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Sample records for 293-epstein-barr virus nuclear

  1. Nuclear entry of DNA viruses

    PubMed Central

    Fay, Nikta; Panté, Nelly

    2015-01-01

    DNA viruses undertake their replication within the cell nucleus, and therefore they must first deliver their genome into the nucleus of their host cells. Thus, trafficking across the nuclear envelope is at the basis of DNA virus infections. Nuclear transport of molecules with diameters up to 39 nm is a tightly regulated process that occurs through the nuclear pore complex (NPC). Due to the enormous diversity of virus size and structure, each virus has developed its own strategy for entering the nucleus of their host cells, with no two strategies alike. For example, baculoviruses target their DNA-containing capsid to the NPC and subsequently enter the nucleus intact, while the hepatitis B virus capsid crosses the NPC but disassembles at the nuclear side of the NPC. For other viruses such as herpes simplex virus and adenovirus, although both dock at the NPC, they have each developed a distinct mechanism for the subsequent delivery of their genome into the nucleus. Remarkably, other DNA viruses, such as parvoviruses and human papillomaviruses, access the nucleus through an NPC-independent mechanism. This review discusses our current understanding of the mechanisms used by DNA viruses to deliver their genome into the nucleus, and further presents the experimental evidence for such mechanisms. PMID:26029198

  2. Viruses Challenge Selectivity Barrier of Nuclear Pores

    PubMed Central

    Labokha, Aksana A.; Fassati, Ariberto

    2013-01-01

    Exchange between the nucleus and the cytoplasm occurs through nuclear pore complexes (NPCs) embedded in the double membrane of the nuclear envelope. NPC permeability barrier restricts the entry of inert molecules larger than 5 nm in diameter but allows facilitated entry of selected cargos, whose size can reach up to 39 nm. The translocation of large molecules is facilitated by nuclear transport receptors (NTRs) that have affinity to proteins of NPC permeability barrier. Viruses that enter the nucleus replicate evolved strategies to overcome this barrier. In this review, we will discuss the functional principles of NPC barrier and nuclear transport machinery, as well as the various strategies viruses use to cross the selective barrier of NPCs. PMID:24084236

  3. Nuclear Proteins Hijacked by Mammalian Cytoplasmic Plus Strand RNA Viruses

    PubMed Central

    Lloyd, Richard E.

    2015-01-01

    Plus strand RNA viruses that replicate in the cytoplasm face challenges in supporting the numerous biosynthetic functions required for replication and propagation. Most of these viruses are genetically simple and rely heavily on co-opting cellular proteins, particularly cellular RNA-binding proteins, into new roles for support of virus infection at the level of virus-specific translation, and building RNA replication complexes. In the course of infectious cycles many nuclear-cytoplasmic shuttling proteins of mostly nuclear distribution are detained in the cytoplasm by viruses and re-purposed for their own gain. Many mammalian viruses hijack a common group of the same factors. This review summarizes recent gains in our knowledge of how cytoplasmic RNA viruses use these co-opted host nuclear factors in new functional roles supporting virus translation and virus RNA replication and common themes employed between different virus groups. PMID:25818028

  4. Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses

    SciTech Connect

    Lloyd, Richard E.

    2015-05-15

    Plus strand RNA viruses that replicate in the cytoplasm face challenges in supporting the numerous biosynthetic functions required for replication and propagation. Most of these viruses are genetically simple and rely heavily on co-opting cellular proteins, particularly cellular RNA-binding proteins, into new roles for support of virus infection at the level of virus-specific translation, and building RNA replication complexes. In the course of infectious cycles many nuclear-cytoplasmic shuttling proteins of mostly nuclear distribution are detained in the cytoplasm by viruses and re-purposed for their own gain. Many mammalian viruses hijack a common group of the same factors. This review summarizes recent gains in our knowledge of how cytoplasmic RNA viruses use these co-opted host nuclear factors in new functional roles supporting virus translation and virus RNA replication and common themes employed between different virus groups. - Highlights: • Nuclear shuttling host proteins are commonly hijacked by RNA viruses to support replication. • A limited group of ubiquitous RNA binding proteins are commonly hijacked by a broad range of viruses. • Key virus proteins alter roles of RNA binding proteins in different stages of virus replication.

  5. Genome Assembly of Citrus Leprosis Virus Nuclear Type Reveals a Close Association with Orchid Fleck Virus

    PubMed Central

    Stone, Andrew; Otero-Colina, Gabriel; Wei, Gang; Choudhary, Nandlal; Achor, Diann; Shao, Jonathan; Levy, Laurene; Nakhla, Mark K.; Hollingsworth, Charla R.; Hartung, John S.; Schneider, William L.

    2013-01-01

    The complete genome of citrus leprosis virus nuclear type (CiLV-N) was identified by small RNA sequencing utilizing leprosis-affected citrus samples collected from the state of Querétaro, Mexico. The nucleotide identity and phylogenetic analysis indicate that CiLV-N is very closely related to orchid fleck virus, which typically infects Cymbidium species. PMID:23887919

  6. Influenza virus mRNA trafficking through host nuclear speckles.

    PubMed

    Mor, Amir; White, Alexander; Zhang, Ke; Thompson, Matthew; Esparza, Matthew; Muñoz-Moreno, Raquel; Koide, Kazunori; Lynch, Kristen W; García-Sastre, Adolfo; Fontoura, Beatriz M A

    2016-01-01

    Influenza A virus is a human pathogen with a genome composed of eight viral RNA segments that replicate in the nucleus. Two viral mRNAs are alternatively spliced. The unspliced M1 mRNA is translated into the matrix M1 protein, while the ion channel M2 protein is generated after alternative splicing. These proteins are critical mediators of viral trafficking and budding. We show that the influenza virus uses nuclear speckles to promote post-transcriptional splicing of its M1 mRNA. We assign previously unknown roles for the viral NS1 protein and cellular factors to an intranuclear trafficking pathway that targets the viral M1 mRNA to nuclear speckles, mediates splicing at these nuclear bodies and exports the spliced M2 mRNA from the nucleus. Given that nuclear speckles are storage sites for splicing factors, which leave these sites to splice cellular pre-mRNAs at transcribing genes, we reveal a functional subversion of nuclear speckles to promote viral gene expression. PMID:27572970

  7. Influenza virus mRNA trafficking through host nuclear speckles.

    PubMed

    Mor, Amir; White, Alexander; Zhang, Ke; Thompson, Matthew; Esparza, Matthew; Muñoz-Moreno, Raquel; Koide, Kazunori; Lynch, Kristen W; García-Sastre, Adolfo; Fontoura, Beatriz M A

    2016-01-01

    Influenza A virus is a human pathogen with a genome composed of eight viral RNA segments that replicate in the nucleus. Two viral mRNAs are alternatively spliced. The unspliced M1 mRNA is translated into the matrix M1 protein, while the ion channel M2 protein is generated after alternative splicing. These proteins are critical mediators of viral trafficking and budding. We show that the influenza virus uses nuclear speckles to promote post-transcriptional splicing of its M1 mRNA. We assign previously unknown roles for the viral NS1 protein and cellular factors to an intranuclear trafficking pathway that targets the viral M1 mRNA to nuclear speckles, mediates splicing at these nuclear bodies and exports the spliced M2 mRNA from the nucleus. Given that nuclear speckles are storage sites for splicing factors, which leave these sites to splice cellular pre-mRNAs at transcribing genes, we reveal a functional subversion of nuclear speckles to promote viral gene expression.

  8. A nuclear export signal in the matrix protein of Influenza A virus is required for efficient virus replication.

    PubMed

    Cao, Shuai; Liu, Xiaoling; Yu, Maorong; Li, Jing; Jia, Xiaojuan; Bi, Yuhai; Sun, Lei; Gao, George F; Liu, Wenjun

    2012-05-01

    The influenza A virus matrix 1 protein (M1) shuttles between the cytoplasm and the nucleus during the viral life cycle and plays an important role in the replication, assembly, and budding of viruses. Here, a leucine-rich nuclear export signal (NES) was identified specifically for the nuclear export of the M1 protein. The predicted NES, designated the Flu-A-M1 NES, is highly conserved among all sequences from the influenza A virus subtype, but no similar NES motifs are found in the M1 sequences of influenza B or C viruses. The biological function of the Flu-A-M1 NES was demonstrated by its ability to translocate an enhanced green fluorescent protein (EGFP)-NES fusion protein from the nucleus to the cytoplasm in transfected cells, compared to the even nuclear and cytoplasmic distribution of EGFP. The translocation of EGFP-NES from the nucleus to the cytoplasm was not inhibited by leptomycin B. NES mutations in M1 caused a nuclear retention of the protein and an increased nuclear accumulation of NEP during transfection. Indeed, as shown by rescued recombinant viruses, the mutation of the NES impaired the nuclear export of M1 and significantly reduced the virus titer compared to titers of wild-type viruses. The NES-defective M1 protein was retained in the nucleus during infection, accompanied by a lowered efficiency of the nuclear export of viral RNPs (vRNPs). In conclusion, M1 nuclear export was specifically dependent on the Flu-A-M1 NES and critical for influenza A virus replication.

  9. Inactivation of nuclear polyhedrosis virus (Baculovirus subgroup A) by monochromatic UV radiation

    SciTech Connect

    Griego, V.M.; Martignoni, M.E.; Claycomb, A.E.

    1985-03-01

    Monochromatic radiation at wavelengths of 290, 300, 310, and 320 nm inactivated occluded nuclear polyhedrosis virus of the Douglas-fir tussock moth, Orgyia pseudotsugata. Data indicate that all of the wavelengths are capable of causing virus inactivation; much greater fluences are needed for virus inactivation as the wavelength increases.

  10. Nuclear trafficking of proteins from RNA viruses: potential target for antivirals?

    PubMed

    Caly, Leon; Wagstaff, Kylie M; Jans, David A

    2012-09-01

    A key aspect of the infectious cycle of many viruses is the transport of specific viral proteins into the host cell nucleus to perturb the antiviral response. Examples include a number of RNA viruses that are significant human pathogens, such as human immunodeficiency virus (HIV)-1, influenza A, dengue, respiratory syncytial virus and rabies, as well agents that predominantly infect livestock, such as Rift valley fever virus and Venezuelan equine encephalitis virus. Inhibiting the nuclear trafficking of viral proteins as a therapeutic strategy offers an attractive possibility, with important recent progress having been made with respect to HIV-1 and dengue. The results validate nuclear protein import as an antiviral target, and suggest the identification and development of nuclear transport inhibitors as a viable therapeutic approach for a range of human and zoonotic pathogenic viruses.

  11. Misdelivery at the Nuclear Pore Complex—Stopping a Virus Dead in Its Tracks

    PubMed Central

    Flatt, Justin W.; Greber, Urs F.

    2015-01-01

    Many viruses deliver their genomes into the host cell’s nucleus before they replicate. While onco-retroviruses and papillomaviruses tether their genomes to host chromatin upon mitotic breakdown of the nuclear envelope, lentiviruses, such as human immunodeficiency virus, adenoviruses, herpesviruses, parvoviruses, influenza viruses, hepatitis B virus, polyomaviruses, and baculoviruses deliver their genomes into the nucleus of post-mitotic cells. This poses the significant challenge of slipping a DNA or RNA genome past the nuclear pore complex (NPC) embedded in the nuclear envelope. Quantitative fluorescence imaging is shedding new light on this process, with recent data implicating misdelivery of viral genomes at nuclear pores as a bottleneck to virus replication. Here, we infer NPC functions for nuclear import of viral genomes from cell biology experiments and explore potential causes of misdelivery, including improper virus docking at NPCs, incomplete translocation, virus-induced stress and innate immunity reactions. We conclude by discussing consequences of viral genome misdelivery for viruses and host cells, and lay out future questions to enhance our understanding of this phenomenon. Further studies into viral genome misdelivery may reveal unexpected aspects about NPC structure and function, as well as aid in developing strategies for controlling viral infections to improve human health. PMID:26226003

  12. Roles of hepatocyte nuclear factors in hepatitis B virus infection.

    PubMed

    Kim, Doo Hyun; Kang, Hong Seok; Kim, Kyun-Hwan

    2016-08-21

    Approximately 350 million people are estimated to be persistently infected with hepatitis B virus (HBV) worldwide. HBV maintains persistent infection by employing covalently closed circular DNA (cccDNA), a template for all HBV RNAs. Chronic hepatitis B (CHB) patients are currently treated with nucleos(t)ide analogs such as lamivudine, adefovir, entecavir, and tenofovir. However, these treatments rarely cure CHB because they are unable to inhibit cccDNA transcription and inhibit only a late stage in the HBV life cycle (the reverse transcription step in the nucleocapsid). Therefore, an understanding of the factors regulating cccDNA transcription is required to stop this process. Among numerous factors, hepatocyte nuclear factors (HNFs) play the most important roles in cccDNA transcription, especially in the generation of viral genomic RNA, a template for HBV replication. Therefore, proper control of HNF function could lead to the inhibition of HBV replication. In this review, we summarize and discuss the current understanding of the roles of HNFs in the HBV life cycle and the upstream factors that regulate HNFs. This knowledge will enable the identification of new therapeutic targets to cure CHB. PMID:27610013

  13. Roles of hepatocyte nuclear factors in hepatitis B virus infection

    PubMed Central

    Kim, Doo Hyun; Kang, Hong Seok; Kim, Kyun-Hwan

    2016-01-01

    Approximately 350 million people are estimated to be persistently infected with hepatitis B virus (HBV) worldwide. HBV maintains persistent infection by employing covalently closed circular DNA (cccDNA), a template for all HBV RNAs. Chronic hepatitis B (CHB) patients are currently treated with nucleos(t)ide analogs such as lamivudine, adefovir, entecavir, and tenofovir. However, these treatments rarely cure CHB because they are unable to inhibit cccDNA transcription and inhibit only a late stage in the HBV life cycle (the reverse transcription step in the nucleocapsid). Therefore, an understanding of the factors regulating cccDNA transcription is required to stop this process. Among numerous factors, hepatocyte nuclear factors (HNFs) play the most important roles in cccDNA transcription, especially in the generation of viral genomic RNA, a template for HBV replication. Therefore, proper control of HNF function could lead to the inhibition of HBV replication. In this review, we summarize and discuss the current understanding of the roles of HNFs in the HBV life cycle and the upstream factors that regulate HNFs. This knowledge will enable the identification of new therapeutic targets to cure CHB. PMID:27610013

  14. Roles of hepatocyte nuclear factors in hepatitis B virus infection

    PubMed Central

    Kim, Doo Hyun; Kang, Hong Seok; Kim, Kyun-Hwan

    2016-01-01

    Approximately 350 million people are estimated to be persistently infected with hepatitis B virus (HBV) worldwide. HBV maintains persistent infection by employing covalently closed circular DNA (cccDNA), a template for all HBV RNAs. Chronic hepatitis B (CHB) patients are currently treated with nucleos(t)ide analogs such as lamivudine, adefovir, entecavir, and tenofovir. However, these treatments rarely cure CHB because they are unable to inhibit cccDNA transcription and inhibit only a late stage in the HBV life cycle (the reverse transcription step in the nucleocapsid). Therefore, an understanding of the factors regulating cccDNA transcription is required to stop this process. Among numerous factors, hepatocyte nuclear factors (HNFs) play the most important roles in cccDNA transcription, especially in the generation of viral genomic RNA, a template for HBV replication. Therefore, proper control of HNF function could lead to the inhibition of HBV replication. In this review, we summarize and discuss the current understanding of the roles of HNFs in the HBV life cycle and the upstream factors that regulate HNFs. This knowledge will enable the identification of new therapeutic targets to cure CHB.

  15. Mutations within the nuclear localization signal of the porcine reproductive and respiratory syndrome virus nucleocapsid protein attenuate virus replication

    SciTech Connect

    Lee, Changhee; Hodgins, Douglas; Calvert, Jay G.; Welch, Siao-Kun W.; Jolie, Rika; Yoo, Dongwan . E-mail: dyoo@uoguelph.ca

    2006-03-01

    Porcine reproductive and respiratory syndrome virus (PRRSV) is an RNA virus replicating in the cytoplasm, but the nucleocapsid (N) protein is specifically localized to the nucleus and nucleolus in virus-infected cells. A 'pat7' motif of 41-PGKK(N/S)KK has previously been identified in the N protein as the functional nuclear localization signal (NLS); however, the biological consequences of N protein nuclear localization are unknown. In the present study, the role of N protein nuclear localization during infection was investigated in pigs using an NLS-null mutant virus. When two lysines at 43 and 44 at the NLS locus were substituted to glycines, the modified NLS with 41-PGGGNKK restricted the N protein to the cytoplasm. This NLS-null mutation was introduced into a full-length infectious cDNA clone of PRRSV. Upon transfection of cells, the NLS-null full-length clone induced cytopathic effects and produced infectious progeny. The NLS-null virus grew to a titer 100-fold lower than that of wild-type virus. To examine the response to NLS-null PRRSV in the natural host, three groups of pigs, consisting of seven animals per group, were intranasally inoculated with wild-type, placebo, or NLS-null virus, and the animals were maintained for 4 weeks. The NLS-null-infected pigs had a significantly shorter mean duration of viremia than wild-type-infected pigs but developed significantly higher titers of neutralizing antibodies. Mutations occurred at the NLS locus in one pig during viremia, and four types of mutations were identified: 41-PGRGNKK, 41-PGGRNKK, and 41-PGRRNKK, and 41-PGKKSKK. Both wild-type and NLS-null viruses persisted in the tonsils for at least 4 weeks, and the NLS-null virus persisting in the tonsils was found to be mutated to either 41-PGRGNKK or 41-PGGRNKK in all pigs. No other mutation was found in the N gene. All types of reversions which occurred during viremia and persistence were able to translocate the mutated N proteins to the nucleus, indicating a

  16. Changes in Nuclear Basic Proteins During Pseudorabies Virus Infection

    PubMed Central

    Stevens, J. G.; Kado-Boll, G. J.; Haven, C. B.

    1969-01-01

    As a preliminary study to investigation of the possible role played by basic proteins in the genetic regulation of virus-infected cells, acid-extractable proteins synthesized during pseudorabies virus infection were investigated. The synthesis of histones was found to decrease in a gradual manner, and arrest was complete by 6 hr after infection. Five virus-induced acid-extractable proteins appeared in nuclei of infected cells after 4 hr of infection. Four of these proteins were virus structural proteins; one was not. All these proteins contained tryptophan and, therefore, were not “classic” histones. PMID:5786178

  17. Nuclear envelope breakdown induced by herpes simplex virus type 1 involves the activity of viral fusion proteins.

    PubMed

    Maric, Martina; Haugo, Alison C; Dauer, William; Johnson, David; Roller, Richard J

    2014-07-01

    Herpesvirus infection reorganizes components of the nuclear lamina usually without loss of integrity of the nuclear membranes. We report that wild-type HSV infection can cause dissolution of the nuclear envelope in transformed mouse embryonic fibroblasts that do not express torsinA. Nuclear envelope breakdown is accompanied by an eight-fold inhibition of virus replication. Breakdown of the membrane is much more limited during infection with viruses that lack the gB and gH genes, suggesting that breakdown involves factors that promote fusion at the nuclear membrane. Nuclear envelope breakdown is also inhibited during infection with virus that does not express UL34, but is enhanced when the US3 gene is deleted, suggesting that envelope breakdown may be enhanced by nuclear lamina disruption. Nuclear envelope breakdown cannot compensate for deletion of the UL34 gene suggesting that mixing of nuclear and cytoplasmic contents is insufficient to bypass loss of the normal nuclear egress pathway.

  18. Sorting of influenza A virus RNA genome segments after nuclear export

    SciTech Connect

    Takizawa, Naoki; Kumakura, Michiko; Takeuchi, Kaoru; Kobayashi, Nobuyuki; Nagata, Kyosuke

    2010-06-05

    The genome of the influenza A virus consists of eight different segments. These eight segments are thought to be sorted selectively in infected cells. However, the cellular compartment where segments are sorted is not known. We examined using temperature sensitive (ts) mutant viruses and cell fusion where segments are sorted in infected cells. Different cells were infected with different ts mutant viruses, and these cells were fused. In fused cells, genome segments are mixed only in the cytoplasm, because M1 prevents their re-import into the nucleus. We made a marker ts53 virus, which has silent mutations in given segments and determined the reassortment frequency on all segments using ts1 and marker ts53. In both co-infected and fused cells, all of marker ts53 segments and ts1 segments were incorporated into progeny virions in a random fashion. These results suggest that influenza virus genome segments are sorted after nuclear export.

  19. 40 CFR 180.1149 - Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... polyhedrosis virus of Anagrapha falcifera; exemption from the requirement of a tolerance. 180.1149 Section 180... Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from the... polyhedrosis virus of Anagrapha falcifera is exempted from the requirement of a tolerance in or on all...

  20. 40 CFR 180.1149 - Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... polyhedrosis virus of Anagrapha falcifera; exemption from the requirement of a tolerance. 180.1149 Section 180... Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from the... polyhedrosis virus of Anagrapha falcifera is exempted from the requirement of a tolerance in or on all...

  1. 40 CFR 180.1149 - Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... polyhedrosis virus of Anagrapha falcifera; exemption from the requirement of a tolerance. 180.1149 Section 180... Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from the... polyhedrosis virus of Anagrapha falcifera is exempted from the requirement of a tolerance in or on all...

  2. 40 CFR 180.1149 - Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... polyhedrosis virus of Anagrapha falcifera; exemption from the requirement of a tolerance. 180.1149 Section 180... Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from the... polyhedrosis virus of Anagrapha falcifera is exempted from the requirement of a tolerance in or on all...

  3. 40 CFR 180.1149 - Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... polyhedrosis virus of Anagrapha falcifera; exemption from the requirement of a tolerance. 180.1149 Section 180... Inclusion bodies of the multi-nuclear polyhedrosis virus of Anagrapha falcifera; exemption from the... polyhedrosis virus of Anagrapha falcifera is exempted from the requirement of a tolerance in or on all...

  4. Herpes simplex virus type 1 protein IE63 affects the nuclear export of virus intron-containing transcripts.

    PubMed Central

    Phelan, A; Dunlop, J; Clements, J B

    1996-01-01

    Using in situ hybridization labelling methods, we have determined that the herpes simplex virus type 1 immediate-early protein IE63 (ICP27) affects the cellular localization of virus transcripts. Intronless transcripts from the IE63, UL38, and UL44 genes are rapidly exported to and accumulate in the cytoplasm throughout infection, in either the presence or absence of IE63 expression. The intron-containing transcripts from the IE110 and UL15 genes, while initially cytoplasmic, are increasingly retained in the nucleus in distinct clumps as infection proceeds, and the clumps colocalize with the redistributed small nuclear ribonucleoprotein particles. Infections with the IE63 mutant virus 27-lacZ demonstrated that in the absence of IE63 expression, nuclear retention of intron-containing transcripts was lost. The nuclear retention of UL15 transcripts, which demonstrated both nuclear and cytoplasmic label, was not as pronounced as that of the IE110 transcripts, and we propose that this is due to the late expression of UL15. Infections with the mutant virus 110C1, in which both introns of IE110 have been precisely removed (R.D. Everett, J. Gen. Virol. 72:651-659, 1991), demonstrated IE110 transcripts in both the nucleus and the cytoplasm; thus, exon definition sequences which regulate viral RNA transport are present in the IE110 transcript. By in situ hybridization a stable population of polyadenylated RNAs was found to accumulate in the nucleus in spots, most of which were separate from the small nuclear ribonucleoprotein particle clumps. The IE63 protein has an involvement, either direct or indirect, in the regulation of nucleocytoplasmic transport of viral transcripts, a function which contrasts with the recently proposed role of herpes simplex virus type 1 Us11 in promoting the nuclear export of partially spliced or unspliced transcripts (J.-J. Diaz, M. Duc Dodon, N. Schaerer-Uthurraly, D. Simonin, K. Kindbeiter, L. Gazzolo, and J.-J. Madjar, Nature [London] 379

  5. Small tumor virus genomes are integrated near nuclear matrix attachment regions in transformed cells.

    PubMed

    Shera, K A; Shera, C A; McDougall, J K

    2001-12-01

    More than 15% of human cancers have a viral etiology. In benign lesions induced by the small DNA tumor viruses, viral genomes are typically maintained extrachromosomally. Malignant progression is often associated with viral integration into host cell chromatin. To study the role of viral integration in tumorigenesis, we analyzed the positions of integrated viral genomes in tumors and tumor cell lines induced by the small oncogenic viruses, including the high-risk human papillomaviruses, hepatitis B virus, simian virus 40, and human T-cell leukemia virus type 1. We show that viral integrations in tumor cells lie near cellular sequences identified as nuclear matrix attachment regions (MARs), while integrations in nonneoplastic cells show no significant correlation with these regions. In mammalian cells, the nuclear matrix functions in gene expression and DNA replication. MARs play varied but poorly understood roles in eukaryotic gene expression. Our results suggest that integrated tumor virus genomes are subject to MAR-mediated transcriptional regulation, providing insight into mechanisms of viral carcinogenesis. Furthermore, the viral oncoproteins serve as invaluable tools for the study of mechanisms controlling cellular growth. Similarly, our demonstration that integrated viral genomes may be subject to MAR-mediated transcriptional effects should facilitate elucidation of fundamental mechanisms regulating eukaryotic gene expression.

  6. First report of citrus leprosis virus nuclear type in Colombia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Citrus leprosis is a difficult viral disease causing significant damage to citrus fruit in South America and Central America. The disease is marked by dramatic lesions on fruit, leaves and stems resulting in unmarketable product. Citrus leprosis virus cytoplasmic types (CiLV-C and CiLV-C2) wer edete...

  7. The R35 residue of the influenza A virus NS1 protein has minimal effects on nuclear localization but alters virus replication through disrupting protein dimerization

    SciTech Connect

    Lalime, Erin N.; Pekosz, Andrew

    2014-06-15

    The influenza A virus NS1 protein has a nuclear localization sequence (NLS) in the amino terminal region. This NLS overlaps sequences that are important for RNA binding as well as protein dimerization. To assess the significance of the NS1 NLS on influenza virus replication, the NLS amino acids were individually mutated to alanines and recombinant viruses encoding these mutations were rescued. Viruses containing NS1 proteins with mutations at R37, R38 and K41 displayed minimal changes in replication or NS1 protein nuclear localization. Recombinant viruses encoding NS1 R35A were not recovered but viruses containing second site mutations at position D39 in addition to the R35A mutation were isolated. The mutations at position 39 were shown to partially restore NS1 protein dimerization but had minimal effects on nuclear localization. These data indicate that the amino acids in the NS1 NLS region play a more important role in protein dimerization compared to nuclear localization. - Highlights: • Mutations were introduced into influenza NS1 NLS1. • NS1 R37A, R38A, K41A viruses had minimal changes in replication and NS1 localization. • Viruses from NS1 R35A rescue all contained additional mutations at D39. • NS1 R35A D39X mutations recover dimerization lost in NS1 R35A mutations. • These results reaffirm the importance of dimerization for NS1 protein function.

  8. Photoreactivation and ultraviolet-enhanced reactivation of ultraviolet-irradiated nuclear polyhedrosis virus by insect cells.

    PubMed

    Witt, D J

    1984-01-01

    The nuclear polyhedrosis virus (Baculovirus) of Galleria mellonella (Pryalidae: Lepidoptera) was used to investigate the capability of cultured insect cells to repair ultraviolet (UV) induced damage in the viral genome. When assayed by the formation of plaques in the cell line TN-368, the survival of the virus was found to decrease linearly with increased ultraviolet exposure. The infectious capacity of UV-irradiated virions was significantly restored after exposing the TN-368 monolayers to either photoreactivation conditions (white fluorescent and black light) or to UV-enhanced reactivation conditions (far ultraviolet radiation). Using both types of repair sequentially resulted in higher reactivation than when either was used alone. These results indicate that pyrimidine dimers are the major factor responsible for inactivation of this virus by UV radiation but that other photolesions not repairable by photoreactivation partially account for the inactivation of the virus. PMID:6365037

  9. Nuclear localization of dengue virus nonstructural protein 5 through its importin alpha/beta-recognized nuclear localization sequences is integral to viral infection.

    PubMed

    Pryor, Melinda J; Rawlinson, Stephen M; Butcher, Rebecca E; Barton, Chenoa L; Waterhouse, Tracey A; Vasudevan, Subhash G; Bardin, Phillip G; Wright, Peter J; Jans, David A; Davidson, Andrew D

    2007-07-01

    Dengue virus nonstructural protein 5 (NS5) is a large multifunctional protein with a central role in viral replication. We previously identified two nuclear localization sequences (NLSs) within the central region of dengue virus type-2 (DENV-2) NS5 ('aNLS' and 'bNLS') that are recognized by the importin alpha/beta and importin beta1 nuclear transporters, respectively. Here, we demonstrate the importance of the kinetics of NS5 nuclear localization to virus production for the first time and show that the aNLS is responsible. Site-specific mutations in the bipartite-type aNLS or bNLS region were introduced into a reporter plasmid encoding green fluorescent protein fused to the N-terminus of DENV-2 NS5, as well as into DENV-2 genomic length complementary DNA. Mutation of basic residues in the highly conserved region of the bNLS did not affect nuclear import of NS5. In contrast, mutations in either basic cluster of the aNLS decreased NS5 nuclear accumulation and reduced virus production, with the greatest reduction observed for mutation of the second cluster (K(387)K(388)K(389)); mutagenesis of both clusters abolished NS5 nuclear import and DENV-2 virus production completely. The latter appeared to relate to the impaired ability of virus lacking nuclear-localizing NS5, as compared with wild-type virus expressing nuclear-localizing NS5, to reduce interleukin-8 production as part of the antiviral response. The results overall indicate that NS5 nuclear localization through the aNLS is integral to viral infection, with significant implications for other flaviviruses of medical importance, such as yellow fever and West Nile viruses.

  10. Characterization of a nuclear localization signal in the foot-and-mouth disease virus polymerase

    SciTech Connect

    Sanchez-Aparicio, Maria Teresa; Rosas, Maria Flora; Sobrino, Francisco

    2013-09-15

    We have experimentally tested whether the MRKTKLAPT sequence in FMDV 3D protein (residues 16 to 24) can act as a nuclear localization signal (NLS). Mutants with substitutions in two basic residues within this sequence, K18E and K20E, were generated. A decreased nuclear localization was observed in transiently expressed 3D and its precursor 3CD, suggesting a role of K18 and K20 in nuclear targeting. Fusion of MRKTKLAPT to the green fluorescence protein (GFP) increased the nuclear localization of GFP, which was not observed when GFP was fused to the 3D mutated sequences. These results indicate that the sequence MRKTKLAPT can be functionally considered as a NLS. When introduced in a FMDV full length RNA replacements K18E and K20E led to production of revertant viruses that replaced the acidic residues introduced (E) by K, suggesting that the presence of lysins at positions 18 and 20 of 3D is essential for virus multiplication. - Highlights: • The FMDV 3D polymerase contains a nuclear localization signal. • Replacements K18E and K20E decrease nuclear localization of 3D and its precursor 3CD. • Fusion of the MRKTKLAPT 3D motif to GFP increases the nuclear localization of GFP. • Replacements K18E and K20E abolish the ability of MRKTKLAPT to relocate GFP. • RNAs harboring replacements K18E and K20E lead to recovery of revertant FMDVs.

  11. Phosphorylation Controls the Nuclear-Cytoplasmic Shuttling of Influenza A Virus Nucleoprotein

    PubMed Central

    Zheng, Weinan; Li, Jing; Wang, Shanshan; Cao, Shuaishuai; Jiang, Jingwen; Chen, Can; Ding, Chan; Qin, Chuan; Ye, Xin; Gao, George F.

    2015-01-01

    ABSTRACT The nucleoprotein (NP) is a major component of the viral ribonucleoprotein (vRNP) complex. During the replication of influenza virus, the vRNP complex undergoes nuclear-cytoplasmic shuttling, during which NP serves as one of the determinants. To date, many phosphorylation sites on NP have been identified, but the biological functions of many of these phosphorylation sites remain unknown. In the present study, the functions of the phosphorylation sites S9, Y10, and Y296 were characterized. These residues are highly conserved, and their phosphorylation was essential for virus growth in cell culture and in a mouse model by regulating the activity of the viral polymerase and the nuclear-cytoplasmic shuttling of NP. The phosphorylation and dephosphorylation of S9 and Y10 controlled nuclear import of NP by affecting the binding affinity between NP and different isoforms of importin-α. In addition, the phosphorylation of Y296 caused nuclear retention of NP by reducing the interaction between NP and CRM1. Furthermore, tyrosine phosphorylation of NP during the early stage of virus infection was ablated when Y296 was mutated to F. However, at later stages of infection, it was weakened by the Y10F mutation. Taken together, the present data indicate that the phosphorylation and dephosphorylation of NP control the shuttling of NP between the nucleus and the cytoplasm during virus replication. IMPORTANCE It is well known that phosphorylation regulates the functions of viral proteins and the life cycle of influenza A virus. As NP is the most abundant protein in the vRNP complex of influenza A virus, several phosphorylation sites on this protein have been identified. However, the functions of these phosphorylation sites were unknown. The present study demonstrates that the phosphorylation status of these sites on NP can mediate its nuclear-cytoplasmic shuttling, which drives the trafficking of vRNP complexes in infected cells. The present data suggest that the

  12. Widespread horizontal gene transfer from double-stranded RNA viruses to eukaryotic nuclear genomes.

    PubMed

    Liu, Huiquan; Fu, Yanping; Jiang, Daohong; Li, Guoqing; Xie, Jiatao; Cheng, Jiasen; Peng, Youliang; Ghabrial, Said A; Yi, Xianhong

    2010-11-01

    Horizontal gene transfer commonly occurs from cells to viruses but rarely occurs from viruses to their host cells, with the exception of retroviruses and some DNA viruses. However, extensive sequence similarity searches in public genome databases for various organisms showed that the capsid protein and RNA-dependent RNA polymerase genes from totiviruses and partitiviruses have widespread homologs in the nuclear genomes of eukaryotic organisms, including plants, arthropods, fungi, nematodes, and protozoa. PCR amplification and sequencing as well as comparative evidence of junction coverage between virus and host sequences support the conclusion that these viral homologs are real and occur in eukaryotic genomes. Sequence comparison and phylogenetic analysis suggest that these genes were likely transferred horizontally from viruses to eukaryotic genomes. Furthermore, we present evidence showing that some of the transferred genes are conserved and expressed in eukaryotic organisms and suggesting that these viral genes are also functional in the recipient genomes. Our findings imply that horizontal transfer of double-stranded RNA viral genes is widespread among eukaryotes and may give rise to functionally important new genes, thus entailing that RNA viruses may play significant roles in the evolution of eukaryotes.

  13. Genome assembly of citurs leprosis virus nuclear type reveals a close association with orchid fleck virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Citrus leprosis is a difficult viral disease causing significant damage to citrus fruit in South America and Central America. The disease is marked by dramatic lesions on fruit, leaves and stems resulting in unmarketable product. Citrus leprosis virus cytoplasmic type (CiLV-C) was detected in states...

  14. Nuclear envelope breakdown induced by herpes simplex virus type 1 involves the activity of viral fusion proteins

    SciTech Connect

    Maric, Martina; Haugo, Alison C.; Dauer, William; Johnson, David; Roller, Richard J.

    2014-07-15

    Herpesvirus infection reorganizes components of the nuclear lamina usually without loss of integrity of the nuclear membranes. We report that wild-type HSV infection can cause dissolution of the nuclear envelope in transformed mouse embryonic fibroblasts that do not express torsinA. Nuclear envelope breakdown is accompanied by an eight-fold inhibition of virus replication. Breakdown of the membrane is much more limited during infection with viruses that lack the gB and gH genes, suggesting that breakdown involves factors that promote fusion at the nuclear membrane. Nuclear envelope breakdown is also inhibited during infection with virus that does not express UL34, but is enhanced when the US3 gene is deleted, suggesting that envelope breakdown may be enhanced by nuclear lamina disruption. Nuclear envelope breakdown cannot compensate for deletion of the UL34 gene suggesting that mixing of nuclear and cytoplasmic contents is insufficient to bypass loss of the normal nuclear egress pathway. - Highlights: • We show that wild-type HSV can induce breakdown of the nuclear envelope in a specific cell system. • The viral fusion proteins gB and gH are required for induction of nuclear envelope breakdown. • Nuclear envelope breakdown cannot compensate for deletion of the HSV UL34 gene.

  15. Non-occluded particles of nuclear polyhedrosis virus infecting Galleria mellonella L.: titration in vivo and in vitro. Brief report.

    PubMed

    Komissarenko, S V; Zherebtsova, E N; Sutugina, L P; Primatchenko, L V

    1979-01-01

    Infectivity of non-occluded particles of nuclear polyhedrosis virus infecting Galleria mellonella L. was determined with bioassay technique in vivo and with plaque assay method in vitro using SCLd 135 (Quiot) established cell culture. A plaque-forming virus unit corresponds to about 50 ID50 for Galleria larvae injected into hemocoel for non-cloned and 0.1-1.0 ID50 for a virus isolate cloned from an infective plaque.

  16. The conserved domain CR2 of Epstein-Barr virus nuclear antigen leader protein is responsible not only for nuclear matrix association but also for nuclear localization.

    PubMed

    Yokoyama, A; Kawaguchi, Y; Kitabayashi, I; Ohki, M; Hirai, K

    2001-01-20

    There is a growing body of evidence for the importance of the nuclear matrix in various nuclear events including gene expression and DNA replication. Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is a nuclear matrix-associated protein that has been suggested to play an important role in EBV-induced transformation. To define the biological significance of the association of EBNA-LP with the nuclear matrix, we mapped the domain of EBNA-LP responsible for nuclear matrix association and investigated the functions of the EBNA-LP mutant mutagenized by substitution of alanines for the cluster of arginine residues in the mapped region. The results of the present study were as follows. (i) Transiently expressed EBNA-LP in COS-7 or BOSC23 cells was associated with the nuclear matrix, similarly to that in EBV-infected B cells. (ii) Mutational analysis of EBNA-LP revealed that a 10-amino acid segment of EBNA-LP is critical for nuclear matrix association of the protein. Interestingly, the identified region overlapped with the region CR2 of EBNA-LP conserved among a subset of primate gammaherpesviruses. The identified segment is referred to as EBNA-LP NMTS (nuclear matrix targeting signal). (iii) The EBNA-LP mutant with the arginine to alanine substitutions in NMTS was no longer localized not only to the nuclear matrix but also to the nucleus. (iv) The EBNA-LP mutant lacked its ability to coactivate EBNA-2-dependent transactivation. These results indicated that EBNA-LP needs to be localized in the nucleus and/or associated with the nuclear matrix through CR2 to elicit its function such as the coactivation of the EBNA-2-dependent transcriptional activation. PMID:11162796

  17. 40 CFR 180.1118 - Spodoptera exigua nuclear polyhedrosis virus; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... virus; exemption from the requirement of a tolerance. 180.1118 Section 180.1118 Protection of... polyhedrosis virus; exemption from the requirement of a tolerance. An exemption from the requirement of a tolerance is established for the microbial pest control agent Spodoptera exigua nuclear polyhedrosis...

  18. 40 CFR 180.1118 - Spodoptera exigua nuclear polyhedrosis virus; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... virus; exemption from the requirement of a tolerance. 180.1118 Section 180.1118 Protection of... polyhedrosis virus; exemption from the requirement of a tolerance. An exemption from the requirement of a tolerance is established for the microbial pest control agent Spodoptera exigua nuclear polyhedrosis...

  19. 40 CFR 180.1118 - Spodoptera exigua nuclear polyhedrosis virus; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... virus; exemption from the requirement of a tolerance. 180.1118 Section 180.1118 Protection of... polyhedrosis virus; exemption from the requirement of a tolerance. An exemption from the requirement of a tolerance is established for the microbial pest control agent Spodoptera exigua nuclear polyhedrosis...

  20. 40 CFR 180.1118 - Spodoptera exigua nuclear polyhedrosis virus; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... virus; exemption from the requirement of a tolerance. 180.1118 Section 180.1118 Protection of... polyhedrosis virus; exemption from the requirement of a tolerance. An exemption from the requirement of a tolerance is established for the microbial pest control agent Spodoptera exigua nuclear polyhedrosis...

  1. 40 CFR 180.1118 - Spodoptera exigua nuclear polyhedrosis virus; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... virus; exemption from the requirement of a tolerance. 180.1118 Section 180.1118 Protection of... polyhedrosis virus; exemption from the requirement of a tolerance. An exemption from the requirement of a tolerance is established for the microbial pest control agent Spodoptera exigua nuclear polyhedrosis...

