scholarly journals The prototype γ-2 herpesvirus nucleocytoplasmic shuttling protein, ORF 57, transports viral RNA through the cellular mRNA export pathway

2005 ◽  
Vol 387 (2) ◽  
pp. 295-308 ◽  
Author(s):  
Ben J. L. WILLIAMS ◽  
James R. BOYNE ◽  
Delyth J. GOODWIN ◽  
Louise ROADEN ◽  
Guillaume M. HAUTBERGUE ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Mammalian Cells ◽  
Mrna Export ◽  
Dominant Negative ◽  
Viral Rna ◽  
Exon Junction Complex ◽  
Independent Manner ◽  
Exon Junction ◽  
Export Pathway ◽  
Rna Export

HVS (herpesvirus saimiri) is the prototype γ-2 herpesvirus. This is a subfamily of herpesviruses gaining importance since the identification of the first human γ-2 herpesvirus, Kaposi's sarcoma-associated herpesvirus. The HVS ORF 57 (open reading frame 57) protein is a multifunctional transregulatory protein homologous with genes identified in all classes of herpesviruses. Recent work has demonstrated that ORF 57 has the ability to bind viral RNA, shuttles between the nucleus and cytoplasm and promotes the nuclear export of viral transcripts. In the present study, we show that ORF 57 shuttles between the nucleus and cytoplasm in a CRM-1 (chromosomal region maintenance 1)-independent manner. ORF 57 interacts with the mRNA export factor REF (RNA export factor) and two other components of the exon junction complex, Y14 and Magoh. The association of ORF 57 with REF stimulates recruitment of the cellular mRNA export factor TAP (Tip-associated protein), and HVS infection triggers the relocalization of REF and TAP from the nuclear speckles to several large clumps within the cell. Using a dominant-negative form of TAP and RNA interference to deplete TAP, we show that it is essential for bulk mRNA export in mammalian cells and is required for ORF 57-mediated viral RNA export. Furthermore, we show that the disruption of TAP reduces viral replication. These results indicate that HVS utilizes ORF 57 to recruit components of the exon junction complex and subsequently TAP to promote viral RNA export through the cellular mRNA export pathway.

scholarly journals Nuclear Export Through Nuclear Envelope Remodeling inSaccharomyces cerevisiae

2017 ◽  
Author(s):  
Baojin Ding ◽  
Anne M. Mirza ◽  
James Ashley ◽  
Vivian Budnik ◽  
Mary Munson
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Export ◽  
Mammalian Cells ◽  
Budding Yeast ◽  
Mrna Export ◽  
Low Frequency ◽  
Nuclear Pore ◽  
Wild Type ◽  
Export Pathway ◽  
Wild Type Yeast

ABSTRACTIn eukaryotes, subsets of exported mRNAs are organized into large ribonucleoprotein (megaRNP) granules. How megaRNPs exit the nucleus is unclear, as their diameters are much larger than the nuclear pore complex (NPC) central channel. We previously identified a non-canonical nuclear export mechanism inDrosophila(Speese et al.,Cell2012) and mammals (Ding et al., in preparation), in which megaRNPs exit the nucleus by budding across nuclear envelope (NE) membranes. Here, we present evidence for a similar pathway in the nucleus of the budding yeast S.cerevisiae, which contain morphologically similar granules bearing mRNAs. Wild-type yeast displayed these granules at very low frequency, but this frequency was dramatically increased when the non-essential NPC protein Nup116 was deleted. These granules were not artifacts of defective NPCs; a mutation in the exportinXPO1(CRM1), in which NPCs are normal, induced similar megaRNP upregulation. We hypothesize that a non-canonical nuclear export pathway, analogous to those observed inDrosophilaand in mammalian cells, exists in yeast, and that this pathway is upregulated for use when NPCs or nuclear export are impaired.SUMMARYDing et al., describe a non-canonical mRNA export pathway in budding yeast similar to that observed inDrosophila. This pathway appears upregulated when the NPC is impaired, nuclear envelope integrity is disrupted, or the export factor Xpo1 (CRM1) is defective.

scholarly journals A heterochromatin-specific RNA export pathway facilitates piRNA production

