adenovirus Late Gene Expression Does Not Require a Rev-Like Nuclear RNA Export Pathway

2000 ◽  
Vol 74 (14) ◽  
pp. 6684-6688 ◽  
Author(s):  
Claudia Rabino ◽  
Anders Aspegren ◽  
Kara Corbin-Lickfett ◽  
Eileen Bridge
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Late Gene Expression ◽  
Export Pathway ◽  
Early Proteins ◽  
Immunodeficiency Virus ◽  
Rna Export ◽  
Nuclear Rna Export ◽  
Export Signal

ABSTRACT Adenovirus late mRNA export is facilitated by viral early proteins of 55 and 34 kDa. The 34-kDa protein contains a leucine-rich nuclear export signal (NES) similar to that of the human immunodeficiency virus Rev protein. It was proposed that the 34-kDa protein might facilitate the export of adenovirus late mRNA through a Rev-like NES-mediated export pathway. We have tested the role of NES-mediated RNA export during adenovirus infection, and we find that it is not essential for the expression of adenovirus late genes.

scholarly journals The Membrane-Anchoring Region of the AcMNPV P74 Protein Is Expendable or Interchangeable with Homologs from Other Species

Viruses ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2416
Author(s):  
María Victoria Nugnes ◽  
Alexandra Marisa Targovnik ◽  
Adrià Mengual-Martí ◽  
María Victoria Miranda ◽  
Carolina Susana Cerrudo ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Virus Infectivity ◽  
Amino Terminal ◽  
Insect Pathogens ◽  
Transmembrane Regions ◽  
Membrane Anchoring ◽  
P74 Gene ◽  
Export Signal

Baculoviruses are insect pathogens that are characterized by assembling the viral dsDNA into two different enveloped virions during an infective cycle: occluded virions (ODVs; immersed in a protein matrix known as occlusion body) and budded virions (BVs). ODVs are responsible for the primary infection in midgut cells of susceptible larvae thanks to the per os infectivity factor (PIF) complex, composed of at least nine essential viral proteins. Among them, P74 is a crucial factor whose activity has been identified as virus-specific. In this work, the p74 gene from AcMNPV was pseudogenized using CRISPR/Cas9 technology and then complemented with wild-type alleles from SeMNPV and HearSNPV species, as well as chimeras combining the P74 amino and carboxyl domains. The results on Spodoptera exigua and Rachiplusia nu larvae showed that an amino terminal sector of P74 (lacking two potential transmembrane regions but possessing a putative nuclear export signal) is sufficient to restore the virus infectivity whether alone or fused to the P74 transmembrane regions of the other evaluated viral species. These results provide novel information about the functional role of P74 and delimit the region on which mutagenesis could be applied to enhance viral activity and, thus, produce better biopesticides.

The Nup98-HoxA9 Leukemogenic Fusion Protein Blocks Xpo1 and Tap-Mediated Nuclear Export

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5228-5228
Author(s):  
Yildirim Dogan ◽  
Sutapa Sahay ◽  
David C Dorn ◽  
Anna Franceschino ◽  
Michael Xu ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Conflicts Of Interest ◽  
Blast Crisis ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Shuttling ◽  
Nuclear Trafficking ◽  
Myeloid Leukemias ◽  
Nuclear Export Receptor ◽  
Export Signal

Abstract Abstract 5228 In leukemia Nup98 fusion genes are found as a product of the reciprocal translocation between Nucleoporin 98 (nup98) and homeobox-cluster genes. Nup98-HoxA9 (NHA9), the product of the t(7;11) translocation, is detected in acute myeloid leukemias (AMLs) and as a secondary abnormality in blast crisis of chronic myeloid leukemia (CML). The role of NHA9 in leukemogenesis is complex and incompletely understood. Here, we show an abrogation of nucleocytoplasmic shuttling of the nuclear export receptor Xpo1 and Tap in NHA9-expressing cells by using retroviral nuclear trafficking as a model. Lentiviral Rev, the prototype for nuclear export signal (NES)-containing proteins is shuttled through the nucleopore by Xpo1. NHA9 sequestered Xpo1 from the nuclear rim into nuclear aggregates resulting in deficient Xpo1-dependent nuclear exit of Rev and its mRNA substrates. Tap is involved in mRNA nucleocytoplasmic shuttling and is also responsible for the nuclear export of D-type retrovirus CTE-mRNAs. By using Tap/CTE-mRNA nuclear export as a model we also found that Tap colocalized in NHA9 nuclear aggregates leading to impaired Tap-mediated nuclear exit of CTE-mRNA substrates. Leukemogenicity of Nup98 fusion proteins may be accounted for in part by defects in Tap and Xpo1-mediated export of their substrates. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The Nuclear Export Signal within the E4orf6 Protein of Adenovirus Type 5 Supports Virus Replication and Cytoplasmic Accumulation of Viral mRNA