  2. Alterations of Nuclear Architecture and Epigenetic Signatures during African Swine Fever Virus Infection

    PubMed Central

    Simões, Margarida; Rino, José; Pinheiro, Inês; Martins, Carlos; Ferreira, Fernando

    2015-01-01

    Viral interactions with host nucleus have been thoroughly studied, clarifying molecular mechanisms and providing new antiviral targets. Considering that African swine fever virus (ASFV) intranuclear phase of infection is poorly understood, viral interplay with subnuclear domains and chromatin architecture were addressed. Nuclear speckles, Cajal bodies, and promyelocytic leukaemia nuclear bodies (PML-NBs) were evaluated by immunofluorescence microscopy and Western blot. Further, efficient PML protein knockdown by shRNA lentiviral transduction was used to determine PML-NBs relevance during infection. Nuclear distribution of different histone H3 methylation marks at lysine’s 9, 27 and 36, heterochromatin protein 1 isoforms (HP1α, HPβ and HPγ) and several histone deacetylases (HDACs) were also evaluated to assess chromatin status of the host. Our results reveal morphological disruption of all studied subnuclear domains and severe reduction of viral progeny in PML-knockdown cells. ASFV promotes H3K9me3 and HP1β foci formation from early infection, followed by HP1α and HDAC2 nuclear enrichment, suggesting heterochromatinization of host genome. Finally, closeness between DNA damage response factors, disrupted PML-NBs, and virus-induced heterochromatic regions were identified. In sum, our results demonstrate that ASFV orchestrates spatio-temporal nuclear rearrangements, changing subnuclear domains, relocating Ataxia Telangiectasia Mutated Rad-3 related (ATR)-related factors and promoting heterochromatinization, probably controlling transcription, repressing host gene expression, and favouring viral replication. PMID:26389938

  3. The Role of Nuclear Antiviral Factors against Invading DNA Viruses: The Immediate Fate of Incoming Viral Genomes

    PubMed Central

    Komatsu, Tetsuro; Nagata, Kyosuke; Wodrich, Harald

    2016-01-01

    In recent years, it has been suggested that host cells exert intrinsic mechanisms to control nuclear replicating DNA viruses. This cellular response involves nuclear antiviral factors targeting incoming viral genomes. Herpes simplex virus-1 (HSV-1) is the best-studied model in this context, and it was shown that upon nuclear entry HSV-1 genomes are immediately targeted by components of promyelocytic leukemia nuclear bodies (PML-NBs) and the nuclear DNA sensor IFI16 (interferon gamma inducible protein 16). Based on HSV-1 studies, together with limited examples in other viral systems, these phenomena are widely believed to be a common cellular response to incoming viral genomes, although formal evidence for each virus is lacking. Indeed, recent studies suggest that the case may be different for adenovirus infection. Here we summarize the existing experimental evidence for the roles of nuclear antiviral factors against incoming viral genomes to better understand cellular responses on a virus-by-virus basis. We emphasize that cells seem to respond differently to different incoming viral genomes and discuss possible arguments for and against a unifying cellular mechanism targeting the incoming genomes of different virus families. PMID:27782081

  4. Characterization of the nuclear localization signal of the hepatitis delta virus antigen

    SciTech Connect

    Alves, Carolina; Freitas, Natalia; Cunha, Celso

    2008-01-05

    The delta antigen (HDAg) is the only protein encoded by the hepatitis delta virus (HDV) RNA genome. The HDAg contains an RNA binding domain, a dimerization domain, and a nuclear localization signal (NLS). The nuclear import of HDV RNPs is thought to be one of the first tasks of the HDAg during the HDV replication cycle. Using c-myc-PK fusions with several regions of the HDAg in transfection assays in Huh7 cells, we found that the HDAg NLS consists of a single stretch of 10 amino acids, EGAPPAKRAR, located in positions 66-75. Deletion and mutation analysis of this region showed that both the acidic glutamic acid residue at position 66 and the basic arginine residue at position 75 are essential for promoting nuclear import.

  5. A cysteine protease encoded by the baculovirus Bombyx mori nuclear polyhedrosis virus.

    PubMed Central

    Ohkawa, T; Majima, K; Maeda, S

    1994-01-01

    Sequence analysis of the BamHI F fragment of the genome of Bombyx mori nuclear polyhedrosis virus (BmNPV) revealed an open reading frame whose deduced amino acid sequence had homology to those of cysteine proteases of the papain superfamily. The putative cysteine protease sequence (BmNPV-CP) was 323 amino acids long and showed 35% identity to a cysteine proteinase precursor from Trypanosoma brucei. Of 36 residues conserved among cathepsins B, H, L, and S and papain, 31 were identical in BmNPV-CP. In order to determine the activity and function of the putative cysteine protease, a BmNPV mutant (BmCysPD) was constructed by homologous recombination of the protease gene with a beta-galactosidase gene cassette. BmCysPD-infected BmN cell extracts were significantly reduced in acid protease activity compared with wild-type virus-infected cell extracts. The cysteine protease inhibitor E-64 [trans-epoxysuccinylleucylamido-(4-guanidino)butane] inhibited wild-type virus-expressed protease activity. Deletion of the cysteine protease gene had no significant effect on viral growth or polyhedron production in BmN cells, indicating that the cysteine protease was not essential for viral replication in vitro. However, B. mori larvae infected with BmCysPD showed symptoms different from those of wild-type BmNPV-infected larvae, e.g., less degradation of the body, including fat body cells, white body surface color due presumably to undegraded epidermal cells, and an increase in the number of polyhedra released into the hemolymph. This is the first report of (i) a virus-encoded protease with activity on general substrates and (ii) evidence that a virus-encoded protease may play a role in degradation of infected larvae to facilitate horizontal transmission of the virus. Images PMID:8083997

  6. Cytoplasmic Utilization of Human Immunodeficiency Virus Type 1 Genomic RNA Is Not Dependent on a Nuclear Interaction with Gag

    PubMed Central

    Hoffmann, Bianca; Ohs, Inga; Blissenbach, Maik; Brandt, Sabine; Tippler, Bettina; Grunwald, Thomas; Überla, Klaus

    2012-01-01

    In some retroviruses, such as Rous sarcoma virus and prototype foamy virus, Gag proteins are known to shuttle between the nucleus and the cytoplasm and are implicated in nuclear export of the viral genomic unspliced RNA (gRNA) for subsequent encapsidation. A similar function has been proposed for human immunodeficiency virus type 1 (HIV-1) Gag based on the identification of nuclear localization and export signals. However, the ability of HIV-1 Gag to transit through the nucleus has never been confirmed. In addition, the lentiviral Rev protein promotes efficient nuclear gRNA export, and previous reports indicate a cytoplasmic interaction between Gag and gRNA. Therefore, functional effects of HIV-1 Gag on gRNA and its usage were explored. Expression of gag in the absence of Rev was not able to increase cytoplasmic gRNA levels of subgenomic, proviral, or lentiviral vector constructs, and gene expression from genomic reporter plasmids could not be induced by Gag provided in trans. Furthermore, Gag lacking the reported nuclear localization and export signals was still able to mediate an efficient packaging process. Although small amounts of Gag were detectable in the nuclei of transfected cells, a Crm1-dependent nuclear export signal in Gag could not be confirmed. Thus, our study does not provide any evidence for a nuclear function of HIV-1 Gag. The encapsidation process of HIV-1 therefore clearly differs from that of Rous sarcoma virus and prototype foamy virus. PMID:22258250

  7. Influenza virus targets the mRNA export machinery and the nuclear pore complex.

    PubMed

    Satterly, Neal; Tsai, Pei-Ling; van Deursen, Jan; Nussenzveig, Daniel R; Wang, Yaming; Faria, Paula A; Levay, Agata; Levy, David E; Fontoura, Beatriz M A

    2007-02-01

    The NS1 protein of influenza A virus is a major virulence factor that is essential for pathogenesis. NS1 functions to impair innate and adaptive immunity by inhibiting host signal transduction and gene expression, but its mechanisms of action remain to be fully elucidated. We show here that NS1 forms an inhibitory complex with NXF1/TAP, p15/NXT, Rae1/mrnp41, and E1B-AP5, which are key constituents of the mRNA export machinery that interact with both mRNAs and nucleoporins to direct mRNAs through the nuclear pore complex. Increased levels of NXF1, p15, or Rae1 revert the mRNA export blockage induced by NS1. Furthermore, influenza virus down-regulates Nup98, a nucleoporin that is a docking site for mRNA export factors. Reduced expression of these mRNA export factors renders cells highly permissive to influenza virus replication, demonstrating that proper levels of key constituents of the mRNA export machinery protect against influenza virus replication. Because Nup98 and Rae1 are induced by interferons, down-regulation of this pathway is likely a viral strategy to promote viral replication. These findings demonstrate previously undescribed influenza-mediated viral-host interactions and provide insights into potential molecular therapies that may interfere with influenza infection.

  8. Optimized expression, solubilization and purification of nuclear inclusion protein b of cardamom mosaic virus.

    PubMed

    Jebasingh, T; Jacob, T; Shah, M; Das, D; Krishnaswamy, S; Usha, R

    2008-04-01

    All RNA viruses encode an RNA-dependent RNA polymerase (RdRP) that is required for replication of the viral genome. Nuclear inclusion b (NIb) gene codes for the RdRp in Potyviridae viruses. In this study, expression, solubilization and purification of NIb protein of Cardamom mosaic virus (CdMV) is reported. The objective of the present study was to express and purify the NIb protein of CdMV on a large scale for structural characterization, as the structure of the RdRp from a plant virus is yet to be determined. However, the expression of NIb protein with hexa-histidine tag in Escherichia coli led to insoluble aggregates. Out of all the approaches [making truncated versions to reduce the size of protein; replacing an amino acid residue likely to be involved in hydrophobic intermolecular interactions with a hydrophilic one; expressing the protein along with chaperones; expression in Origami cells for proper disulphide bond formation, in E. coli as a fusion with maltose-binding protein (MBP) and in Nicotiana tabacum] to obtain the RdRp in a soluble form, only expression in E. coli as a fusion with MBP and its expression in N. tabacum were successful. The NIb expressed in plant or as a fusion with MBP in E. coli can be scaled up for further work.

  9. Transcriptional Regulation of BK Virus by Nuclear Factor of Activated T Cells▿

    PubMed Central

    Jordan, Joslynn A.; Manley, Kate; Dugan, Aisling S.; O'Hara, Bethany A.; Atwood, Walter J.

    2010-01-01

    The human polyomavirus BK virus (BKV) is a common virus for which 80 to 90% of the adult population is seropositive. BKV reactivation in immunosuppressed patients or renal transplant patients is the primary cause of polyomavirus-associated nephropathy (PVN). Using the Dunlop strain of BKV, we found that nuclear factor of activated T cells (NFAT) plays an important regulatory role in BKV infection. Luciferase reporter assays and chromatin immunoprecipitation assays demonstrated that NFAT4 bound to the viral promoter and regulated viral transcription and infection. The mutational analysis of the NFAT binding sites demonstrated complex functional interactions between NFAT, c-fos, c-jun, and the p65 subunit of NF-κB that together influence promoter activity and viral growth. These data indicate that NFAT is required for BKV infection and is involved in a complex regulatory network that both positively and negatively influences promoter activity and viral infection. PMID:19955309

  10. Nuclear dynamics of influenza A virus ribonucleoproteins revealed by live-cell imaging studies

    SciTech Connect

    Loucaides, Eva M.; Kirchbach, Johann C. von; Foeglein, Agnes; Sharps, Jane; Fodor, Ervin; Digard, Paul

    2009-11-10

    The negative sense RNA genome of influenza A virus is transcribed and replicated in the nuclei of infected cells by the viral RNA polymerase. Only four viral polypeptides are required but multiple cellular components are potentially involved. We used fluorescence recovery after photobleaching (FRAP) to characterise the dynamics of GFP-tagged viral ribonucleoprotein (RNP) components in living cells. The nucleoprotein (NP) displayed very slow mobility that significantly increased on formation of transcriptionally active RNPs. Conversely, single or dimeric polymerase subunits showed fast nuclear dynamics that decreased upon formation of heterotrimers, suggesting increased interaction of the full polymerase complex with a relatively immobile cellular component(s). Treatment with inhibitors of cellular transcription indicated that in part, this reflected an interaction with cellular RNA polymerase II. Analysis of mutated influenza virus polymerase complexes further suggested that this was through an interaction between PB2 and RNA Pol II separate from PB2 cap-binding activity.

  11. Mutations within the conserved NS1 nuclear export signal lead to inhibition of influenza A virus replication

    PubMed Central

    2014-01-01

    Background The influenza A virus NS1 protein is a virulence factor and an antagonist of host cell innate immune responses. During virus infection NS1 protein has several functions both in the nucleus and in the cytoplasm and its intracellular localization is regulated by one or two nuclear localization signals (NLS) and a nuclear export signal (NES). Methods In order to investigate the role of NS1 NES in intracellular localization, virus life cycle and host interferon responses, we generated recombinant A/Udorn/72 viruses harboring point mutations in the NES sequence. Results NS1 NES was found to be inactivated by several of the mutations resulting in nuclear retention of NS1 at late stages of infection confirming that this sequence is a bona fide functional NES. Some of the mutant viruses showed reduced growth properties in cell culture, inability to antagonize host cell interferon production and increased p-IRF3 levels, but no clear correlation between these phenotypes and NS1 localization could be made. Impaired activation of Akt phosphorylation by the replication-deficient viruses indicates possible disruption of NS1-p85β interaction by mutations in the NES region. Conclusion We conclude that mutations within the NS1 NES result in impairment of several NS1 functions which extends further from the NES site being only involved in regulating the nuclear-cytoplasmic trafficking of NS1. PMID:25023993

  12. Nuclear factor 1 family members mediate repression of the BK virus late promoter.

    PubMed

    Kraus, R J; Shadley, L; Mertz, J E

    2001-08-15

    BK virus (BKV) is a member of the polyoma virus family that is ubiquitous in humans. Its 5-kb DNA genome consists of a bidirectional promoter region situated between two temporally regulated coding regions. We mapped the transcription initiation site of the major late promoter (MLP) of the archetype strain BKV(WW) to nt 185. We found that it lies within the sequence TGGN6GCCA, a binding site for members of the nuclear factor 1 (NF1) family of transcription factors. Competition electrophoretic mobility shift and immunoshift assays confirmed that NF1 factors present in nuclear extracts of HeLa and CV-1 cells bind to the BKV-MLP. Because BKV(WW) grew poorly in tissue culture and failed to express detectable levels of RNA in vitro, SV40-BKV chimeric viruses were constructed to investigate the transcriptional function of this NF-1 binding site. These sequence-specific factors repressed transcription in a cell-free system when template copy number was low. This repression could be relieved by the addition in trans of oligonucleotides containing wild-type, but not mutated, NF1-binding site sequences. SV40-BKV chimeric viruses defective in this NF1-binding site overproduced late RNA at early, but not late, times after transfection of CV-1 cells. Finally, transient expression in 293 cells of cDNAs encoding the family members NF1-A4, NF1-C2, and NF1-X2 specifically repressed transcription from the BKV late promoter approximately 3-, 10-, and 10-fold, respectively, in a DNA binding-dependent manner. We conclude that some members of the NF1 family of transcription factors can act as sequence-specific cellular repressors of the BKV-MLP. We propose that titration of these and other cellular repressors by viral genome amplification may be responsible in part for the replication-dependent component of the early-to-late switch in BKV gene expression.

  13. Vaccinia virus protein N2 is a nuclear IRF3 inhibitor that promotes virulence.

    PubMed

    Ferguson, Brian J; Benfield, Camilla T O; Ren, Hongwei; Lee, Vivian H; Frazer, Gordon L; Strnadova, Pavla; Sumner, Rebecca P; Smith, Geoffrey L

    2013-09-01

    Vaccinia virus (VACV) expresses many proteins that are non-essential for virus replication but promote virulence by inhibiting components of the host immune response to infection. These immunomodulators include a family of proteins that have, or are predicted to have, a structure related to the B-cell lymphoma (Bcl)-2 protein. Five members of the VACV Bcl-2 family (N1, B14, A52, F1 and K7) have had their crystal structure solved, others have been characterized and a function assigned (C6, A46), and others are predicted to be Bcl-2 proteins but are uncharacterized hitherto (N2, B22, C1). Data presented here show that N2 is a nuclear protein that is expressed early during infection and inhibits the activation of interferon regulatory factor (IRF)3. Consistent with its nuclear localization, N2 inhibits IRF3 downstream of the TANK-binding kinase (TBK)-1 and after IRF3 translocation into the nucleus. A mutant VACV strain Western Reserve lacking the N2L gene (vΔN2) showed normal replication and spread in cultured cells compared to wild-type parental (vN2) and revertant (vN2-rev) viruses, but was attenuated in two murine models of infection. After intranasal infection, the vΔN2 mutant induced lower weight loss and signs of illness, and virus was cleared more rapidly from the infected tissue. In the intradermal model of infection, vΔN2 induced smaller lesions that were resolved more rapidly. In summary, the N2 protein is an intracellular virulence factor that inhibits IRF3 activity in the nucleus.

  14. Murine Leukemia Virus Uses TREX Components for Efficient Nuclear Export of Unspliced Viral Transcripts

    PubMed Central

    Sakuma, Toshie; Tonne, Jason M.; Ikeda, Yasuhiro

    2014-01-01

    Previously we reported that nuclear export of both unspliced and spliced murine leukemia virus (MLV) transcripts depends on the nuclear export factor (NXF1) pathway. Although the mRNA export complex TREX, which contains Aly/REF, UAP56, and the THO complex, is involved in the NXF1-mediated nuclear export of cellular mRNAs, its contribution to the export of MLV mRNA transcripts remains poorly understood. Here, we studied the involvement of TREX components in the export of MLV transcripts. Depletion of UAP56, but not Aly/REF, reduced the level of both unspliced and spliced viral transcripts in the cytoplasm. Interestingly, depletion of THO components, including THOC5 and THOC7, affected only unspliced viral transcripts in the cytoplasm. Moreover, the RNA immunoprecipitation assay showed that only the unspliced viral transcript interacted with THOC5. These results imply that MLV requires UAP56, THOC5 and THOC7, in addition to NXF1, for nuclear export of viral transcripts. Given that naturally intronless mRNAs, but not bulk mRNAs, require THOC5 for nuclear export, it is plausible that THOC5 plays a key role in the export of unspliced MLV transcripts. PMID:24618812

  15. Verdinexor, a Novel Selective Inhibitor of Nuclear Export, Reduces Influenza A Virus Replication In Vitro and In Vivo

    PubMed Central

    Perwitasari, Olivia; Johnson, Scott; Yan, Xiuzhen; Howerth, Elizabeth; Shacham, Sharon; Landesman, Yosef; Baloglu, Erkan; McCauley, Dilara; Tamir, Sharon; Tompkins, S. Mark

    2014-01-01

    ABSTRACT Influenza is a global health concern, causing death, morbidity, and economic losses. Chemotherapeutics that target influenza virus are available; however, rapid emergence of drug-resistant strains is common. Therapeutic targeting of host proteins hijacked by influenza virus to facilitate replication is an antiviral strategy to reduce the development of drug resistance. Nuclear export of influenza virus ribonucleoprotein (vRNP) from infected cells has been shown to be mediated by exportin 1 (XPO1) interaction with viral nuclear export protein tethered to vRNP. RNA interference screening has identified XPO1 as a host proinfluenza factor where XPO1 silencing results in reduced influenza virus replication. The Streptomyces metabolite XPO1 inhibitor leptomycin B (LMB) has been shown to limit influenza virus replication in vitro; however, LMB is toxic in vivo, which makes it unsuitable for therapeutic use. In this study, we tested the anti-influenza virus activity of a new class of orally available small-molecule selective inhibitors of nuclear export, specifically, the XPO1 antagonist KPT-335 (verdinexor). Verdinexor was shown to potently and selectively inhibit vRNP export and effectively inhibited the replication of various influenza virus A and B strains in vitro, including pandemic H1N1 virus, highly pathogenic H5N1 avian influenza virus, and the recently emerged H7N9 strain. In vivo, prophylactic and therapeutic administration of verdinexor protected mice against disease pathology following a challenge with influenza virus A/California/04/09 or A/Philippines/2/82-X79, as well as reduced lung viral loads and proinflammatory cytokine expression, while having minimal toxicity. These studies show that verdinexor acts as a novel anti-influenza virus therapeutic agent. IMPORTANCE Antiviral drugs represent important means of influenza virus control. However, substantial resistance to currently approved influenza therapeutic drugs has developed. New antiviral

  16. Generalized Immunoassay for Autographa californica Nuclear Polyhedrosis Virus Infectivity In Vitro

    PubMed Central

    Volkman, Loy E.; Goldsmith, Phyllis A.

    1982-01-01

    A quantitative in vitro immunoassay for the infectivity of Autographa californica nuclear polyhedrosis virus was developed and performed in six different lepidopteran cell lines. The assay was not dependent upon cytopathic effect or polyhedron production, but rather upon viral antigen production and its recognition in a peroxidase-antiperoxidase staining procedure. The importance of using such an assay for accurately assessing infectivity in cell lines which produce polyhedra inefficiently was demonstrated. Differences among the cell lines in sensitivity to viral infection were clearly shown. Differences in the time required to produce infectious progeny were also noted among cells of the same cell line. Images PMID:16346059

  17. Surface localization of the nuclear receptor CAR in influenza A virus-infected cells

    SciTech Connect

    Takahashi, Tadanobu; Moriyama, Yusuke; Ikari, Akira; Sugatani, Junko; Suzuki, Takashi; Miwa, Masao

    2008-04-11

    Constitutive active/androstane receptor CAR is a member of the nuclear receptors which regulate transcription of xenobiotic metabolism enzymes. CAR is usually localized in the cytosol and nucleus. Here, we found that CAR was localized at the cell surface of influenza A virus (IAV)-infected cells. Additionally, we demonstrated that expression of a viral envelope glycoprotein, either hemagglutinin (HA) or neuraminidase (NA), but not viral nucleoprotein (NP), was responsible for this localization. This report is the first demonstration of CAR at the surface of tissue culture cells, and suggests that CAR may exert the IAV infection mechanism.

  18. The Pseudorabies Virus DNA Polymerase Accessory Subunit UL42 Directs Nuclear Transport of the Holoenzyme.

    PubMed

    Wang, Yi-Ping; Du, Wen-Juan; Huang, Li-Ping; Wei, Yan-Wu; Wu, Hong-Li; Feng, Li; Liu, Chang-Ming

    2016-01-01

    Pseudorabies virus (PRV) DNA replication occurs in the nuclei of infected cells and requires the viral DNA polymerase. The PRV DNA polymerase comprises a catalytic subunit, UL30, and an accessory subunit, UL42, that confers processivity to the enzyme. Its nuclear localization is a prerequisite for its enzymatic function in the initiation of viral DNA replication. However, the mechanisms by which the PRV DNA polymerase holoenzyme enters the nucleus have not been determined. In this study, we characterized the nuclear import pathways of the PRV DNA polymerase catalytic and accessory subunits. Immunofluorescence analysis showed that UL42 localizes independently in the nucleus, whereas UL30 alone predominantly localizes in the cytoplasm. Intriguingly, the localization of UL30 was completely shifted to the nucleus when it was coexpressed with UL42, demonstrating that nuclear transport of UL30 occurs in an UL42-dependent manner. Deletion analysis and site-directed mutagenesis of the two proteins showed that UL42 contains a functional and transferable bipartite nuclear localization signal (NLS) at amino acids 354-370 and that K(354), R(355), and K(367) are important for the NLS function, whereas UL30 has no NLS. Coimmunoprecipitation assays verified that UL42 interacts with importins α3 and α4 through its NLS. In vitro nuclear import assays demonstrated that nuclear accumulation of UL42 is a temperature- and energy-dependent process and requires both importins α and β, confirming that UL42 utilizes the importin α/β-mediated pathway for nuclear entry. In an UL42 NLS-null mutant, the UL42/UL30 heterodimer was completely confined to the cytoplasm when UL42 was coexpressed with UL30, indicating that UL30 utilizes the NLS function of UL42 for its translocation into the nucleus. Collectively, these findings suggest that UL42 contains an importin α/β-mediated bipartite NLS that transports the viral DNA polymerase holoenzyme into the nucleus in an in vitro expression

  19. Asymmetric Arginine dimethylation of Epstein-Barr virus nuclear antigen 2 promotes DNA targeting

    SciTech Connect

    Gross, Henrik; Barth, Stephanie; Mamiani, Alfredo; Zimber-Strobl, Ursula; West, Michelle J.; Kremmer, Elisabeth; Graesser, Friedrich A.

    2010-02-20

    The Epstein-Barr virus (EBV) growth-transforms B-lymphocytes. The virus-encoded nuclear antigen 2 (EBNA2) is essential for transformation and activates gene expression by association with DNA-bound transcription factors such as RBPJkappa (CSL/CBF1). We have previously shown that EBNA2 contains symmetrically dimethylated Arginine (sDMA) residues. Deletion of the RG-repeat results in a reduced ability of the virus to immortalise B-cells. We now show that the RG repeat also contains asymmetrically dimethylated Arginines (aDMA) but neither non-methylated (NMA) Arginines nor citrulline residues. We demonstrate that only aDMA-containing EBNA2 is found in a complex with DNA-bound RBPJkappa in vitro and preferentially associates with the EBNA2-responsive EBV C, LMP1 and LMP2A promoters in vivo. Inhibition of methylation in EBV-infected cells results in reduced expression of the EBNA2-regulated viral gene LMP1, providing additional evidence that methylation is a prerequisite for DNA-binding by EBNA2 via association with the transcription factor RBPJkappa.

  20. Semipermissive replication of a nuclear polyhedrosis virus of Autographa californica in a gypsy moth cell line

    SciTech Connect

    McClintock, J.T.; Dougherty, E.M.; Weiner, R.M.

    1986-01-01

    Several gypsy moth cell lines have been previously described as nonpermissive for the multiple-embedded nuclear polyhedrosis virus of Autographa californica (AcMNPV). In this report, the authors demonstrate the semipermissive infection of a gypsy moth cell line, IPLB-LD-652Y, with AcMNPV. IPLB-LD-652Y cells infected with AcMNPV produced classic cytopathic effects but failed to yield infectious progeny virus. Results of experiments employing DNA-DNA dot hybridization suggested that AcMNPV DNA synthesis was initiated from 8 to 12 h postinfection (p.i.), continued at a maximum rate from 12 to 20 h p.i., and declined from 20 to 36 h p.i. The rate of AcMNPV DNA synthesis approximated that observed in the permissive TN-368 cell line. AcMNPV-infected IPLB-LD-652Y cells, pulse-labeled with (/sup 35/S)methionine at various time intervals p.i. and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, revealed four virus-induced proteins, one novel to the semipermissive system and three early ..cap alpha.. proteins, synthesized from 1 to 20 h p.i. Thereafter, both host and viral protein synthesis was completely suppressed. These results suggest that AcMNPV adsorbed, penetrated, and initiated limited macromolecular synthesis in the semipermissive gypsy moth cell line. However, the infection cycle was restricted during the early phase of AcMNPV replication.

  1. [Identification of prototype foamy virus Bel1 nuclear localization signal and its corresponding importins].

    PubMed

    Ma, Qing-Lin; Yu, Miao; Luo, Di; Tan, Juan; Qiao, Wen-Tao

    2014-07-01

    Bel1, a transactivator of prototype foamy virus (PFV), plays pivotal roles in the replication of PFV. Previous studies have shown that Bel1 bears a nuclear localization signal (NLS), but its amino acid sequence remains unclear and the corresponding importins have not been identified. In this report, we inserted various fragments of Bel1 into an EGFP-GST fusion protein and investigated their subcellular localization by fluorescence microscopy. We found that the 215PRQKRPR221 fragment could direct nuclear localization, which accords with the consensus sequence K(K/R)X(K/R) of monopartite NLS. Point mutation experiments revealed that K218, R219, and R221 are essential for the nuclear localization of Bel1. The results of the GST-pulldown showed that the Bel1 fragment with residues 215-223, which bears the NLS, interacts with KPNA1, KPNA6, and KPNA7. This result suggests that KPNA1, KPNA6, and KPNA7 maybe involved in Bel1 nuclear translocation.

  2. Varicella-zoster virus induces the formation of dynamic nuclear capsid aggregates

    SciTech Connect

    Lebrun, Marielle; Thelen, Nicolas; Thiry, Marc; Riva, Laura; Ote, Isabelle; Condé, Claude; Vandevenne, Patricia; Di Valentin, Emmanuel; Bontems, Sébastien; Sadzot-Delvaux, Catherine

    2014-04-15

    The first step of herpesviruses virion assembly occurs in the nucleus. However, the exact site where nucleocapsids are assembled, where the genome and the inner tegument are acquired, remains controversial. We created a recombinant VZV expressing ORF23 (homologous to HSV-1 VP26) fused to the eGFP and dually fluorescent viruses with a tegument protein additionally fused to a red tag (ORF9, ORF21 and ORF22 corresponding to HSV-1 UL49, UL37 and UL36). We identified nuclear dense structures containing the major capsid protein, the scaffold protein and maturing protease, as well as ORF21 and ORF22. Correlative microscopy demonstrated that the structures correspond to capsid aggregates and time-lapse video imaging showed that they appear prior to the accumulation of cytoplasmic capsids, presumably undergoing the secondary egress, and are highly dynamic. Our observations suggest that these structures might represent a nuclear area important for capsid assembly and/or maturation before the budding at the inner nuclear membrane. - Highlights: • We created a recombinant VZV expressing the small capsid protein fused to the eGFP. • We identified nuclear dense structures containing capsid and procapsid proteins. • Correlative microscopy showed that the structures correspond to capsid aggregates. • Procapsids and partial capsids are found within the aggregates of WT and eGFP-23 VZV. • FRAP and FLIP experiments demonstrated that they are dynamic structures.

  3. Binding of nuclear factors to functional domains of the duck hepatitis B virus enhancer.

    PubMed Central

    Lilienbaum, A; Crescenzo-Chaigne, B; Sall, A A; Pillot, J; Elfassi, E

    1993-01-01

    We have analyzed the structures, relative organization, and activities of binding sites for nuclear factors in the duck hepatitis B virus (duck HBV) enhancer. DNase I footprinting analysis and mobility shift assays demonstrate that this enhancer of 192 bp contains at least three binding sites for transcription factors: one for hepatocyte-adipocyte C/EBP, a second for the liver-specific transactivator hepatocyte nuclear factor 1 HNF-1, and a third for a factor, called F3, which binds to a DNA sequence bearing some resemblance to that for the ubiquitous factor EF-C. Analysis of transcriptional activity reveals that oligonucleotides corresponding to the individual binding sites, inserted upstream from a heterologous promoter, display very weak enhancer activity, whereas the enhancer encompassing these three sites displays very high activity. Analysis of duck HBV enhancer mutants indicates that the deletion of any of these sites leads to a modification of transcriptional enhancer activity. The hepatocyte nuclear factor 1 binding site is crucial, since an internal deletion of 14 bp abolishes the activity. The C/EBP site can act as repressor, and the F3 site is required for full activity. Comparative analysis reveals that the nuclear factors are similar to those bound to the human HBV enhancer but that the organization of their binding sites in the duck HBV enhancer is different. Images PMID:8371357

  4. DNA hybridization assay for detection of gypsy moth nuclear polyhedrosis virus in infected gypsy moth (Lymantria dispar L. ) larvae

    SciTech Connect

    Keating, S.T.; Burand, J.P.; Elkinton, J.S. )

    1989-11-01

    Radiolabeled Lymantria dispar nuclear polyhedrosis virus DNA probes were used in a DNA hybridization assay to detect the presence of viral DNA in extracts from infected larvae. Total DNA was extracted from larvae, bound to nitrocellulose filters, and assayed for the presence of viral DNA by two methods: slot-blot vacuum filtration and whole-larval squashes. The hybridization results were closely correlated with mortality observed in reared larvae. Hybridization of squashes of larvae frozen 4 days after receiving the above virus treatments also produced accurate measures of the incidence of virus infection.

  5. Insights into the nuclear export of murine leukemia virus intron-containing RNA

    PubMed Central

    Pessel-Vivares, Lucie; Houzet, Laurent; Lainé, Sébastien; Mougel, Marylène

    2015-01-01

    The retroviral genome consists of an intron-containing transcript that has essential cytoplasmic functions in the infected cell. This viral transcript can escape splicing, circumvent the nuclear checkpoint mechanisms and be transported to the cytoplasm by hijacking the host machinery. Once in the cytoplasm, viral unspliced RNA acts as mRNA to be translated and as genomic RNA to be packaged into nascent viruses. The murine leukemia virus (MLV) is among the first retroviruses discovered and is classified as simple Retroviridae due to its minimal encoding capacity. The oncogenic and transduction abilities of MLV are extensively studied, whereas surprisingly the crucial step of its nuclear export has remained unsolved until 2014. Recent work has revealed the recruitment by MLV of the cellular NXF1/Tap-dependent pathway for export. Unconventionally, MLV uses of Tap to export both spliced and unspliced viral RNAs. Unlike other retroviruses, MLV does not harbor a unique RNA signal for export. Indeed, multiple sequences throughout the MLV genome appear to promote export of the unspliced MLV RNA. We review here the current understanding of the export mechanism and highlight the determinants that influence MLV export. As the molecular mechanism of MLV export is elucidated, we will gain insight into the contribution of the export pathway to the cytoplasmic fate of the viral RNA. PMID:26158194

  6. Structural determination of importin alpha in complex with beak and feather disease virus capsid nuclear localization signal

    SciTech Connect

    Patterson, Edward I.; Dombrovski, Andrew K.; Swarbrick, Crystall M.D.; Raidal, Shane R.; Forwood, Jade K.

    2013-09-06

    Highlights: •Circovirus capsid proteins contain large nuclear localization signals (NLS). •A method of nuclear import has not been elucidated. •Beak and feather disease virus (BFDV) capsid NLS was crystallized with importin α. •The structure showed BFDV NLS binding to the major site of importin α. •Result shows implications for mechanism of nuclear transport for all circoviruses. -- Abstract: Circoviruses represent a rapidly increasing genus of viruses that infect a variety of vertebrates. Replication requires shuttling viral molecules into the host cell nucleus, a process facilitated by capsid-associated protein (Cap). Whilst a nuclear localization signal (NLS) has been shown to mediate nuclear translocation, the mode of nuclear transport remains to be elucidated. To better understand this process, beak and feather disease virus (BFDV) Cap NLS was crystallized with nuclear import receptor importin-α (Impα). Diffraction yielded structural data to 2.9 Å resolution, and the binding site on both Impα and BFDV Cap NLS were well resolved. The binding mechanism for the major site is likely conserved across circoviruses as supported by the similarity of NLSs in circovirus Caps. This finding illuminates a crucial step for infection of host cells by this viral family, and provides a platform for rational drug design against the binding interface.

  7. Super-resolution imaging of nuclear import of adeno-associated virus in live cells

    PubMed Central

    Kelich, Joseph M; Ma, Jiong; Dong, Biao; Wang, Qizhao; Chin, Mario; Magura, Connor M; Xiao, Weidong; Yang, Weidong

    2015-01-01

    Adeno-associated virus (AAV) has been developed as a promising human gene therapy vector. Particularly, recombinant AAV vector (rAAV) achieves its transduction of host cells by crossing at least three physiological barriers including plasma membrane, endosomal membrane, and nuclear envelope (NE). So far, the AAV transduction mechanism has not been explored thoroughly at the single viral particle level. In this study, we employed high-speed super-resolution single-point edge-excitation sub-diffraction (SPEED) microscopy to map the events of single rAAV2 particles infecting live human cells with an unprecedented spatiotemporal resolution of 9–12 nm and 2–20 ms. Data reveal that rAAV2 particles are imported through nuclear pore complexes (NPCs) rather than nuclear membrane budding into the nucleus. Moreover, approximately 17% of the rAAV2 molecules starting from the cytoplasm successfully transverse the NPCs to reach the nucleoplasm, revealing that the NPCs act as a strict selective step for AAV delivery. This study lastly suggests a new pathway to improve AAV vectors for human gene therapy. PMID:26665132

  8. Super-resolution imaging of nuclear import of adeno-associated virus in live cells.

    PubMed

    Kelich, Joseph M; Ma, Jiong; Dong, Biao; Wang, Qizhao; Chin, Mario; Magura, Connor M; Xiao, Weidong; Yang, Weidong

    2015-01-01

    Adeno-associated virus (AAV) has been developed as a promising human gene therapy vector. Particularly, recombinant AAV vector (rAAV) achieves its transduction of host cells by crossing at least three physiological barriers including plasma membrane, endosomal membrane, and nuclear envelope (NE). So far, the AAV transduction mechanism has not been explored thoroughly at the single viral particle level. In this study, we employed high-speed super-resolution single-point edge-excitation sub-diffraction (SPEED) microscopy to map the events of single rAAV2 particles infecting live human cells with an unprecedented spatiotemporal resolution of 9-12 nm and 2-20 ms. Data reveal that rAAV2 particles are imported through nuclear pore complexes (NPCs) rather than nuclear membrane budding into the nucleus. Moreover, approximately 17% of the rAAV2 molecules starting from the cytoplasm successfully transverse the NPCs to reach the nucleoplasm, revealing that the NPCs act as a strict selective step for AAV delivery. This study lastly suggests a new pathway to improve AAV vectors for human gene therapy. PMID:26665132

  9. Influenza A viruses escape from MxA restriction at the expense of efficient nuclear vRNP import.

    PubMed

    Götz, Veronika; Magar, Linda; Dornfeld, Dominik; Giese, Sebastian; Pohlmann, Anne; Höper, Dirk; Kong, Byung-Whi; Jans, David A; Beer, Martin; Haller, Otto; Schwemmle, Martin

    2016-01-01

    To establish a new lineage in the human population, avian influenza A viruses (AIV) must overcome the intracellular restriction factor MxA. Partial escape from MxA restriction can be achieved when the viral nucleoprotein (NP) acquires the critical human-adaptive amino acid residues 100I/V, 283P, and 313Y. Here, we show that introduction of these three residues into the NP of an avian H5N1 virus renders it genetically unstable, resulting in viruses harboring additional single mutations, including G16D. These substitutions restored genetic stability yet again yielded viruses with varying degrees of attenuation in mammalian and avian cells. Additionally, most of the mutant viruses lost the capacity to escape MxA restriction, with the exception of the G16D virus. We show that MxA escape is linked to attenuation by demonstrating that the three substitutions promoting MxA escape disturbed intracellular trafficking of incoming viral ribonucleoprotein complexes (vRNPs), thereby resulting in impaired nuclear import, and that the additional acquired mutations only partially compensate for this import block. We conclude that for adaptation to the human host, AIV must not only overcome MxA restriction but also an associated block in nuclear vRNP import. This inherent difficulty may partially explain the frequent failure of AIV to become pandemic. PMID:26988202

  10. Influenza A viruses escape from MxA restriction at the expense of efficient nuclear vRNP import

    PubMed Central

    Götz, Veronika; Magar, Linda; Dornfeld, Dominik; Giese, Sebastian; Pohlmann, Anne; Höper, Dirk; Kong, Byung-Whi; Jans, David A.; Beer, Martin; Haller, Otto; Schwemmle, Martin

    2016-01-01

    To establish a new lineage in the human population, avian influenza A viruses (AIV) must overcome the intracellular restriction factor MxA. Partial escape from MxA restriction can be achieved when the viral nucleoprotein (NP) acquires the critical human-adaptive amino acid residues 100I/V, 283P, and 313Y. Here, we show that introduction of these three residues into the NP of an avian H5N1 virus renders it genetically unstable, resulting in viruses harboring additional single mutations, including G16D. These substitutions restored genetic stability yet again yielded viruses with varying degrees of attenuation in mammalian and avian cells. Additionally, most of the mutant viruses lost the capacity to escape MxA restriction, with the exception of the G16D virus. We show that MxA escape is linked to attenuation by demonstrating that the three substitutions promoting MxA escape disturbed intracellular trafficking of incoming viral ribonucleoprotein complexes (vRNPs), thereby resulting in impaired nuclear import, and that the additional acquired mutations only partially compensate for this import block. We conclude that for adaptation to the human host, AIV must not only overcome MxA restriction but also an associated block in nuclear vRNP import. This inherent difficulty may partially explain the frequent failure of AIV to become pandemic. PMID:26988202

  11. Efficient large-scale protein production of larvae and pupae of silkworm by Bombyx mori nuclear polyhedrosis virus bacmid system.