2019 ◽  
Author(s):  
Mostafa F. ElMaghraby ◽  
Peter Refsing Andersen ◽  
Florian Pühringer ◽  
Katharina Meixner ◽  
Thomas Lendl ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Mrna Export ◽  
Surveillance Systems ◽  
Rna Surveillance ◽  
Export Pathway ◽  
Transposon Silencing ◽  
Nuclear Rna ◽  
Rna Export ◽  
Non Coding Rnas ◽  
Pirna Pathway

PIWI-interacting RNAs (piRNAs) guide transposon silencing in animals. The 22-30nt piRNAs are processed in the cytoplasm from long non-coding RNAs. How piRNA precursors, which often lack RNA processing hallmarks of export-competent transcripts, achieve nuclear export is unknown. Here, we uncover the RNA export pathway specific for piRNA precursors in theDrosophilagermline. This pathway requires Nxf3-Nxt1, a variant of the hetero-dimeric mRNA export receptor Nxf1-Nxt1. Nxf3 interacts with UAP56, a nuclear RNA helicase essential for mRNA export, and CG13741/Bootlegger, which recruits Nxf3-Nxt1 and UAP56 to heterochromatic piRNA source loci. Upon RNA cargo binding, Nxf3 achieves nuclear export via the exportin Crm1, and accumulates together with Bootlegger in peri-nuclear nuage, suggesting that after export, Nxf3-Bootlegger delivers precursor transcripts to the piRNA processing sites. Our findings indicate that the piRNA pathway bypasses nuclear RNA surveillance systems to achieve export of heterochromatic, unprocessed transcripts to the cytoplasm, a strategy also exploited by retroviruses.

scholarly journals Roles for RNA export factor, Nxt1, in ensuring muscle integrity and normal RNA expression in Drosophila

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Kevin van der Graaf ◽  
Katia Jindrich ◽  
Robert Mitchell ◽  
Helen White-Cooper
Keyword(s):  
Mrna Export ◽  
Rna Stability ◽  
Muscle Degeneration ◽  
Loss Of Function ◽  
Cellular Functions ◽  
Partial Loss ◽  
Export Pathway ◽  
Pathway Gene ◽  
Rna Export

Abstract The mRNA export pathway is responsible for the transport of mRNAs from the nucleus to the cytoplasm, and thus is essential for protein production and normal cellular functions. A partial loss of function allele of the mRNA export factor Nxt1 in Drosophila shows reduced viability and sterility. A previous study has shown that the male fertility defect is due to a defect in transcription and RNA stability, indicating the potential for this pathway to be implicated in processes beyond the known mRNA transport function. Here we investigate the reduced viability of Nxt1 partial loss of function mutants, and describe a defect in growth and maintenance of the larval muscles, leading to muscle degeneration. RNA-seq revealed reduced expression of a set of mRNAs, particularly from genes with long introns in Nxt1 mutant carcass. We detected differential expression of circRNA, and significantly fewer distinct circRNAs expressed in the mutants. Despite the widespread defects in gene expression, muscle degeneration was rescued by increased expression of the costamere component tn (abba) in muscles. This is the first report of a role for the RNA export pathway gene Nxt1 in the maintenance of muscle integrity. Our data also links the mRNA export pathway to a specific role in the expression of mRNA and circRNA from common precursor genes, in vivo.

scholarly journals Adenovirus Ubiquitin-Protein Ligase Stimulates Viral Late mRNA NuclearExport

2007 ◽  
Vol 81 (2) ◽  
pp. 575-587 ◽  
Author(s):  
Jennifer L. Woo ◽  
Arnold J. Berk
Keyword(s):  
Protein Synthesis ◽  
Nuclear Export ◽  
Mrna Export ◽  
Dominant Negative ◽  
Cellular Proteins ◽  
Mutant Form ◽  
Ubiquitin Protein Ligase ◽  
Mrn Complex ◽  
Early Protein ◽  
Cullin 5