2000 ◽  
Vol 74 (2) ◽  
pp. 764-772 ◽  
Author(s):  
Silke Weigel ◽  
Matthias Dobbelstein
Keyword(s):  
Nuclear Export ◽  
Late Phase ◽  
Nuclear Export Signal ◽  
Adenovirus Type ◽  
Adenovirus Infection ◽  
Viral Mrna ◽  
Viral Dna ◽  
Cytoplasmic Accumulation ◽  
Adenovirus Type 5 ◽  
Export Signal

ABSTRACT 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.

scholarly journals The Adenovirus Type 5 E1B-55K Oncoprotein Actively Shuttles in Virus-Infected Cells, Whereas Transport of E4orf6 Is Mediated by a CRM1-Independent Mechanism

2001 ◽  
Vol 75 (12) ◽  
pp. 5677-5683 ◽  
Author(s):  
Tanja Dosch ◽  
Florian Horn ◽  
Grit Schneider ◽  
Friedrich Krätzer ◽  
Thomas Dobner ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Mrna Export ◽  
Nuclear Export Signal ◽  
Adenovirus Type ◽  
Alternative Mechanism ◽  
Leptomycin B ◽  
Infected Cells ◽  
Adenovirus Type 5 ◽  
Export Signal

ABSTRACT The E1B-55K and E4orf6 proteins of adenovirus type 5 are involved in viral mRNA export. Here we demonstrate that adenovirus infection does not inhibit the function of the E1B-55K nuclear export signal and that E1B-55K also shuttles in infected cells. Even during virus infection, E1B-55K was exported by the leptomycin B-sensitive CRM1 pathway, whereas E4orf6 transport appeared to be mediated by an alternative mechanism. Our results strengthen the potential role of E1B-55K as the “driving force” for adenoviral late mRNA export.

scholarly journals Nucleocytoplasmic Distribution of Human RNA-editing Enzyme ADAR1 Is Modulated by Double-stranded RNA-binding Domains, a Leucine-rich Export Signal, and a Putative Dimerization Domain

2002 ◽  
Vol 13 (11) ◽  
pp. 3822-3835 ◽  
Author(s):  
Alexander Strehblow ◽  
Martina Hallegger ◽  
Michael F. Jantsch
Keyword(s):  
Rna Editing ◽  
Nuclear Localization ◽  
Nuclear Export ◽  
Rna Binding ◽  
Nuclear Export Signal ◽  
Nuclear Accumulation ◽  
Double Stranded Rna ◽  
Cytoplasmic Accumulation ◽  
Editing Enzyme ◽  
Export Signal

The human RNA-editing enzyme adenosine deaminase that acts on RNA (ADAR1) is expressed in two versions. A longer 150-kDa protein is interferon inducible and can be found both in the nucleus and cytoplasm. An amino-terminally truncated 110-kDa version, in contrast, is constitutively expressed and predominantly nuclear. In the absence of transcription, however, the shorter protein is also cytoplasmic and thus displays the hallmarks of a shuttling protein. The nuclear localization signal (NLS) of human hsADAR1 is atypical and overlaps with its third double-stranded RNA-binding domain (dsRBD). Herein, we identify regions in hsADAR1 that interfere with nuclear localization and mediate cytoplasmic accumulation. We show that interferon-inducible hsADAR1 contains a Crm1-dependent nuclear export signal in its amino terminus. Most importantly, we demonstrate that the first dsRBD of hsADAR1 interferes with nuclear localization of a reporter construct containing dsRBD3 as an active NLS. The same effect can be triggered by several other, but not all dsRBDs. Active RNA binding of either the inhibitory dsRBD1 or the NLS bearing dsRBD3 is required for cytoplasmic accumulation. Furthermore, hsADAR1's dsRBD1 has no effect on other NLSs, suggesting RNA-mediated cross talk between dsRBDs, possibly leading to masking of the NLS. A model, incorporating these findings is presented. Finally, we identify a third region located in the C terminus of hsADAR1 that also interferes with nuclear accumulation of this protein.

Sign in / Sign up

Export Citation Format

Share Document