    PubMed

    Motohashi, Tomoko; Shimojima, Tsukasa; Fukagawa, Tatsuo; Maenaka, Katsumi; Park, Enoch Y

    2005-01-21

    Silkworm is one of the most attractive hosts for large-scale production of eukaryotic proteins as well as recombinant baculoviruses for gene transfer to mammalian cells. The bacmid system of Autographa californica nuclear polyhedrosis virus (AcNPV) has already been established and widely used. However, the AcNPV does not have a potential to infect silkworm. We developed the first practical Bombyx mori nuclear polyhedrosis virus bacmid system directly applicable for the protein expression of silkworm. By using this system, the green fluorescence protein was successfully expressed in silkworm larvae and pupae not only by infection of its recombinant virus but also by direct injection of its bacmid DNA. This method provides the rapid protein production in silkworm as long as 10 days, is free from biohazard, thus will be a powerful tool for the future production factory of recombinant eukaryotic proteins and baculoviruses. PMID:15596136

  12. Basic Amino Acid Mutations in the Nuclear Localization Signal of Hibiscus Chlorotic Ringspot Virus p23 Inhibit Virus Long Distance Movement

    PubMed Central

    Gao, Ruimin; Wong, Sek-Man

    2013-01-01

    The p23 is a unique protein in the Hibiscus chlorotic ringspot virus which belongs to Family Tombusviridae Genus Carmovirus. Our previous results showed that the p23 is indispensable for host-specific replication and is localized in the nucleus with a novel nuclear localization signal. To investigate additional function(s) of p23, mutations of basic amino acids lysine (K), arginine (R) and histidine (H) that abolish its nuclear localization, were introduced into a biologically active full-length cDNA clone p223 of HCRSV for testing its effects on virus replication and virus movement in vivo. Primer-specific reverse transcription-PCR was conducted to detect gene transcript level of p23 and viral coat protein separately. Virus replication and its coat protein expression were detected by fluorescent in situ hybridization and Western blot, respectively. The effect of p23 was further confirmed by using artificial microRNA inoculation-mediated silencing. Results showed that the two mutants were able to replicate in protoplasts but unable to move from inoculated leaves to newly emerged leaves. Both the p23 and the CP genes of HCRSV were detected in the newly emerged leaves of infected plants but CP was not detected by Western blot and no symptom was observed on those leaves at 19 days post inoculation. This study demonstrates that when p23 is prevented from entering the nucleus, it results in restriction of virus long distance movement which in turn abrogates symptom expression in the newly emerged leaves. We conclude that the p23 protein of HCRSV is required for virus long distance movement. PMID:24019944

  13. Nuclear import of influenza B virus nucleoprotein: Involvement of an N-terminal nuclear localization signal and a cleavage-protection motif

    SciTech Connect

    Wanitchang, Asawin; Narkpuk, Jaraspim; Jongkaewwattana, Anan

    2013-08-15

    The nucleoprotein of influenza B virus (BNP) shares several characteristics with its influenza A virus counterpart (ANP), including localization in the host's nucleus. However, while the nuclear localization signal(s) (NLS) of ANP are well characterized, little is known about those of BNP. In this study, we showed that the fusion protein bearing the BNP N-terminus fused with GFP (N70–GFP) is exclusively nuclear, and identified a highly conserved KRXR motif spanning residues 44–47 as a putative NLS. In addition, we demonstrated that residues 3–15 of BNP, though not an NLS, are also crucial for nuclear import. Results from mutational analyses of N70–GFP and the full-length BNP suggest that this region may be required for protection of the N-terminus from proteolytic cleavage. Altogether, we propose that the N-terminal region of BNP contains the NLS and cleavage-protection motif, which together drive its nuclear localization. - Highlights: • The N-terminal region of BNP is required for nuclear accumulation. • The conserved motif at position 44–47 is a putative nuclear localization signal. • The first 15 amino acids of BNP may function as a cleavage-protection motif. • BNP may get access to the nucleus via a mechanism distinct from ANP.

  14. [Quantitative determination of the infectivity of nuclear polyhedrosis virus DNA on honeycomb moth (Galleria mellonella) larvae].

    PubMed

    Gorbunova, E E; Makarova, N I; Shchelkunova, G A

    1980-01-01

    The possibility of using honeycomb moth larvae for titration of nuclear polyhedrosis virus (NPV) infectious DNA and determinations of transfection effectiveness was studied. Honeycomb moth larvae were shown to be a sensitive system for NPV DNA titration. DEAE-dextran used as a protector increased NPV DNA infectivity 1000-fold, LD50 in this instance being 2 X 10(8) molecules per larva. The method of NPV DNA infectivity determinations by the number of larvae with polyhedreae in the fatty tissue is more sensitive than infectivity determinations by the number of dead larvae and permits titrations of low DNA concentrations. The curve of DNA titration in the presence of DEAE-dextran by the number of larvae with polyhedrae in the fatty tissue allows to quantitate native DNA within the range of 0.01 to 5 micrograms/ml.

  15. Respiratory syncytial virus M2-1 protein induces the activation of nuclear factor kappa B

    SciTech Connect

    Reimers, Kerstin . E-mail: reimers.kerstin@mh-hannover.de; Buchholz, Katja; Werchau, Hermann

    2005-01-20

    Respiratory syncytial virus (RSV) induces the production of a number of cytokines and chemokines by activation of nuclear factor kappa B (NF-{kappa}B). The activation of NF-{kappa}B has been shown to depend on viral replication in the infected cells. In this study, we demonstrate that expression of RSV M2-1 protein, a transcriptional processivity and anti-termination factor, is sufficient to activate NF-{kappa}B in A549 cells. Electromobility shift assays show increased NF-{kappa}B complexes in the nuclei of M2-1-expressing cells. M2-1 protein is found in nuclei of M2-1-expressing cells and in RSV-infected cells. Co-immunoprecipitations of nuclear extracts of M2-1-expressing cells and of RSV-infected cells revealed an association of M2-1 with Rel A protein. Furthermore, the activation of NF-{kappa}B depends on the C-terminus of the RSV M2-1 protein, as shown by NF-{kappa}B-induced gene expression of a reporter gene construct.

  16. Nuclear Translocation of Crk Adaptor Proteins by the Influenza A Virus NS1 Protein

    PubMed Central

    Ylösmäki, Leena; Fagerlund, Riku; Kuisma, Inka; Julkunen, Ilkka; Saksela, Kalle

    2016-01-01

    The non-structural protein-1 (NS1) of many influenza A strains, especially those of avian origin, contains an SH3 ligand motif, which binds tightly to the cellular adaptor proteins Crk (Chicken tumor virus number 10 (CT10) regulator of kinase) and Crk-like adapter protein (CrkL). This interaction has been shown to potentiate NS1-induced activation of the phosphatidylinositol 3-kinase (PI3K), but additional effects on the host cell physiology may exist. Here we show that NS1 can induce an efficient translocation of Crk proteins from the cytoplasm into the nucleus, which results in an altered pattern of nuclear protein tyrosine phosphorylation. This was not observed using NS1 proteins deficient in SH3 binding or engineered to be exclusively cytoplasmic, indicating a physical role for NS1 as a carrier in the nuclear translocation of Crk. These data further emphasize the role of Crk proteins as host cell interaction partners of NS1, and highlight the potential for host cell manipulation gained by a viral protein simply via acquiring a short SH3 binding motif. PMID:27092521

  17. Making sense of nuclear localization: A zinc-finger protein encoded by a cytoplasmically replicating plant RNA virus acts a transcription factor

    PubMed Central

    Lukhovitskaya, Nina I.; Gushchin, Vladimir A.; Solovyev, Andrey G.; Savenkov, Eugene I.

    2013-01-01

    Recent studies have uncovered numerous nucleus-localized proteins encoded by plant RNA viruses. Whereas for some of these viruses nuclear (or, more specifically, nucleolar) passage of the proteins is needed for the virus movement within the plant or suppression of host defense, the nuclear function of these proteins remains largely unknown. Recently, the situation has been clarified for one group of plant RNA viruses, the Carlaviruses. Being positive-stranded RNA viruses, carlaviruses multiply exclusively in the cytoplasm. Chrysanthemum virus B (CVB, a carlavirus) encodes a zinc-finger protein p12 targeted to the nucleus in a nuclear localization signal-dependent manner. In a recent work, we demonstrated that p12 directly interacts with chromatin and plant promoters, thus, acts as a eukaryotic transcription factor (TF) and activates expression of a host TF involved in regulation of cell size and proliferation to favor virus infection. Therefore our studies identified a novel nuclear stage of in CVB infection involving modulation of host gene expression and plant development. Whereas it is well established that any RNA virus actively replicating in the cell causes changes in the transcriptome, our study expanded this view by showing that some positive-stranded RNA viruses can directly manipulate host transcription by encoding eukaryotic TFs. PMID:23759549

  18. [The effect of fodder on the susceptibility of gypsy moth (Lymantria dispar L.) to nuclear polyhedrosis virus].

    PubMed

    Bakhvalov, S A; Bakhvalova, V N

    2009-01-01

    Gypsy moth (Lymantria dispar L.) growing on different feeding substrates was shown to affect their susceptibility to nuclear polyhedrosis virus (NPV). The insects feeding on birch leaves had the lowest sensitivity to NPV than those on willow leaves, but the insects growing on pine needles showed the highest susceptibility. The sensitivity of the gypsy moths on willow leaves was higher than that of the gypsy moths on birch leaves and lower than that of those on pine needles. At the same time, it did not differ from that of the caterpillars on artificial feeding. The virus polyhedrons formed in the caterpillars on birch or willow leaves were more than those on another fodder.

  19. Host Protein Moloney Leukemia Virus 10 (MOV10) Acts as a Restriction Factor of Influenza A Virus by Inhibiting the Nuclear Import of the Viral Nucleoprotein

    PubMed Central

    Zhang, Junsong; Huang, Feng; Tan, Likai; Bai, Chuan; Chen, Bing; Liu, Jun; Liang, Juanran; Liu, Chao; Zhang, Shaoying; Lu, Gen; Chen, Yuan

    2016-01-01

    ABSTRACT The viral ribonucleoprotein (vRNP) complex of influenza A viruses (IAVs) contains an RNA-dependent RNA polymerase complex (RdRp) and nucleoprotein (NP) and is the functional unit for viral RNA transcription and replication. The vRNP complex is an important determinant of virus pathogenicity and host adaptation, implying that its function can be affected by host factors. In our study, we identified host protein Moloney leukemia virus 10 (MOV10) as an inhibitor of IAV replication, since depletion of MOV10 resulted in a significant increase in virus yield. MOV10 inhibited the polymerase activity in a minigenome system through RNA-mediated interaction with the NP subunit of vRNP complex. Importantly, we found that the interaction between MOV10 and NP prevented the binding of NP to importin-α, resulting in the retention of NP in the cytoplasm. Both the binding of MOV10 to NP and its inhibitory effect on polymerase activity were independent of its helicase activity. These results suggest that MOV10 acts as an anti-influenza virus factor through specifically inhibiting the nuclear transportation of NP and subsequently inhibiting the function of the vRNP complex. IMPORTANCE The interaction between the influenza virus vRNP complex and host factors is a major determinant of viral tropism and pathogenicity. Our study identified MOV10 as a novel host restriction factor for the influenza virus life cycle since it inhibited the viral growth rate. Conversely, importin-α has been shown as a determinant for influenza tropism and a positive regulator for viral polymerase activity in mammalian cells but not in avian cells. MOV10 disrupted the interaction between NP and importin-α, suggesting that MOV10 could also be an important host factor for influenza virus transmission and pathogenicity. Importantly, as an interferon (IFN)-inducible protein, MOV10 exerted a novel mechanism for IFNs to inhibit the replication of influenza viruses. Furthermore, our study potentially

  20. Simulated acid rain reduces the susceptibility of the European pine sawfly (Neodiprion sertifer) to its nuclear polyhedrosis virus.

    PubMed

    Neuvonen, S; Saikkonen, K; Haukioja, E

    1990-06-01

    The study dealt with the effect of simulated acid rain (both H(2)SO(4) and HNO(3); acidities of pH 4 and pH 3) on the susceptibility of the larvae of Neodiprion sertifer to its nuclear polyhedrosis virus. Scots pines growing in a subarctic area with low ambient pollution levels were irrigated with simulated acid rain during two summers. Neodiprion larvae fed with foliage from the experimental trees were infected with a dilute virus suspension. The acid treatment of host trees had a significant effect on the proportion of virus-treated larvae alive 16 days after the virus application: there were almost no differences between the controls and the pH 4 irrigation group, but on the needles of pH 3-treated trees larval survival was twice as high as with other treatments. The direct spraying of acid water on the needles before they were fed to the larvae did not significantly affect the survival of virus infected larvae. Our results suggest that acid rain may reduce the susceptibility of Neodiprion larvae to virus disease via changes in the quality of pine foliage.

  1. The ORF012 Gene of Marek's Disease Virus Type 1 Produces a Spliced Transcript and Encodes a Novel Nuclear Phosphoprotein Essential for Virus Growth

    PubMed Central

    Schippers, Timo; Jarosinski, Keith

    2014-01-01

    ABSTRACT Marek's disease virus (MDV), an alphaherpesvirus, is the causative agent of a lethal disease in chickens characterized by generalized nerve inflammation and rapid lymphoma development. The extensive colinearity of the MDV genome with those of related herpesviruses has eased functional characterization of many MDV genes. However, MDV carries a number of unique open reading frames (ORFs) that have not yet been investigated regarding their coding potentials and the functions of their products. Among these unique ORFs are two putative ORFs, ORF011 and ORF012, which are found at the extreme left end of the MDV unique long region. Using reverse transcriptase PCR, we showed that ORF011 and ORF012 are not individual genes but form a single gene through mRNA splicing of a small intron, resulting in the novel ORF012. We generated an ORF012-null virus using an infectious clone of MDV strain RB-1B. The deletion virus had a marked growth defect in vitro and could not be passaged in cultured cells, suggesting an essential role for the ORF012 product in virus replication. Further studies revealed that protein 012 (p012) localized to the nucleus in transfected and infected cells, and we identified by site-directed mutagenesis and green fluorescent protein (GFP) reporter fusion assays a nuclear localization signal (NLS) that was mapped to a 23-amino-acid sequence at the protein's C terminus. Nuclear export was blocked using leptomycin B, suggesting a potential role for p012 as a nuclear/cytoplasmic shuttling protein. Finally, p012 is phosphorylated at multiple residues, a modification that could possibly regulate its subcellular distribution. IMPORTANCE Marek's disease virus (MDV) causes a devastating oncogenic disease in chickens with high morbidity and mortality. The costs for disease prevention reach several billion dollars annually. The functional investigation of MDV genes is necessary to understand its complex replication cycle, which eventually could help us to

  2. Transfection of influenza A virus nuclear export protein induces the expression of tumor necrosis factor alpha.

    PubMed

    Lara-Sampablo, Alejandra; Flores-Alonso, Juan Carlos; De Jesús-Ortega, Nereyda; Santos-López, Gerardo; Vallejo-Ruiz, Verónica; Rosas-Murrieta, Nora; Reyes-Carmona, Sandra; Herrera-Camacho, Irma; Reyes-Leyva, Julio

    2014-06-24

    Influenza A virus genomic segments eight codes for non-structural 1 (NS1) protein that is involved in evasion of innate antiviral response, and nuclear export protein (NEP) that participates in the export of viral ribonucleoprotein (RNP) complexes, transcription and replication. Tumor necrosis factor alpha (TNF-α) is highly expressed during influenza virus infections and is considered an anti-infective cytokine. NS1 and NEP proteins were overexpressed and their role on TNF-α expression was evaluated. Both TNF-α mRNA and protein increased in cells transfected with NEP but not with NS1. We further investigate if NS1 or NEP regulates the activity of TNF-α promoter. In the presence of NEP the activity of TNF-α promoter increased significantly compared with the control (83.5±2.9 vs. 30.9±2.8, respectively; p=0.001). This effect decreased 15-fold when the TNF-α promoter distal region was deleted, suggesting the involvement of mitogen-activated protein kinases (MAPK) and NF-kB response elements. This was corroborated by testing the effect produced on TNF-α promoter by the treatment with Raf/MEK/ERK (U0126), NF-kB (Bay-11-7082) and PI3K (Ly294-002) cell signaling inhibitors. Treatment with U0126 and Bay-117082 reduced the activity of TNF-α promoter mediated by NEP (41.5±3.2, 70% inhibition; and 80.6±7.4, 35% inhibition, respectively) compared to mock-treated control. The results suggest a new role for NEP protein that participates in the transcriptional regulation of human TNF-α expression.

  3. Expression of the IE1 transactivator of Autographa californica nuclear polyhedrosis virus during viral infection.

    PubMed

    Choi, J; Guarino, L A

    1995-05-10

    The immediate-early IE1 protein of Autographa californica nuclear polyhedrosis virus (AcMNPV) is an important regulator of viral gene transcription. To provide a tool for further analysis of the expression and function of IE1, a polyclonal antiserum was raised against IE1 expressed in bacteria. Immunoblot analysis of infected cell lysates was used to monitor the accumulation of IE1 throughout the viral life cycle. When extracts were prepared in the presence of phosphatase inhibitors, only one protein band was detected on SDS-polyacrylamide gels. However, in the absence of phosphatase inhibitors, at least four distinct electrophoretic species were detected. Mobility shift assays were conducted using an enhancer DNA probe and whole cell extracts prepared at different times postinfection. Results indicated that the enhancer-binding activity of IE1 increased from 4 to 72 hr postinfection. DNA-protein complexes formed with infected cell extracts migrated more slowly than those formed with transfected cell extracts. This effect was more pronounced with extracts prepared in the presence of phosphatase inhibitors. Supershift experiments with IE1 antiserum confirmed that IE1 was a component of DNA-protein complexes in both transfected and infected cell extracts. A titration experiment was done to determine the minimal amounts of IE1 required for activation of the 39k promoter in the presence and absence of a cis-linked enhancer element. These analyses indicated that the intracellular levels of IE1 are not sufficient for enhancer-independent activation of the 39k promoter during the early phase of viral infection. Quantitative immunoblots revealed that the amount of IE1 in budded virus was less than 0.68 mole per mole of viral DNA, suggesting that IE1 is not a structural protein of AcNPV.

  4. Hepatitis C virus suppresses Hepatocyte Nuclear Factor 4 alpha, a key regulator of hepatocellular carcinoma.

    PubMed

    Vallianou, Ioanna; Dafou, Dimitra; Vassilaki, Niki; Mavromara, Penelope; Hadzopoulou-Cladaras, Margarita

    2016-09-01

    Hepatitis C Virus (HCV) infection presents with a disturbed lipid profile and can evolve to hepatic steatosis and hepatocellular carcinoma (HCC). Hepatocyte Nuclear Factor 4 alpha (HNF4α) is the most abundant transcription factor in the liver, a key regulator of hepatic lipid metabolism and a critical determinant of Epithelial to Mesenchymal Transition and hepatic development. We have previously shown that transient inhibition of HNF4α initiates transformation of immortalized hepatocytes through a feedback loop consisting of miR-24, IL6 receptor (IL6R), STAT3, miR-124 and miR-629, suggesting a central role of HNF4α in HCC. However, the role of HNF4α in Hepatitis C Virus (HCV)-related hepatocarcinoma has not been evaluated and remains controversial. In this study, we provide strong evidence suggesting that HCV downregulates HNF4α expression at both transcriptional and translational levels. The observed decrease of HNF4α expression correlated with the downregulation of its downstream targets, HNF1α and MTP. Ectopic overexpression of HCV proteins also exhibited an inhibitory effect on HNF4α levels. The inhibition of HNF4α expression by HCV appeared to be mediated at transcriptional level as HCV proteins suppressed HNF4α gene promoter activity. HCV also up-regulated IL6R, activated STAT3 protein phosphorylation and altered the expression of acute phase genes. Furthermore, as HCV triggered the loss of HNF4α a consequent change of miR-24, miR-629 or miR-124 was observed. Our findings demonstrated that HCV-related HCC could be mediated through HNF4α-microRNA deregulation implying a possible role of HNF4α in HCV hepatocarcinogenesis. HCV inhibition of HNF4α could be sustained to promote HCC. PMID:27477312

  5. Early, Active, and Specific Localization of Herpes Simplex Virus Type 1 gM to Nuclear Membranes▿

    PubMed Central

    Zhang, Jie; Nagel, Claus-Henning; Sodeik, Beate; Lippé, Roger

    2009-01-01

    Thirteen different glycoproteins are incorporated into mature herpes simplex virus type 1 (HSV-1) virions. Five of them play important roles during entry, while others intervene during egress of the virus. Although HSV-1 gM is not essential in cell culture, its deletion reduces viral yields and promotes syncytium formation. Furthermore, gM is conserved among herpesviruses, is essential for several of them, and can redirect the gD and gH/gL viral glycoproteins from the cell surface to the trans-Golgi network, where gM presumably modulates final capsid envelopment. Late in infection, gM reaches the nuclear envelope and decorates perinuclear virions. This process seemingly requires UL31 and UL34 and occurs when several markers of the trans-Golgi network have relocalized to the nucleus. However, the precise mechanism of gM nuclear targeting is unclear. We now report that gM is quickly and specifically targeted to nuclear membranes in a virus-dependent manner. This occurs prior to the HSV-1-induced reorganization of the trans-Golgi network and before gM enters the secretory pathway. The presence of a high-mannose glycosylation pattern on gM further corroborated these findings. While gM was targeted to the inner nuclear membrane early in infection, its partners gD, gH, gN, VP22, UL31, and UL34 did not colocalize with gM. These data suggest that nuclear gM fulfills an early nuclear function that is independent of its known interaction partners and its function in viral egress. PMID:19812164

  6. Characterization of baculovirus Autographa californica multiple nuclear polyhedrosis virus infection in mammalian cells.

    PubMed

    Kitajima, Masayuki; Hamazaki, Hiroyuki; Miyano-Kurosaki, Naoko; Takaku, Hiroshi

    2006-05-01

    The baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) is used as a vector in many gene therapy studies. Wild-type AcMNPV infects many mammalian cell types in vitro, but does not replicate. We investigated the dynamics of AcMNPV genomic DNA in infected mammalian cells and used flow cytometric analysis to demonstrate that recombinant baculovirus containing a cytomegalovirus immediate early promoter/enhancer with green fluorescent protein (GFP) expressed high levels of GFP in Huh-7 cells, but not B16, Raw264.7, or YAC-1 cells. The addition of butyrate, a deacetylase inhibitor, markedly enhanced the percentage of GFP-expressing Huh-7 and B16 cells, but not Raw264.7 and YAC-1 cells. The addition of 5-aza-2'-deoxycytidine, a DNA methylation inhibitor, had no enhancing effect. Polymerase chain reaction analysis using AcMNPV-gp64-specific primers indicated that AcMNPV infected not only Huh-7 and B16 cells, but also Raw264.7 and YAC-1 cells in vitro. The genomic DNA was detected in Huh-7 and B16 cells 96 h after infection. Genomic AcMNPV DNA in YAC-1 cells was not transported to the nucleus. Luciferase assay indicated that AcMNPV p35 gene mRNA and p35 promoter activity were clearly expressed only in Huh-7 and B16 cells. These results suggest that viral genomic DNA expression is restricted by different host cell factors, such as degradation, deacetylation, and inhibition of nuclear transport, depending on the mammalian cell type. PMID:16545777

  7. Inhibition of hepatitis B virus (HBV) by LNA-mediated nuclear interference with HBV DNA transcription

    SciTech Connect

    Sun, Zhen; Xiang, Wenqing; Guo, Yajuan; Chen, Zhi; Liu, Wei; Lu, Daru

    2011-06-10

    Highlights: {yields} LNA-modified oligonucleotides can pass through the plasma membrane of cultured cells even without using transfection machinery. {yields} LNA-modified oligonucleotides passed efficiently across the cell membrane, and lipid-coating facilitated translocation from the cytoplasm to the nucleus. {yields} LNA-oligonucleotide designed to target nuclear HBV DNA efficiently suppresses HBV replication and transcription in cultured hepatic cells. -- Abstract: Silencing target genes with small regulatory RNAs is widely used to investigate gene function and therapeutic drug development. Recently, triplex-based approaches have provided another attractive means to achieve targeted gene regulation and gene manipulation at the molecular and cellular levels. Nuclear entry of oligonucleotides and enhancement of their affinity to the DNA targets are key points of such approaches. In this study, we developed lipid-based transport of a locked-nucleic-acid (LNA)-modified oligonucleotide for hepatitis B virus (HBV) DNA interference in human hepatocytes expressing HBV genomic DNA. In these cells, the LNA-modified oligonucleotides passed efficiently across the cell membrane, and lipid-coating facilitated translocation from the cytoplasm to the nucleus. The oligonucleotide specifically targeting HBV DNA clearly interfered with HBV DNA transcription as shown by a block in pregenomic RNA (pgRNA) production. The HBV DNA-targeted oligonucleotide suppressed HBV DNA replication and HBV protein production more efficiently than small interfering RNAs directed to the pgRNA. These results demonstrate that fusion with lipid can carry LNA-modified oligonucleotides to the nucleus where they regulate gene expression. Interfering with HBV DNA transcription by LNA-modified oligonucleotides has strong potential as a new strategy for HBV inhibition.

  8. Characterization of baculovirus Autographa californica multiple nuclear polyhedrosis virus infection in mammalian cells

    SciTech Connect

    Kitajima, Masayuki; Hamazaki, Hiroyuki; Miyano-Kurosaki, Naoko; Takaku, Hiroshi . E-mail: hiroshi.takaku@it-chiba.ac.jp

    2006-05-05

    The baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) is used as a vector in many gene therapy studies. Wild-type AcMNPV infects many mammalian cell types in vitro, but does not replicate. We investigated the dynamics of AcMNPV genomic DNA in infected mammalian cells and used flow cytometric analysis to demonstrate that recombinant baculovirus containing a cytomegalovirus immediate early promoter/enhancer with green fluorescent protein (GFP) expressed high levels of GFP in Huh-7 cells, but not B16, Raw264.7, or YAC-1 cells. The addition of butyrate, a deacetylase inhibitor, markedly enhanced the percentage of GFP-expressing Huh-7 and B16 cells, but not Raw264.7 and YAC-1 cells. The addition of 5-aza-2'-deoxycytidine, a DNA methylation inhibitor, had no enhancing effect. Polymerase chain reaction analysis using AcMNPV-gp64-specific primers indicated that AcMNPV infected not only Huh-7 and B16 cells, but also Raw264.7 and YAC-1 cells in vitro. The genomic DNA was detected in Huh-7 and B16 cells 96 h after infection. Genomic AcMNPV DNA in YAC-1 cells was not transported to the nucleus. Luciferase assay indicated that AcMNPV p35 gene mRNA and p35 promoter activity were clearly expressed only in Huh-7 and B16 cells. These results suggest that viral genomic DNA expression is restricted by different host cell factors, such as degradation, deacetylation, and inhibition of nuclear transport, depending on the mammalian cell type.

  9. The bovine immunodeficiency virus Rev protein: identification of a novel nuclear import pathway and nuclear export signal among retroviral Rev/Rev-like proteins.

    PubMed

    Gomez Corredor, Andrea; Archambault, Denis

    2012-05-01

    The Rev protein is essential for the replication of lentiviruses. Rev is a shuttling protein that transports unspliced and partially spliced lentiviral RNAs from the nucleus to the cytoplasm via the nucleopore. To transport these RNAs, the human immunodeficiency virus type 1 (HIV-1) Rev uses the karyopherin β family importin β and CRM1 proteins that interact with the Rev nuclear localization signal (NLS) and nuclear exportation signal (NES), respectively. Recently, we reported the presence of new types of bipartite NLS and nucleolar localization signal (NoLS) in the bovine immunodeficiency virus (BIV) Rev protein. Here we report the characterization of the nuclear import and export pathways of BIV Rev. By using an in vitro nuclear import assay, we showed that BIV Rev is transported into the nucleus by a cytosolic and energy-dependent importin α/β classical pathway. Results from glutathione S-transferase (GST) pulldown assays that showed the binding of BIV Rev with importins α3 and α5 were in agreement with those from the nuclear import assay. We also identified a leptomycin B-sensitive NES in BIV Rev, which indicates that the protein is exported via CRM1 like HIV-1 Rev. Mutagenesis experiments showed that the BIV Rev NES maps between amino acids 109 to 121 of the protein. Remarkably, the BIV Rev NES was found to be of the cyclic AMP (cAMP)-dependent protein kinase inhibitor (PKI) type instead of the HIV-1 Rev type. In summary, our data showed that the nuclear import mechanism of BIV Rev is novel among Rev proteins characterized so far in lentiviruses.

  10. Evidence for an inhibitory feedback loop regulating simian virus 40 large T-antigen fusion protein nuclear transport.

    PubMed Central

    Seydel, U; Jans, D A

    1996-01-01

    Nuclear protein import is central to eukaryotic cell function. It is dependent on ATP, temperature and cytosolic factors, and requires specific targeting sequences called nuclear localization signals (NLSs). Nuclear import kinetics was studied in vitro using digitonin-permeabilized cells of the HTC rat hepatoma cell line and a fluorescently labelled beta-galactosidase fusion protein carrying amino acids 111-135 of the simian virus 40 large T-antigen (T-ag), including the NLS. Nuclear accumulation was rapid, reaching steady-state after about 80 min at 37 degrees C (t1/2 at about 17 min). Surprisingly, maximal nuclear concentration was found to be directly proportional to the concentration of the cytosolic extract and of cytoplasmic T-ag protein. Neither preincubation of cells for 1 h at 37 degrees C before the addition of T-ag protein nor the addition of fresh transport medium after 1 h and continuation of the incubation for another hour affected the maximal nuclear concentration. If cells were allowed to accumulate T-ag protein for 1 h before the addition of fresh transport medium containing different concentrations of T-ag protein and incubated for a further hour, the maximal nuclear concentration did not change unless the concentration of T-ag protein in the second transport mixture exceeded that in the first, in which case the nuclear concentration increased. Nuclear import of T-ag thus appeared (i) to be strictly unidirectional over 2 h at 37 degrees C and (ii) to be regulated by an inhibitory feedback loop, whereby the cytosolic concentration of protein appears to determine directly the precise end point of nuclear accumulation. This study represents the first characterization of this previously undescribed mechanism of regulation of nuclear protein import. PMID:8670127

  11. Ribosome Protein L4 is essential for Epstein–Barr Virus Nuclear Antigen 1 function

    PubMed Central

    Shen, Chih-Lung; Liu, Cheng-Der; You, Ren-In; Ching, Yung-Hao; Liang, Jun; Ke, Liangru; Chen, Ya-Lin; Chen, Hong-Chi; Hsu, Hao-Jen; Liou, Je-Wen; Kieff, Elliott; Peng, Chih-Wen

    2016-01-01

    Epstein–Barr Virus (EBV) Nuclear Antigen 1 (EBNA1)-mediated origin of plasmid replication (oriP) DNA episome maintenance is essential for EBV-mediated tumorigenesis. We have now found that EBNA1 binds to Ribosome Protein L4 (RPL4). RPL4 shRNA knockdown decreased EBNA1 activation of an oriP luciferase reporter, EBNA1 DNA binding in lymphoblastoid cell lines, and EBV genome number per lymphoblastoid cell line. EBV infection increased RPL4 expression and redistributed RPL4 to cell nuclei. RPL4 and Nucleolin (NCL) were a scaffold for an EBNA1-induced oriP complex. The RPL4 N terminus cooperated with NCL-K429 to support EBNA1 and oriP-mediated episome binding and maintenance, whereas the NCL C-terminal K380 and K393 induced oriP DNA H3K4me2 modification and promoted EBNA1 activation of oriP-dependent transcription. These observations provide new insights into the mechanisms by which EBV uses NCL and RPL4 to establish persistent B-lymphoblastoid cell infection. PMID:26858444

  12. Identifying the RNA polymerases that synthesize specific transcripts of the Autographa californica nuclear polyhedrosis virus.

    PubMed

    Huh, N E; Weaver, R F

    1990-01-01

    Nuclear run-on assays carried out in the presence and absence of the RNA polymerase II inhibitor, alpha-amanitin, were used to determine the exact timing of the switch from inhibitor-sensitive transcription catalysed by host RNA polymerase II, to inhibitor-resistant transcription catalysed by the baculovirus-induced RNA polymerase. These studies revealed that the onset of alpha-amanitin-resistant transcription is just after 6 h post-infection, simultaneous with the beginning of the late phase of infection. They also showed that transcripts from the p26 gene in the HindIII Q/P region and the p35 gene in the HindIII K/Q region of the viral genome are synthesized by the host RNA polymerase II both early and late in infection. On the other hand, transcripts of the p10 gene in the HindIII Q/P region and the gamma transcripts in the HindIII K region are synthesized by the alpha-amanitin-resistant, virus-induced RNA polymerase late in infection. PMID:2106003

  13. Direct effects of recombinant nuclear polyhedrosis viruses on selected nontarget organisms.

    PubMed

    Heinz, K M; McCutchen, B F; Herrmann, R; Parrella, M P; Hammock, B D

    1995-04-01

    A limitation to effective field use of naturally occurring nuclear polyhedrosis viruses (NPVs) is the slow rate at which they kill their host. In making NPVs a more attractive pest management tool, this problem has been addressed by modifying NPVs genetically to express insecticidal proteins resulting in substantial increases in their speed of action. One concern associated with these recombinant NPVs, however, is their effects on nontarget insects associated with pests targeted for control by applications of NPVs. Our studies evaluated the direct effects of wild-type Autographa californica NPV (AcNPV) and a recombinant AcNPV (AcAaIT) on three insects beneficial to production agriculture. The recombinant NPV expresses an insect-selective neurotoxin, AaIT, which was isolated from the scorpion, Androctonus australis Hector. Two generalist predators, Chysoperla carnea Stephens and Orius insidiosus (Say), were not adversely affected by feeding on larvae of Heliothis virescens (F.) infected with AcAaIT. Similarly, no adverse effects were detected in the honey bee, Apis mellifera L., when injected with wild-type or recombinant NPVs. Results from this study may provide a foundation upon which potential risks associated with genetically engineered NPVs may be evaluated on a limited scale in greenhouse or field experiments. PMID:7722081

  14. Modulation of Epstein–Barr Virus Nuclear Antigen 2-dependent transcription by protein arginine methyltransferase 5

    SciTech Connect

    Liu, Cheng-Der; Cheng, Chi-Ping; Fang, Jia-Shih; Chen, Ling-Chih; Zhao, Bo; Kieff, Elliott; Peng, Chih-Wen

    2013-01-18

    Highlights: ► Catalytic active PRMT5 substantially binds to the EBNA2 RG domain. ► PRMT5 augments the EBNA2-dependent transcription. ► PRMT5 triggers the symmetric dimethylation of the EBNA2 RG domain. ► PRMT5 enhances the promoter occupancy of EBNA2 on its target promoters. -- Abstract: Epstein–Barr Virus Nuclear Antigen (EBNA) 2 features an Arginine–Glycine repeat (RG) domain at amino acid positions 335–360, which is a known target for protein arginine methyltransferaser 5 (PRMT5). In this study, we performed protein affinity pull-down assays to demonstrate that endogenous PRMT5 derived from lymphoblastoid cells specifically associated with the protein bait GST-E2 RG. Transfection of a plasmid expressing PRMT5 induced a 2.5- to 3-fold increase in EBNA2-dependent transcription of both the LMP1 promoter in AKATA cells, which contain the EBV genome endogenously, and a Cp-Luc reporter plasmid in BJAB cells, which are EBV negative. Furthermore, we showed that there was a 2-fold enrichment of EBNA2 occupancy in target promoters in the presence of exogenous PRMT5. Taken together, we show that PRMT5 triggers the symmetric dimethylation of EBNA2 RG domain to coordinate with EBNA2-mediated transcription. This modulation suggests that PRMT5 may play a role in latent EBV infection.

  15. Multiple early transcripts and splicing of the Autographa californica nuclear polyhedrosis virus IE-1 gene.

    PubMed Central

    Chisholm, G E; Henner, D J

    1988-01-01

    The immediate-early IE-1 gene of Autographa californica nuclear polyhedrosis virus was cloned, and its nucleotide sequence was determined. Sequence analysis indicated that this gene would encode a protein of 582 amino acids with a predicted molecular weight of 66,822. Analysis of IE-1 gene expression during baculovirus infection identified two transcripts. One, 1.9 kilobases (kb), was expressed at constant steady-state levels throughout infection, whereas the other, 2.1 kb, was expressed only early in infection. Analysis of IE-1 cDNA clones demonstrated that the 2.1-kb transcript contained the entire 1.9-kb transcript (exon 1) plus an additional 5' end (exon 0). Genomic Southern analysis placed the exon 0 sequences on the EcoRI B fragment, 4 kilobase pairs upstream of exon 1. Sequencing of the upstream region identified an open reading frame whose 5' end was identical to the exon 0 sequences in the cDNAs. Examination of the genomic DNA sequences around the exon-exon junction revealed sequences similar to published consensus splice acceptor and donor sequences. This is the first example of splicing of any viral transcript during baculovirus infection. Images PMID:3043024

  16. Nuclear Trafficking of the Rabies Virus Interferon Antagonist P-Protein Is Regulated by an Importin-Binding Nuclear Localization Sequence in the C-Terminal Domain.

    PubMed

    Rowe, Caitlin L; Wagstaff, Kylie M; Oksayan, Sibil; Glover, Dominic J; Jans, David A; Moseley, Gregory W

    2016-01-01

    Rabies virus P-protein is expressed as five isoforms (P1-P5) which undergo nucleocytoplasmic trafficking important to roles in immune evasion. Although nuclear import of P3 is known to be mediated by an importin (IMP)-recognised nuclear localization sequence in the N-terminal region (N-NLS), the mechanisms underlying nuclear import of other P isoforms in which the N-NLS is inactive or has been deleted have remained unresolved. Based on the previous observation that mutation of basic residues K214/R260 of the P-protein C-terminal domain (P-CTD) can result in nuclear exclusion of P3, we used live cell imaging, protein interaction analysis and in vitro nuclear transport assays to examine in detail the nuclear trafficking properties of this domain. We find that the effect of mutation of K214/R260 on P3 is largely dependent on nuclear export, suggesting that nuclear exclusion of mutated P3 involves the P-CTD-localized nuclear export sequence (C-NES). However, assays using cells in which nuclear export is pharmacologically inhibited indicate that these mutations significantly inhibit P3 nuclear accumulation and, importantly, prevent nuclear accumulation of P1, suggestive of effects on NLS-mediated import activity in these isoforms. Consistent with this, molecular binding and transport assays indicate that the P-CTD mediates IMPα2/IMPβ1-dependent nuclear import by conferring direct binding to the IMPα2/IMPβ1 heterodimer, as well as to a truncated form of IMPα2 lacking the IMPβ-binding autoinhibitory domain (ΔIBB-IMPα2), and IMPβ1 alone. These properties are all dependent on K214 and R260. This provides the first evidence that P-CTD contains a genuine IMP-binding NLS, and establishes the mechanism by which P-protein isoforms other than P3 can be imported to the nucleus. These data underpin a refined model for P-protein trafficking that involves the concerted action of multiple NESs and IMP-binding NLSs, and highlight the intricate regulation of P

  17. Nuclear Trafficking of the Rabies Virus Interferon Antagonist P-Protein Is Regulated by an Importin-Binding Nuclear Localization Sequence in the C-Terminal Domain

    PubMed Central

    Rowe, Caitlin L.; Wagstaff, Kylie M.; Oksayan, Sibil; Glover, Dominic J.

    2016-01-01

    Rabies virus P-protein is expressed as five isoforms (P1-P5) which undergo nucleocytoplasmic trafficking important to roles in immune evasion. Although nuclear import of P3 is known to be mediated by an importin (IMP)-recognised nuclear localization sequence in the N-terminal region (N-NLS), the mechanisms underlying nuclear import of other P isoforms in which the N-NLS is inactive or has been deleted have remained unresolved. Based on the previous observation that mutation of basic residues K214/R260 of the P-protein C-terminal domain (P-CTD) can result in nuclear exclusion of P3, we used live cell imaging, protein interaction analysis and in vitro nuclear transport assays to examine in detail the nuclear trafficking properties of this domain. We find that the effect of mutation of K214/R260 on P3 is largely dependent on nuclear export, suggesting that nuclear exclusion of mutated P3 involves the P-CTD-localized nuclear export sequence (C-NES). However, assays using cells in which nuclear export is pharmacologically inhibited indicate that these mutations significantly inhibit P3 nuclear accumulation and, importantly, prevent nuclear accumulation of P1, suggestive of effects on NLS-mediated import activity in these isoforms. Consistent with this, molecular binding and transport assays indicate that the P-CTD mediates IMPα2/IMPβ1-dependent nuclear import by conferring direct binding to the IMPα2/IMPβ1 heterodimer, as well as to a truncated form of IMPα2 lacking the IMPβ-binding autoinhibitory domain (ΔIBB-IMPα2), and IMPβ1 alone. These properties are all dependent on K214 and R260. This provides the first evidence that P-CTD contains a genuine IMP-binding NLS, and establishes the mechanism by which P-protein isoforms other than P3 can be imported to the nucleus. These data underpin a refined model for P-protein trafficking that involves the concerted action of multiple NESs and IMP-binding NLSs, and highlight the intricate regulation of P

  18. Nuclear Trafficking of the Rabies Virus Interferon Antagonist P-Protein Is Regulated by an Importin-Binding Nuclear Localization Sequence in the C-Terminal Domain.

    PubMed

    Rowe, Caitlin L; Wagstaff, Kylie M; Oksayan, Sibil; Glover, Dominic J; Jans, David A; Moseley, Gregory W

    2016-01-01

    Rabies virus P-protein is expressed as five isoforms (P1-P5) which undergo nucleocytoplasmic trafficking important to roles in immune evasion. Although nuclear import of P3 is known to be mediated by an importin (IMP)-recognised nuclear localization sequence in the N-terminal region (N-NLS), the mechanisms underlying nuclear import of other P isoforms in which the N-NLS is inactive or has been deleted have remained unresolved. Based on the previous observation that mutation of basic residues K214/R260 of the P-protein C-terminal domain (P-CTD) can result in nuclear exclusion of P3, we used live cell imaging, protein interaction analysis and in vitro nuclear transport assays to examine in detail the nuclear trafficking properties of this domain. We find that the effect of mutation of K214/R260 on P3 is largely dependent on nuclear export, suggesting that nuclear exclusion of mutated P3 involves the P-CTD-localized nuclear export sequence (C-NES). However, assays using cells in which nuclear export is pharmacologically inhibited indicate that these mutations significantly inhibit P3 nuclear accumulation and, importantly, prevent nuclear accumulation of P1, suggestive of effects on NLS-mediated import activity in these isoforms. Consistent with this, molecular binding and transport assays indicate that the P-CTD mediates IMPα2/IMPβ1-dependent nuclear import by conferring direct binding to the IMPα2/IMPβ1 heterodimer, as well as to a truncated form of IMPα2 lacking the IMPβ-binding autoinhibitory domain (ΔIBB-IMPα2), and IMPβ1 alone. These properties are all dependent on K214 and R260. This provides the first evidence that P-CTD contains a genuine IMP-binding NLS, and establishes the mechanism by which P-protein isoforms other than P3 can be imported to the nucleus. These data underpin a refined model for P-protein trafficking that involves the concerted action of multiple NESs and IMP-binding NLSs, and highlight the intricate regulation of P

  19. A nuclear fraction of turnip crinkle virus capsid protein is important for elicitation of the host resistance response.