ABSTRACT Theadenovirus type 5 (Ad5) E1B-55K and E4orf6 proteins are required together to stimulate viral late nuclear mRNA export to the cytoplasm and to restrict host cell nuclear mRNA export during the late phase of infection. Previous studies have shown that these two viral proteins interact with the cellular proteins elongins B and C, cullin 5, RBX1, and additional cellular proteins to form an E3 ubiquitin-protein ligase that polyubiquitinates p53 and probably one or more subunits of the MRE11-RAD50-NBS1 (MRN) complex, directing their proteasomal degradation. The MRN complex is required for cellular DNA double-strand break repair and induction of the DNA damage response by adenovirus infection. To determine if the ability of E1B-55K and E4orf6 to stimulate viral late mRNA nuclear export requires the ubiquitin-protein ligase activity of this viral ubiquitin-protein ligase complex, we designed and expressed a dominant-negative mutant form of cullin 5 in HeLa cells before infection with wild-type Ad5 or the E1B-55K null mutant dl1520. The dominant-negative cullin 5 protein stabilized p53 and the MRN complex, indicating that it inhibited the viral ubiquitin-protein ligase but had no effect on viral early mRNA synthesis, early protein synthesis, or viral DNA replication. However, expression of the dominant-negative cullin 5 protein caused a decrease in viral late protein synthesis and viral nuclear mRNA export similar to the phenotype produced by mutations in E1B-55K. We conclude that the stimulation of adenovirus late mRNA nuclear export by E1B-55K and E4orf6 results from the ubiquitin-protein ligase activity of the adenovirus ubiquitin-protein ligase complex.

scholarly journals Mutational Definition of Functional Domains within the Rev Homolog Encoded by Human Endogenous Retrovirus K

2000 ◽  
Vol 74 (20) ◽  
pp. 9353-9361 ◽  
Author(s):  
Hal P. Bogerd ◽  
Heather L. Wiegand ◽  
Jin Yang ◽  
Bryan R. Cullen
Keyword(s):  
Nuclear Export ◽  
Biological Activities ◽  
Endogenous Retrovirus ◽  
Viral Rna ◽  
Human Endogenous Retrovirus ◽  
Nuclear Rna ◽  
Rna Export ◽  
Nuclear Rna Export ◽  
Hiv 1 ◽  
Rev Protein

ABSTRACT Nuclear export of the incompletely spliced mRNAs encoded by several complex retroviruses, including human immunodeficiency virus type 1 (HIV-1), is dependent on a virally encoded adapter protein, termed Rev in HIV-1, that directly binds both to a cis-acting viral RNA target site and to the cellular Crm1 export factor. Human endogenous retrovirus K, a family of ancient endogenous retroviruses that is not related to the exogenous retrovirus HIV-1, was recently shown to also encode a Crm1-dependent nuclear RNA export factor, termed K-Rev. Although HIV-1 Rev and K-Rev display little sequence identity, they share the ability not only to bind to Crm1 and to RNA but also to form homomultimers and shuttle between nucleus and cytoplasm. We have used mutational analysis to identify sequences in the 105-amino-acid K-Rev protein required for each of these distinct biological activities. While mutations in K-Rev that inactivate any one of these properties also blocked K-Rev-dependent nuclear RNA export, several K-Rev mutants were comparable to wild type when assayed for any of these individual activities yet nevertheless defective for RNA export. Although several nonfunctional K-Rev mutants acted as dominant negative inhibitors of K-Rev-, but not HIV-1 Rev-, dependent RNA export, these were not defined by their inability to bind to Crm1, as is seen with HIV-1 Rev. In total, this analysis suggests a functional architecture for K-Rev that is similar to, but distinct from, that described for HIV-1 Rev and raises the possibility that viral RNA export mediated by the ∼25 million-year-old K-Rev protein may require an additional cellular cofactor that is not required for HIV-1 Rev function.

scholarly journals Mammalian UPF3A and UPF3B activate NMD independently of their EJC binding

2021 ◽  
Author(s):  
Zhongxia Yi ◽  
René M Arvola ◽  
Sean Myers ◽  
Corinne N Dilsavor ◽  
Rabab Abu Alhasan ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Active Role ◽  
Exon Junction Complex ◽  
Human Cell Lines ◽  
Wild Type ◽  
Human Colorectal Cancer ◽  
Independent Manner ◽  
Exon Junction ◽  
Nonsense Mediated Mrna Decay ◽  
Hct116 Cells