    PubMed

    Kang, Sung-Hwan; Qu, Feng; Morris, T Jack

    2015-12-01

    The N-terminal 25 amino acids (AAs) of turnip crinkle virus (TCV) capsid protein (CP) are recognized by the resistance protein HRT to trigger a hypersensitive response (HR) and systemic resistance to TCV infection. This same region of TCV CP also contains a motif that interacts with the transcription factor TIP, as well as a nuclear localization signal (NLS). However, it is not yet known whether nuclear localization of TCV CP is needed for the induction of HRT-mediated HR and resistance. Here we present new evidence suggesting a tight correlation between nuclear inclusions formed by CP and the manifestation of HR. We show that a fraction of TCV CP localized to cell nuclei to form discrete inclusion-like structures, and a mutated CP (R6A) known to abolish HR failed to form nuclear inclusions. Notably, TIP-CP interaction augments the inclusion-forming activity of CP by tethering inclusions to the nuclear membrane. This TIP-mediated augmentation is also critical for HR resistance, as another CP mutant (R8A) known to elicit a less restrictive HR, though still self-associated into nuclear inclusions, failed to direct inclusions to the nuclear membrane due to its inability to interact with TIP. Finally, exclusion of CP from cell nuclei abolished induction of HR. Together, these results uncovered a strong correlation between nuclear localization and nuclear inclusion formation by TCV CP and induction of HR, and suggest that CP nuclear inclusions could be the key trigger of the HRT-dependent, yet TIP-reinforced, resistance to TCV.

  20. A nuclear fraction of turnip crinkle virus capsid protein is important for elicitation of the host resistance response.

    PubMed

    Kang, Sung-Hwan; Qu, Feng; Morris, T Jack

    2015-12-01

    The N-terminal 25 amino acids (AAs) of turnip crinkle virus (TCV) capsid protein (CP) are recognized by the resistance protein HRT to trigger a hypersensitive response (HR) and systemic resistance to TCV infection. This same region of TCV CP also contains a motif that interacts with the transcription factor TIP, as well as a nuclear localization signal (NLS). However, it is not yet known whether nuclear localization of TCV CP is needed for the induction of HRT-mediated HR and resistance. Here we present new evidence suggesting a tight correlation between nuclear inclusions formed by CP and the manifestation of HR. We show that a fraction of TCV CP localized to cell nuclei to form discrete inclusion-like structures, and a mutated CP (R6A) known to abolish HR failed to form nuclear inclusions. Notably, TIP-CP interaction augments the inclusion-forming activity of CP by tethering inclusions to the nuclear membrane. This TIP-mediated augmentation is also critical for HR resistance, as another CP mutant (R8A) known to elicit a less restrictive HR, though still self-associated into nuclear inclusions, failed to direct inclusions to the nuclear membrane due to its inability to interact with TIP. Finally, exclusion of CP from cell nuclei abolished induction of HR. Together, these results uncovered a strong correlation between nuclear localization and nuclear inclusion formation by TCV CP and induction of HR, and suggest that CP nuclear inclusions could be the key trigger of the HRT-dependent, yet TIP-reinforced, resistance to TCV. PMID:26299399

  1. Fusion between perinuclear virions and the outer nuclear membrane requires the fusogenic activity of herpes simplex virus gB.

    PubMed

    Wright, Catherine C; Wisner, Todd W; Hannah, Brian P; Eisenberg, Roselyn J; Cohen, Gary H; Johnson, David C

    2009-11-01

    Herpesviruses cross nuclear membranes (NMs) in two steps, as follows: (i) capsids assemble and bud through the inner NM into the perinuclear space, producing enveloped virus particles, and (ii) the envelopes of these virus particles fuse with the outer NM. Two herpes simplex virus (HSV) glycoproteins, gB and gH (the latter, likely complexed as a heterodimer with gL), are necessary for the second step of this process. Mutants lacking both gB and gH accumulate in the perinuclear space or in herniations (membrane vesicles derived from the inner NM). Both gB and gH/gL are also known to act directly in fusing the virion envelope with host cell membranes during HSV entry into cells, i.e., both glycoproteins appear to function directly in different aspects of the membrane fusion process. We hypothesized that HSV gB and gH/gL also act directly in the membrane fusion that occurs during virus egress from the nucleus. Previous studies of the role of gB and gH/gL in nuclear egress involved HSV gB and gH null mutants that could potentially also possess gross defects in the virion envelope. Here, we produced recombinant HSV-expressing mutant forms of gB with single amino acid substitutions in the hydrophobic "fusion loops." These fusion loops are thought to play a direct role in membrane fusion by insertion into cellular membranes. HSV recombinants expressing gB with any one of four fusion loop mutations (W174R, W174Y, Y179K, and A261D) were unable to enter cells. Moreover, two of the mutants, W174Y and Y179K, displayed reduced abilities to mediate HSV cell-to-cell spread, and W174R and A261D exhibited no spread. All mutant viruses exhibited defects in nuclear egress, enveloped virions accumulated in herniations and in the perinuclear space, and fewer enveloped virions were detected on cell surfaces. These results support the hypothesis that gB functions directly to mediate the fusion between perinuclear virus particles and the outer NM.

  2. Nuclear import of prototype foamy virus transactivator Bel1 is mediated by KPNA1, KPNA6 and KPNA7

    PubMed Central

    Duan, Jihui; Tang, Zhiqin; Mu, Hong; Zhang, Guojun

    2016-01-01

    Bel1, a transactivator of the prototype foamy virus (PFV), plays pivotal roles in the replication of PFV. Previous studies have demonstrated that Bel1 bears a nuclear localization signal (NLS); however, its amino acid sequence remains unclear and the corresponding adaptor importins have not yet been identified. In this study, we inserted various fragments of Bel1 into an EGFP-GST fusion protein and investigated their subcellular localization by fluorescence microscopy. We found that the 215PRQKRPR221 fragment, which accords with the consensus sequence K(K/R)X(K/R) of the monopartite NLS, directed the nuclear translocation of Bel1. Point mutation experiments revealed that K218, R219 and R221 were essential for the nuclear localization of Bel1. The results of GST pull-down assay revealed that the Bel1 peptide 215-221, which bears the NLS, interacted with the nucleocytoplasmic transport receptors, karyopherin alpha 1 (importin alpha 5) (KPNA1), karyopherin alpha 6 (importin alpha 7) (KPNA6) and karyopherin alpha 7 (importin alpha 8) (KPNA7). Finally, in vitro nuclear import assays demonstrated that KPNA1, KPNA6 or KPNA7, along with other necessary nuclear factors, caused Bel1 to localize to the nucleus. Thus, the findings of our study indicate that KPNA1, KPNA6 and KPNA7 are involved in Bel1 nuclear distribution. PMID:27277550

  3. A mechanism for negative gene regulation in Autographa californica multinucleocapsid nuclear polyhedrosis virus

    USGS Publications Warehouse

    Leisy, D.J.; Rasmussen, C.; Owusu, E.O.; Rohrmann, G.F.

    1997-01-01

    The Autographa californica multinucleocapsid nuclear polyhedrosis virus (AcMNPV) ie-1 gene product (IE-1) is thought to play a central role in stimulating early viral transcription. IE-1 has been demonstrated to activate several early viral gene promoters and to negatively regulate the promoters of two other AcMNPV regulatory genes, ie-0 and ie-2. Our results indicate that IE-1 negatively regulates the expression of certain genes by binding directly, or as part of a complex, to promoter regions containing a specific IE-1-binding motif (5'-ACBYGTAA-3') near their mRNA start sites. The IE-1 binding motif was also found within the palindromic sequences of AcMNPV homologous repeat (hr) regions that have been shown to bind IE-1. The role of this IE-1 binding motif in the regulation of the ie-2 and pe-38 promoters was examined by introducing mutations in these promoters in which the central 6 bp were replaced with Bg/II sites. GUS reporter constructs containing ie-2 and pe-38 promoter fragments with and without these specific mutations were cotransfected into Sf9 cells with various amounts of an ie-1-containing plasmid (ple-1). Comparisons of GUS expression produced by the mutant and wild-type constructs demonstrated that the IE-1 binding motif mediated a significant decrease in expression from the ie-2 and pe-38 promoters in response to increasing pIe-1 concentrations. Electrophoretic mobility shift assays with pIe-1-transfected cell extracts and supershift assays with IE-1- specific antiserum demonstrated that IE-1 binds to promoter fragments containing the IE-1 binding motif but does not bind to promoter fragments lacking this motif.

  4. Adaptive mutation in nuclear export protein allows stable transgene expression in a chimaeric influenza A virus vector.

    PubMed

    Kuznetsova, Irina; Shurygina, Anna-Polina; Wolf, Brigitte; Wolschek, Markus; Enzmann, Florian; Sansyzbay, Abylay; Khairullin, Berik; Sandybayev, Nurlan; Stukova, Marina; Kiselev, Oleg; Egorov, Andrej; Bergmann, Michael

    2014-02-01

    The development of influenza virus vectors with long insertions of foreign sequences remains difficult due to the small size and instable nature of the virus. Here, we used the influenza virus inherent property of self-optimization to generate a vector stably expressing long transgenes from the NS1 protein ORF. This was achieved by continuous selection of bright fluorescent plaques of a GFP-expressing vector during multiple passages in mouse B16f1 cells. The newly generated vector acquired stability in IFN-competent cell lines and in vivo in murine lungs. Although improved vector fitness was associated with the appearance of four coding mutations in the polymerase (PB2), haemagglutinin and non-structural (NS) segments, the stability of the transgene expression was dependent primarily on the single mutation Q20R in the nuclear export protein (NEP). Importantly, a longer insert, such as a cassette of 1299 nt encoding two Mycobacterium tuberculosis Esat6 and Ag85A proteins, could substitute for the GFP transgene. Thus, the inherent property of the influenza virus to adapt can also be used to adjust a vector backbone to give stable expression of long transgenes. PMID:24222196

  5. Characterization of the nuclear export signal of human T-cell lymphotropic virus type 1 Rex reveals that nuclear export is mediated by position-variable hydrophobic interactions.

    PubMed Central

    Kim, F J; Beeche, A A; Hunter, J J; Chin, D J; Hope, T J

    1996-01-01

    We previously determined that amino acids 64 to 120 of human T-cell lymphotropic virus type 1 (HTLV-1) Rex can restore the function of an effector domain mutant of human immunodeficiency virus type 1 (HIV-1) Rev (T. J. Hope, B. L. Bond, D. McDonald, N. P. Klein, and T. G. Parslow, J. Virol. 65:6001-6007, 1991). In this report, we (i) identify and characterize a position-independent 17-amino-acid region of HTLV-1 Rex that fully complements HIV-1 Rev effector domain mutants and (ii) show that this 17-amino-acid region and specific hydrophobic substitutions can serve as nuclear export signals. Mutagenesis studies revealed that four leucines within the minimal region were essential for function. Alignment of the minimal Rex region with the HIV-1 Rev effector domain suggested that the position of some of the conserved leucines is flexible. We found two of the leucines could each occupy one of two positions within the context of the full-length HTLV-1 Rex protein and maintain function. The idea of flexibility within the Rex effector domain was confirmed and extended by identifying functional substitutions by screening a library of effector domain mutants in which the two regions of flexibility were randomized. Secondly, the functional roles of the minimal Rex effector domain and hydrophobic substitutions were independently confirmed by demonstrating that these effector domains could serve as nuclear export signals when conjugated with bovine serum albumin. Nuclear export of the wild-type Rex conjugates was temperature dependent and sensitive to wheat germ agglutinin and was blocked by a 20-fold excess of unlabeled conjugates. Together, these studies reveal that position-variable hydrophobic interactions within the HTLV-1 Rex effector domain mediate nuclear export function. PMID:8756672

  6. Intra-nuclear localization of two envelope proteins, gB and gD, of herpes simplex virus.

    PubMed

    Stannard, L M; Himmelhoch, S; Wynchank, S

    1996-01-01

    The envelopes of herpes simplex virus (HSV) particles are acquired from the inner nuclear membrane (INM) of the infected cell and virus-coded glycoproteins are present in the envelope of mature virions. Our ultrastructural study examined the process of virus envelopment and the targeting of two major viral glycoproteins, gB and gD, to the INM in HSV-infected human embryonic fibroblasts. It was shown that envelopment and transport of virus particles from the nucleus is facilitated by the formation of a dynamic tubulo-reticulum arising from the INM. Capsids were assembled in the nucleus and collected within INM tubules which protruded into the perinuclear space and thence into the cisternae of the endoplasmic reticulum (ER). Envelopment occurred by constriction and fusion of the tubular channel walls, releasing enveloped virions into the ER. Transport to the cell surface took place in membrane-bound compartments and probably followed the normal secretory pathway through the Golgi apparatus. Immunogold probes, tagged with specific monoclonal antibodies, were used to localize gB and gD during the process of virus maturation. Cytoplasmic membranes were not labelled, but probes bound inside the nucleus, mainly at sites of virus assembly. Labelling occurred on the nucleoplasmic side of the INM which surrounded capsids in the process of envelopment, but not on the outside of that membrane, although characteristic gB glycoprotein spikes were labelled on the envelopes of extracellular virus particles and on virions in trans-Golgi transport vesicles just prior to their release from the infected cell. gB was not detected on the surface of enveloped virions in the perinuclear space, or the cisternae of the ER or cis-Golgi, which suggests that the specific epitope was masked during that stage of intracellular processing. gD probes bound to virion envelopes and also to the tegument region of some particles found in both perinuclear and extracellular sites. We postulate the

  7. Nuclear import of Maize fine streak virus proteins in Drosophila S2 cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Maize fine streak virus (MFSV) is a member of the genus Nucleorhabdovirus, family Rhabdoviridae and is transmitted by the leafhopper Graminella nigrifons. The virus replicates in both its plant host and in its insect vector. Nucleorhabdoviruses replicate in the nucleus and assemble at the inner nu...

  8. Inhibition of the host translation shutoff response by herpes simplex virus 1 triggers nuclear envelope-derived autophagy.

    PubMed

    Radtke, Kerstin; English, Luc; Rondeau, Christiane; Leib, David; Lippé, Roger; Desjardins, Michel

    2013-04-01

    Macroautophagy is a cellular pathway that degrades intracellular pathogens and contributes to antigen presentation. Herpes simplex virus 1 (HSV-1) infection triggers both macroautophagy and an additional form of autophagy that uses the nuclear envelope as a source of membrane. The present study constitutes the first in-depth analysis of nuclear envelope-derived autophagy (NEDA). We established LC3a as a marker that allowed us to distinguish between NEDA and macroautophagy in both immunofluorescence and flow cytometry. NEDA was observed in many different cell types, indicating that it is a general response to HSV-1 infection. This autophagic pathway is known to depend on the viral protein γ34.5, which can inhibit macroautophagy via binding to beclin-1. Using mutant viruses, we were able to show that binding of beclin-1 by γ34.5 had no effect on NEDA, demonstrating that NEDA is regulated differently than macroautophagy. Instead, NEDA was triggered in response to γ34.5 binding to protein phosphatase 1α, an interaction used by the virus to prevent host cells from shutting off protein translation. NEDA was not triggered when late viral protein production was inhibited with acyclovir or hippuristanol, indicating that the accumulation of these proteins might stress infected cells. Interestingly, expression of the late viral protein gH was sufficient to rescue NEDA in the context of infection with a virus that otherwise does not support strong late viral protein expression. We argue that NEDA is a cellular stress response triggered late during HSV-1 infection and might compensate for the viral alteration of the macroautophagic response.

  9. Nuclear localization of a double-stranded RNA-binding protein encoded by the vaccinia virus E3L gene.

    PubMed

    Yuwen, H; Cox, J H; Yewdell, J W; Bennink, J R; Moss, B

    1993-08-01

    We produced a B cell hybridoma (TW2.3) from vaccinia virus-infected mice that secreted a monoclonal antibody (MAb) reactive with a 25-kDA early viral protein that was localized by laser scanning confocal microscopy to the nucleus and cytoplasmic viral factory regions of infected cells. By cell-free translation of mRNA selected by hybridization to a complete library of vaccinia virus DNA fragments, the immunoreactive polypeptide was mapped to open reading frame E3L. The RNA start site of an early promoter was located 26 nucleotides upstream of the first methionine codon of E3L. Evidence was obtained that translation initiation occurs in vivo and in vitro at both the first and second methionine codons to produce major and minor polypeptides of 25 and 19 kDa, respectively. Both polypeptides bound double-stranded RNA, confirming the recent report of H.-W. Chang, J. C. Watson, and B. L. Jacobs (Proc. Natl. Acad. Sci. USA 89, 4825-4829, 1992). Other vaccinia virus proteins were not required for the nuclear localization of the E3L protein, since MAb TW2.3 bound to the nuclei of uninfected cells that were transfected with the E3L gene under the control of the SV40 early promoter. We also demonstrated that the E3L protein can bind to nuclei of aldehyde fixed and detergent permeabilized uninfected cells. This binding was abrogated by treatment of the cells with RNase but not DNase. The nuclear and cytoplasmic locations of the double-stranded RNA binding protein are consistent with multiple functions in the vaccinia virus infectious cycle.

  10. Brevipalpus mites (Acari: Tenuipalpidae): vectors of invasive, non-systemic cytoplasmic and nuclear viruses in plants.

    PubMed

    Rodrigues, Jose Carlos Verle; Childers, Carl C

    2013-02-01

    Multi-directional interactions occur among plant hosts, Brevipalpus mites and the plant viruses they transmit. Such interactions should be considered when evaluating the severity of a disease such as citrus leprosis. The current understanding of Brevipalpus-transmitted viruses relies on the capability of the vector to transmit the disease, the persistence of the virus in the host plant and the ability of the disease to spread. Previously, we discussed the Citrus leprosis virus (CiLV) and its importance and spread over the past decade into new areas of South and Central America, most recently into southern Mexico and Belize. Here, we address key questions to better understand the biology of the mite vector, fitness costs, and the peculiarities of Brevipalpus mite reproduction, virus survival, transmissibility and spread, and the expansion of the host plant range of Brevipalpus species vectoring the disease.

  11. Brevipalpus mites (Acari: Tenuipalpidae): vectors of invasive, non-systemic cytoplasmic and nuclear viruses in plants.

    PubMed

    Rodrigues, Jose Carlos Verle; Childers, Carl C

    2013-02-01

    Multi-directional interactions occur among plant hosts, Brevipalpus mites and the plant viruses they transmit. Such interactions should be considered when evaluating the severity of a disease such as citrus leprosis. The current understanding of Brevipalpus-transmitted viruses relies on the capability of the vector to transmit the disease, the persistence of the virus in the host plant and the ability of the disease to spread. Previously, we discussed the Citrus leprosis virus (CiLV) and its importance and spread over the past decade into new areas of South and Central America, most recently into southern Mexico and Belize. Here, we address key questions to better understand the biology of the mite vector, fitness costs, and the peculiarities of Brevipalpus mite reproduction, virus survival, transmissibility and spread, and the expansion of the host plant range of Brevipalpus species vectoring the disease. PMID:23203501

  12. A model for the dynamic nuclear/nucleolar/cytoplasmic trafficking of the porcine reproductive and respiratory syndrome virus (PRRSV) nucleocapsid protein based on live cell imaging

    SciTech Connect

    You, Jae-Hwan; Howell, Gareth; Pattnaik, Asit K.; Osorio, Fernando A.; Hiscox, Julian A.

    2008-08-15

    Porcine reproductive and respiratory syndrome virus (PRRSV), an arterivirus, in common with many other positive strand RNA viruses, encodes a nucleocapsid (N) protein which can localise not only to the cytoplasm but also to the nucleolus in virus-infected cells and cells over-expressing N protein. The dynamic trafficking of positive strand RNA virus nucleocapsid proteins and PRRSV N protein in particular between the cytoplasm and nucleolus is unknown. In this study live imaging of permissive and non-permissive cell lines, in conjunction with photo-bleaching (FRAP and FLIP), was used to investigate the trafficking of fluorescent labeled (EGFP) PRRSV-N protein. The data indicated that EGFP-PRRSV-N protein was not permanently sequestered to the nucleolus and had equivalent mobility to cellular nucleolar proteins. Further the nuclear import of N protein appeared to occur faster than nuclear export, which may account for the observed relative distribution of N protein between the cytoplasm and the nucleolus.

  13. Small molecule and peptide-mediated inhibition of Epstein-Barr virus nuclear antigen 1 dimerization

    SciTech Connect

    Kim, Sun Young; Song, Kyung-A; Kieff, Elliott; Kang, Myung-Soo

    2012-07-27

    Highlights: Black-Right-Pointing-Pointer Evidence that targeting EBNA1 dimer, an EBV onco-antigen, can be achievable. Black-Right-Pointing-Pointer A small molecule and a peptide as EBNA1 dimerization inhibitors identified. Black-Right-Pointing-Pointer Both inhibitors associated with EBNA1 and blocked EBNA1 DNA binding activity. Black-Right-Pointing-Pointer Also, prevented its dimerization, and repressed viral gene transcription. -- Abstract: Latent Epstein-Barr virus (EBV) infection is associated with human B cell lymphomas and certain carcinomas. EBV episome persistence, replication, and gene expression are dependent on EBV-encoded nuclear antigen 1 (EBNA1)'s DNA binding domain (DBD)/dimerization domain (DD)-mediated sequence-specific DNA binding activity. Homodimerization of EBNA1 is essential for EBNA1 DNA binding and transactivation. In this study, we characterized a novel small molecule EBNA1 inhibitor EiK1, screened from the previous high throughput screening (HTS). The EiK1 compound specifically inhibited the EBNA1-dependent, OriP-enhanced transcription, but not EBNA1-independent transcription. A Surface Plasmon Resonance Biacore assay revealed that EiK1 associates with EBNA1 amino acid 459-607 DBD/DD. Consistent with the SPR data, in vitro gel shift assays showed that EiK1 suppressed the activity of EBNA1 binding to the cognate familial repeats (FR) sequence, but not control RBP-J{kappa} binding to the J{kappa} site. Subsequently, a cross-linker-mediated in vitro multimerization assay and EBNA1 homodimerization-dependent yeast two-hybrid assay showed that EiK1 significantly inhibited EBNA1 dimerization. In an attempt to identify more highly specific peptide inhibitors, small peptides encompassing the EBNA1 DBD/DD were screened for inhibition of EBNA1 DBD-mediated DNA binding function. The small peptide P85, covering EBNA1 a.a. 560-574, significantly blocked EBNA1 DNA binding activity in vitro, prevented dimerization in vitro and in vivo, associated with

  14. Effects of Deletion and Overexpression of the Autographa californica Nuclear Polyhedrosis Virus FP25K Gene on Synthesis of Two Occlusion-Derived Virus Envelope Proteins and Their Transport into Virus-Induced Intranuclear Membranes

    PubMed Central

    Rosas-Acosta, Germán; Braunagel, Sharon C.; Summers, Max D.

    2001-01-01

    Partial deletions within Autographa californica open reading frame 61 (FP25K) alter the expression and accumulation profile of several viral proteins and the transport of occlusion-derived virus (ODV)-E66 to intranuclear membranes during infection (S. C. Braunagel et al., J. Virol. 73:8559–8570, 1999). Here we show the effects of a full deletion and overexpression of FP25K on the transport and expression of two ODV envelope proteins, ODV-E66 (E66) and ODV-E25 (E25). Deletion and overexpression of FP25K substantially altered the levels of expression of E66 during infection. Compared with cells infected with wild-type (wt) virus, the levels of E66 were reduced fivefold in cells infected with a viral mutant lacking FP25K (ΔFP25K) and were slightly increased in cells infected with a viral mutant overexpressing FP25K (FP25Kpolh). In contrast, no significant changes were observed in the levels of E25 among wt-, ΔFP25K-, and FP25Kpolh-infected cells. The changes observed in the levels of E66 among the different viral mutants were not accompanied by changes in either the time of synthesis, membrane association, protein turnover, or steady-state transcript abundance. Deletion of FP25K also substantially altered the transport and localization of E66 during infection. In cells infected with the ΔFP25K mutant virus, E66 accumulated in localized regions at the nuclear periphery and the outer nuclear membrane and did not traffic to intranuclear membranes. In contrast, in cells infected with the FP25Kpolh mutant virus E66 trafficked to intranuclear membranes. For comparison, E25 was normally transported to intranuclear membranes in both ΔFP25K- and FP25Kpolh-infected cells. Altogether these studies suggest that FP25K affects the synthesis of E66 at a posttranscriptional level, probably by altering the translation of E66; additionally, the block in transport of E66 at the nuclear envelope in ΔFP25K-infected cells suggests that the pathway of E66 trafficking to the inner

  15. Functional Characterization of Nuclear Localization and Export Signals in Hepatitis C Virus Proteins and Their Role in the Membranous Web

    PubMed Central

    Levin, Aviad; Neufeldt, Christopher J.; Pang, Daniel; Wilson, Kristen; Loewen-Dobler, Darci; Joyce, Michael A.; Wozniak, Richard W.; Tyrrell, D. Lorne J

    2014-01-01

    The hepatitis C virus (HCV) is a positive strand RNA virus of the Flavivirus family that replicates in the cytoplasm of infected hepatocytes. Previously, several nuclear localization signals (NLS) and nuclear export signals (NES) have been identified in HCV proteins, however, there is little evidence that these proteins travel into the nucleus during infection. We have recently shown that nuclear pore complex (NPC) proteins (termed nucleoporins or Nups) are present in the membranous web and are required during HCV infection. In this study, we identify a total of 11 NLS and NES sequences in various HCV proteins. We show direct interactions between HCV proteins and importin α5 (IPOA5/kapα1), importin β3 (IPO5/kap β3), and exportin 1 (XPO1/CRM1) both in-vitro and in cell culture. These interactions can be disrupted using peptides containing the specific NLS or NES sequences of HCV proteins. Moreover, using a synchronized infection system, we show that these peptides inhibit HCV infection during distinct phases of the HCV life cycle. The inhibitory effects of these peptides place them in two groups. The first group binds IPOA5 and inhibits infection during the replication stage of HCV life cycle. The second group binds IPO5 and is active during both early replication and early assembly. This work delineates the entire life cycle of HCV and the active involvement of NLS sequences during HCV replication and assembly. Given the abundance of NLS sequences within HCV proteins, our previous finding that Nups play a role in HCV infection, and the relocation of the NLS double-GFP reporter in HCV infected cells, this work supports our previous hypothesis that NPC-like structures and nuclear transport factors function in the membranous web to create an environment conducive to viral replication. PMID:25485706

  16. Functional characterization of nuclear localization and export signals in hepatitis C virus proteins and their role in the membranous web.

    PubMed

    Levin, Aviad; Neufeldt, Christopher J; Pang, Daniel; Wilson, Kristen; Loewen-Dobler, Darci; Joyce, Michael A; Wozniak, Richard W; Tyrrell, D Lorne J

    2014-01-01

    The hepatitis C virus (HCV) is a positive strand RNA virus of the Flavivirus family that replicates in the cytoplasm of infected hepatocytes. Previously, several nuclear localization signals (NLS) and nuclear export signals (NES) have been identified in HCV proteins, however, there is little evidence that these proteins travel into the nucleus during infection. We have recently shown that nuclear pore complex (NPC) proteins (termed nucleoporins or Nups) are present in the membranous web and are required during HCV infection. In this study, we identify a total of 11 NLS and NES sequences in various HCV proteins. We show direct interactions between HCV proteins and importin α5 (IPOA5/kapα1), importin β3 (IPO5/kap β3), and exportin 1 (XPO1/CRM1) both in-vitro and in cell culture. These interactions can be disrupted using peptides containing the specific NLS or NES sequences of HCV proteins. Moreover, using a synchronized infection system, we show that these peptides inhibit HCV infection during distinct phases of the HCV life cycle. The inhibitory effects of these peptides place them in two groups. The first group binds IPOA5 and inhibits infection during the replication stage of HCV life cycle. The second group binds IPO5 and is active during both early replication and early assembly. This work delineates the entire life cycle of HCV and the active involvement of NLS sequences during HCV replication and assembly. Given the abundance of NLS sequences within HCV proteins, our previous finding that Nups play a role in HCV infection, and the relocation of the NLS double-GFP reporter in HCV infected cells, this work supports our previous hypothesis that NPC-like structures and nuclear transport factors function in the membranous web to create an environment conducive to viral replication.

  17. Enhancement of CD8(+) T-cell memory by removal of a vaccinia virus nuclear factor-κB inhibitor.

    PubMed

    Ren, Hongwei; Ferguson, Brian J; Maluquer de Motes, Carlos; Sumner, Rebecca P; Harman, Laura E R; Smith, Geoffrey L

    2015-05-01

    Factors influencing T-cell responses are important for vaccine development but are incompletely understood. Here, vaccinia virus (VACV) protein N1 is shown to impair the development of both effector and memory CD8(+) T cells and this correlates with its inhibition of nuclear factor-κB (NF-κB) activation. Infection with VACVs that either have the N1L gene deleted (vΔN1) or contain a I6E mutation (vN1.I6E) that abrogates its inhibition of NF-κB resulted in increased central and memory CD8(+) T-cell populations, increased CD8(+) T-cell cytotoxicity and lower virus titres after challenge. Furthermore, CD8(+) memory T-cell function was increased following infection with vN1.I6E, with more interferon-γ production and greater protection against VACV infection following passive transfer to naive mice, compared with CD8(+) T cells from mice infected with wild-type virus (vN1.WT). This demonstrates the importance of NF-κB activation within infected cells for long-term CD8(+) T-cell memory and vaccine efficacy. Further, it provides a rationale for deleting N1 from VACV vectors to enhance CD8(+) T-cell immunogenicity, while simultaneously reducing virulence to improve vaccine safety.

  18. Domains involved in calcineurin phosphatase inhibition and nuclear localisation in the African swine fever virus A238L protein

    SciTech Connect

    Abrams, Charles C.; Chapman, Dave A.G.; Silk, Rhiannon; Liverani, Elisabetta; Dixon, Linda K.

    2008-05-10

    The African swine fever virus A238L protein inhibits calcineurin phosphatase activity and activation of NF-{kappa}B and p300 co-activator. An 82 amino acid domain containing residues 157 to 238 at the C-terminus of A238L was expressed in E. coli and purified. This purified A238L fragment acted as a potent inhibitor of calcineurin phosphatase in vitro with an IC{sub 50} of approximately 70 nM. Two putative nuclear localisation signals were identified between residues 80 to 86 (NLS-1) and between residues 203 to 207 overlapping with the N-terminus of the calcineurin docking motif (NLS-2). Mutation of these motifs independently did not reduce nuclear localisation compared to the wild type A238L protein, whereas mutation of both motifs significantly reduced nuclear localisation of A238L. Mutation of the calcineurin docking motif resulted in a dramatic increase in the nuclear localisation of A238L provided an intact NLS was present. We propose that binding of calcineurin to A238L masks NLS-2 contributing to the cytoplasmic retention of A238L.

  19. Latency Entry of Herpes Simplex Virus 1 Is Determined by the Interaction of Its Genome with the Nuclear Environment.

    PubMed

    Maroui, Mohamed Ali; Callé, Aleth; Cohen, Camille; Streichenberger, Nathalie; Texier, Pascale; Takissian, Julie; Rousseau, Antoine; Poccardi, Nolwenn; Welsch, Jérémy; Corpet, Armelle; Schaeffer, Laurent; Labetoulle, Marc; Lomonte, Patrick

    2016-09-01

    Herpes simplex virus 1 (HSV-1) establishes latency in trigeminal ganglia (TG) sensory neurons of infected individuals. The commitment of infected neurons toward the viral lytic or latent transcriptional program is likely to depend on both viral and cellular factors, and to differ among individual neurons. In this study, we used a mouse model of HSV-1 infection to investigate the relationship between viral genomes and the nuclear environment in terms of the establishment of latency. During acute infection, viral genomes show two major patterns: replication compartments or multiple spots distributed in the nucleoplasm (namely "multiple-acute"). Viral genomes in the "multiple-acute" pattern are systematically associated with the promyelocytic leukemia (PML) protein in structures designated viral DNA-containing PML nuclear bodies (vDCP-NBs). To investigate the viral and cellular features that favor the acquisition of the latency-associated viral genome patterns, we infected mouse primary TG neurons from wild type (wt) mice or knock-out mice for type 1 interferon (IFN) receptor with wt or a mutant HSV-1, which is unable to replicate due to the synthesis of a non-functional ICP4, the major virus transactivator. We found that the inability of the virus to initiate the lytic program combined to its inability to synthesize a functional ICP0, are the two viral features leading to the formation of vDCP-NBs. The formation of the "multiple-latency" pattern is favored by the type 1 IFN signaling pathway in the context of neurons infected by a virus able to replicate through the expression of a functional ICP4 but unable to express functional VP16 and ICP0. Analyses of TGs harvested from HSV-1 latently infected humans showed that viral genomes and PML occupy similar nuclear areas in infected neurons, eventually forming vDCP-NB-like structures. Overall our study designates PML protein and PML-NBs to be major cellular components involved in the control of HSV-1 latency, probably

  20. Latency Entry of Herpes Simplex Virus 1 Is Determined by the Interaction of Its Genome with the Nuclear Environment

    PubMed Central

    Cohen, Camille; Streichenberger, Nathalie; Texier, Pascale; Takissian, Julie; Rousseau, Antoine; Poccardi, Nolwenn; Welsch, Jérémy; Corpet, Armelle; Schaeffer, Laurent; Labetoulle, Marc; Lomonte, Patrick

    2016-01-01

    Herpes simplex virus 1 (HSV-1) establishes latency in trigeminal ganglia (TG) sensory neurons of infected individuals. The commitment of infected neurons toward the viral lytic or latent transcriptional program is likely to depend on both viral and cellular factors, and to differ among individual neurons. In this study, we used a mouse model of HSV-1 infection to investigate the relationship between viral genomes and the nuclear environment in terms of the establishment of latency. During acute infection, viral genomes show two major patterns: replication compartments or multiple spots distributed in the nucleoplasm (namely “multiple-acute”). Viral genomes in the “multiple-acute” pattern are systematically associated with the promyelocytic leukemia (PML) protein in structures designated viral DNA-containing PML nuclear bodies (vDCP-NBs). To investigate the viral and cellular features that favor the acquisition of the latency-associated viral genome patterns, we infected mouse primary TG neurons from wild type (wt) mice or knock-out mice for type 1 interferon (IFN) receptor with wt or a mutant HSV-1, which is unable to replicate due to the synthesis of a non-functional ICP4, the major virus transactivator. We found that the inability of the virus to initiate the lytic program combined to its inability to synthesize a functional ICP0, are the two viral features leading to the formation of vDCP-NBs. The formation of the “multiple-latency” pattern is favored by the type 1 IFN signaling pathway in the context of neurons infected by a virus able to replicate through the expression of a functional ICP4 but unable to express functional VP16 and ICP0. Analyses of TGs harvested from HSV-1 latently infected humans showed that viral genomes and PML occupy similar nuclear areas in infected neurons, eventually forming vDCP-NB-like structures. Overall our study designates PML protein and PML-NBs to be major cellular components involved in the control of HSV-1 latency

  1. Latency Entry of Herpes Simplex Virus 1 Is Determined by the Interaction of Its Genome with the Nuclear Environment.

    PubMed

    Maroui, Mohamed Ali; Callé, Aleth; Cohen, Camille; Streichenberger, Nathalie; Texier, Pascale; Takissian, Julie; Rousseau, Antoine; Poccardi, Nolwenn; Welsch, Jérémy; Corpet, Armelle; Schaeffer, Laurent; Labetoulle, Marc; Lomonte, Patrick

    2016-09-01

    Herpes simplex virus 1 (HSV-1) establishes latency in trigeminal ganglia (TG) sensory neurons of infected individuals. The commitment of infected neurons toward the viral lytic or latent transcriptional program is likely to depend on both viral and cellular factors, and to differ among individual neurons. In this study, we used a mouse model of HSV-1 infection to investigate the relationship between viral genomes and the nuclear environment in terms of the establishment of latency. During acute infection, viral genomes show two major patterns: replication compartments or multiple spots distributed in the nucleoplasm (namely "multiple-acute"). Viral genomes in the "multiple-acute" pattern are systematically associated with the promyelocytic leukemia (PML) protein in structures designated viral DNA-containing PML nuclear bodies (vDCP-NBs). To investigate the viral and cellular features that favor the acquisition of the latency-associated viral genome patterns, we infected mouse primary TG neurons from wild type (wt) mice or knock-out mice for type 1 interferon (IFN) receptor with wt or a mutant HSV-1, which is unable to replicate due to the synthesis of a non-functional ICP4, the major virus transactivator. We found that the inability of the virus to initiate the lytic program combined to its inability to synthesize a functional ICP0, are the two viral features leading to the formation of vDCP-NBs. The formation of the "multiple-latency" pattern is favored by the type 1 IFN signaling pathway in the context of neurons infected by a virus able to replicate through the expression of a functional ICP4 but unable to express functional VP16 and ICP0. Analyses of TGs harvested from HSV-1 latently infected humans showed that viral genomes and PML occupy similar nuclear areas in infected neurons, eventually forming vDCP-NB-like structures. Overall our study designates PML protein and PML-NBs to be major cellular components involved in the control of HSV-1 latency, probably

  2. Influenza A Virus Polymerase Recruits the RNA Helicase DDX19 to Promote the Nuclear Export of Viral mRNAs

    PubMed Central

    Diot, Cédric; Fournier, Guillaume; Dos Santos, Mélanie; Magnus, Julie; Komarova, Anastasia; van der Werf, Sylvie; Munier, Sandie; Naffakh, Nadia

    2016-01-01

    Enhancing the knowledge of host factors that are required for efficient influenza A virus (IAV) replication is essential to address questions related to pathogenicity and to identify targets for antiviral drug development. Here we focused on the interplay between IAV and DExD-box RNA helicases (DDX), which play a key role in cellular RNA metabolism by remodeling RNA-RNA or RNA-protein complexes. We performed a targeted RNAi screen on 35 human DDX proteins to identify those involved in IAV life cycle. DDX19 was a major hit. In DDX19-depleted cells the accumulation of viral RNAs and proteins was delayed, and the production of infectious IAV particles was strongly reduced. We show that DDX19 associates with intronless, unspliced and spliced IAV mRNAs and promotes their nuclear export. In addition, we demonstrate an RNA-independent association between DDX19 and the viral polymerase, that is modulated by the ATPase activity of DDX19. Our results provide a model in which DDX19 is recruited to viral mRNAs in the nucleus of infected cells to enhance their nuclear export. Information gained from this virus-host interaction improves the understanding of both the IAV replication cycle and the cellular function of DDX19. PMID:27653209

  3. Nuclear import of prototype foamy virus transactivator Bel1 is mediated by KPNA1, KPNA6 and KPNA7.

    PubMed

    Duan, Jihui; Tang, Zhiqin; Mu, Hong; Zhang, Guojun

    2016-08-01

    Bel1, a transactivator of the prototype foamy virus (PFV), plays pivotal roles in the replication of PFV. Previous studies have demonstrated that Bel1 bears a nuclear localization signal (NLS); however, its amino acid sequence remains unclear and the corresponding adaptor importins have not yet been identified. In this study, we inserted various fragments of Bel1 into an EGFP-GST fusion protein and investigated their subcellular localization by fluorescence microscopy. We found that the 215PRQKRPR221 fragment, which accords with the consensus sequence K(K/R)X(K/R) of the monopartite NLS, directed the nuclear translocation of Bel1. Point mutation experiments revealed that K218, R219 and R221 were essential for the nuclear localization of Bel1. The results of GST pull-down assay revealed that the Bel1 peptide 215-221, which bears the NLS, interacted with the nucleocytoplasmic transport receptors, karyopherin alpha 1 (importin alpha 5) (KPNA1), karyopherin alpha 6 (importin alpha 7) (KPNA6) and karyopherin alpha 7 (importin alpha 8) (KPNA7). Finally, in vitro nuclear import assays demonstrated that KPNA1, KPNA6 or KPNA7, along with other necessary nuclear factors, caused Bel1 to localize to the nucleus. Thus, the findings of our study indicate that KPNA1, KPNA6 and KPNA7 are involved in Bel1 nuclear distribution. PMID:27277550

  4. Data for increase of Lymantria dispar male survival after topical application of single-stranded RING domain fragment of IAP-3 gene of its nuclear polyhedrosis virus.

    PubMed

    Oberemok, Volodymyr V; Laikova, Kateryna V; Zaitsev, Aleksei S; Gushchin, Vladimir A; Skorokhod, Oleksii A

    2016-06-01

    This data article is related to the research article entitled "The RING for gypsy moth control: topical application of fragment of its nuclear polyhedrosis virus anti-apoptosis gene as insecticide" [1]. This article reports on significantly higher survival of gypsy moth Lymantria dispar male individuals in response to topical application of single-stranded DNA, based on RING (really interesting new gene) domain fragment of LdMNPV (L. dispar multicapsid nuclear polyhedrosis virus) IAP-3 (inhibitor of apoptosis) gene and acted as DNA insecticide. PMID:27054151

  5. Data for increase of Lymantria dispar male survival after topical application of single-stranded RING domain fragment of IAP-3 gene of its nuclear polyhedrosis virus

    PubMed Central

    Oberemok, Volodymyr V.; Laikova, Kateryna V.; Zaitsev, Aleksei S.; Gushchin, Vladimir A.; Skorokhod, Oleksii A.

    2016-01-01

    This data article is related to the research article entitled “The RING for gypsy moth control: topical application of fragment of its nuclear polyhedrosis virus anti-apoptosis gene as insecticide” [1]. This article reports on significantly higher survival of gypsy moth Lymantria dispar male individuals in response to topical application of single-stranded DNA, based on RING (really interesting new gene) domain fragment of LdMNPV (L. dispar multicapsid nuclear polyhedrosis virus) IAP-3 (inhibitor of apoptosis) gene and acted as DNA insecticide. PMID:27054151

  6. Human Heat shock protein 40 (Hsp40/DnaJB1) promotes influenza A virus replication by assisting nuclear import of viral ribonucleoproteins.

    PubMed

    Batra, Jyoti; Tripathi, Shashank; Kumar, Amrita; Katz, Jacqueline M; Cox, Nancy J; Lal, Renu B; Sambhara, Suryaprakash; Lal, Sunil K

    2016-01-01

    A unique feature of influenza A virus (IAV) life cycle is replication of the viral genome in the host cell nucleus. The nuclear import of IAV genome is an indispensable step in establishing virus infection. IAV nucleoprotein (NP) is known to mediate the nuclear import of viral genome via its nuclear localization signals. Here, we demonstrate that cellular heat shock protein 40 (Hsp40/DnaJB1) facilitates the nuclear import of incoming IAV viral ribonucleoproteins (vRNPs) and is important for efficient IAV replication. Hsp40 was found to interact with NP component of IAV RNPs during early stages of infection. This interaction is mediated by the J domain of Hsp40 and N-terminal region of NP. Drug or RNAi mediated inhibition of Hsp40 resulted in reduced nuclear import of IAV RNPs, diminished viral polymerase function and attenuates overall viral replication. Hsp40 was also found to be required for efficient association between NP and importin alpha, which is crucial for IAV RNP nuclear translocation. These studies demonstrate an important role for cellular chaperone Hsp40/DnaJB1 in influenza A virus life cycle by assisting nuclear trafficking of viral ribonucleoproteins. PMID:26750153

  7. Human Heat shock protein 40 (Hsp40/DnaJB1) promotes influenza A virus replication by assisting nuclear import of viral ribonucleoproteins

    PubMed Central

    Batra, Jyoti; Tripathi, Shashank; Kumar, Amrita; Katz, Jacqueline M.; Cox, Nancy J.; Lal, Renu B.; Sambhara, Suryaprakash; Lal, Sunil K.

    2016-01-01

    A unique feature of influenza A virus (IAV) life cycle is replication of the viral genome in the host cell nucleus. The nuclear import of IAV genome is an indispensable step in establishing virus infection. IAV nucleoprotein (NP) is known to mediate the nuclear import of viral genome via its nuclear localization signals. Here, we demonstrate that cellular heat shock protein 40 (Hsp40/DnaJB1) facilitates the nuclear import of incoming IAV viral ribonucleoproteins (vRNPs) and is important for efficient IAV replication. Hsp40 was found to interact with NP component of IAV RNPs during early stages of infection. This interaction is mediated by the J domain of Hsp40 and N-terminal region of NP. Drug or RNAi mediated inhibition of Hsp40 resulted in reduced nuclear import of IAV RNPs, diminished viral polymerase function and attenuates overall viral replication. Hsp40 was also found to be required for efficient association between NP and importin alpha, which is crucial for IAV RNP nuclear translocation. These studies demonstrate an important role for cellular chaperone Hsp40/DnaJB1 in influenza A virus life cycle by assisting nuclear trafficking of viral ribonucleoproteins. PMID:26750153

  8. Replication of Syngrapha falcifera Multiple-Nuclear Polyhedrosis Virus-D in Different Insect Cells

    NASA Astrophysics Data System (ADS)

    Khalid Nessr Alhag, Sadeq; Xin, Peng Jian

    Six insect cell lines were tested for susceptibility to Syngrapha falcifera multiple nucleocapsid nucleopolyhedrovirus-D (SfaMNPV-D) infection by use of a typical endpoint assay procedure. Cell lines from Trichoplusia ni (Tn5B1-4), (L105-clone), Spodoptera litura (SL-ZSU-1), Spodoptera frugiperda (IPLB-SF-21), Pieris rapaeb (Pr-E-HNU9) and Helicoverpa zea (BCIRL-HZ-AM1) in 96-well tissue culture plates were infected with dilutions of extra cellular virus suspensions of (SfaMNPV-D). Each cell/virus combination was incubated at temperatures 27°C and wells were scored for positive infection at 2 to 4 day intervals. The resulting data were analyzed by Reed and Muench method, providing virus titers for each combination of virus, cell line. The results were categorized by accuracy and by rapidity of maximum titer. Virus titer of Tn5B-4 was higher than other cell lines TCID50 8.7x108, the lowest level detected in infected was in (Pr-E-HNU9) cells TCID50 2.4x108. No Virions or polyhedral inclusion bodies were detected in infected SL-ZSU-1 cells.

  9. Dynamic Response of IFI16 and Promyelocytic Leukemia Nuclear Body Components to Herpes Simplex Virus 1 Infection

    PubMed Central

    2015-01-01

    ABSTRACT Intrinsic immunity is an aspect of antiviral defense that operates through diverse mechanisms at the intracellular level through a wide range of constitutively expressed cellular proteins. In the case of herpesviruses, intrinsic resistance involves the repression of viral gene expression during the very early stages of infection, a process that is normally overcome by viral tegument and/or immediate-early proteins. Thus, the balance between cellular repressors and virus-counteracting proteins determines whether or not a cell becomes productively infected. One aspect of intrinsic resistance to herpes simplex virus 1 (HSV-1) is conferred by components of promyelocytic leukemia nuclear bodies (PML NBs), which respond to infection by accumulating at sites that are closely associated with the incoming parental HSV-1 genomes. Other cellular proteins, including IFI16, which has been implicated in sensing pathogen DNA and initiating signaling pathways that lead to an interferon response, also respond to viral genomes in this manner. Here, studies of the dynamics of the response of PML NB components and IFI16 to invading HSV-1 genomes demonstrated that this response is extremely rapid, occurring within the first hour after addition of the virus, and that human Daxx (hDaxx) and IFI16 respond more rapidly than PML. In the absence of HSV-1 regulatory protein ICP0, which counteracts the recruitment process, the newly formed, viral-genome-induced PML NB-like foci can fuse with existing PML NBs. These data are consistent with a model involving viral genome sequestration into such structures, thereby contributing to the low probability of initiation of lytic infection in the absence of ICP0. IMPORTANCE Herpesviruses have intimate interactions with their hosts, with infection leading either to the productive lytic cycle or to a quiescent infection in which viral gene expression is suppressed while the viral genome is maintained in the host cell nucleus. Whether a cell

  10. Autoselection of cytoplasmic yeast virus like elements encoding toxin/antitoxin systems involves a nuclear barrier for immunity gene expression.

    PubMed

    Kast, Alene; Voges, Raphael; Schroth, Michael; Schaffrath, Raffael; Klassen, Roland; Meinhardt, Friedhelm

    2015-05-01

    Cytoplasmic virus like elements (VLEs) from Kluyveromyces lactis (Kl), Pichia acaciae (Pa) and Debaryomyces robertsiae (Dr) are extremely A/T-rich (>75%) and encode toxic anticodon nucleases (ACNases) along with specific immunity proteins. Here we show that nuclear, not cytoplasmic expression of either immunity gene (PaORF4, KlORF3 or DrORF5) results in transcript fragmentation and is insufficient to establish immunity to the cognate ACNase. Since rapid amplification of 3' ends (RACE) as well as linker ligation of immunity transcripts expressed in the nucleus revealed polyadenylation to occur along with fragmentation, ORF-internal poly(A) site cleavage due to the high A/T content is likely to prevent functional expression of the immunity genes. Consistently, lowering the A/T content of PaORF4 to 55% and KlORF3 to 46% by gene synthesis entirely prevented transcript cleavage and permitted functional nuclear expression leading to full immunity against the respective ACNase toxin. Consistent with a specific adaptation of the immunity proteins to the cognate ACNases, cross-immunity to non-cognate ACNases is neither conferred by PaOrf4 nor KlOrf3. Thus, the high A/T content of cytoplasmic VLEs minimizes the potential of functional nuclear recruitment of VLE encoded genes, in particular those involved in autoselection of the VLEs via a toxin/antitoxin principle. PMID:25973601

  11. A ΩXaV motif in the Rift Valley fever virus NSs protein is essential for degrading p62, forming nuclear filaments and virulence.

    PubMed

    Cyr, Normand; de la Fuente, Cynthia; Lecoq, Lauriane; Guendel, Irene; Chabot, Philippe R; Kehn-Hall, Kylene; Omichinski, James G

    2015-05-12

    Rift Valley fever virus (RVFV) is a single-stranded RNA virus capable of inducing fatal hemorrhagic fever in humans. A key component of RVFV virulence is its ability to form nuclear filaments through interactions between the viral nonstructural protein NSs and the host general transcription factor TFIIH. Here, we identify an interaction between a ΩXaV motif in NSs and the p62 subunit of TFIIH. This motif in NSs is similar to ΩXaV motifs found in nucleotide excision repair (NER) factors and transcription factors known to interact with p62. Structural and biophysical studies demonstrate that NSs binds to p62 in a similar manner as these other factors. Functional studies in RVFV-infected cells show that the ΩXaV motif is required for both nuclear filament formation and degradation of p62. Consistent with the fact that the RVFV can be distinguished from other Bunyaviridae-family viruses due to its ability to form nuclear filaments in infected cells, the motif is absent in the NSs proteins of other Bunyaviridae-family viruses. Taken together, our studies demonstrate that p62 binding to NSs through the ΩXaV motif is essential for degrading p62, forming nuclear filaments and enhancing RVFV virulence. In addition, these results show how the RVFV incorporates a simple motif into the NSs protein that enables it to functionally mimic host cell proteins that bind the p62 subunit of TFIIH. PMID:25918396

  12. A ΩXaV motif in the Rift Valley fever virus NSs protein is essential for degrading p62, forming nuclear filaments and virulence

    PubMed Central

    Cyr, Normand; de la Fuente, Cynthia; Lecoq, Lauriane; Guendel, Irene; Chabot, Philippe R.; Kehn-Hall, Kylene; Omichinski, James G.

    2015-01-01

    Rift Valley fever virus (RVFV) is a single-stranded RNA virus capable of inducing fatal hemorrhagic fever in humans. A key component of RVFV virulence is its ability to form nuclear filaments through interactions between the viral nonstructural protein NSs and the host general transcription factor TFIIH. Here, we identify an interaction between a ΩXaV motif in NSs and the p62 subunit of TFIIH. This motif in NSs is similar to ΩXaV motifs found in nucleotide excision repair (NER) factors and transcription factors known to interact with p62. Structural and biophysical studies demonstrate that NSs binds to p62 in a similar manner as these other factors. Functional studies in RVFV-infected cells show that the ΩXaV motif is required for both nuclear filament formation and degradation of p62. Consistent with the fact that the RVFV can be distinguished from other Bunyaviridae-family viruses due to its ability to form nuclear filaments in infected cells, the motif is absent in the NSs proteins of other Bunyaviridae-family viruses. Taken together, our studies demonstrate that p62 binding to NSs through the ΩXaV motif is essential for degrading p62, forming nuclear filaments and enhancing RVFV virulence. In addition, these results show how the RVFV incorporates a simple motif into the NSs protein that enables it to functionally mimic host cell proteins that bind the p62 subunit of TFIIH. PMID:25918396

  13. Transient induction of a nuclear antigen unrelated to Epstein-Barr nuclear antigen in cells of two human B-lymphoma lines converted by Epstein-Barr virus.

    PubMed

    Fresen, K O; zur Hausen, H

    1977-01-01

    Infection of cells of the Epstein-Barr virus (EBV)-negative human B-lymphoma lines BJAB and Ramos with EBV preparations from P3HR-1 or B 95-8 cells converted these cells to EBV genome carriers expressing Epstein-Barr nuclear antigen (EBNA) in almost 100% of these cells. Induction of these cells as well as of clones from P3HR-1 EBV-converted BJAB cells with iododeoxyuridine, aminopterin, and hypoxanthine resulted in the appearance of a nuclear antigen in about 1-6% of the cells 1-4 days after induction. The antigen is different from known EBV-induced antigens like EBNA, viral capsid antigen (VCA) or the D- and R-subspecificities of the early antigen (EA) complex. It is demonstrated by indirect immunofluorescence and inactivated after acetone fixation. The antigen was not detectable after induction of uninfected BJAB and Ramos cells nor has it been found in noninduced or induced P3HR-1 and Raji cells. Thus, it appears that EBV-infection mediates the expression of this antigen, for which the name TINA (transiently induced nuclear antigen) is suggested. Sera reacting against TINA generally contained high antibody titers against EBV-induced EA. Only a limited number of highly EA-reactive sera, however, were also positive for TINA. Among 200 sera tested thus far, TINA reactivity was most frequently observed in sera of patients with nasopharyngeal carcinoma (7 out of 28), in sera of the only two patients with immunoblastoma tested and occasionally in sera from patients with Hodgkin's disease and chronic lymphatic leukemia. Among 70 sera from nontumor patients, TINA reactivity was observed three times: two patients suffered from "chronic" infectious mononucleosis, the other revealed persistent splenomegaly. PMID:189313

  14. Cell cycle regulation of human immunodeficiency virus type 1 integration in T cells: antagonistic effects of nuclear envelope breakdown and chromatin condensation

    SciTech Connect

    Mannioui, Abdelkrim . E-mail: karim.mannioui@chu-stlouis.fr; Schiffer, Cecile . E-mail: cecile.schiffer@voila.fr; Felix, Nathalie . E-mail: nathalie.felix@chu-stlouis.fr

    2004-11-10

    We examined the influence of mitosis on the kinetics of human immunodeficiency virus type 1 integration in T cells. Single-round infection of cells arrested in G1b or allowed to synchronously proceed through division showed that mitosis delays virus integration until 18-24 h postinfection, whereas integration reaches maximum levels by 15 h in G1b-arrested cells. Subcellular fractionation of metaphase-arrested cells indicated that, while nuclear envelope disassembly facilitates docking of viral DNA to chromatin, chromosome condensation directly antagonizes and therefore delays integration. As a result of the balance between the two effects, virus integration efficiency is eventually up to threefold greater in dividing cells. At the single-cell level, using a green fluorescent protein-expressing reporter virus, we found that passage through mitosis leads to prominent asymmetric segregation of the viral genome in daughter cells without interfering with provirus expression.

  15. The NS2 Proteins of Parvovirus Minute Virus of Mice Are Required for Efficient Nuclear Egress of Progeny Virions in Mouse Cells

    PubMed Central

    Eichwald, Virginie; Daeffler, Laurent; Klein, Michèle; Rommelaere, Jean; Salomé, Nathalie

    2002-01-01

    The small nonstructural NS2 proteins of parvovirus minute virus of mice (MVMp) were previously shown to interact with the nuclear export receptor Crm1. We report here the analysis of two MVM mutant genomic clones generating NS2 proteins that are unable to interact with Crm1 as a result of amino acid substitutions within their nuclear export signal (NES) sequences. Upon transfection of human and mouse cells, the MVM-NES21 and MVM-NES22 mutant genomic clones were proficient in synthesis of the four virus-encoded proteins. While the MVM-NES22 clone was further able to produce infectious mutant virions, no virus could be recovered from cells transfected with the MVM-NES21 clone. Whereas the defect of MVM-NES21 appeared to be complex, the phenotype of MVM-NES22 could be traced back to a novel distinct NS2 function. Infection of mouse cells with the MVM-NES22 mutant led to stronger nuclear retention not only of the NS2 proteins but also of infectious progeny MVM particles. This nuclear sequestration correlated with a severe delay in the release of mutant virions in the medium and with prolonged survival of the infected cell populations compared with wild-type virus-treated cultures. This defect could explain, at least in part, the small size of the plaques generated by the MVM-NES22 mutant when assayed on mouse indicator cells. Altogether, our data indicate that the interaction of MVMp NS2 proteins with the nuclear export receptor Crm1 plays a critical role at a late stage of the parvovirus life cycle involved in release of progeny viruses. PMID:12239307

  16. Epstein-Barr nuclear antigen 1 mediates a DNA loop within the latent replication origin of Epstein-Barr virus.

    PubMed

    Frappier, L; O'Donnell, M

    1991-12-01

    Epstein-Barr virus-encoded nuclear antigen 1 (EBNA-1) binds and activates the viral latent origin of DNA replication, oriP. We have used electron microscopy to examine the assembly of EBNA-1 onto oriP. The oriP region consists of two essential elements separated by approximately 1 kilobase pair of DNA. One element contains 20 tandom EBNA-1 binding sites [called the family of repeats (FR)] and serves to activate initiation of replication at the dyad symmetry (DS) element, which contains 4 EBNA-1 binding sites. Titration of homogeneous EBNA-1 produced in baculovirus (bEBNA-1) onto oriP DNA showed an order to the assembly of bEBNA-1 onto oriP. At low concentrations, bEBNA-1 was located exclusively on the FR element. As the level of bEBNA-1 was raised, a loop between the FR and DS elements became the most prevalent DNA-protein complex. These data suggest protein-mediated DNA looping may play a role in activating latent-phase replication of the Epstein-Barr virus.

  17. Herpes simplex virus 2 UL13 protein kinase disrupts nuclear lamins

    SciTech Connect

    Cano-Monreal, Gina L.; Wylie, Kristine M.; Cao, Feng; Tavis, John E.; Morrison, Lynda A.

    2009-09-15

    Herpesviruses must cross the inner nuclear membrane and underlying lamina to exit the nucleus. HSV-1 US3 and PKC can phosphorylate lamins and induce their dispersion but do not elicit all of the phosphorylated lamin species produced during infection. UL13 is a serine threonine protein kinase conserved among many herpesviruses. HSV-1 UL13 phosphorylates US3 and thereby controls UL31 and UL34 nuclear rim localization, indicating a role in nuclear egress. Here, we report that HSV-2 UL13 alone induced conformational changes in lamins A and C and redistributed lamin B1 from the nuclear rim to intranuclear granular structures. HSV-2 UL13 directly phosphorylated lamins A, C, and B1 in vitro, and the lamin A1 tail domain. HSV-2 infection recapitulated the lamin alterations seen upon expression of UL13 alone, and other alterations were also observed, indicating that additional viral and/or cellular proteins cooperate with UL13 to alter lamins during HSV-2 infection to allow nuclear egress.

  18. Characterization of a nuclear localization signal in the C-terminus of the adeno-associated virus Rep68/78 proteins

    SciTech Connect

    Cassell, Geoffrey D.; Weitzman, Matthew D. . E-mail: weitzman@salk.edu

    2004-10-01

    Adeno-associated virus (AAV) replicates in the nucleus of infected cells, and therefore multiple nuclear import events are required for productive infection. We analyzed nuclear import of the viral Rep proteins and characterized a nuclear localization signal (NLS) in the C-terminus. We demonstrate that basic residues in this region constitute an NLS that is transferable and mediates interaction with the nuclear import receptor importin {alpha} in vitro. Mutant Rep proteins are predominantly cytoplasmic and are severely compromised for interactions with importin {alpha}, but retain their enzymatic functions in vitro. Interestingly, mutations of the NLS had significantly less effect on importin {alpha} interaction and replication in the context of Rep78 than when incorporated into the Rep68 protein. Together, our results demonstrate that a bipartite NLS exists in the shared part of Rep68 and Rep78, and suggest that an alternate entry mechanism may also contribute to nuclear localization of the Rep78 protein.

  19. Functional study of hot pepper 26S proteasome subunit RPN7 induced by Tobacco mosaic virus from nuclear proteome analysis

    SciTech Connect

    Lee, Boo-Ja; Kwon, Sun Jae; Kim, Sung-Kyu; Kim, Ki-Jeong; Park, Chang-Jin; Kim, Young-Jin; Park, Ohkmae K.; Paek, Kyung-Hee . E-mail: khpaek95@korea.ac.kr

    2006-12-15

    Two-dimensional gel electrophoresis (2-DE) was applied for the screening of Tobacco mosaic virus (TMV)-induced hot pepper (Capsicum annuum cv. Bugang) nuclear proteins. From differentially expressed protein spots, we acquired the matched peptide mass fingerprint (PMF) data, analyzed by MALDI-TOF MS, from the non-redundant hot pepper EST protein FASTA database using the VEMS 2.0 software. Among six identified nuclear proteins, the hot pepper 26S proteasome subunit RPN7 (CaRPN7) was subjected to further study. The level of CaRPN7 mRNA was specifically increased during incompatible TMV-P{sub 0} interaction, but not during compatible TMV-P{sub 1.2} interaction. When CaRPN7::GFP fusion protein was targeted in onion cells, the nuclei had been broken into pieces. In the hot pepper leaves, cell death was exacerbated and genomic DNA laddering was induced by Agrobacterium-mediated transient overexpression of CaPRN7. Thus, this report presents that the TMV-induced CaRPN7 may be involved in programmed cell death (PCD) in the hot pepper plant.

  20. Involvement of the UL24 protein in herpes simplex virus 1-induced dispersal of B23 and in nuclear egress

    SciTech Connect

    Lymberopoulos, Maria H.; Bourget, Amelie; Abdeljelil, Nawel Ben; Pearson, Angela

    2011-04-10

    UL24 of herpes simplex virus 1 (HSV-1) is widely conserved within the Herpesviridae family. Herein, we tested the hypothesis that UL24, which we have previously shown to induce the redistribution of nucleolin, also affects the localization of the nucleolar protein B23. We found that HSV-1-induced dispersal of B23 was dependent on UL24. The conserved N-terminal portion of UL24 was sufficient to induce the redistribution of B23 in transient transfection assays. Mutational analysis revealed that the endonuclease motif of UL24 was important for B23 dispersal in both transfected and infected cells. Nucleolar protein relocalization during HSV-1 infection was also observed in non-immortalized cells. Analysis of infected cells by electron microscopy revealed a decrease in the ratio of cytoplasmic versus nuclear viral particles in cells infected with a UL24-deficient strain compared to KOS-infected cells. Our results suggest that UL24 promotes nuclear egress of nucleocapsids during HSV-1 infection, possibly though effects on nucleoli.

  1. Nucleocytoplasmic shuttling of the rabies virus P protein requires a nuclear localization signal and a CRM1-dependent nuclear export signal

    SciTech Connect

    Pasdeloup, David; Poisson, Nicolas; Raux, Helene; Gaudin, Yves; Ruigrok, Rob W.H. . E-mail: danielle.blondel@vms.cnrs-gif.fr

    2005-04-10

    Rabies virus P protein is a co-factor of the viral RNA polymerase. It has been shown previously that P mRNA directs the synthesis of four N-terminally truncated P products P2, P3, P4, and P5 due to translational initiation by a leaky scanning mechanism at internal Met codons. Whereas P and P2 are located in the cytoplasm, P3, P4, and P5 are found in the nucleus. Here, we have analyzed the molecular basis of the subcellular localization of these proteins. Using deletion mutants fused to GFP protein, we show the presence of a nuclear localization signal (NLS) in the C-terminal part of P (172-297). This domain contains a short lysine-rich stretch ({sup 211}KKYK{sup 214}) located in close proximity with arginine 260 as revealed by the crystal structure of P. We demonstrate the critical role of lysine 214 and arginine 260 in NLS activity. In the presence of Leptomycin B, P is retained in the nucleus indicating that it contains a CRM1-dependent nuclear export signal (NES). The subcellular distribution of P deletion mutants indicates that the domain responsible for export is the amino-terminal part of the protein. The use of fusion proteins that have amino terminal fragments of P fused to {beta}-galactosidase containing the NLS of SV40 T antigen allows us to identify a NES between residues 49 and 58. The localization of NLS and NES determines the cellular distribution of the P gene products.

  2. A nuclear localization of the infectious haematopoietic necrosis virus NV protein is necessary for optimal viral growth

    USGS Publications Warehouse

    Choi, M.K.; Moon, C.H.; Ko, M.S.; Lee, U.-H.; Cho, W.J.; Cha, S.J.; Do, J.W.; Heo, G.J.; Jeong, S.G.; Hahm, Y.S.; Harmache, A.; Bremont, M.; Kurath, G.; Park, J.-W.

    2011-01-01

    The nonvirion (NV) protein of infectious hematopoietic necrosis virus (IHNV) has been previously reported to be essential for efficient growth and pathogenicity of IHNV. However, little is known about the mechanism by which the NV supports the viral growth. In this study, cellular localization of NV and its role in IHNV growth in host cells was investigated. Through transient transfection in RTG-2 cells of NV fused to green fluorescent protein (GFP), a nuclear localization of NV was demonstrated. Deletion analyses showed that the 32EGDL35 residues were essential for nuclear localization of NV protein, and fusion of these 4 amino acids to GFP directed its transport to the nucleus. We generated a recombinant IHNV, rIHNV-NV-ΔEGDL in which the 32EGDL35 was deleted from the NV. rIHNVs with wild-type NV (rIHNV-NV) or with the NV gene replaced with GFP (rIHNV-ΔNV-GFP) were used as controls. RTG-2 cells infected with rIHNV-ΔNV-GFP and rIHNV-NV-ΔEGDL yielded 12- and 5-fold less infectious virion, respectively, than wild type rIHNV-infected cells at 48 h post-infection (p.i.). While treatment with poly I:C at 24 h p.i. did not inhibit replication of wild-type rIHNVs, replication rates of rIHNV-ΔNV-GFP and rIHNV-NV-ΔEGDL were inhibited by poly I:C. In addition, both rIHNV-ΔNV and rIHNV-NV-ΔEGDL induced higher levels of expressions of both IFN1 and Mx1 than wild-type rIHNV. These data suggest that the IHNV NV may support the growth of IHNV through inhibition of the INF system and the amino acid residues of 32EGDL35 responsible for nuclear localization are important for the inhibitory activity of NV.

  3. Two Domains of the V Protein of Virulent Canine Distemper Virus Selectively Inhibit STAT1 and STAT2 Nuclear Import▿

    PubMed Central

    Röthlisberger, Anne; Wiener, Dominique; Schweizer, Matthias; Peterhans, Ernst; Zurbriggen, Andreas; Plattet, Philippe

    2010-01-01

    Canine distemper virus (CDV) causes in dogs a severe systemic infection, with a high frequency of demyelinating encephalitis. Among the six genes transcribed by CDV, the P gene encodes the polymerase cofactor protein (P) as well as two additional nonstructural proteins, C and V; of these V was shown to act as a virulence factor. We investigated the molecular mechanisms by which the P gene products of the neurovirulent CDV A75/17 strain disrupt type I interferon (IFN-α/β)-induced signaling that results in the establishment of the antiviral state. Using recombinant knockout A75/17 viruses, the V protein was identified as the main antagonist of IFN-α/β-mediated signaling. Importantly, immunofluorescence analysis illustrated that the inhibition of IFN-α/β-mediated signaling correlated with impaired STAT1/STAT2 nuclear import, whereas the phosphorylation state of these proteins was not affected. Coimmunoprecipitation assays identified the N-terminal region of V (VNT) responsible for STAT1 targeting, which correlated with its ability to inhibit the activity of the IFN-α/β-mediated antiviral state. Conversely, while the C-terminal domain of V (VCT) could not function autonomously, when fused to VNT it optimally interacted with STAT2 and subsequently efficiently suppressed the IFN-α/β-mediated signaling pathway. The latter result was further supported by a single mutation at position 110 within the VNT domain of CDV V protein, resulting in a mutant that lost STAT1 binding while retaining a partial STAT2 association. Taken together, our results identified the CDV VNT and VCT as two essential modules that complement each other to interfere with the antiviral state induced by IFN-α/β-mediated signaling. Hence, our experiments reveal a novel mechanism of IFN-α/β evasion among the morbilliviruses. PMID:20427537

  4. Picornaviruses and nuclear functions: targeting a cellular compartment distinct from the replication site of a positive-strand RNA virus

    PubMed Central

    Flather, Dylan; Semler, Bert L.

    2015-01-01

    The compartmentalization of DNA replication and gene transcription in the nucleus and protein production in the cytoplasm is a defining feature of eukaryotic cells. The nucleus functions to maintain the integrity of the nuclear genome of the cell and to control gene expression based on intracellular and environmental signals received through the cytoplasm. The spatial separation of the major processes that lead to the expression of protein-coding genes establishes the necessity of a transport network to allow biomolecules to translocate between these two regions of the cell. The nucleocytoplasmic transport network is therefore essential for regulating normal cellular functioning. The Picornaviridae virus family is one of many viral families that disrupt the nucleocytoplasmic trafficking of cells to promote viral replication. Picornaviruses contain positive-sense, single-stranded RNA genomes and replicate in the cytoplasm of infected cells. As a result of the limited coding capacity of these viruses, cellular proteins are required by these intracellular parasites for both translation and genomic RNA replication. Being of messenger RNA polarity, a picornavirus genome can immediately be translated upon entering the cell cytoplasm. However, the replication of viral RNA requires the activity of RNA-binding proteins, many of which function in host gene expression, and are consequently localized to the nucleus. As a result, picornaviruses disrupt nucleocytoplasmic trafficking to exploit protein functions normally localized to a different cellular compartment from which they translate their genome to facilitate efficient replication. Furthermore, picornavirus proteins are also known to enter the nucleus of infected cells to limit host-cell transcription and down-regulate innate antiviral responses. The interactions of picornavirus proteins and host-cell nuclei are extensive, required for a productive infection, and are the focus of this review. PMID:26150805

  5. The human immunodeficiency virus type 1 Rev protein shuttles between the cytoplasm and nuclear compartments.

    PubMed Central

    Kalland, K H; Szilvay, A M; Brokstad, K A; Saetrevik, W; Haukenes, G

    1994-01-01

    A retroviral regulatory protein, Rev (regulator of virion protein expression), is made in cells infected by human immunodeficiency virus (HIV). Rev is essential for the completion of the retroviral life cycle and interacts with the host cell at some posttranscriptional step in order to express the incompletely spliced HIV mRNAs from which HIV structural proteins are translated. Neither the host cell components nor the mechanisms responsible for this important regulation have been defined. We now report that Rev is a nucleocytoplasmic shuttle protein which is continuously transported between the cytoplasm, the nucleoli, and nucleoplasmic speckles enriched in RNA splicing and processing factors. The results show that Rev has the potential to interfere specifically with the splicing of the HIV pre-mRNA in the nucleoplasm and, next, guide such mRNAs to the cytoplasm for translation. Images PMID:7935458

  6. Nuclear Sensing of Viral DNA, Epigenetic Regulation of Herpes Simplex Virus Infection, and Innate Immunity

    PubMed Central

    Knipe, David M.

    2015-01-01

    Herpes simplex virus (HSV) undergoes a lytic infection in epithelial cells and a latent infection in neuronal cells, and epigenetic mechanisms play a major role in the differential gene expression under the two conditions. Herpes viron DNA is not associated with histones but is rapidly loaded with heterochromatin upon entry into the cell. Viral proteins promote reversal of the epigenetic silencing in epithelial cells while the viral latency-associated transcript promotes additional heterochromatin in neuronal cells. The cellular sensors that initiate the chromatinization of foreign DNA have not been fully defined. IFI16 and cGAS are both essential for innate sensing of HSV DNA, and new evidence shows how they work together to initiate innate signaling. IFI16 also plays a role in the heterochromatinization of HSV DNA, and this review will examine how IFI16 integrates epigenetic regulation and innate sensing of foreign viral DNA to show how these two responses are related. PMID:25742715

  7. Adenovirus E1A Targets the DREF Nuclear Factor To Regulate Virus Gene Expression, DNA Replication, and Growth

    PubMed Central

    Radko, Sandi; Koleva, Maria; James, Kris M. D.; Jung, Richard; Mymryk, Joe S.

    2014-01-01

    ABSTRACT The adenovirus E1A gene is the first gene expressed upon viral infection. E1A remodels the cellular environment to maximize permissivity for viral replication. E1A is also the major transactivator of viral early gene expression and a coregulator of a large number of cellular genes. E1A carries out its functions predominantly by binding to cellular regulatory proteins and altering their activities. The unstructured nature of E1A enables it to bind to a large variety of cellular proteins and form new molecular complexes with novel functions. The C terminus of E1A is the least-characterized region of the protein, with few known binding partners. Here we report the identification of cellular factor DREF (ZBED1) as a novel and direct binding partner of E1A. Our studies identify a dual role for DREF in the viral life cycle. DREF contributes to activation of gene expression from all viral promoters early in infection. Unexpectedly, it also functions as a growth restriction factor for adenovirus as knockdown of DREF enhances virus growth and increases viral genome copy number late in the infection. We also identify DREF as a component of viral replication centers. E1A affects the subcellular distribution of DREF within PML bodies and enhances DREF SUMOylation. Our findings identify DREF as a novel E1A C terminus binding partner and provide evidence supporting a role for DREF in viral replication. IMPORTANCE This work identifies the putative transcription factor DREF as a new target of the E1A oncoproteins of human adenovirus. DREF was found to primarily localize with PML nuclear bodies in uninfected cells and to relocalize into virus replication centers during infection. DREF was also found to be SUMOylated, and this was enhanced in the presence of E1A. Knockdown of DREF reduced the levels of viral transcripts detected at 20 h, but not at 40 h, postinfection, increased overall virus yield, and enhanced viral DNA replication. DREF was also found to localize to

  8. Nuclear localization of dengue virus (DENV) 1-4 non-structural protein 5; protection against all 4 DENV serotypes by the inhibitor Ivermectin.

    PubMed

    Tay, M Y F; Fraser, J E; Chan, W K K; Moreland, N J; Rathore, A P; Wang, C; Vasudevan, S G; Jans, D A

    2013-09-01

    Infection by one of the 4 distinct serotypes of dengue virus (DENV) threatens >40% of the world's population, with no efficacious vaccine or antiviral agent currently available. DENV replication through the virus-encoded nonstructural protein (NS) 5 protein occurs in the infected cell cytoplasm, but NS5 from DENV2 has thus far been shown to localize strongly in the nucleus throughout infection. Here we use specific antibodies cross-reactive with NS5 from DENV1-4 to demonstrate nuclear localization of NS5 from all DENV serotypes for the first time in both infected as well as transfected cells, although to differing extents. The small-molecule inhibitor Ivermectin was inhibitory towards both DENV 1 and 2 NS5 interaction with its nuclear transporter importin α/β in vitro, and protected against infection from DENV1-4. Ivermectin thus has potential in the clinical setting as a dengue antiviral.

  9. Oxadiazols: a New Class of Rationally Designed Anti-Human Immunodeficiency Virus Compounds Targeting the Nuclear Localization Signal of the Viral Matrix Protein

    PubMed Central

    Haffar, Omar; Dubrovsky, Larisa; Lowe, Richard; Berro, Reem; Kashanchi, Fatah; Godden, Jeffrey; Vanpouille, Christophe; Bajorath, Jürgen; Bukrinsky, Michael

    2005-01-01

    Despite recent progress in anti-human immunodeficiency virus (HIV) therapy, drug toxicity and emergence of drug-resistant isolates during long-term treatment of HIV-infected patients necessitate the search for new targets that can be used to develop novel antiviral agents. One such target is the process of nuclear translocation of the HIV preintegration complex. Previously we described a class of arylene bis(methylketone) compounds that inhibit HIV-1 nuclear import by targeting the nuclear localization signal (NLS) in the matrix protein (MA). Here we report a different class of MA NLS-targeting compounds that was selected using computer-assisted drug design. The leading compound from this group, ITI-367, showed potent anti-HIV activity in cultures of T lymphocytes and macrophages and also inhibited HIV-1 replication in ex vivo cultured lymphoid tissue. The virus carrying inactivating mutations in MA NLS was resistant to ITI-367. Analysis by real-time PCR demonstrated that the compound specifically inhibited nuclear import of viral DNA, measured by two-long terminal repeat circle formation. Evidence of the existence of this mechanism was provided by immunofluorescent microscopy, using fluorescently labeled HIV-1, which demonstrated retention of the viral DNA in the cytoplasm of drug-treated macrophages. Compounds inhibiting HIV-1 nuclear import may be attractive candidates for further development. PMID:16189005

  10. Identification of two independent transcriptional activation domains in the Autographa californica multicapsid nuclear polyhedrosis virus IE1 protein.

    PubMed

    Slack, J M; Blissard, G W

    1997-12-01

    The Autographa californica multicapsid nuclear polyhedrosis virus immediate-early protein, IE1, is a 582-amino-acid phosphoprotein that regulates the transcription of early viral genes. Deletion of N-terminal regions of IE1 in previous studies (G. R. Kovacs, J. Choi, L. A. Guarino, and M. D. Summers, J. Virol. 66:7429-7437, 1992) resulted in the loss of transcriptional activation, suggesting that this region may contain an acidic activation domain. To identify independently functional transcriptional activation domains, we developed a heterologous system in which potential regulatory domains were fused with a modified Escherichia coli Lac repressor protein that contains a nuclear localization signal (NLacR). Transcriptional activation by the resulting NLacR-IE1 chimeras was measured with a basal baculovirus early promoter containing optimized Lac repressor binding sites (lac operators). Chimeras containing IE1 peptides dramatically activated transcription of the basal promoter only when lac operator sequences were present. In addition, transcriptional activation by NLacR-IE1 chimeras was allosterically regulated by the lactose analog, isopropyl-beta-D-thiogalactopyranoside (IPTG). For a more detailed analysis of IE1 regulatory domains, the M1 to T266 N-terminal portion of IE1 was subdivided (on the basis of average amino acid charge) into five smaller regions which were fused in various combinations to NLacR. Regions M1 to N125 and A168 to G222 were identified as independent transcriptional activation domains. Some NLacR-IE1 chimeras exhibited retarded migration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels. As with wild-type IE1, this aberrant gel mobility was associated with phosphorylation. Mapping studies with the NLacR-IE1 chimeras indicate that the M1 to A168 region of IE1 is necessary for this phosphorylation-associated effect.

  11. Complete genome sequence and integrated protein localization and interaction map for alfalfa dwarf virus, which combines properties of both cytoplasmic and nuclear plant rhabdoviruses

    SciTech Connect

    Bejerman, Nicolás; Giolitti, Fabián; Breuil, Soledad de; Trucco, Verónica; Nome, Claudia; Lenardon, Sergio; Dietzgen, Ralf G.

    2015-09-15

    Summary: We have determined the full-length 14,491-nucleotide genome sequence of a new plant rhabdovirus, alfalfa dwarf virus (ADV). Seven open reading frames (ORFs) were identified in the antigenomic orientation of the negative-sense, single-stranded viral RNA, in the order 3′-N-P-P3-M-G-P6-L-5′. The ORFs are separated by conserved intergenic regions and the genome coding region is flanked by complementary 3′ leader and 5′ trailer sequences. Phylogenetic analysis of the nucleoprotein amino acid sequence indicated that this alfalfa-infecting rhabdovirus is related to viruses in the genus Cytorhabdovirus. When transiently expressed as GFP fusions in Nicotiana benthamiana leaves, most ADV proteins accumulated in the cell periphery, but unexpectedly P protein was localized exclusively in the nucleus. ADV P protein was shown to have a homotypic, and heterotypic nuclear interactions with N, P3 and M proteins by bimolecular fluorescence complementation. ADV appears unique in that it combines properties of both cytoplasmic and nuclear plant rhabdoviruses. - Highlights: • The complete genome of alfalfa dwarf virus is obtained. • An integrated localization and interaction map for ADV is determined. • ADV has a genome sequence similarity and evolutionary links with cytorhabdoviruses. • ADV protein localization and interaction data show an association with the nucleus. • ADV combines properties of both cytoplasmic and nuclear plant rhabdoviruses.

  12. Nuclear sensing of viral DNA, epigenetic regulation of herpes simplex virus infection, and innate immunity

    SciTech Connect

    Knipe, David M.

    2015-05-15

    Herpes simplex virus (HSV) undergoes a lytic infection in epithelial cells and a latent infection in neuronal cells, and epigenetic mechanisms play a major role in the differential gene expression under the two conditions. HSV viron DNA is not associated with histones but is rapidly loaded with heterochromatin upon entry into the cell. Viral proteins promote reversal of the epigenetic silencing in epithelial cells while the viral latency-associated transcript promotes additional heterochromatin in neuronal cells. The cellular sensors that initiate the chromatinization of foreign DNA have not been fully defined. IFI16 and cGAS are both essential for innate sensing of HSV DNA, and new evidence shows how they work together to initiate innate signaling. IFI16 also plays a role in the heterochromatinization of HSV DNA, and this review will examine how IFI16 integrates epigenetic regulation and innate sensing of foreign viral DNA to show how these two responses are related. - Highlights: • HSV lytic and latent gene expression is regulated differentially by epigenetic processes. • The sensors of foreign DNA have not been defined fully. • IFI16 and cGAS cooperate to sense viral DNA in HSV-infected cells. • IFI16 plays a role in both innate sensing of HSV DNA and in restricting its expression.