Nonsense-mediated mRNA decay (NMD) is governed by the three conserved factors - UPF1, UPF2 and UPF3. While all three are required for NMD in yeast, UPF3B is dispensable for NMD in mammals, with its paralog UPF3A suggested to only weakly activate or even repress NMD due to its weaker binding to the exon junction complex (EJC). Here we characterize the UPF3B-dependent and -independent NMD in human cell lines knocked-out of one or both UPF3 paralogs. We show that in human colorectal cancer HCT116 cells, EJC-mediated NMD can operate in UPF3B-dependent and -independent manner. While UPF3A is almost completely dispensable for NMD in wild-type cells, it strongly activates EJC-mediated NMD in cells lacking UPF3B. Surprisingly, this major NMD branch can operate in UPF3-independent manner questioning the idea that UPF3 is needed to bridge UPF proteins to the EJC during NMD. Complementation studies in UPF3 knockout cells further show that EJC-binding domain of UPF3 paralogs is not essential for NMD. Instead, the conserved mid domain of UPF3B, previously shown to engage with ribosome release factors, is required for its full NMD activity. Altogether, UPF3 plays a more active role in NMD than simply being a bridge between the EJC and the UPF complex.

scholarly journals The Naturally Occurring Variant of Estrogen Receptor (ER) ERΔE7 Suppresses Estrogen-Dependent Transcriptional Activation by Both Wild-Type ERα and ERβ

Endocrinology ◽  
2003 ◽  
Vol 144 (7) ◽  
pp. 2967-2976 ◽  
Author(s):  
Juana M. García Pedrero ◽  
Pedro Zuazua ◽  
Carlos Martínez-Campa ◽  
Pedro S. Lazo ◽  
Sofía Ramos
Keyword(s):  
Estrogen Receptor ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Activation Function ◽  
Inhibitory Effect ◽  
Dominant Negative ◽  
Wild Type ◽  
Independent Manner ◽  
Naturally Occurring ◽  
Estrogen Response

Abstract We have isolated and functionally characterized the exon 7-skipped variant (ERΔE7) of estrogen receptor (ER)α, which has emerged as the predominant variant expressed in multiple normal and tumoral tissues. However, to date no function has been established for this variant in mammalian cells. ERΔE7 exhibits a negligible ability to bind ligands, insensitivity to allosteric modulation by estrogen and antiestrogens, and loss of estrogen-dependent interaction with p160 coactivators such as SRC-1 and AIB1. ERΔE7 is able to form heterodimers with both ERα and ERβ in a ligand-independent manner. Transient expression experiments in HeLa cells show that increasing amounts of ERΔE7 result in a progressive inhibition of the estrogen-dependent transcriptional activation by both wild-type ERα and ERβ on estrogen response element-driven promoters. The inhibitory effect of ERΔE7 is due to the inhibition of binding of wild-type receptors to their responsive elements. Surprisingly, the activation function (AF)-1-dependent transactivation triggered by epithelial growth factor and phorbol-12-myristate-13-acetate is also abolished in ERΔE7 despite AF1 integrity, suggesting a cross-talk between AF1 and AF2 regions of the receptor. These results indicate that the naturally occurring variant ERΔE7 is a dominant negative receptor that, when expressed at high levels relative to wild-type ERs, might have profound effects on several estrogen-dependent functions.

scholarly journals A Day in the Life of the Exon Junction Complex

Biomolecules ◽  
2020 ◽  
Vol 10 (6) ◽  
pp. 866 ◽  
Author(s):  
Lena P. Schlautmann ◽  
Niels H. Gehring
Keyword(s):  
Mrna Export ◽  
Mrna Splicing ◽  
Exon Junction Complex ◽  
Chronological Order ◽  
Exon Junction ◽  
Messenger Ribonucleoprotein ◽  
Associated Proteins ◽  
Core Components ◽  
Nonsense Mediated Mrna Decay ◽  
Exon Junctions

The exon junction complex (EJC) is an abundant messenger ribonucleoprotein (mRNP) component that is assembled during splicing and binds to mRNAs upstream of exon-exon junctions. EJCs accompany the mRNA during its entire life in the nucleus and the cytoplasm and communicate the information about the splicing process and the position of introns. Specifically, the EJC’s core components and its associated proteins regulate different steps of gene expression, including pre-mRNA splicing, mRNA export, translation, and nonsense-mediated mRNA decay (NMD). This review summarizes the most important functions and main protagonists in the life of the EJC. It also provides an overview of the latest findings on the assembly, composition and molecular activities of the EJC and presents them in the chronological order, in which they play a role in the EJC’s life cycle.

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