  13. Nucleolin is important for Epstein–Barr virus nuclear antigen 1-mediated episome binding, maintenance, and transcription

    PubMed Central

    Chen, Ya-Lin; Liu, Cheng-Der; Cheng, Chi-Ping; Zhao, Bo; Hsu, Hao-Jen; Shen, Chih-Long; Chiu, Shu-Jun; Kieff, Elliott; Peng, Chih-wen

    2014-01-01

    Epstein–Barr virus (EBV) nuclear antigen 1 (EBNA1) is essential for EBV episome maintenance, replication, and transcription. These effects are mediated by EBNA1 binding to cognate oriP DNA, which comprise 20 imperfect copies of a 30-bp dyad symmetry enhancer and an origin for DNA replication. To identify cell proteins essential for these EBNA1 functions, EBNA1 associated cell proteins were immune precipitated and analyzed by liquid chromatography-tandem mass spectrometry. Nucleolin (NCL) was identified to be EBNA1 associated. EBNA1's N-terminal 100 aa and NCL's RNA-binding domains were critical for EBNA1/NCL interaction. Lentivirus shRNA-mediated NCL depletion substantially reduced EBNA1 recruitment to oriP DNA, EBNA1-dependent transcription of an EBV oriP luciferase reporter, and EBV genome maintenance in lymphoblastoid cell lines. NCL RNA-binding domain K429 was critical for ATP and EBNA1 binding. NCL overexpression increased EBNA1 binding to oriP and transcription, whereas NCL K429A was deficient. Moreover, NCL silencing impaired lymphoblastoid cell line growth. These experiments reveal a surprisingly critical role for NCL K429 in EBNA1 episome maintenance and transcription, which may be a target for therapeutic intervention. PMID:24344309

  14. Induction of natural killer cell-dependent antitumor immunity by the Autographa californica multiple nuclear polyhedrosis virus.

    PubMed

    Kitajima, Masayuki; Abe, Takayuki; Miyano-Kurosaki, Naoko; Taniguchi, Masaru; Nakayama, Toshinori; Takaku, Hiroshi

    2008-02-01

    Wild-type Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) infects a variety of mammalian cell types in vitro, but does not replicate in these cells. We investigated the effects of AcMNPV in the induction of the immune response and tumor metastasis in mice. After intravenous injection, AcMNPV was taken up by the liver and spleen, and preferentially infected dendritic cells (DCs) and B cells in the spleen; costimulatory molecules CD40, CD80, and CD86 were upregulated in the DCs. The hepatic mononuclear cells (MNCs) in these animals were highly cytotoxic to natural killer (NK)-sensitive YAC-1 and B16 melanoma cells, but not to NK-resistant EL4 cells. Intravenous injection of AcMNPV-induced NK cell proliferation in the liver and spleen, and enhanced antitumor immunity in mice with B16 liver metastases. Furthermore, such treatment increased the survival of C57BL/6, J alpha 281 (-/-), and interferon (IFN)-gamma (-/-) mice that were previously injected with B16 tumor cells. AcMNPV injection did not enhance the survival of NK cell-depleted mice. Moreover, one AcMNPV treatment effectively prolonged survival in a B16 liver metastasis model, and was equivalent to five treatments with recombinant interleukin-12 (IL-12) protein. These findings suggest that AcMNPV efficiently stimulates NK cell-mediated antitumor immunity. PMID:18059370

  15. Regulation of a nuclear export signal by an adjacent inhibitory sequence: the effector domain of the influenza virus NS1 protein.

    PubMed

    Li, Y; Yamakita, Y; Krug, R M

    1998-04-28

    In the cell nucleus the NS1 protein of influenza A virus inhibits both pre-mRNA splicing and the nuclear export of mRNAs. Both the RNA-binding and effector domains of the protein are required for these nuclear functions. Here we demonstrate that the NS1 protein has a latent nuclear export signal (NES) that is located at the amino end of the effector domain. In uninfected, transfected cells the NS1 protein is localized in the nucleus because the NES is specifically inhibited by the adjacent amino acid sequence in the effector domain. Substitution of alanine residues for specific amino acids in the adjacent sequence abrogates its inhibitory activity, thereby unmasking the NES and causing the full-length NS1 protein to be localized to the cytoplasm. In contrast to uninfected cells, a substantial amount of the NS1 protein in influenza virus-infected cells is located in the cytoplasm. Consequently, the NES of these NS1 protein molecules is unmasked in infected cells, indicating that the NS1 protein most likely carries out functions in the cytoplasm as well as the nucleus. A dramatically different localization of the NS1 protein occurs in cells that are infected by a virus encoding an NS1 protein lacking the NES: the shortened NS1 protein molecules are almost totally in the nucleus. Because the NES of the full-length NS1 protein is unmasked in infected but not uninfected cells, it is likely that this unmasking results from a specific interaction of another virus-specific protein with the NS1 protein.

  16. Epstein-Barr virus nuclear antigen 3C regulated genes in lymphoblastoid cell lines.

    PubMed

    Zhao, Bo; Mar, Jessica C; Maruo, Seiji; Lee, Sungwook; Gewurz, Benjamin E; Johannsen, Eric; Holton, Kristina; Rubio, Renee; Takada, Kenzo; Quackenbush, John; Kieff, Elliott

    2011-01-01

    EBV nuclear antigen 3C (EBNA3C) is an essential transcription factor for EBV transformed lymphoblast cell line (LCL) growth. To identify EBNA3C-regulated genes in LCLs, microarrays were used to measure RNA abundances in each of three different LCLs that conditionally express EBNA3C fused to a 4-OH-Tamoxifen-dependent estrogen receptor hormone binding domain (EBNA3CHT). At least three RNAs were assayed for each EBNA3CHT LCL under nonpermissive conditions, permissive conditions, and nonpermissive conditions with wild-type EBNA3C transcomplementation. Using a two-way ANOVA model of EBNA3C levels, we identified 550 regulated genes that were at least 1.5-fold up- or down-regulated with false discovery rates < 0.01. EBNA3C-regulated genes overlapped significantly with genes regulated by EBNA2 and EBNA3A consistent with coordinated effects on cell gene transcription. Of the 550 EBNA3C-regulated genes, 106 could be placed in protein networks. A seeded Bayesian network analysis of the 80 most significant EBNA3C-regulated genes suggests that RAC1, LYN, and TNF are upstream of other EBNA3C-regulated genes. Gene set enrichment analysis found enrichment for MAP kinase signaling, cytokine-cytokine receptor interactions, JAK-STAT signaling, and cell adhesion molecules, implicating these pathways in EBNA3C effects on LCL growth or survival. EBNA3C significantly up-regulated the CXCL12 ligand and its CXCR4 receptor and increased LCL migration. CXCL12 up-regulation depended on EBNA3C's interaction with the cell transcription factor, RBPJ, which is essential for LCL growth. EBNA3C also up-regulated MYC 1.3-fold and down-regulated CDKN2A exons 2 and 3, shared by p16 and p14, 1.4-fold, with false discovery rates < 5 × 10(-4).

  17. Amphibian transcription factor IIIA proteins contain a sequence element functionally equivalent to the nuclear export signal of human immunodeficiency virus type 1 Rev.

    PubMed

    Fridell, R A; Fischer, U; Lührmann, R; Meyer, B E; Meinkoth, J L; Malim, M H; Cullen, B R

    1996-04-01

    The human immunodeficiency virus type 1 (HIV-1) Rev protein is required for nuclear export of late HIV-1 mRNAs. This function is dependent on the mutationally defined Rev activation domain, which also forms a potent nuclear export signal. Transcription factor IIIA (TFIIIA) binds to 5S rRNA transcripts and this interaction has been proposed to play a role in the efficient nuclear export of 5S rRNA in amphibian oocytes. Here it is reported that amphibian TFIIIA proteins contain a sequence element with homology to the Rev activation domain that effectively substitutes for this domain in inducing the nuclear export of late HIV-1 mRNAs. It is further demonstrated that this TFIIIA sequence element functions as a protein nuclear export signal in both human cells and frog oocytes. Thus, this shared protein motif may play an analogous role in mediating the nuclear export of both late HIV-1 RNAs and 5S rRNA transcripts. PMID:8610146

  18. A Novel Antiviral Target Structure Involved in the RNA Binding, Dimerization, and Nuclear Export Functions of the Influenza A Virus Nucleoprotein

    PubMed Central

    Yamada, Kazunori; Kondoh, Yasumitsu; Hikono, Hirokazu; Osada, Hiroyuki; Tomii, Kentaro; Saito, Takehiko; Aida, Yoko

    2015-01-01

    Developing antiviral therapies for influenza A virus (IAV) infection is an ongoing process because of the rapid rate of antigenic mutation and the emergence of drug-resistant viruses. The ideal strategy is to develop drugs that target well-conserved, functionally restricted, and unique surface structures without affecting host cell function. We recently identified the antiviral compound, RK424, by screening a library of 50,000 compounds using cell-based infection assays. RK424 showed potent antiviral activity against many different subtypes of IAV in vitro and partially protected mice from a lethal dose of A/WSN/1933 (H1N1) virus in vivo. Here, we show that RK424 inhibits viral ribonucleoprotein complex (vRNP) activity, causing the viral nucleoprotein (NP) to accumulate in the cell nucleus. In silico docking analysis revealed that RK424 bound to a small pocket in the viral NP. This pocket was surrounded by three functionally important domains: the RNA binding groove, the NP dimer interface, and nuclear export signal (NES) 3, indicating that it may be involved in the RNA binding, oligomerization, and nuclear export functions of NP. The accuracy of this binding model was confirmed in a NP-RK424 binding assay incorporating photo-cross-linked RK424 affinity beads and in a plaque assay evaluating the structure-activity relationship of RK424. Surface plasmon resonance (SPR) and pull-down assays showed that RK424 inhibited both the NP-RNA and NP-NP interactions, whereas size exclusion chromatography showed that RK424 disrupted viral RNA-induced NP oligomerization. In addition, in vitro nuclear export assays confirmed that RK424 inhibited nuclear export of NP. The amino acid residues comprising the NP pocket play a crucial role in viral replication and are highly conserved in more than 7,000 NP sequences from avian, human, and swine influenza viruses. Furthermore, we found that the NP pocket has a surface structure different from that of the pocket in host molecules. Taken

  19. A Novel Antiviral Target Structure Involved in the RNA Binding, Dimerization, and Nuclear Export Functions of the Influenza A Virus Nucleoprotein.

    PubMed

    Kakisaka, Michinori; Sasaki, Yutaka; Yamada, Kazunori; Kondoh, Yasumitsu; Hikono, Hirokazu; Osada, Hiroyuki; Tomii, Kentaro; Saito, Takehiko; Aida, Yoko

    2015-07-01

    Developing antiviral therapies for influenza A virus (IAV) infection is an ongoing process because of the rapid rate of antigenic mutation and the emergence of drug-resistant viruses. The ideal strategy is to develop drugs that target well-conserved, functionally restricted, and unique surface structures without affecting host cell function. We recently identified the antiviral compound, RK424, by screening a library of 50,000 compounds using cell-based infection assays. RK424 showed potent antiviral activity against many different subtypes of IAV in vitro and partially protected mice from a lethal dose of A/WSN/1933 (H1N1) virus in vivo. Here, we show that RK424 inhibits viral ribonucleoprotein complex (vRNP) activity, causing the viral nucleoprotein (NP) to accumulate in the cell nucleus. In silico docking analysis revealed that RK424 bound to a small pocket in the viral NP. This pocket was surrounded by three functionally important domains: the RNA binding groove, the NP dimer interface, and nuclear export signal (NES) 3, indicating that it may be involved in the RNA binding, oligomerization, and nuclear export functions of NP. The accuracy of this binding model was confirmed in a NP-RK424 binding assay incorporating photo-cross-linked RK424 affinity beads and in a plaque assay evaluating the structure-activity relationship of RK424. Surface plasmon resonance (SPR) and pull-down assays showed that RK424 inhibited both the NP-RNA and NP-NP interactions, whereas size exclusion chromatography showed that RK424 disrupted viral RNA-induced NP oligomerization. In addition, in vitro nuclear export assays confirmed that RK424 inhibited nuclear export of NP. The amino acid residues comprising the NP pocket play a crucial role in viral replication and are highly conserved in more than 7,000 NP sequences from avian, human, and swine influenza viruses. Furthermore, we found that the NP pocket has a surface structure different from that of the pocket in host molecules. Taken

  20. A Novel Antiviral Target Structure Involved in the RNA Binding, Dimerization, and Nuclear Export Functions of the Influenza A Virus Nucleoprotein.

    PubMed

    Kakisaka, Michinori; Sasaki, Yutaka; Yamada, Kazunori; Kondoh, Yasumitsu; Hikono, Hirokazu; Osada, Hiroyuki; Tomii, Kentaro; Saito, Takehiko; Aida, Yoko

    2015-07-01

    Developing antiviral therapies for influenza A virus (IAV) infection is an ongoing process because of the rapid rate of antigenic mutation and the emergence of drug-resistant viruses. The ideal strategy is to develop drugs that target well-conserved, functionally restricted, and unique surface structures without affecting host cell function. We recently identified the antiviral compound, RK424, by screening a library of 50,000 compounds using cell-based infection assays. RK424 showed potent antiviral activity against many different subtypes of IAV in vitro and partially protected mice from a lethal dose of A/WSN/1933 (H1N1) virus in vivo. Here, we show that RK424 inhibits viral ribonucleoprotein complex (vRNP) activity, causing the viral nucleoprotein (NP) to accumulate in the cell nucleus. In silico docking analysis revealed that RK424 bound to a small pocket in the viral NP. This pocket was surrounded by three functionally important domains: the RNA binding groove, the NP dimer interface, and nuclear export signal (NES) 3, indicating that it may be involved in the RNA binding, oligomerization, and nuclear export functions of NP. The accuracy of this binding model was confirmed in a NP-RK424 binding assay incorporating photo-cross-linked RK424 affinity beads and in a plaque assay evaluating the structure-activity relationship of RK424. Surface plasmon resonance (SPR) and pull-down assays showed that RK424 inhibited both the NP-RNA and NP-NP interactions, whereas size exclusion chromatography showed that RK424 disrupted viral RNA-induced NP oligomerization. In addition, in vitro nuclear export assays confirmed that RK424 inhibited nuclear export of NP. The amino acid residues comprising the NP pocket play a crucial role in viral replication and are highly conserved in more than 7,000 NP sequences from avian, human, and swine influenza viruses. Furthermore, we found that the NP pocket has a surface structure different from that of the pocket in host molecules. Taken

  1. The Hepatitis C Virus-Induced Membranous Web and Associated Nuclear Transport Machinery Limit Access of Pattern Recognition Receptors to Viral Replication Sites

    PubMed Central

    Neufeldt, Christopher J.; Joyce, Michael A.; Van Buuren, Nicholas; Levin, Aviad; Kirkegaard, Karla; Gale Jr., Michael; Tyrrell, D. Lorne J.; Wozniak, Richard W.

    2016-01-01

    Hepatitis C virus (HCV) is a positive-strand RNA virus of the Flaviviridae family and a major cause of liver disease worldwide. HCV replicates in the cytoplasm, and the synthesis of viral proteins induces extensive rearrangements of host cell membranes producing structures, collectively termed the membranous web (MW). The MW contains the sites of viral replication and assembly, and we have identified distinct membrane fractions derived from HCV-infected cells that contain replication and assembly complexes enriched for viral RNA and infectious virus, respectively. The complex membrane structure of the MW is thought to protect the viral genome limiting its interactions with cytoplasmic pattern recognition receptors (PRRs) and thereby preventing activation of cellular innate immune responses. Here we show that PRRs, including RIG-I and MDA5, and ribosomes are excluded from viral replication and assembly centers within the MW. Furthermore, we present evidence that components of the nuclear transport machinery regulate access of proteins to MW compartments. We show that the restricted assess of RIG-I to the MW can be overcome by the addition of a nuclear localization signal sequence, and that expression of a NLS-RIG-I construct leads to increased immune activation and the inhibition of viral replication. PMID:26863439

  2. The Hepatitis C Virus-Induced Membranous Web and Associated Nuclear Transport Machinery Limit Access of Pattern Recognition Receptors to Viral Replication Sites.

    PubMed

    Neufeldt, Christopher J; Joyce, Michael A; Van Buuren, Nicholas; Levin, Aviad; Kirkegaard, Karla; Gale, Michael; Tyrrell, D Lorne J; Wozniak, Richard W

    2016-02-01

    Hepatitis C virus (HCV) is a positive-strand RNA virus of the Flaviviridae family and a major cause of liver disease worldwide. HCV replicates in the cytoplasm, and the synthesis of viral proteins induces extensive rearrangements of host cell membranes producing structures, collectively termed the membranous web (MW). The MW contains the sites of viral replication and assembly, and we have identified distinct membrane fractions derived from HCV-infected cells that contain replication and assembly complexes enriched for viral RNA and infectious virus, respectively. The complex membrane structure of the MW is thought to protect the viral genome limiting its interactions with cytoplasmic pattern recognition receptors (PRRs) and thereby preventing activation of cellular innate immune responses. Here we show that PRRs, including RIG-I and MDA5, and ribosomes are excluded from viral replication and assembly centers within the MW. Furthermore, we present evidence that components of the nuclear transport machinery regulate access of proteins to MW compartments. We show that the restricted assess of RIG-I to the MW can be overcome by the addition of a nuclear localization signal sequence, and that expression of a NLS-RIG-I construct leads to increased immune activation and the inhibition of viral replication. PMID:26863439

  3. Expression of epstein-barr virus encoded nuclear antigen 1 in benign and malignant tissues harbouring EBV.

    PubMed Central

    Oudejans, J J; Dukers, D F; Jiwa, N M; van den Brule, A J; Grässer, F A; de Bruin, P C; Horstman, A; Vos, W; van Gorp, J; Middeldorp, J M; Meijer, C J

    1996-01-01

    AIMS: To determine levels of expression of Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) in benign and malignant tissues harbouring EBV in relation to EBNA1 promoter usage. METHODS: Expression of EBNA1 was investigated by means of immunohistochemistry using a mixture of two EBNA1 specific monoclonal antibodies, 1H4-1 and 2B4-1. The presence of EBV was detected by EBER1/2 RNA in situ hybridisation. Detection of promoter specific EBNA1 transcripts was by RT-PCR analysis. RESULTS: EBNA1 positive cells were detected in all 20 EBV associated B cell lymphomas, 18 of which had arisen in immunocompromised patients; in eight of nine EBV associated T cell lymphomas; in 11 of 27 EBV positive cases of Hodgkin's disease; and in reactive lymphoid tissue harbouring EBV, including four cases of infectious mononucleosis. A diffuse EBNA1 staining pattern was observed in most of the EBV associated B cell lymphomas and was comparable with the EBER1/2 staining pattern. In the T cell lymphomas the number of EBNA1 positive cells was usually considerably less than the number of EBER1/2 positive ones. RT-PCR analysis revealed that in tumours with restricted EBNA1 expression-that is, T cell lymphomas and Hodgkin's disease lesions, EBNA1 transcripts were usually generated only by the F/Q promoter, whereas in B cell lymphomas EBNA1 transcripts were usually generated by both the C/W and F/Q promoters. CONCLUSIONS: EBNA1 is expressed in all types of tissue harbouring EBV, but the level of expression varies greatly. This may be the result of differential promoter usage. Images PMID:8944608

  4. Influenza Virus Infection Induces the Nuclear Relocalization of the Hsp90 Co-Chaperone p23 and Inhibits the Glucocorticoid Receptor Response

    PubMed Central

    Chase, Geoffrey; dos Santos Afonso, Emmanuel; Picard, Didier; Schwemmle, Martin; Naffakh, Nadia

    2011-01-01

    The genomic RNAs of influenza A viruses are associated with the viral polymerase subunits (PB1, PB2, PA) and nucleoprotein (NP), forming ribonucleoprotein complexes (RNPs). Transcription/replication of the viral genome occurs in the nucleus of infected cells. A role for Hsp90 in nuclear import and assembly of newly synthetized RNA-polymerase subunits has been proposed. Here we report that the p23 cochaperone of Hsp90, which plays a major role in glucocorticoid receptor folding and function, associates with influenza virus polymerase. We show that p23 is not essential for viral multiplication in cultured cells but relocalizes to the nucleus in influenza virus-infected cells, which may alter some functions of p23 and Hsp90. Moreover, we show that influenza virus infection inhibits glucocorticoid receptor-mediated gene transactivation, and that this negative effect can occur through a p23-independent pathway. Viral-induced inhibition of the glucocorticoid receptor response might be of significant importance regarding the physiopathology of influenza infections in vivo. PMID:21853119

  5. Nuclear Localization of the C1 Factor (Host Cell Factor) in Sensory Neurons Correlates with Reactivation of Herpes Simplex Virus from Latency

    NASA Astrophysics Data System (ADS)

    Kristie, Thomas M.; Vogel, Jodi L.; Sears, Amy E.

    1999-02-01

    After a primary infection, herpes simplex virus is maintained in a latent state in neurons of sensory ganglia until complex stimuli reactivate viral lytic replication. Although the mechanisms governing reactivation from the latent state remain unknown, the regulated expression of the viral immediate early genes represents a critical point in this process. These genes are controlled by transcription enhancer complexes whose assembly requires and is coordinated by the cellular C1 factor (host cell factor). In contrast to other tissues, the C1 factor is not detected in the nuclei of sensory neurons. Experimental conditions that induce the reactivation of herpes simplex virus in mouse model systems result in rapid nuclear localization of the protein, indicating that the C1 factor is sequestered in these cells until reactivation signals induce a redistribution of the protein. The regulated localization suggests that C1 is a critical switch determinant of the viral lytic-latent cycle.

  6. Inhibition of human immunodeficiency virus type 1 (HIV-1) nuclear import via Vpr-Importin {alpha} interactions as a novel HIV-1 therapy

    SciTech Connect

    Suzuki, Tatsunori; Yamamoto, Norio; Nonaka, Mizuho; Hashimoto, Yoshie; Matsuda, Go; Takeshima, Shin-nosuke; Matsuyama, Megumi; Igarashi, Tatsuhiko; Miura, Tomoyuki; Tanaka, Rie; Kato, Shingo; Aida, Yoko

    2009-03-20

    The development of multidrug-resistant viruses compromises the efficacy of anti-human immunodeficiency virus (HIV) therapy and limits treatment options. Therefore, new targets that can be used to develop novel antiviral agents need to be identified. One such target is the interaction between Vpr, one of the accessory gene products of HIV-1 and Importin {alpha}, which is crucial, not only for the nuclear import of Vpr, but also for HIV-1 replication in macrophages. We have identified a potential parent compound, hematoxylin, which suppresses Vpr-Importin {alpha} interaction, thereby inhibiting HIV-1 replication in a Vpr-dependent manner. Analysis by real-time PCR demonstrated that hematoxylin specifically inhibited nuclear import step of pre-integration complex. Thus, hematoxylin is a new anti-HIV-1 inhibitor that targets the nuclear import of HIV-1 via the Vpr-Importin {alpha} interaction, suggesting that a specific inhibitor of the interaction between viral protein and the cellular factor may provide a new strategy for HIV-1 therapy.

  7. Effects of the nuclear localization of the N(pro) protein of classical swine fever virus on its virulence in pigs.

    PubMed

    Li, Yongfeng; Shen, Liang; Sun, Yuan; Wang, Xiao; Li, Chao; Huang, Junhua; Chen, Jianing; Li, Lianfeng; Zhao, Bibo; Luo, Yuzi; Li, Su; Qiu, Hua-Ji

    2014-12-01

    The N(pro) protein of classical swine fever virus (CSFV) is localized in the cytoplasm and nucleus. However, it is unknown whether the nuclear localization of N(pro) correlates with the virulence of CSFV in the host. Previously, we showed that the N(pro) protein fused with interferon regulatory factor 3 (IRF3) was present only in the cytoplasm. Here, we generated and evaluated a recombinant CSFV vSM-IRF3 harboring the IRF3 gene inserted into the N(pro) gene of the highly virulent CSFV Shimen strain. Compared to the even nuclear and cytoplasmic distribution of the enhanced green fluorescent protein (EGFP)-N(pro) fusion expressed by the recombinant CSFV EGFP-CSFV, vSM-IRF3 expressed an IRF3-N(pro) fusion protein that only was localized in the cytoplasm. vSM-IRF3 was markedly attenuated in vitro and in vivo, and the inoculated pigs were completely protected from lethal CSFV challenge, whereas the parental virus as well as EGFP-CSFV exhibited a typical virulent phenotype. Taken together, the nuclear localization of N(pro) plays a significant role in the CSFV replication and virulence.

  8. Herpesvirus gB-induced fusion between the virion envelope and outer nuclear membrane during virus egress is regulated by the viral US3 kinase.

    PubMed

    Wisner, Todd W; Wright, Catherine C; Kato, Akihisa; Kawaguchi, Yasushi; Mou, Fan; Baines, Joel D; Roller, Richard J; Johnson, David C

    2009-04-01

    Herpesvirus capsids collect along the inner surface of the nuclear envelope and bud into the perinuclear space. Enveloped virions then fuse with the outer nuclear membrane (NM). We previously showed that herpes simplex virus (HSV) glycoproteins gB and gH act in a redundant fashion to promote fusion between the virion envelope and the outer NM. HSV mutants lacking both gB and gH accumulate enveloped virions in herniations, vesicles that bulge into the nucleoplasm. Earlier studies had shown that HSV mutants lacking the viral serine/threonine kinase US3 also accumulate herniations. Here, we demonstrate that HSV gB is phosphorylated in a US3-dependent manner in HSV-infected cells, especially in a crude nuclear fraction. Moreover, US3 directly phosphorylated the gB cytoplasmic (CT) domain in in vitro assays. Deletion of gB in the context of a US3-null virus did not add substantially to defects in nuclear egress. The majority of the US3-dependent phosphorylation of gB involved the CT domain and amino acid T887, a residue present in a motif similar to that recognized by US3 in other proteins. HSV recombinants lacking gH and expressing either gB substitution mutation T887A or a gB truncated at residue 886 displayed substantial defects in nuclear egress. We concluded that phosphorylation of the gB CT domain is important for gB-mediated fusion with the outer NM. This suggested a model in which the US3 kinase is incorporated into the tegument layer (between the capsid and envelope) in HSV virions present in the perinuclear space. By this packaging, US3 might be brought close to the gB CT tail, leading to phosphorylation and triggering fusion between the virion envelope and the outer NM.

  9. Evaluation of the functional involvement of human immunodeficiency virus type 1 integrase in nuclear import of viral cDNA during acute infection.

    PubMed

    Ikeda, Tamako; Nishitsuji, Hironori; Zhou, Xin; Nara, Nobuo; Ohashi, Takashi; Kannagi, Mari; Masuda, Takao

    2004-11-01

    Nuclear import of viral cDNA is a critical step for establishing the proviral state of human immunodeficiency virus type 1 (HIV-1). The contribution of HIV-1 integrase (IN) to the nuclear import of viral cDNA is controversial, partly due to a lack of identification of its bona fide nuclear localization signal. In this study, to address this putative function of HIV-1 IN, the effects of mutations at key residues for viral cDNA recognition (PYNP at positions 142 to 145, K156, K159, and K160) were evaluated in the context of viral replication. During acute infection, some mutations (N144Q, PYNP>KL, and KKK>AAA) severely reduced viral gene expression to less than 1% the wild-type (WT) level. None of the mutations affected the synthesis of viral cDNA. Meanwhile, the levels of integrated viral cDNA produced by N144Q, PYNP>KL, and KKK>AAA mutants were severely reduced to less than 1% the WT level. Quantitative PCR analysis of viral cDNA in nuclei and fluorescence in situ hybridization analysis showed that these mutations significantly reduced the level of viral cDNA accumulation in nuclei. Further analysis revealed that IN proteins carrying the N144Q, PYNP>KL, and KKK>AAA mutations showed severely reduced binding to viral cDNA but kept their karyophilic properties. Taken together, these results indicate that mutations that reduced the binding of IN to viral cDNA resulted in severe impairment of virus infectivity, most likely by affecting the nuclear import of viral cDNA that proceeds integration. These results suggest that HIV-1 IN may be one of the critical constituents for the efficient nuclear import of viral cDNA.

  10. Epstein-Barr Virus Nuclear Antigen 3 (EBNA3) Proteins Regulate EBNA2 Binding to Distinct RBPJ Genomic Sites

    PubMed Central

    Wang, Anqi; Welch, Rene; Zhao, Bo; Ta, Tram; Keleş, Sündüz

    2015-01-01

    ABSTRACT Latent infection of B lymphocytes by Epstein-Barr virus (EBV) in vitro results in their immortalization into lymphoblastoid cell lines (LCLs); this latency program is controlled by the EBNA2 viral transcriptional activator, which targets promoters via RBPJ, a DNA binding protein in the Notch signaling pathway. Three other EBNA3 proteins (EBNA3A, EBNA3B, and EBNA3C) interact with RBPJ to regulate cell gene expression. The mechanism by which EBNAs regulate different genes via RBPJ remains unclear. Our chromatin immunoprecipitation with deep sequencing (ChIP-seq) analysis of the EBNA3 proteins analyzed in concert with prior EBNA2 and RBPJ data demonstrated that EBNA3A, EBNA3B, and EBNA3C bind to distinct, partially overlapping genomic locations. Although RBPJ interaction is critical for EBNA3A and EBNA3C growth effects, only 30 to 40% of EBNA3-bound sites colocalize with RBPJ. Using LCLs conditional for EBNA3A or EBNA3C activity, we demonstrate that EBNA2 binding at sites near EBNA3A- or EBNA3C-regulated genes is specifically regulated by the respective EBNA3. To investigate EBNA3 binding specificity, we identified sequences and transcription factors enriched at EBNA3A-, EBNA3B-, and EBNA3C-bound sites. This confirmed the prior observation that IRF4 is enriched at EBNA3A- and EBNA3C-bound sites and revealed IRF4 enrichment at EBNA3B-bound sites. Using IRF4-negative BJAB cells, we demonstrate that IRF4 is essential for EBNA3C, but not EBNA3A or EBNA3B, binding to specific sites. These results support a model in which EBNA2 and EBNA3s compete for distinct subsets of RBPJ sites to regulate cell genes and where EBNA3 subset specificity is determined by interactions with other cell transcription factors. IMPORTANCE Epstein-Barr virus (EBV) latent gene products cause human cancers and transform B lymphocytes into immortalized lymphoblastoid cell lines in vitro. EBV nuclear antigens (EBNAs) and membrane proteins constitutively activate pathways important for

  11. Nuclear localization of dengue virus nonstructural protein 5 does not strictly correlate with efficient viral RNA replication and inhibition of type I interferon signaling.

    PubMed

    Kumar, Anil; Bühler, Sandra; Selisko, Barbara; Davidson, Andrew; Mulder, Klaas; Canard, Bruno; Miller, Sven; Bartenschlager, Ralf

    2013-04-01

    Dengue virus (DENV) is an important human pathogen, especially in the tropical and subtropical parts of the world, causing considerable morbidity and mortality. DENV replication occurs in the cytoplasm; however, a high proportion of nonstructural protein 5 (NS5), containing methyltransferase (MTase) and RNA-dependent RNA polymerase (RdRp) activities, accumulates in the nuclei of infected cells. The present study investigates the impact of nuclear localization of NS5 on its known functions, including viral RNA replication and subversion of the type I interferon response. By using a mutation analysis approach, we identified the most critical residues within the αβ nuclear localization signal (αβNLS), which are essential for the nuclear accumulation of this protein. Although we observed an overall correlation between reduced nuclear accumulation of NS5 and impaired RNA replication, we identified one mutant with drastically reduced amounts of nuclear NS5 and virtually unaffected RNA replication, arguing that nuclear localization of NS5 does not correlate strictly with DENV replication, at least in cell culture. Because NS5 plays an important role in blocking interferon signaling via STAT-2 (signal transducer and activator of transcription 2) degradation, the abilities of the NLS mutants to block this pathway were investigated. All mutants were able to degrade STAT-2, with accordingly similar type I interferon resistance phenotypes. Since the NLS is contained within the RdRp domain, the MTase and RdRp activities of the mutants were determined by using recombinant full-length NS5. We found that the C-terminal region of the αβNLS is a critical functional element of the RdRp domain required for polymerase activity. These results indicate that efficient DENV RNA replication requires only minimal, if any, nuclear NS5, and they identify the αβNLS as a structural element required for proper RdRp activity. PMID:23408610

  12. Genome-wide analysis of host-chromosome binding sites for Epstein-Barr Virus Nuclear Antigen 1 (EBNA1)

    PubMed Central

    2010-01-01

    The Epstein-Barr Virus (EBV) Nuclear Antigen 1 (EBNA1) protein is required for the establishment of EBV latent infection in proliferating B-lymphocytes. EBNA1 is a multifunctional DNA-binding protein that stimulates DNA replication at the viral origin of plasmid replication (OriP), regulates transcription of viral and cellular genes, and tethers the viral episome to the cellular chromosome. EBNA1 also provides a survival function to B-lymphocytes, potentially through its ability to alter cellular gene expression. To better understand these various functions of EBNA1, we performed a genome-wide analysis of the viral and cellular DNA sites associated with EBNA1 protein in a latently infected Burkitt lymphoma B-cell line. Chromatin-immunoprecipitation (ChIP) combined with massively parallel deep-sequencing (ChIP-Seq) was used to identify cellular sites bound by EBNA1. Sites identified by ChIP-Seq were validated by conventional real-time PCR, and ChIP-Seq provided quantitative, high-resolution detection of the known EBNA1 binding sites on the EBV genome at OriP and Qp. We identified at least one cluster of unusually high-affinity EBNA1 binding sites on chromosome 11, between the divergent FAM55 D and FAM55B genes. A consensus for all cellular EBNA1 binding sites is distinct from those derived from the known viral binding sites, suggesting that some of these sites are indirectly bound by EBNA1. EBNA1 also bound close to the transcriptional start sites of a large number of cellular genes, including HDAC3, CDC7, and MAP3K1, which we show are positively regulated by EBNA1. EBNA1 binding sites were enriched in some repetitive elements, especially LINE 1 retrotransposons, and had weak correlations with histone modifications and ORC binding. We conclude that EBNA1 can interact with a large number of cellular genes and chromosomal loci in latently infected cells, but that these sites are likely to represent a complex ensemble of direct and indirect EBNA1 binding sites. PMID

  13. Yellow mosaic symptom caused by the nuclear shuttle protein gene of mungbean yellow mosaic virus is associated with single-stranded DNA accumulation and mesophyll spread of the virus.

    PubMed

    Kuruba, B L; Buvani, A P; Veluthambi, K

    2016-01-01

    Mungbean yellow mosaic virus-[India:Vigna] (MYMV-[IN:Vig]), a blackgram isolate of MYMV, causes yellow mosaic disease in blackgram and mungbean. Two variable DNA-B components, KA22 and KA27, cause distinct symptoms in blackgram [V. mungo (L.) Hepper] with the same DNA-A component. KA22 + DNA-A-agroinoculated blackgram plants displayed yellow mosaic symptom and accumulated high levels of viral single-stranded (ss) DNA. KA27 + DNA-A-agroinoculated blackgram plants displayed severe stunting symptom and accumulated very low levels of viral ssDNA. However, in mungbean [V. radiata (L.) Wilczek], KA27 + DNA-A caused yellow mosaic symptom and a high level of viral ssDNA accumulated. Swapping of KA27 DNA-B with the nuclear shuttle protein gene (NSP) of KA22 DNA-B (KA27xKA22 NSP) caused yellow mosaic symptom in blackgram, suggesting that KA22 NSP is the determinant of yellow mosaic symptom. Interestingly, KA27xKA22 NSP-infected blackgram plants accumulated high levels of viral ssDNA, comparable to that of KA22 DNA-B infection, suggesting that the KA22 NSP is responsible for accumulation of high levels of viral ssDNA. MYMV distribution was studied in blackgram and mungbean plants by leaf tissue hybridization, which showed mesophyll spread of the virus in KA22-infected blackgram leaflets and in KA27-infected mungbean leaflets, both of which displayed yellow mosaic symptom. However, the virus did not accumulate in the mesophyll in the case of KA27-infected blackgram leaflets. Interestingly, the swapped KA27xKA22 NSP-infected blackgram leaflets showed mesophyll accumulation of the virus, suggesting that KA22 NSP determines its mesophyll spread. PMID:27640432

  14. Yellow mosaic symptom caused by the nuclear shuttle protein gene of mungbean yellow mosaic virus is associated with single-stranded DNA accumulation and mesophyll spread of the virus.

    PubMed

    Kuruba, B L; Buvani, A P; Veluthambi, K

    2016-01-01

    Mungbean yellow mosaic virus-[India:Vigna] (MYMV-[IN:Vig]), a blackgram isolate of MYMV, causes yellow mosaic disease in blackgram and mungbean. Two variable DNA-B components, KA22 and KA27, cause distinct symptoms in blackgram [V. mungo (L.) Hepper] with the same DNA-A component. KA22 + DNA-A-agroinoculated blackgram plants displayed yellow mosaic symptom and accumulated high levels of viral single-stranded (ss) DNA. KA27 + DNA-A-agroinoculated blackgram plants displayed severe stunting symptom and accumulated very low levels of viral ssDNA. However, in mungbean [V. radiata (L.) Wilczek], KA27 + DNA-A caused yellow mosaic symptom and a high level of viral ssDNA accumulated. Swapping of KA27 DNA-B with the nuclear shuttle protein gene (NSP) of KA22 DNA-B (KA27xKA22 NSP) caused yellow mosaic symptom in blackgram, suggesting that KA22 NSP is the determinant of yellow mosaic symptom. Interestingly, KA27xKA22 NSP-infected blackgram plants accumulated high levels of viral ssDNA, comparable to that of KA22 DNA-B infection, suggesting that the KA22 NSP is responsible for accumulation of high levels of viral ssDNA. MYMV distribution was studied in blackgram and mungbean plants by leaf tissue hybridization, which showed mesophyll spread of the virus in KA22-infected blackgram leaflets and in KA27-infected mungbean leaflets, both of which displayed yellow mosaic symptom. However, the virus did not accumulate in the mesophyll in the case of KA27-infected blackgram leaflets. Interestingly, the swapped KA27xKA22 NSP-infected blackgram leaflets showed mesophyll accumulation of the virus, suggesting that KA22 NSP determines its mesophyll spread.

  15. The Herpes Simplex Virus Protein pUL31 Escorts Nucleocapsids to Sites of Nuclear Egress, a Process Coordinated by Its N-Terminal Domain

    PubMed Central

    Nagel, Claus-Henning; Binz, Anne; Sodeik, Beate; Bauerfeind, Rudolf; Bailer, Susanne M.

    2015-01-01

    Progeny capsids of herpesviruses leave the nucleus by budding through the nuclear envelope. Two viral proteins, the membrane protein pUL34 and the nucleo-phosphoprotein pUL31 form the nuclear egress complex that is required for capsid egress out of the nucleus. All pUL31 orthologs are composed of a diverse N-terminal domain with 1 to 3 basic patches and a conserved C-terminal domain. To decipher the functions of the N-terminal domain, we have generated several Herpes simplex virus mutants and show here that the N-terminal domain of pUL31 is essential with basic patches being critical for viral propagation. pUL31 and pUL34 entered the nucleus independently of each other via separate routes and the N-terminal domain of pUL31 was required to prevent their premature interaction in the cytoplasm. Unexpectedly, a classical bipartite nuclear localization signal embedded in this domain was not required for nuclear import of pUL31. In the nucleus, pUL31 associated with the nuclear envelope and newly formed capsids. Viral mutants lacking the N-terminal domain or with its basic patches neutralized still associated with nucleocapsids but were unable to translocate them to the nuclear envelope. Replacing the authentic basic patches with a novel artificial one resulted in HSV1(17+)Lox-UL31-hbpmp1mp2, that was viable but delayed in nuclear egress and compromised in viral production. Thus, while the C-terminal domain of pUL31 is sufficient for the interaction with nucleocapsids, the N-terminal domain was essential for capsid translocation to sites of nuclear egress and a coordinated interaction with pUL34. Our data indicate an orchestrated sequence of events with pUL31 binding to nucleocapsids and escorting them to the inner nuclear envelope. We propose a common mechanism for herpesviral nuclear egress: pUL31 is required for intranuclear translocation of nucleocapsids and subsequent interaction with pUL34 thereby coupling capsid maturation with primary envelopment. PMID:26083367

  16. The Herpes Simplex Virus Protein pUL31 Escorts Nucleocapsids to Sites of Nuclear Egress, a Process Coordinated by Its N-Terminal Domain.

    PubMed

    Funk, Christina; Ott, Melanie; Raschbichler, Verena; Nagel, Claus-Henning; Binz, Anne; Sodeik, Beate; Bauerfeind, Rudolf; Bailer, Susanne M

    2015-06-01

    Progeny capsids of herpesviruses leave the nucleus by budding through the nuclear envelope. Two viral proteins, the membrane protein pUL34 and the nucleo-phosphoprotein pUL31 form the nuclear egress complex that is required for capsid egress out of the nucleus. All pUL31 orthologs are composed of a diverse N-terminal domain with 1 to 3 basic patches and a conserved C-terminal domain. To decipher the functions of the N-terminal domain, we have generated several Herpes simplex virus mutants and show here that the N-terminal domain of pUL31 is essential with basic patches being critical for viral propagation. pUL31 and pUL34 entered the nucleus independently of each other via separate routes and the N-terminal domain of pUL31 was required to prevent their premature interaction in the cytoplasm. Unexpectedly, a classical bipartite nuclear localization signal embedded in this domain was not required for nuclear import of pUL31. In the nucleus, pUL31 associated with the nuclear envelope and newly formed capsids. Viral mutants lacking the N-terminal domain or with its basic patches neutralized still associated with nucleocapsids but were unable to translocate them to the nuclear envelope. Replacing the authentic basic patches with a novel artificial one resulted in HSV1(17+)Lox-UL31-hbpmp1mp2, that was viable but delayed in nuclear egress and compromised in viral production. Thus, while the C-terminal domain of pUL31 is sufficient for the interaction with nucleocapsids, the N-terminal domain was essential for capsid translocation to sites of nuclear egress and a coordinated interaction with pUL34. Our data indicate an orchestrated sequence of events with pUL31 binding to nucleocapsids and escorting them to the inner nuclear envelope. We propose a common mechanism for herpesviral nuclear egress: pUL31 is required for intranuclear translocation of nucleocapsids and subsequent interaction with pUL34 thereby coupling capsid maturation with primary envelopment. PMID:26083367

  17. Distinct roles of enhancer nuclear factor 1 (NF1) sites in plasmacytoma and osteopetrosis induction by Akv1-99 murine leukemia virus

    SciTech Connect

    Sorensen, Karina Dalsgaard; Sorensen, Annette Balle; Quintanilla-Martinez, Leticia; Kunder, Sandra; Schmidt, Joerg; Pedersen, Finn Skou . E-mail: fsp@mb.au.dk

    2005-04-10

    Murine leukemia viruses (MLVs) can be lymphomagenic and bone pathogenic. In this work, the possible roles of two distinct proviral enhancer nuclear factor 1 (NF1) binding sites in osteopetrosis and tumor induction by B-lymphomagenic Akv1-99 MLV were investigated. Akv1-99 and mutants either with NF1 site 1, NF1 site 2 or both sites disrupted induced tumors (plasma cell proliferations by histopathology) with remarkably similar incidence and mean latency in inbred NMRI mice. Clonal immunoglobulin gene rearrangement detection, by Southern analysis, confirmed approximately half of the tumors induced by each virus to be plasmacytomas while the remaining lacked detectable clonally rearranged Ig genes and were considered polyclonal; a demonstration that enhancer NF1 sites are dispensable for plasmacytoma induction by Akv1-99. In contrast, X-ray analysis revealed significant differences in osteopetrosis induction by the four viruses strongly indicating that NF1 site 2 is critical for viral bone pathogenicity, whereas NF1 site 1 is neutral or moderately inhibitory. In conclusion, enhancer NF1 sites are major determinants of osteopetrosis induction by Akv1-99 without significant influence on viral oncogenicity.

  18. Baculovirus replication: characterization of DNA and proteins synthesized by a nuclear polyhedrosis virus of Lymantria dispar, the gypsy moth, in a homologous cell line

    SciTech Connect

    McClintock, J.T.

    1985-01-01

    A multiple-embedded nuclear polyhedrosis virus (NPV) of the gypsy moth, Lymantria dispar (LdMNPV), is used for biological control. However, LdMNPV has low natural virulence and a long infection cycle in relation to other NPVs. Therefore, the replicative cycle of LdMNPV was investigated using a homologous cell line, IPLB-LD-652Y. Based on analyses of virus growth curves LdMNPV nonoccluded virus and polyhedral inclusion bodies appeared approximately 20 and 50 hr postinfection (p.i.), respectively. LdMNPV polypeptides, identified by autoradiography of (/sup 35/S)-methionine labeled fractions in SDS-PAGE, were synthesized in sequential phases: (1) an early ..cap alpha.. phase of replication (4 polypeptides from 4 to 12 hr p.i.), (2) an intermediate ..beta.. phase (20 polypeptides from 12 to 24 hr p.i.), and a late ..gamma.. phase (4 polypeptides from 24 to 28 hr p.i.). In infected cells at least four polypeptides were post-translational cleaved and/or modified. Pulse-labeling with (/sup 3/H)-mannose, (/sup 3/H)-N-acetyl-glucosamine or (/sup 32/P)-monosodium phosphate revealed several viral polypeptides which were glycosylated and/or phosphorylated. DNA:DNA dot hybridization experiments suggested that LdMNPV DNA synthesis was initiated between 12 to 16 hr p.i., increasing significantly thereafter.

  19. The RING for gypsy moth control: Topical application of fragment of its nuclear polyhedrosis virus anti-apoptosis gene as insecticide.

    PubMed

    Oberemok, Volodymyr V; Laikova, Kateryna V; Zaitsev, Aleksei S; Gushchin, Vladimir A; Skorokhod, Oleksii A

    2016-07-01

    Numerous studies suggest a cellular origin for the Lymantria dispar multicapsid nuclear polyhedrosis virus (LdMNPV) anti-apoptosis genes IAPs, thus opening a possibility to use the fragments of these genes for modulation of host metabolism. We report here the strong insecticidal and metabolic effect of single-stranded antisense DNA fragment from RING (really interesting new gene) domain of gypsy moth LdMNPV IAP-3 gene: specifically, on reduction of biomass (by 35%) and survival of L. dispar caterpillars. The treatment with this DNA fragment leads to a significantly higher mortality rates of female insects (1.7 fold) accompanied with the signs of apoptosis. Additionally, we show increased expression of host IAP-1, caspase-4 and gelsolin genes in eggs laid by survived females treated with RING DNA fragment accompanied with calcium and magnesium imbalance, indicating that the strong stress reactions and metabolic effects are not confined to treated insects but likely led to apoptosis in eggs too. The proposed new approach for insect pest management, which can be considered as advancement of "microbial pesticides", is based on the application of the specific virus DNA, exploiting the knowledge about virus-pest interactions and putting it to the benefit of mankind.

  20. The RING for gypsy moth control: Topical application of fragment of its nuclear polyhedrosis virus anti-apoptosis gene as insecticide.

    PubMed

    Oberemok, Volodymyr V; Laikova, Kateryna V; Zaitsev, Aleksei S; Gushchin, Vladimir A; Skorokhod, Oleksii A

    2016-07-01

    Numerous studies suggest a cellular origin for the Lymantria dispar multicapsid nuclear polyhedrosis virus (LdMNPV) anti-apoptosis genes IAPs, thus opening a possibility to use the fragments of these genes for modulation of host metabolism. We report here the strong insecticidal and metabolic effect of single-stranded antisense DNA fragment from RING (really interesting new gene) domain of gypsy moth LdMNPV IAP-3 gene: specifically, on reduction of biomass (by 35%) and survival of L. dispar caterpillars. The treatment with this DNA fragment leads to a significantly higher mortality rates of female insects (1.7 fold) accompanied with the signs of apoptosis. Additionally, we show increased expression of host IAP-1, caspase-4 and gelsolin genes in eggs laid by survived females treated with RING DNA fragment accompanied with calcium and magnesium imbalance, indicating that the strong stress reactions and metabolic effects are not confined to treated insects but likely led to apoptosis in eggs too. The proposed new approach for insect pest management, which can be considered as advancement of "microbial pesticides", is based on the application of the specific virus DNA, exploiting the knowledge about virus-pest interactions and putting it to the benefit of mankind. PMID:27265824

  1. Herpes simplex virus glycoproteins gB and gH function in fusion between the virion envelope and the outer nuclear membrane.

    PubMed

    Farnsworth, Aaron; Wisner, Todd W; Webb, Michael; Roller, Richard; Cohen, Gary; Eisenberg, Roselyn; Johnson, David C

    2007-06-12

    Herpesviruses must traverse the nuclear envelope to gain access to the cytoplasm and, ultimately, to exit cells. It is believed that herpesvirus nucleocapsids enter the perinuclear space by budding through the inner nuclear membrane (NM). To reach the cytoplasm these enveloped particles must fuse with the outer NM and the unenveloped capsids then acquire a second envelope in the trans-Golgi network. Little is known about the process by which herpesviruses virions fuse with the outer NM. Here we show that a herpes simplex virus (HSV) mutant lacking both the two putative fusion glycoproteins gB and gH failed to cross the nuclear envelope. Enveloped virions accumulated in the perinuclear space or in membrane vesicles that bulged into the nucleoplasm (herniations). By contrast, mutants lacking just gB or gH showed only minor or no defects in nuclear egress. We concluded that either HSV gB or gH can promote fusion between the virion envelope and the outer NM. It is noteworthy that fusion associated with HSV entry requires the cooperative action of both gB and gH, suggesting that the two types of fusion (egress versus entry) are dissimilar processes.

  2. Inhibitory function of adapter-related protein complex 2 alpha 1 subunit in the process of nuclear translocation of human immunodeficiency virus type 1 genome

    SciTech Connect

    Kitagawa, Yukiko; Kameoka, Masanori Shoji-Kawata, Sanae; Iwabu, Yukie; Mizuta, Hiroyuki; Tokunaga, Kenzo; Fujino, Masato; Natori, Yukikazu; Yura, Yoshiaki; Ikuta, Kazuyoshi

    2008-03-30

    The transfection of human cells with siRNA against adapter-related protein complex 2 alpha 1 subunit (AP2{alpha}) was revealed to significantly up-regulate the replication of human immunodeficiency virus type 1 (HIV-1). This effect was confirmed by cell infection with vesicular stomatitis virus G protein-pseudotyped HIV-1 as well as CXCR4-tropic and CCR5-tropic HIV-1. Viral adsorption, viral entry and reverse transcription processes were not affected by cell transfection with siRNA against AP2{alpha}. In contrast, viral nuclear translocation as well as the integration process was significantly up-regulated in cells transfected with siRNA against AP2{alpha}. Confocal fluorescence microscopy revealed that a subpopulation of AP2{alpha} was not only localized in the cytoplasm but was also partly co-localized with lamin B, importin {beta} and Nup153, implying that AP2{alpha} negatively regulates HIV-1 replication in the process of nuclear translocation of viral DNA in the cytoplasm or the perinuclear region. We propose that AP2{alpha} may be a novel target for disrupting HIV-1 replication in the early stage of the viral life cycle.

  3. Tap and Dbp5, but not Gag, are involved in DR-mediated nuclear export of unspliced Rous sarcoma virus RNA

    SciTech Connect

    LeBlanc, Jason J.; Uddowla, Sabena; Abraham, Benjamin; Clatterbuck, Sarah; Beemon, Karen L. . E-mail: KLB@jhu.edu

    2007-07-05

    All retroviruses must circumvent cellular restrictions on the export of unspliced RNAs from the nucleus. While the unspliced RNA export pathways for HIV and Mason-Pfizer monkey virus are well characterized, that of Rous sarcoma virus (RSV) is not. We have previously reported that the RSV direct repeat (DR) elements are involved in the cytoplasmic accumulation of unspliced viral RNA. Here, using fluorescent in situ hybridization (FISH), we demonstrate that unspliced viral RNAs bearing a single point mutation (G8863C) in the DR exhibit a restricted cellular localization in and around the nucleus. In contrast, wild type unspliced viral RNA had a diffuse localization throughout the nucleus and cytoplasm. Since the RSV Gag protein has a transient localization in the nucleus, we examined the effect of Gag over-expression on a DR-mediated reporter construct. While Gag did not enhance DR-mediated nuclear export, the dominant-negative expression of two cellular export factors, Tap and Dbp5, inhibited expression of the same reporter construct. Furthermore, FISH studies using the dominant-negative Dbp5 demonstrated that unspliced wild type RSV RNA was retained within the nucleus. Taken together, these results further implicate the DR in nuclear RNA export through interactions with Tap and Dbp5.

  4. Large scale in vivo risk assessment of bovine viral diarrhea virus (BVDV) transmission through transfer of bovine embryos produced via somatic cell nuclear transfer (SCNT).

    PubMed

    Gregg, K; Gosch, G; Guerra, T; Chen, S H; Xiang, T; Broek, D; Bruner, B; Polejaeva, I

    2010-10-15

    The objective was to use the bovine viral diarrhea virus (BVDV) as a model to assess the risk of infectious disease transmission in the system of in vitro embryo production and transfer via somatic cell nuclear transfer (SCNT) technology. The risks of BVDV transmission in the SCNT embryo production were previously evaluated. In that in vitro study, following standard operating procedures (SOP), including pre-nuclear transfer donor cell testing, oocyte decontamination and virus-free cell and embryo culture conditions, SCNT embryos produced were free of detectable viral RNA. The current study focused on the evaluation of the potential risk of disease transmission from SCNT embryos to recipients, and the risk of producing persistently infected animals via SCNT embryo transfer. Blood samples were collected from 553 recipients of SCNT embryos and 438 cloned calves and tested for the presence of BVDV viral RNA via a sensitive real time PCR method. All samples tested were negative. These results, in conjunction with the previous in vitro study, confirmed that the established SCNT embryo production and transfer system is safe and presents no detectable risk of disease transmission.

  5. Interference of fisetin with targets of the nuclear factor-κB signal transduction pathway activated by Epstein-Barr virus encoded latent membrane protein 1.

    PubMed

    Li, Rong; Liang, Hong-Ying; Li, Ming-Yong; Lin, Chun-Yan; Shi, Meng-Jie; Zhang, Xiu-Juan

    2014-01-01

    Fisetin is an effective compound extracted from lacquer which has been used in the treatment of various diseases. Preliminary data indicate that it also exerts specific anti-cancer effects. However, the manner in which fisetin regulates cancer growth remains unknown. In this study, we elucidated interference of fisetin with targets of the nuclear factorκB signal transduction pathway activated by Epstein-Barr virus encoding latent membrane protein 1 (LMP1)in nasopharyngeal carcinoma (NPC) cells, Results showed that fisetin inhibited the survival rate of CNE-LMP1 cells and NF-κB activation caused by LMP1. Fisetin also suppressed nuclear translocation of NF-κB (p65) and IκBα phosphorylation, while inhibiting CyclinD1, all key targets of the NF-κB signal transduction pathway. It was suggested that interference effects of fisetin with signal transduction activated by LMP1 encoded by the Epstein-Barr virus may play an important role in its anticancer potential.

  6. The nuclear and adherent junction complex component protein ubinuclein negatively regulates the productive cycle of Epstein-Barr virus in epithelial cells.

    PubMed

    Gruffat, Henri; Lupo, Julien; Morand, Patrice; Boyer, Véronique; Manet, Evelyne

    2011-01-01

    The Epstein-Barr Virus (EBV) productive cycle is initiated by the expression of the viral trans-activator EB1 (also called Zebra, Zta, or BZLF1), which belongs to the basic leucine zipper transcription factor family. We have previously identified the cellular NACos (nuclear and adherent junction complex components) protein ubinuclein (Ubn-1) as a partner for EB1, but the function of this complex has never been studied. Here, we have evaluated the consequences of this interaction on the EBV productive cycle and find that Ubn-1 overexpression represses the EBV productive cycle whereas Ubn-1 downregulation by short hairpin RNA (shRNA) increases virus production. By a chromatin immunoprecipitation (ChIP) assay, we show that Ubn-1 blocks EB1-DNA interaction. We also show that in epithelial cells, relocalization and sequestration of Ubn-1 to the tight junctions of nondividing cells allow increased activation of the productive cycle. We propose a model in which Ubn-1 is a modulator of the EBV productive cycle: in proliferating epithelial cells, Ubn-1 is nuclear and inhibits activation of the productive cycle, whereas in differentiated cells, Ubn-1 is sequestrated to tight junctions, thereby allowing EB1 to fully function in the nucleus.

  7. The Nuclear and Adherent Junction Complex Component Protein Ubinuclein Negatively Regulates the Productive Cycle of Epstein-Barr Virus in Epithelial Cells▿

    PubMed Central

    Gruffat, Henri; Lupo, Julien; Morand, Patrice; Boyer, Véronique; Manet, Evelyne

    2011-01-01

    The Epstein-Barr Virus (EBV) productive cycle is initiated by the expression of the viral trans-activator EB1 (also called Zebra, Zta, or BZLF1), which belongs to the basic leucine zipper transcription factor family. We have previously identified the cellular NACos (nuclear and adherent junction complex components) protein ubinuclein (Ubn-1) as a partner for EB1, but the function of this complex has never been studied. Here, we have evaluated the consequences of this interaction on the EBV productive cycle and find that Ubn-1 overexpression represses the EBV productive cycle whereas Ubn-1 downregulation by short hairpin RNA (shRNA) increases virus production. By a chromatin immunoprecipitation (ChIP) assay, we show that Ubn-1 blocks EB1-DNA interaction. We also show that in epithelial cells, relocalization and sequestration of Ubn-1 to the tight junctions of nondividing cells allow increased activation of the productive cycle. We propose a model in which Ubn-1 is a modulator of the EBV productive cycle: in proliferating epithelial cells, Ubn-1 is nuclear and inhibits activation of the productive cycle, whereas in differentiated cells, Ubn-1 is sequestrated to tight junctions, thereby allowing EB1 to fully function in the nucleus. PMID:21084479

  8. Recruitment of activated IRF-3 and CBP/p300 to herpes simplex virus ICP0 nuclear foci: Potential role in blocking IFN-{beta} induction

    SciTech Connect

    Melroe, Gregory T.; Silva, Lindsey; Schaffer, Priscilla A.; Knipe, David M. . E-mail: david_knipe@hms.harvard.edu

    2007-04-10

    The host innate response to viral infection includes the production of interferons, which is dependent on the coordinated activity of multiple transcription factors. Herpes simplex virus 1 (HSV-1) has been shown to block efficient interferon expression by multiple mechanisms. We and others have demonstrated that HSV-1 can inhibit the transcription of genes promoted by interferon regulatory factor-3 (IRF-3), including interferon beta (IFN-{beta}), and that the immediate-early ICP0 protein is sufficient for this function. However, the exact mechanism by which ICP0 blocks IRF-3 activity has yet to be determined. Unlike some other viral proteins that inhibit IRF-3 activity, ICP0 does not appear to affect phosphorylation and dimerization of IRF-3. Here, we show that a portion of activated IRF-3 co-localizes with nuclear foci containing ICP0 at early times after virus infection. Co-localization to ICP0-containing foci is also seen with the IRF-3-binding partners and transcriptional co-activators, CBP and p300. In addition, using immunoprecipitation of infected cell lysates, we can immunoprecipitate a complex containing ICP0, IRF-3, and CBP. Thus we hypothesize that ICP0 recruits activated IRF-3 and CBP/p300 to nuclear structures, away from the host chromatin. This leads to the inactivation and accelerated degradation of IRF-3, resulting in reduced transcription of IFN-{beta} and an inhibition of the host response. Therefore, ICP0 provides an example of how viruses can block IFN-{beta} induction by sequestration of important transcription factors essential for the host response.

  9. Recruitment of Activated IRF-3 and CBP/p300 to Herpes Simplex Virus ICP0 Nuclear Foci: Potential Role in Blocking IFN-β Induction

    PubMed Central

    Melroe, Gregory T.; Silva, Lindsey; Schaffer, Priscilla A.; Knipe, David M.

    2007-01-01

    The host innate response to viral infection includes the production of interferons, which is dependent on the coordinated activity of multiple transcription factors. Herpes simplex virus 1 (HSV-1) has been shown to block efficient interferon expression by multiple mechanisms. We and others have demonstrated that HSV-1 can inhibit the transcription of genes promoted by Interferon Regulatory Factor-3 (IRF-3), including interferon beta (IFN-β), and that the immediate-early ICP0 protein is sufficient for this function. However, the exact mechanism by which ICP0 blocks IRF-3 activity has yet to be determined. Unlike some other viral proteins that inhibit IRF-3 activity, ICP0 does not appear to affect phosphorylation and dimerization of IRF-3. Here, we show that a portion of activated IRF-3 co-localizes with nuclear foci containing ICP0 at early times after virus infection. Co-localization to ICP0-containing foci is also seen with the IRF-3-binding partners and transcriptional co-activators, CBP and p300. In addition, using immunoprecipitation of infected cell lysates, we can immunoprecipitate a complex containing ICP0, IRF-3, and CBP. Thus we hypothesize that ICP0 recruits activated IRF-3 and CBP/p300 to nuclear structures, away from the host chromatin. This leads to the inactivation and accelerated degradation of IRF-3, resulting in reduced transcription of IFN-β and an inhibition of the host response. Therefore, ICP0 provides an example of how viruses can block IFN-β induction by sequestration of important transcription factors essential for the host response. PMID:17126870

  10. Conserved amino acids of the human immunodeficiency virus type 2 Vpx nuclear localization signal are critical for nuclear targeting of the viral preintegration complex in non-dividing cells

    SciTech Connect

    Belshan, Michael; Mahnke, Lisa A.; Ratner, Lee . E-mail: lratner@im.wustl.edu

    2006-03-01

    The HIV-2 viral accessory protein Vpx is related to, but distinct from the Vpr protein of HIV-1. Vpx is packaged into virions and as a component of the viral preintegration complex (PIC) is required for efficient virus replication in non-dividing cells. We have previously reported that the minimal transferable region of Vpx that contained karyophilic properties was aa 65 to 72. Analysis of Vpx sequences from various HIV-2/SIV strains reveals that this region contains highly conserved amino acids, including two basic residues (K68, R70) and three tyrosines (Y66, Y69, Y71). Here, we demonstrate that mutation of the basic or tyrosine residues abolishes PIC nuclear import in arrested cells as assessed by PCR detection of viral integration. Examination of cell-free virus by Western blot indicated that all mutant proteins were incorporated into virions, suggesting that the lack of replication in arrested cells was not due to a loss of Vpx in target cells. Together, these studies map critical residues of the Vpx nuclear localization signal that are required for efficient infection of non-dividing cells.

  11. Heterogeneous nuclear ribonuclear protein K interacts with Sindbis virus nonstructural proteins and viral subgenomic mRNA

    SciTech Connect

    Burnham, Andrew J.; Gong, Lei; Hardy, Richard W.

    2007-10-10

    Alphaviruses are a group of arthropod-borne human and animal pathogens that can cause epidemics of significant public health and economic consequence. Alphavirus RNA synthesis requires four virally encoded nonstructural proteins and probably a number of cellular proteins. Using comparative two-dimensional electrophoresis we were able to identify proteins enriched in cytoplasmic membrane fractions containing viral RNA synthetic complexes following infection with Sindbis virus. Our studies demonstrated the following: (i) the host protein hnRNP K is enriched in cytoplasmic membrane fractions following Sindbis virus infection, (ii) viral nonstructural proteins co-immunoprecipitate with hnRNP K, (iii) nsP2 and hnRNP K co-localize in the cytoplasm of Sindbis virus infected cells, (iv) Sindbis virus subgenomic mRNA, but not genomic RNA co-immunoprecipitates with hnRNP K, (v) viral RNA does not appear to be required for the interaction of hnRNP K with the nonstructural proteins. Potential functions of hnRNP K during virus replication are discussed.

  12. BZLF1, an Epstein-Barr virus immediate-early protein, induces p65 nuclear translocation while inhibiting p65 transcriptional function

    SciTech Connect

    Morrison, Thomas E.; Kenney, Shannon C. . E-mail: shann@med.unc.edu

    2004-10-25

    We have previously demonstrated that the Epstein-Barr virus immediate-early BZLF1 protein interacts with, and is inhibited by, the NF-{kappa}B family member p65. However, the effects of BZLF1 on NF-{kappa}B activity have not been intensively studied. Here we show that BZLF1 inhibits p65-dependent gene expression. BZLF1 inhibited the ability of IL-1, as well as transfected p65, to activate the expression of two different NF-{kappa}B-responsive genes, ICAM-1 and I{kappa}B-{alpha}. BZLF1 also reduced the constitutive level of I{kappa}B-{alpha} protein in HeLa and A549 cells, and increased the amount of nuclear NF-{kappa}B to a similar extent as tumor necrosis factor-alpha (TNF-{alpha}) treatment. In spite of this BZLF1-associated increase in the nuclear form of NF-{kappa}B, BZLF1 did not induce binding of NF-{kappa}B to NF-{kappa}B responsive promoters (as determined by chromatin immunoprecipitation assay) in vivo, although TNF-{alpha} treatment induced NF-{kappa}B binding as expected. Overexpression of p65 dramatically inhibited the lytic replication cycle of EBV in 293-EBV cells, confirming that NF-{kappa}B also inhibits BZLF1 transcriptional function. Our results are consistent with a model in which BZLF1 inhibits the transcriptional function of p65, resulting in decreased transcription of I{kappa}B-{alpha}, decreased expression of I{kappa}B-{alpha} protein, and subsequent translocation of NF-{kappa}B to the nucleus. This nuclear translocation of NF-{kappa}B may promote viral latency by negatively regulating BZLF1 transcriptional activity. In situations where p65 activity is limiting in comparison to BZLF1, the ability of BZLF1 to inhibit p65 transcriptional function may protect the virus from the host immune system during the lytic form of infection.

  13. A high-throughput screening system targeting the nuclear export pathway via the third nuclear export signal of influenza A virus nucleoprotein.

    PubMed

    Kakisaka, Michinori; Mano, Takafumi; Aida, Yoko

    2016-06-01

    Two classes of antiviral drugs, M2 channel inhibitors and neuraminidase (NA) inhibitors, are currently approved for the treatment of influenza; however, the development of resistance against these agents limits their efficacy. Therefore, the identification of new targets and the development of new antiviral drugs against influenza are urgently needed. The third nuclear export signal (NES3) of nucleoprotein (NP) is the most important for viral replication among seven NESs encoded by four viral proteins, NP, M1, NS1, and NS2. NP-NES3 is critical for the nuclear export of NP, and targeting NP-NES3 is therefore a promising strategy that may lead to the development of antiviral drugs. However, a high-throughput screening (HTS) system to identify inhibitors of NP nuclear export has not been established. Here, we developed a novel HTS system to evaluate the inhibitory effects of compounds on the nuclear export pathway mediated by NP-NES3 using a MDCK cell line stably expressing NP-NES3 fused to a green fluorescent protein from aequorea coerulescens (AcGFP-NP-NES3) and a cell imaging analyzer. This HTS system was used to screen a 9600-compound library, leading to the identification of several hit compounds with inhibitory activity against the nuclear export of AcGFP-NP-NES3. The present HTS system provides a useful strategy for the identification of inhibitors targeting the nuclear export of NP via its NES3 sequence. PMID:26948263

  14. A high-throughput screening system targeting the nuclear export pathway via the third nuclear export signal of influenza A virus nucleoprotein.

    PubMed

    Kakisaka, Michinori; Mano, Takafumi; Aida, Yoko

    2016-06-01

    Two classes of antiviral drugs, M2 channel inhibitors and neuraminidase (NA) inhibitors, are currently approved for the treatment of influenza; however, the development of resistance against these agents limits their efficacy. Therefore, the identification of new targets and the development of new antiviral drugs against influenza are urgently needed. The third nuclear export signal (NES3) of nucleoprotein (NP) is the most important for viral replication among seven NESs encoded by four viral proteins, NP, M1, NS1, and NS2. NP-NES3 is critical for the nuclear export of NP, and targeting NP-NES3 is therefore a promising strategy that may lead to the development of antiviral drugs. However, a high-throughput screening (HTS) system to identify inhibitors of NP nuclear export has not been established. Here, we developed a novel HTS system to evaluate the inhibitory effects of compounds on the nuclear export pathway mediated by NP-NES3 using a MDCK cell line stably expressing NP-NES3 fused to a green fluorescent protein from aequorea coerulescens (AcGFP-NP-NES3) and a cell imaging analyzer. This HTS system was used to screen a 9600-compound library, leading to the identification of several hit compounds with inhibitory activity against the nuclear export of AcGFP-NP-NES3. The present HTS system provides a useful strategy for the identification of inhibitors targeting the nuclear export of NP via its NES3 sequence.

  15. Identification of a purified complement-fixing antigen as the Epstein-Barr-virus determined nuclear antigen (EBNA) by its binding to metaphase chromosomes.

    PubMed

    Ohno, S; Luka, J; Lindahl, T; Klein, G

    1977-04-01

    A soluble complement-fixing antigen carried by Epstein-Barr virus (EBV)-transformed human cells has been previously extracted from cell nuclei and purified by DNA-cellulose chromatography [Luka, J., Siegert, W. & Klein, G. (1977) J. Virol., in press]. On addition of this antigen to methanol/acetic acid-fixed metaphase chrmosomes, followed by exposure to human sera containing antibodies against the EBV-determined nuclear antigen (EBNA), brilliant positive staining was obtained by anti-complement immunofluorescence. There was no staining after exposure to EBV-negative sera. Moreover, a nuclear protein fraction, prepared from an EBV-negative cell line in an analogous fashion, failed to induce the staining reaction. These data identify the soluble purified antigen as the EBV-determined nuclear antigen. The purified antigen has a molecular weight of 174,000 +/- 15,000, as determined by sucrose gradient centrifugation and gel filtration experiments. In neutral buffers containing 0.5-1.0 M NaCl, the antigen dissociates into a form of approximately one-half the original molecular weight with retained complement-fixing activity. This "monomer" has a molecular weight of 98,000 +/- 8,000. PMID:67603

  16. [Detection of the nuclear polyhedrosis virus DNA in samples from eggs and caterpillars at different stages of the gypsy moth Lymantria dispar (L.) population dynamics].

    PubMed

    Bakhvalov, S A; Martem'ianov, V V; Bakhvalova, V N; Morozova, O V

    2012-01-01

    The nuclear polyhedrosis virus (NPV) DNA was detected in samples from eggs and caterpillars of the gypsy moth collected in natural populations of the Western Siberia and Ural by means of PCR with primers corresponding to the polyhedrin gene. According to censuring data, the gypsy moth populations of Western Siberia were at the depression stage. The NPV DNA detection frequencies in eggs (8.6 +/- 4.8% - 13.6 +/- 5.2%) and caterpillars (21.0 +/- 6.3% - 22.2 +/- 6.7%) were not significantly differed. In the Urals, collection of the insects was performed in their gradation focus at the phase of maximal abundance. The DNA detection rate in eggs (11.4 +/- 5.0%) was confidently (p < 0.001) lower than in caterpillars (59.8 +/- 5.6%). Consequently, variations of the NPV infection prevalence during ontogenesis of Lymantria dispar (L.) was associated with the gradation cycle of the insect population dynamics.

  17. Expression dynamics and ultrastructural localization of epitope-tagged Abutilon mosaic virus nuclear shuttle and movement proteins in Nicotiana benthamiana cells

    SciTech Connect

    Kleinow, Tatjana; Tanwir, Fariha; Kocher, Cornelia; Krenz, Bjoern; Wege, Christina; Jeske, Holger

    2009-09-01

    The geminivirus Abutilon mosaic virus (AbMV) encodes two proteins which are essential for viral spread within plants. The nuclear shuttle protein (NSP) transfers viral DNA between the nucleus and cytoplasm, whereas the movement protein (MP) facilitates transport between cells through plasmodesmata and long-distance via phloem. An inducible overexpression system for epitope-tagged NSP and MP in plants yielded unprecedented amounts of both proteins. Western blots revealed extensive posttranslational modification and truncation for MP, but not for NSP. Ultrastructural examination of Nicotiana benthamiana tissues showed characteristic nucleopathic alterations, including fibrillar rings, when epitope-tagged NSP and MP were simultaneously expressed in leaves locally infected with an AbMV DNA A in which the coat protein gene was replaced by a green fluorescent protein encoding gene. Immunogold labelling localized NSP in the nucleoplasm and in the fibrillar rings. MP appeared at the cell periphery, probably the plasma membrane, and plasmodesmata.

  18. Enhanced responsiveness to nuclear factor kappa B contributes to the unique phenotype of simian immunodeficiency virus variant SIVsmmPBj14.

    PubMed Central

    Dollard, S C; Gummuluru, S; Tsang, S; Fultz, P N; Dewhurst, S

    1994-01-01

    Infection with a variant of simian immunodeficiency virus, SIVsmmPBj14, leads to severe acute disease in macaques. This study was designed to investigate the functional significance of previously described mutations in the viral long terminal repeat (LTR) and to elucidate their contribution to the unique phenotype of SIVsmmPBj14. LTR-directed transcription was measured by using luciferase reporter constructs that were transiently transfected into cultured cells. In a wide range of cell types, the basal transcriptional activity of the LTR from SIVsmmPBj14 was found to be 2- to 4.5-fold higher than that of an LTR from a non-acutely pathogenic strain. These LTRs differ by five point mutations and a 22-bp duplication in SIVsmmPBj14, which includes a nuclear factor kappa B (NF kappa B) site. Transcriptional differences between these LTRs were further enhanced by two- to threefold upon treatment of cells with phorbol ester or tumor necrosis factor alpha or by cotransfection with plasmids expressing NF kappa B subunits. Mutagenesis studies, and the use of a reporter construct containing an enhancerless promoter, indicate that these transcriptional effects are due principally to the 22-bp sequence duplication and the NF kappa B site contained within it. Finally, infectious virus stocks that were isogenic except for the LTR were generated. The LTR from SIVsmmPBj14 was found to confer an increase in the kinetics of virus replication in cultured cells. Inclusion of this LTR in recombinant SIVs also resulted in a two- to threefold rise in the extent of cellular proliferation that was induced in quiescent simian peripheral blood mononuclear cells. These studies are consistent with the hypothesis that LTR mutations assist SIVsmmPBj14 in responding efficiently to cellular stimulation and allow it to replicate to high titers during the acute phase of viral infection. Images PMID:7966569

  19. Light-Activated Nuclear Translocation of Adeno-Associated Virus Nanoparticles Using Phytochrome B for Enhanced, Tunable, and Spatially Programmable Gene Delivery.

    PubMed

    Gomez, Eric J; Gerhardt, Karl; Judd, Justin; Tabor, Jeffrey J; Suh, Junghae

    2016-01-26

    Gene delivery vectors that are activated by external stimuli may allow improved control over the location and the degree of gene expression in target populations of cells. Light is an attractive stimulus because it does not cross-react with cellular signaling networks, has negligible toxicity, is noninvasive, and can be applied in space and time with unparalleled precision. We used the previously engineered red (R)/far-red (FR) light-switchable protein phytochrome B (PhyB) and its R light dependent interaction partner phytochrome interacting factor 6 (PIF6) from Arabidopsis thaliana to engineer an adeno-associated virus (AAV) platform whose gene delivery efficiency is controlled by light. Upon exposure to R light, AAV engineered to display PIF6 motifs on the capsid bind to PhyB tagged with a nuclear localization sequence (NLS), resulting in significantly increased translocation of viruses into the host cell nucleus and overall gene delivery efficiency. By modulating the ratio of R to FR light, the gene delivery efficiency can be tuned to as little as 35% or over 600% of the unengineered AAV. We also demonstrate spatial control of gene delivery using projected patterns of codelivered R and FR light. Overall, our successful use of light-switchable proteins in virus capsid engineering extends these important optogenetic tools into the adjacent realm of nucleic acid delivery and enables enhanced, tunable, and spatially controllable regulation of viral gene delivery. Our current light-triggered viral gene delivery prototype may be broadly useful for genetic manipulation of cells ex vivo or in vivo in transgenic model organisms, with the ultimate prospect of achieving dose- and site-specific gene expression profiles for either therapeutic (e.g., regenerative medicine) or fundamental discovery research efforts.

  20. Light-Activated Nuclear Translocation of Adeno-Associated Virus Nanoparticles Using Phytochrome B for Enhanced, Tunable, and Spatially Programmable Gene Delivery.

    PubMed

    Gomez, Eric J; Gerhardt, Karl; Judd, Justin; Tabor, Jeffrey J; Suh, Junghae

    2016-01-26

    Gene delivery vectors that are activated by external stimuli may allow improved control over the location and the degree of gene expression in target populations of cells. Light is an attractive stimulus because it does not cross-react with cellular signaling networks, has negligible toxicity, is noninvasive, and can be applied in space and time with unparalleled precision. We used the previously engineered red (R)/far-red (FR) light-switchable protein phytochrome B (PhyB) and its R light dependent interaction partner phytochrome interacting factor 6 (PIF6) from Arabidopsis thaliana to engineer an adeno-associated virus (AAV) platform whose gene delivery efficiency is controlled by light. Upon exposure to R light, AAV engineered to display PIF6 motifs on the capsid bind to PhyB tagged with a nuclear localization sequence (NLS), resulting in significantly increased translocation of viruses into the host cell nucleus and overall gene delivery efficiency. By modulating the ratio of R to FR light, the gene delivery efficiency can be tuned to as little as 35% or over 600% of the unengineered AAV. We also demonstrate spatial control of gene delivery using projected patterns of codelivered R and FR light. Overall, our successful use of light-switchable proteins in virus capsid engineering extends these important optogenetic tools into the adjacent realm of nucleic acid delivery and enables enhanced, tunable, and spatially controllable regulation of viral gene delivery. Our current light-triggered viral gene delivery prototype may be broadly useful for genetic manipulation of cells ex vivo or in vivo in transgenic model organisms, with the ultimate prospect of achieving dose- and site-specific gene expression profiles for either therapeutic (e.g., regenerative medicine) or fundamental discovery research efforts. PMID:26618393

  1. The open reading frame VI product of Cauliflower mosaic virus is a nucleocytoplasmic protein: its N terminus mediates its nuclear export and formation of electron-dense viroplasms.

    PubMed

    Haas, Muriel; Geldreich, Angèle; Bureau, Marina; Dupuis, Laurence; Leh, Véronique; Vetter, Guillaume; Kobayashi, Kappei; Hohn, Thomas; Ryabova, Lyubov; Yot, Pierre; Keller, Mario

    2005-03-01

    The Cauliflower mosaic virus (CaMV) open reading frame VI product (P6) is essential for the viral infection cycle. It controls translation reinitiation of the viral polycistronic RNAs and forms cytoplasmic inclusion bodies (viroplasms) where virus replication and assembly occur. In this study, the mechanism involved in viroplasm formation was investigated by in vitro and in vivo experiments. Far protein gel blot assays using a collection of P6 deletion mutants demonstrated that the N-terminal alpha-helix of P6 mediates interaction between P6 molecules. Transient expression in tobacco (Nicotiana tabacum) BY-2 cells of full-length P6 and P6 mutants fused to enhanced green fluorescent protein revealed that viroplasms are formed at the periphery of the nucleus and that the N-terminal domain of P6 is an important determinant in this process. Finally, this study led to the unexpected finding that P6 is a nucleocytoplasmic shuttle protein and that its nuclear export is mediated by a Leu-rich sequence that is part of the alpha-helix domain implicated in viroplasm formation. The discovery that P6 can localize to the nucleus opens new prospects for understanding yet unknown roles of this viral protein in the course of the CaMV infection cycle.

  2. The open reading frame VI product of Cauliflower mosaic virus is a nucleocytoplasmic protein: its N terminus mediates its nuclear export and formation of electron-dense viroplasms.

    PubMed

    Haas, Muriel; Geldreich, Angèle; Bureau, Marina; Dupuis, Laurence; Leh, Véronique; Vetter, Guillaume; Kobayashi, Kappei; Hohn, Thomas; Ryabova, Lyubov; Yot, Pierre; Keller, Mario

    2005-03-01

    The Cauliflower mosaic virus (CaMV) open reading frame VI product (P6) is essential for the viral infection cycle. It controls translation reinitiation of the viral polycistronic RNAs and forms cytoplasmic inclusion bodies (viroplasms) where virus replication and assembly occur. In this study, the mechanism involved in viroplasm formation was investigated by in vitro and in vivo experiments. Far protein gel blot assays using a collection of P6 deletion mutants demonstrated that the N-terminal alpha-helix of P6 mediates interaction between P6 molecules. Transient expression in tobacco (Nicotiana tabacum) BY-2 cells of full-length P6 and P6 mutants fused to enhanced green fluorescent protein revealed that viroplasms are formed at the periphery of the nucleus and that the N-terminal domain of P6 is an important determinant in this process. Finally, this study led to the unexpected finding that P6 is a nucleocytoplasmic shuttle protein and that its nuclear export is mediated by a Leu-rich sequence that is part of the alpha-helix domain implicated in viroplasm formation. The discovery that P6 can localize to the nucleus opens new prospects for understanding yet unknown roles of this viral protein in the course of the CaMV infection cycle. PMID:15746075

  3. Epstein–Barr virus nuclear antigen 3C interact with p73: Interplay between a viral oncoprotein and cellular tumor suppressor

    SciTech Connect

    Sahu, Sushil Kumar; Mohanty, Suchitra; Kumar, Amit; Kundu, Chanakya N.; Verma, Subhash C.; Choudhuri, Tathagata

    2014-01-05

    The p73 protein has structural and functional homology with the tumor suppressor p53, which plays an important role in cell cycle regulation, apoptosis, and DNA repair. The p73 locus encodes both a tumor suppressor (TAp73) and a putative oncogene (ΔNp73). p73 May play a significant role in p53-deficient lymphomas infected with Epstein–Barr virus (EBV). EBV produces an asymptomatic infection in the majority of the global population, but it is associated with several human B-cell malignancies. The EBV-encoded Epstein–Barr virus nuclear antigen 3C (EBNA3C) is thought to disrupt the cell cycle checkpoint by interacting directly with p53 family proteins. Doxorubicin, a commonly used chemotherapeutic agent, induces apoptosis through p53 and p73 signaling such that the lowΔNp73 level promotes the p73-mediated intrinsic pathway of apoptosis. In this report, we investigated the mechanism by which EBV infection counters p73α-induced apoptosis through EBNA3C. - Highlights: • EBV-encoded EBNA3C suppresses doxorubicin-induced apoptosis in B-cell lymphomas. • EBNA3C binds to p73 to suppress its apoptotic effect. • EBNA3C maintains latency by regulating downstream mitochondrial pathways.

  4. Redefining the Epstein-Barr virus-encoded nuclear antigen EBNA-1 gene promoter and transcription initiation site in group I Burkitt lymphoma cell lines.

    PubMed Central

    Schaefer, B C; Strominger, J L; Speck, S H

    1995-01-01

    The Epstein-Barr virus-encoded nuclear antigen EBNA-1 gene promoter for the restricted Epstein-Barr virus (EBV) latency program operating in group I Burkitt lymphoma (BL) cell lines was previously identified incorrectly. Here we present evidence from RACE (rapid amplification of cDNA ends) cloning, reverse transcription-PCR, and S1 nuclease analyses, which demonstrates that the EBNA-1 gene promoter in group I BL cell lines is located in the viral BamHI Q fragment, immediately upstream of two low-affinity EBNA-1 binding sites. Transcripts initiated from this promoter, referred to as Qp, have the previously reported Q/U/K exon splicing pattern. Qp is active in group I BL cell lines but not in group III BL cell lines or in EBV immortalized B-lymphoblastoid cell lines. In addition, transient transfection of Qp-driven reporter constructs into both an EBV-negative BL cell line and a group I BL cell line gave rise to correctly initiated transcripts. Inspection of Qp revealed that it is a TATA-less promoter whose architecture is similar to the promoters of housekeeping genes, suggesting that Qp may be a default promoter which ensures EBNA-1 expression in cells that cannot run the full viral latency program. Elucidation of the genetic mechanism responsible for the EBNA-1-restricted program of EBV latency is an essential step in understanding control of viral latency in EBV-associated tumors. Images Fig. 1 Fig. 3 Fig. 4 PMID:7479841

  5. Single-stranded DNA fragments of insect-specific nuclear polyhedrosis virus act as selective DNA insecticides for gypsy moth control.

    PubMed

    Oberemok, Volodymyr V; Skorokhod, Oleksii A

    2014-07-01

    This paper focuses on the DNA insecticides as a novel preparation against gypsy moth (Lymantria dispar) based on DNA fragments of the anti-apoptotic gene of its nuclear polyhedrosis virus. It was found that the external application of a solution with two single-stranded DNA fragments from BIR and RING domains of LdMNPV (L.dispar multicapsid nuclear polyhedrosis virus) IAP-3 (inhibitor of apoptosis) gene induces a significantly higher mortality of gypsy moth caterpillars in comparison with the application of the control solutions. This effect does not depend on the infection of caterpillars with LdMNPV. The results also show that DNA insecticides based on LdMNPV IAP-3 gene fragments can be selective in action, and at least are not harmful to tobacco hornworm (Manduca sexta) and black cutworm (Agrotis ipsilon). Part of the gypsy moth genome cloned with the fragments of BIR and RING domains of LdMNPV IAP-3 gene as primers, has an overlap with the corresponding part of the LdMNPV IAP-3 gene and L.dispar IAP-1 mRNA for an inhibitor of apoptosis protein with the high cover by query, allows assuming that we cloned a part of gypsy moth anti-apoptosis gene. This finding gives the grounding that proposed here DNA insecticides might act through the blocking of the mechanisms involved in post transcriptional expression of insect anti-apoptosis genes. The results show the insecticidal potential of the viral genome fragments that can be used to create safe and relatively fast-acting DNA insecticides to control the quantity of gypsy moth populations, important task for forestry and agriculture.

  6. Kaposi's sarcoma-associated herpesvirus noncoding polyadenylated nuclear RNA interacts with virus- and host cell-encoded proteins and suppresses expression of genes involved in immune modulation.

    PubMed

    Rossetto, Cyprian C; Pari, Gregory S

    2011-12-01

    During lytic infection, Kaposi's sarcoma-associated herpesvirus (KSHV) expresses a polyadenylated nuclear RNA (PAN RNA). This noncoding RNA (ncRNA) is localized to the nucleus and is the most abundant viral RNA during lytic infection; however, to date, the role of PAN RNA in the virus life cycle is unknown. Many examples exist where ncRNAs have a defined key regulatory function controlling gene expression by various mechanisms. Our goal for this study was to identify putative binding partners for PAN RNA in an effort to elucidate a possible function for the transcript in KSHV infection. We employed an in vitro affinity protocol where PAN RNA was used as bait for factors present in BCBL-1 cell nuclear extract to show that PAN RNA interacts with several virus- and host cell-encoded factors, including histones H1 and H2A, mitochondrial and cellular single-stranded binding proteins (SSBPs), and interferon regulatory factor 4 (IRF4). RNA chromatin immunoprecipitation (ChIP) assays confirmed that PAN RNA interacted with these factors in the infected cell environment. A luciferase reporter assay showed that PAN RNA expression interfered with the ability of IRF4/PU.1 to activate the interleukin-4 (IL-4) promoter, strongly suggesting a role for PAN RNA in immune modulation. Since the proteomic screen and functional data suggested a role in immune responses, we investigated if constitutive PAN RNA expression could affect other genes involved in immune responses. PAN RNA expression decreased expression of gamma interferon, interleukin-18, alpha interferon 16, and RNase L. These data strongly suggest that PAN RNA interacts with viral and cellular proteins and can function as an immune modulator. PMID:21957289

  7. Single-stranded DNA fragments of insect-specific nuclear polyhedrosis virus act as selective DNA insecticides for gypsy moth control.

    PubMed

    Oberemok, Volodymyr V; Skorokhod, Oleksii A

    2014-07-01

    This paper focuses on the DNA insecticides as a novel preparation against gypsy moth (Lymantria dispar) based on DNA fragments of the anti-apoptotic gene of its nuclear polyhedrosis virus. It was found that the external application of a solution with two single-stranded DNA fragments from BIR and RING domains of LdMNPV (L.dispar multicapsid nuclear polyhedrosis virus) IAP-3 (inhibitor of apoptosis) gene induces a significantly higher mortality of gypsy moth caterpillars in comparison with the application of the control solutions. This effect does not depend on the infection of caterpillars with LdMNPV. The results also show that DNA insecticides based on LdMNPV IAP-3 gene fragments can be selective in action, and at least are not harmful to tobacco hornworm (Manduca sexta) and black cutworm (Agrotis ipsilon). Part of the gypsy moth genome cloned with the fragments of BIR and RING domains of LdMNPV IAP-3 gene as primers, has an overlap with the corresponding part of the LdMNPV IAP-3 gene and L.dispar IAP-1 mRNA for an inhibitor of apoptosis protein with the high cover by query, allows assuming that we cloned a part of gypsy moth anti-apoptosis gene. This finding gives the grounding that proposed here DNA insecticides might act through the blocking of the mechanisms involved in post transcriptional expression of insect anti-apoptosis genes. The results show the insecticidal potential of the viral genome fragments that can be used to create safe and relatively fast-acting DNA insecticides to control the quantity of gypsy moth populations, important task for forestry and agriculture. PMID:25052520

  8. Relocalization of nuclear ALY proteins to the cytoplasm by the tomato bushy stunt virus P19 pathogenicity protein.

    PubMed

    Uhrig, Joachim F; Canto, Tomas; Marshall, David; MacFarlane, Stuart A

    2004-08-01

    The P19 protein of tomato bushy stunt virus (TBSV) is a multifunctional pathogenicity determinant involved in suppression of posttranscriptional gene silencing, virus movement, and symptom induction. Here, we report that P19 interacts with the conserved RNA-binding domain of an as yet uncharacterized family of plant ALY proteins that, in animals, are involved in export of RNAs from the nucleus and transcriptional coactivation. We show that the four ALY proteins encoded by the Arabidopsis genome and two ALY proteins from Nicotiana benthamiana are localized to the nucleus. Moreover, and in contrast to animal ALY, all but one of the proteins are also in the nucleolus, with distinct subnuclear localizations. Infection of plants by TBSV or expression of P19 from Agrobacterium results in relocation of three of the six ALY proteins from the nucleus to the cytoplasm demonstrating specific targeting of the ALY proteins by P19. The differential effects on subcellular localization indicate that, in plants, the various ALY proteins may have different functions. Interaction with and relocalization of ALY is prevented by mutation of P19 at residues previously shown to be important for P19 function in plants. Down-regulation of expression of two N. benthamiana ALY genes by virus-induced gene silencing did not interfere with posttranscriptional gene silencing. Targeting of ALY proteins during TBSV infection may therefore be related to functions of P19 in addition to its silencing suppression activity. PMID:15299117

  9. Promiscuous trans activation of gene expression by an Epstein-Barr virus-encoded early nuclear protein.

    PubMed Central

    Lieberman, P M; O'Hare, P; Hayward, G S; Hayward, S D

    1986-01-01

    We identified an Epstein-Barr virus (EBV) gene product which functions in transient-expression assays as a nonspecific trans activator. In Vero cells, cotransfection of the BglII J DNA fragment of EBV together with recombinant constructs containing the bacterial chloramphenicol acetyltransferase (CAT) gene gave up to a 100-fold increased expression of CAT activity over that in cells transfected with the recombinant CAT constructs alone. The BglII J fragment acted promiscuously, in that increased CAT synthesis was observed regardless of whether the promoter sequences driving the CAT gene were of EBV, simian virus 40, adenovirus, or herpes simplex virus origin. Cleavage of cloned BglII-J plasmid DNA before transfection revealed that activation was dependent upon the presence of an intact BMLF1 open reading frame. This was confirmed with subclones of BglII-J and with hybrid promoter-open reading frame constructs. This region of the genome is also present in the rearranged P3HR-1-defective DNA species, and defective DNA clones containing these sequences produced a similar activation of CAT expression in cotransfection experiments. The heterogeneous 45-60-kilodalton polypeptide product of BMLF1 may play an important regulatory role in expression of lytic-cycle proteins in EBV-infected lymphocytes. Images PMID:3018281

  10. The intergenic region of maize streak virus contains a GC-rich element that activates rightward transcription and binds maize nuclear factors.

    PubMed

    Fenoll, C; Schwarz, J J; Black, D M; Schneider, M; Howell, S H

    1990-12-01

    Maize streak virus (MSV) is transcribed bidirectionally from an intergenic region and rightward transcription produces an RNA that encodes the coat protein. The intergenic region contains promoter elements required for rightward transcription including an upstream activating sequence (UAS) which endows the promoter with full activity in a maize transient expression system. The UAS contains two GC-rich repeats (GC boxes) and a long inverted repeat or hairpin with a loop harboring a TAATATTAC sequence common to all geminiviruses. Deletions through the UAS demonstrated the presence of an element, called the rightward promoter element (rpe1), which is responsible for transcriptional activation. Rpe1 includes the two GC-rich boxes, which are similar in sequence to Sp1 binding sites in mammalian cells, but not the conserved hairpin loop. Rpe1 binds maize nuclear factors in vitro and the characteristics of the binding interaction have been determined by 1) binding competition with oligonucleotides, 2) methidiumpropyl-EDTA footprinting and 3) methylation interference assays. Binding of maize nuclear factors to the UAS generates two major bands, slow and fast migrating bands, in gel retardation assays. Footprinting and factor titration data suggest that the fast bands arise by the binding of factors to one GC box while the slow bands are generated by factors binding to both boxes. The data further indicate that the factors bind to the two GC-rich boxes with little cooperativity and bind on opposite faces of the DNA helix.

  11. Clustered basic amino acids of the small sendai virus C protein Y1 are critical to its RAN GTPase-mediated nuclear localization.

    PubMed

    Irie, Takashi; Yoshida, Asuka; Sakaguchi, Takemasa

    2013-01-01

    The Sendai virus (SeV) C proteins are shown to exert multiple functions during the course of infection. Perhaps reflecting their many functions, they occur at multiple sites of the cell. In this study, we focused on the nuclear-localizing ability of the smaller C protein, Y1, and found that this translocation is mediated by Ran GTPase but not by passive diffusion, and that basic residues within the 149-157 amino acid region are critical for that. The mechanism of inhibition of interferon (IFN)-signaling seemed to differ between the C and Y1 proteins, since deletion of 12 C-terminal amino acids resulted in a loss of the function for the C but not for the Y1 protein. The ability of Y1 mutants to inhibit IFN-α-induced, ISRE-driven expression of a reporter gene almost paralleled with that to localize in the nucleus. These results suggest that nuclear localization of the Y1 protein might be important for the inhibitory effect on type-I IFN-stimulated gene expression.

  12. Methamphetamine activates nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and induces human immunodeficiency virus (HIV) transcription in human microglial cells.

    PubMed

    Wires, Emily S; Alvarez, David; Dobrowolski, Curtis; Wang, Yun; Morales, Marisela; Karn, Jonathan; Harvey, Brandon K

    2012-10-01

    Human immunodeficiency virus (HIV) primarily infects glial cells in the central nervous system (CNS). Recent evidence suggests that HIV-infected individuals who abuse drugs such as methamphetamine (METH) have higher viral loads and experience more severe neurological complications than HIV-infected individuals who do not abuse drugs. The aim of this study was to determine the effect of METH on HIV expression from the HIV long terminal repeat (LTR) promoter and on an HIV integrated provirus in microglial cells, the primary host cells for HIV in the CNS. Primary human microglial cells immortalized with SV40 T antigen (CHME-5 cells) were cotransfected with an HIV LTR reporter and the HIV Tat gene, a key regulator of viral replication and gene expression, and exposed to METH. Our results demonstrate that METH treatment induced LTR activation, an effect potentiated in the presence of Tat. We also found that METH increased the nuclear translocation of the nuclear factor kappa B (NF-κB), a key cellular transcriptional regulator of the LTR promoter, and the activity of an NF-κB-specific reporter plasmid in CHME-5 cells. The presence of a dominant-negative regulator of NF-κB blocked METH-related activation of the HIV LTR. Furthermore, treatment of HIV-latently infected CHME-5 (CHME-5/HIV) cells with METH induced HIV expression and nuclear translocation of the p65 subunit of NF-κB. These results suggest that METH can stimulate HIV gene expression in microglia cells through activation of the NF-κB signaling pathway. This mechanism may outline the initial biochemical events leading to the observed increased neurodegeneration in HIV-positive individuals who use METH.

  13. JC virus inclusions in progressive multifocal leukoencephalopathy: scaffolding promyelocytic leukemia nuclear bodies grow with cell cycle transition through an S-to-G2-like state in enlarging oligodendrocyte nuclei.

    PubMed

    Shishido-Hara, Yukiko; Yazawa, Takuya; Nagane, Motoo; Higuchi, Kayoko; Abe-Suzuki, Shiho; Kurata, Morito; Kitagawa, Masanobu; Kamma, Hiroshi; Uchihara, Toshiki

    2014-05-01

    In progressive multifocal leukoencephalopathy, JC virus-infected oligodendroglia display 2 distinct patterns of intranuclear viral inclusions: full inclusions in which progeny virions are present throughout enlarged nuclei and dot-shaped inclusions in which virions are clustered in subnuclear domains termed "promyelocytic leukemia nuclear bodies" (PML-NBs). Promyelocytic leukemia nuclear bodies may serve a scaffolding role in viral progeny production. We analyzed the formation process of intranuclear viral inclusions by morphometry and assessed PML-NB alterations in the brains of 2 patients with progressive multifocal leukoencephalopathy. By immunohistochemistry, proliferating cell nuclear antigen was most frequently detected in smaller nuclei; cyclin A was detected in larger nuclei. This suggests an S-to-G2 cell cycle transition in infected cells associated with nuclear enlargement. Sizes of PML-NBs were variable, but they were usually either small speckles 200 to 400 nm in diameter or distinct spherical shells with a diameter of 1 μm or more. By confocal microscopy, JC virus capsid proteins were associated with both small and large PML-NBs, but disruption of large PML-NBs was observed by ground-state depletion fluorescence nanoscopy. Clusters of progeny virions were also detected by electron microscopy. Our data suggest that, in progressive multifocal leukoencephalopathy, JC virus produces progeny virions in enlarging oligodendrocyte nuclei in association with growing PML-NBs and with cell cycle transition through an S-to-G2-like state.

  14. Human T cell leukaemia virus type 2 tax protein mediates CC-chemokine expression in peripheral blood mononuclear cells via the nuclear factor kappa B canonical pathway.

    PubMed

    Barrios, C S; Castillo, L; Zhi, H; Giam, C-Z; Beilke, M A

    2014-01-01

    Retroviral co-infections with human immunodeficiency virus type-1 (HIV-1) and human T cell leukaemia virus type 1 (HTLV-1) or type 2 (HTLV-2) are prevalent in many areas worldwide. It has been observed that HIV-1/HTLV-2 co-infections are associated with slower rates of CD4(+) T cell decline and delayed progression to AIDS. This immunological benefit has been linked to the ability of Tax2, the transcriptional activating protein of HTLV-2, to induce the expression of macrophage inflammatory protein (MIP)-1α/CCL3, MIP-1β/CCL4 and regulated upon activation normal T cell expressed and secreted (RANTES)/CCL5 and to down-regulate the expression of the CCR5 co-receptor in peripheral blood mononuclear cells (PBMCs). This study aimed to assess the role of Tax2-mediated activation of the nuclear factor kappa B (NF-κB) signalling pathway on the production of the anti-viral CC-chemokines MIP-1α, MIP-1β and RANTES. Recombinant Tax1 and Tax2 proteins, or proteins expressed via adenoviral vectors used to infect cells, were tested for their ability to activate the NF-κB pathway in cultured PBMCs in the presence or absence of NF-κB pathway inhibitors. Results showed a significant release of MIP-1α, MIP-1β and RANTES by PBMCs after the activation of p65/RelA and p50. The secretion of these CC-chemokines was significantly reduced (P < 0·05) by canonical NF-κB signalling inhibitors. In conclusion, Tax2 protein may promote innate anti-viral immune responses through the activation of the canonical NF-κB pathway.

  15. Mutations in the nuclear localization signal of nsP2 influencing RNA synthesis, protein expression and cytotoxicity of Semliki Forest virus.

    PubMed

    Tamm, Kristi; Merits, Andres; Sarand, Inga

    2008-03-01

    The cytotoxicity of Semliki Forest virus (SFV) infection is caused partly by the non-structural protein nsP2, an essential component of the SFV replicase complex. Due to the presence of a nuclear localization signal (NLS), nsP2 also localizes in the nucleus of infected cells. The present study analysed recombinant SFV replicons and genomes with various deletions or substitutions in the NLS, or with a proline-to-glycine mutation at position 718 of nsP2 (P718G). Deletion of one or two arginine residues from the NLS or substitution of two of the arginines with aspartic acid resulted in a virus with a temperature-sensitive phenotype, and substitution of all three arginines was lethal. Thus, most of the introduced mutations severely affected nsP2 functioning in viral replication; in addition, they inhibited the ability of SFV to induce translational shut-off and kill infected cells. SFV replicons with a P718G mutation or replacement of the NLS residues (648)RRR(650) with RDD were found to be the least cytotoxic. Corresponding replicons expressed non-structural proteins at normal levels, but had severely reduced genomic RNA synthesis and were virtually unable to replicate and transcribe co-electroporated helper RNA. The non-cytotoxic phenotype was maintained in SFV full-length genomes harbouring the corresponding mutations; however, during a single cycle of cell culture, these were converted to a cytotoxic phenotype, probably due to the accumulation of compensatory mutations. PMID:18272758

  16. A Genome-Wide Integrative Genomic Study Localizes Genetic Factors Influencing Antibodies against Epstein-Barr Virus Nuclear Antigen 1 (EBNA-1)

    PubMed Central

    Rubicz, Rohina; Yolken, Robert; Drigalenko, Eugene; Carless, Melanie A.; Dyer, Thomas D.; Bauman, Lara; Melton, Phillip E.; Kent, Jack W.; Harley, John B.; Curran, Joanne E.; Johnson, Matthew P.; Cole, Shelley A.; Almasy, Laura; Moses, Eric K.; Dhurandhar, Nikhil V.; Kraig, Ellen; Blangero, John; Leach, Charles T.; Göring, Harald H. H.

    2013-01-01

    Infection with Epstein-Barr virus (EBV) is highly prevalent worldwide, and it has been associated with infectious mononucleosis and severe diseases including Burkitt lymphoma, Hodgkin lymphoma, nasopharyngeal lymphoma, and lymphoproliferative disorders. Although EBV has been the focus of extensive research, much still remains unknown concerning what makes some individuals more sensitive to infection and to adverse outcomes as a result of infection. Here we use an integrative genomics approach in order to localize genetic factors influencing levels of Epstein Barr virus (EBV) nuclear antigen-1 (EBNA-1) IgG antibodies, as a measure of history of infection with this pathogen, in large Mexican American families. Genome-wide evidence of both significant linkage and association was obtained on chromosome 6 in the human leukocyte antigen (HLA) region and replicated in an independent Mexican American sample of large families (minimum p-value in combined analysis of both datasets is 1.4×10−15 for SNPs rs477515 and rs2516049). Conditional association analyses indicate the presence of at least two separate loci within MHC class II, and along with lymphocyte expression data suggest genes HLA-DRB1 and HLA-DQB1 as the best candidates. The association signals are specific to EBV and are not found with IgG antibodies to 12 other pathogens examined, and therefore do not simply reveal a general HLA effect. We investigated whether SNPs significantly associated with diseases in which EBV is known or suspected to play a role (namely nasopharyngeal lymphoma, Hodgkin lymphoma, systemic lupus erythematosus, and multiple sclerosis) also show evidence of associated with EBNA-1 antibody levels, finding an overlap only for the HLA locus, but none elsewhere in the genome. The significance of this work is that a major locus related to EBV infection has been identified, which may ultimately reveal the underlying mechanisms by which the immune system regulates infection with this pathogen

  17. Proper use of serum antibody titres against Epstein-Barr virus in nasopharyngeal carcinoma: IgA/virus capsid antigen for diagnosis and EBV-related nuclear antigen-2 for follow-up.

    PubMed

    Shimakage, M; Dezawa, T; Chatani, M

    2000-01-01

    Sera from patients with nasopharyngeal carcinoma (NPC) show high titres of IgA antibodies to Epstein-Barr viral capsid antigen (IgA/VCA). We reported previously that the serum titres for Epstein-Barr virus-related nuclear antigen-2 (EBNA2) correlated with NPC patients' prognosis. To investigate which is better for diagnosing NPC and predicting patient prognosis, the titration of serum IgA/VCA or EBNA2, we examined the same serum titres. Sixteen cases of NPC in which serum EBNA2 antibody titres had been tested, were investigated for the serum IgA/VCA antibody titres before and after radiation treatment. All NPC cases showed positive reactions with indirect immunofluorescence staining, and the median titre was 252. Twelve normal controls, 5 mesopharyngeal carcinoma patients, 4 hypopharyngeal carcinoma patients, 4 laryngeal carcinoma patients and 6 malignant lymphoma were also examined, but they showed negative or relatively low titres. A follow-up study revealed that IgA/VCA titres remained mostly stable. These results indicate a close relationship between IgA/VCA and NPC, however, prognosis correlated better with EBNA2 titres than with IgA/VCA titres.

  18. Notch1, Notch2, and Epstein-Barr virus-encoded nuclear antigen 2 signaling differentially affects proliferation and survival of Epstein-Barr virus-infected B cells.

    PubMed

    Kohlhof, Hella; Hampel, Franziska; Hoffmann, Reinhard; Burtscher, Helmut; Weidle, Ulrich H; Hölzel, Michael; Eick, Dirk; Zimber-Strobl, Ursula; Strobl, Lothar J

    2009-05-28

    The canonical mode of transcriptional activation by both the Epstein-Barr viral protein, Epstein-Barr virus-encoded nuclear antigen 2 (EBNA2), and an activated Notch receptor (Notch-IC) requires their recruitment to RBPJ, suggesting that EBNA2 uses the Notch pathway to achieve B-cell immortalization. To gain further insight into the biologic equivalence between Notch-IC and EBNA2, we performed a genome-wide expression analysis, revealing that Notch-IC and EBNA2 exhibit profound differences in the regulation of target genes. Whereas Notch-IC is more potent in regulating genes associated with differentiation and development, EBNA2 is more potent in inducing viral and cellular genes involved in proliferation, survival, and chemotaxis. Because both EBNA2 and Notch-IC induced the expression of cell cycle-associated genes, we analyzed whether Notch1-IC or Notch2-IC can replace EBNA2 in B-cell immortalization. Although Notch-IC could drive quiescent B cells into the cell cycle, B-cell immortalization was not maintained, partially due to an increased apoptosis rate in Notch-IC-expressing cells. Expression analysis revealed that both EBNA2 and Notch-IC induced the expression of proapoptotic genes, but only in EBNA2-expressing cells were antiapoptotic genes strongly up-regulated. These findings suggest that Notch signaling in B cells and B-cell lymphomas is only compatible with proliferation if pathways leading to antiapototic signals are active. PMID:19339697

  19. Identification and Molecular Characterization of Nuclear Citrus leprosis virus, a Member of the Proposed Dichorhavirus Genus Infecting Multiple Citrus Species in Mexico.

    PubMed

    Roy, Avijit; Stone, Andrew L; Shao, Jonathan; Otero-Colina, Gabriel; Wei, Gang; Choudhary, Nandlal; Achor, Diann; Levy, Laurene; Nakhla, Mark K; Hartung, John S; Schneider, William L; Brlansky, Ronald H

    2015-04-01

    Citrus leprosis is one of the most destructive diseases of Citrus spp. and is associated with two unrelated virus groups that produce particles primarily in either the cytoplasm or nucleus of infected plant cells. Symptoms of leprosis, including chlorotic spots surrounded by yellow haloes on leaves and necrotic spots on twigs and fruit, were observed on leprosis-affected mandarin and navel sweet orange trees in the state of Querétaro, Mexico. Serological and molecular assays showed that the cytoplasmic types of Citrus leprosis virus (CiLV-C) often associated with leprosis symptomatic tissues were absent. However, using transmission electron microscopy, bullet-shaped rhabdovirus-like virions were observed in the nuclei and cytoplasm of the citrus leprosis-infected leaf tissues. An analysis of small RNA populations from symptomatic tissue was carried out to determine the genome sequence of the rhabdovirus-like particles observed in the citrus leprosis samples. The complete genome sequence showed that the nuclear type of CiLV (CiLV-N) present in the samples consisted of two negative-sense RNAs: 6,268-nucleotide (nt)-long RNA1 and 5,847-nt-long RNA2, excluding the poly(A) tails. CiLV-N had a genome organization identical to that of Orchid fleck virus (OFV), with the exception of shorter 5' untranslated regions in RNA1 (53 versus 205 nt) and RNA2 (34 versus 182 nt). Phylogenetic trees constructed with the amino acid sequences of the nucleocapsid (N) and glycoproteins (G) and the RNA polymerase (L protein) showed that CiLV-N clusters with OFV. Furthermore, phylogenetic analyses of N protein established CiLV-N as a member of the proposed genus Dichorhavirus. Reverse-transcription polymerase chain reaction primers for the detection of CiLV-N were designed based on the sequence of the N gene and the assay was optimized and tested to detect the presence of CiLV-N in both diseased and symptom-free plants.

  20. A theoretical study of rotational diffusion models for rod-shaped viruses. The influence of motion on 31P nuclear magnetic resonance lineshapes and transversal relaxation.

    PubMed Central

    Magusin, P C; Hemminga, M A

    1993-01-01

    Information about the interaction between nucleic acids and coat proteins in intact virus particles may be obtained by studying the restricted backbone dynamics of the incapsulated nucleic acids using 31P nuclear magnetic resonance (NMR) spectroscopy. In this article, simulations are carried out to investigate how reorientation of a rod-shaped virus particle as a whole and isolated nucleic acid motions within the virion influence the 31P NMR lineshape and transversal relaxation dominated by the phosphorus chemical shift anisotropy. Two opposite cases are considered on a theoretical level. First, isotropic rotational diffusion is used as a model for mobile nucleic acids that are loosely or partially bound to the protein coat. The effect of this type of diffusion on lineshape and transversal relaxation is calculated by solving the stochastic Liouville equation by an expansion in spherical functions. Next, uniaxial rotational diffusion is assumed to represent the mobility of phosphorus in a virion that rotates as a rigid rod about its length axis. This type of diffusion is approximated by an exchange process among discrete sites. As turns out from these simulations, the amplitude and the frequency of the motion can only be unequivocally determined from experimental data by a combined analysis of the lineshape and the transversal relaxation. In the fast motional region both the isotropic and the uniaxial diffusion model predict the same transversal relaxation as the Redfield theory. For very slow motion, transversal relaxation resembles the nonexponential relaxation as observed for water molecules undergoing translational diffusion in a magnetic field gradient. In this frequency region T2e is inversely proportional to the cube root of the diffusion coefficient. In addition to the isotropic and uniaxial diffusion models, a third model is presented, in which fast restricted nucleic acid backbone motions dominating the lineshape are superimposed on a slow rotation of the

  1. Epstein-Barr virus nuclear protein 3C modulates transcription through interaction with the sequence-specific DNA-binding protein J kappa.

    PubMed Central

    Robertson, E S; Grossman, S; Johannsen, E; Miller, C; Lin, J; Tomkinson, B; Kieff, E

    1995-01-01

    The Epstein-Barr virus (EBV) nuclear protein 3C (EBNA 3C) is essential for EBV-mediated transformation of primary B lymphocytes, is turned on by EBNA 2, and regulates transcription of some of the viral and cellular genes which are regulated by EBNA 2. EBNA 2 is targeted to response elements by binding to the DNA sequence-specific, transcriptional repressor protein J kappa. We now show that EBNA 3C also binds to J kappa. EBNA 3C causes J kappa to not bind DNA or EBNA 2. J kappa DNA binding activity in EBV-transformed lymphoblastoid cells is consequently reduced. More than 10% of the EBNA 3C coimmunoprecipitated with J kappa from extracts of non-EBV-infected B lymphoblasts that had been stably converted to EBNA 3C expression. EBNA 3C in nuclear extracts from these cells (or in vitro-translated EBNA 3C) prevented J kappa from interacting with a high-affinity DNA binding site. Under conditions of transient overexpression in B lymphoblasts, EBNA 2 and EBNA 3C associated with J kappa and less EBNA 2 associated with J kappa when EBNA 3C was coexpressed in the same cell. EBNA 3C had no effect on the activity of a -512/+40 LMP1 promoter-CAT reporter construct that has two upstream J kappa sites, but it did inhibit EBNA 2 transactivation of this promoter. These data are compatible with a role for EBNA 3C as a "feedback" down modulator of EBNA 2-mediated transactivation. EBNA 3C could, in theory, also activate transcription by inhibiting the interaction of the J kappa repressor with its cognate DNA. The interaction of two viral transcriptional regulators with the same cell protein may reflect an unusually high level of complexity or stringency in target gene regulation. PMID:7707539

  2. Role of cellular caspases, nuclear factor-kappa B and interferon regulatory factors in Bluetongue virus infection and cell fate

    PubMed Central

    2010-01-01

    Background Bluetongue virus (BTV) infection causes haemorrhagic disease in ruminants and induces cell death. The pathogenesis in animals and in cell culture has been linked to BTV-induced apoptosis. Results In this report, we investigated BTV-induced apoptosis in cell culture in depth and show that both extrinsic (caspase-8 activation) and intrinsic (caspase-9 activation) pathways play roles in BTV apoptosis. Further, by using chemical inhibitors and knock-out cell lines, we show that these pathways act independently of each other in BTV infected cells. In addition to activation of caspase-8, -9 and executioner caspase-3, we also identified that BTV infection causes the activation of caspase-7, which results in the cleavage of poly (ADP-ribose) polymerase (PARP). BTV-induced cell death appears to be due to apoptosis rather than necrosis, as the HMBG-1 was not translocated from the nucleus. We also examined if NF-κB response is related to BTV-induced apoptosis as in reovirus. Our data suggests that NF-κB response is not linked to the induction of apoptosis. It is controlled by the degradation of only IκBα but not IκBβ, resulting in a rapid transient response during BTV infection. This was supported using an NF-κB dependent luciferase reporter gene assay, which demonstrated early response, that appeared to be suppressed by the late stage of BTV replication. Furthermore, virus titres were higher in the presence of NF-κB inhibitor (SN50), indicating that NF-κB has a role in initiating an antiviral environment. In addition, we show that BTV infection induces the translocation of interferon regulatory factors (IRF-3 and IRF-7) into the nucleus. The induction of IRF responses, when measured by IRF dependent luciferase reporter gene assay, revealed that the IRF responses, like NF-κB response, were also at early stage of infection and mirrored the timing of NF-κB induction. Conclusion BTV triggers a wide range of caspase activities resulting in cell apoptosis

  3. Human herpes virus-6 increases HIV-1 expression in co-infected T cells via nuclear factors binding to the HIV-1 enhancer.

    PubMed Central

    Ensoli, B; Lusso, P; Schachter, F; Josephs, S F; Rappaport, J; Negro, F; Gallo, R C; Wong-Staal, F

    1989-01-01

    Human Herpes virus-6 (HHV-6) can co-infect with HIV-1 human CD4+ T-cells, leading to accelerated cell death, and factors in HHV-6-infected cells stimulate HIV-1 LTR directed gene expression. In this study, we have examined the mechanism of HIV-1 activation by HHV-6 and localized the cis-acting sequences of HIV-1 LTR responsive to trans-activation. Increased HIV-1 LTR directed gene expression is obtained in HIV-1 infected cells co-infected with HHV-6, or in HHV-6 infected cells co-transfected with the HIV-1 tat gene. Parallel increases of HIV-1-specific transcripts are seen by in situ hybridization in HHV-6/HIV-1 doubly infected cells as compared to single HIV-1 infection. Similarly, infection by HHV-6 increases the steady-state level of HIV-1 LTR mRNA that parallels CAT enzymatic activity, suggesting a transcriptional and/or post-transcriptional activation. Sequences necessary for HIV-1 LTR activation by HHV-6 are distinct from those required for that tat response and map to a region of the HIV-1 LTR from -103 to -48. The HIV-1 enhancer sequence (-105 to -80) is sufficient to confer HHV-6 inducibility to a heterologous promoter, and nuclear protein(s) activated or induced by HHV-6 infection specifically bind to the NF kappa B motifs of the HIV-1 enhancer region.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:2573513

  4. The nuclear localization of the Arabidopsis transcription factor TIP is blocked by its interaction with the coat protein of Turnip crinkle virus

    SciTech Connect

    Ren Tao; Qu Feng; Morris, T. Jack . E-mail: jmorris@unlnotes.unl.edu

    2005-01-20

    We have previously reported that TIP, an Arabidopsis protein, interacts with the coat protein (CP) of Turnip crinkle virus (TCV) in yeast cells and that this interaction correlated with the resistance response in the TCV-resistant Arabidopsis ecotype Dijon-17. TIP was also able to activate transcription of reporter genes in yeast cells, suggesting that it is likely a transcription factor. We have now verified the physical interaction between TIP and TCV CP in vitro and showed that CP mutants unable to interact with TIP in yeast cells bind TIP with much lower affinity in vitro. Secondly, we have performed gel shift experiments demonstrating that TIP does not bind to DNA in a sequence-specific manner. The subcellular localization of TIP was also investigated by transiently expressing green fluorescence protein (GFP)-tagged TIP in Nicotiana benthamiana plant cells, which showed that GFP-tagged TIP localizes primarily to nuclei. Significantly, co-expression of TCVCP and GFP-TIP prevented the nuclear localization of TIP. Together, these results suggest that TIP might be a transcription factor involved in regulating the defense response of Arabidopsis to TCV and that its normal role is compromised by interaction with the invading viral CP.

  5. Nuclear Export of Human Hepatitis B Virus Core Protein and Pregenomic RNA Depends on the Cellular NXF1-p15 Machinery

    PubMed Central

    Yang, Ching-Chun; Huang, Er-Yi; Li, Hung-Cheng; Su, Pei-Yi; Shih, Chiaho

    2014-01-01

    Hepatitis B virus (HBV) core protein (HBc) can shuttle between nucleus and cytoplasm. Cytoplasm-predominant HBc is clinically associated with severe liver inflammation. Previously, we found that HBc arginine-rich domain (ARD) can associate with a host factor NXF1 (TAP) by coimmunoprecipitation. It is well known that NXF1-p15 heterodimer can serve as a major export receptor of nuclear mRNA as a ribonucleoprotein complex (RNP). In the NXF1-p15 pathway, TREX (transcription/export) complex plays an important role in coupling nuclear pre-mRNA processing with mRNA export in mammalian cells. Here, we tested the hypothesis whether HBc and HBV specific RNA can be exported via the TREX and NXF1-p15 mediated pathway. We demonstrated here that HBc can physically and specifically associate with TREX components, and the NXF1-p15 export receptor by coimmunoprecipitation. Accumulation of HBc protein in the nucleus can be induced by the interference with TREX and NXF1-p15 mediated RNA export machinery. HBV transcripts encodes a non-spliced 3.5 kb pregenomic RNA (pgRNA) which can serve as a template for reverse transcription. Cytoplasmic HBV pgRNA appeared to be reduced by siRNA treatment specific for the NXF1-p15 complex by quantitative RT-qPCR and Northern blot analyses. This result suggests that the pgRNA was also exported via the NXF1-p15 machinery. We entertain the hypothesis that HBc protein can be exported as an RNP cargo via the mRNA export pathway by hijacking the TREX and NXF1-p15 complex. In our current and previous studies, HBc is not required for pgRNA accumulation in the cytoplasm. Furthermore, HBc ARD can mediate nuclear export of a chimeric protein containing HBc ARD in a pgRNA-independent manner. Taken together, it suggests that while both pgRNA and HBc protein exports are dependent on NXF1-p15, they are using the same export machinery in a manner independent of each other. PMID:25360769

  6. Epstein-Barr virus nuclear antigen 3C targets p53 and modulates its transcriptional and apoptotic activities

    SciTech Connect

    Yi Fuming; Saha, Abhik; Murakami, Masanao; Kumar, Pankaj; Knight, Jason S.; Cai Qiliang; Choudhuri, Tathagata; Robertson, Erle S.

    2009-06-05

    The p53 tumor suppressor gene is one of the most commonly mutated genes in human cancers and the corresponding encoded protein induces apoptosis or cell-cycle arrest at the G1/S checkpoint in response to DNA damage. To date, previous studies have shown that antigens encoded by human tumor viruses such as SV40 large T antigen, adenovirus E1A and HPV E6 interact with p53 and disrupt its functional activity. In a similar fashion, we now show that EBNA3C, one of the EBV latent antigens essential for the B-cell immortalization in vitro, interacts directly with p53. Additionally, we mapped the interaction of EBNA3C with p53 to the C-terminal DNA-binding and the tetramerization domain of p53, and the region of EBNA3C responsible for binding to p53 was mapped to the N-terminal domain of EBNA3C (residues 130-190), previously shown to interact with a number of important cell-cycle components, specifically SCF{sup Skp2}, cyclin A, and cMyc. Furthermore, we demonstrate that EBNA3C substantially represses the transcriptional activity of p53 in luciferase based reporter assays, and rescues apoptosis induced by ectopic p53 expression in SAOS-2 (p53{sup -/-}) cells. Interestingly, we also show that the DNA-binding ability of p53 is diminished in the presence of EBNA3C. Thus, the interaction between the p53 and EBNA3C provides new insights into the mechanism(s) by which the EBNA3C oncoprotein can alter cellular gene expression in EBV associated human cancers.

  7. Nuclear receptor REV-ERBα mediates circadian sensitivity to mortality in murine vesicular stomatitis virus-induced encephalitis.

    PubMed

    Gagnidze, Khatuna; Hajdarovic, Kaitlyn H; Moskalenko, Marina; Karatsoreos, Ilia N; McEwen, Bruce S; Bulloch, Karen

    2016-05-17

    Certain components and functions of the immune system, most notably cytokine production and immune cell migration, are under circadian regulation. Such regulation suggests that circadian rhythms may have an effect on disease onset, progression, and resolution. In the vesicular stomatitis virus (VSV)-induced encephalitis model, the replication, caudal penetration, and survivability of intranasally applied VSV depends on both innate and adaptive immune mechanisms. In the current study, we investigated the effect of circadian time of infection on the progression and outcome of VSV-induced encephalitis and demonstrated a significant decrease in the survival rate in mice infected at the start of the rest cycle, zeitgeber time 0 (ZT0). The lower survival rate in these mice was associated with higher levels of circulating chemokine (C-C motif) ligand 2 (CCL2), a greater number of peripherally derived immune cells accumulating in the olfactory bulb (OB), and increased production of proinflammatory cytokines, indicating an immune-mediated pathology. We also found that the acrophase of molecular circadian clock component REV-ERBα mRNA expression in the OB coincides with the start of the active cycle, ZT12, when VSV infection results in a more favorable outcome. This result led us to hypothesize that REV-ERBα may mediate the circadian effect on survival following VSV infection. Blocking REV-ERBα activity before VSV administration resulted in a significant increase in the expression of CCL2 and decreased survival in mice infected at the start of the active cycle. These data demonstrate that REV-ERBα-mediated inhibition of CCL2 expression during viral-induced encephalitis may have a protective effect. PMID:27143721

  8. Cooperativity among Rev-Associated Nuclear Export Signals Regulates HIV-1 Gene Expression and Is a Determinant of Virus Species Tropism

    PubMed Central

    Aligeti, Mounavya; Behrens, Ryan T.; Pocock, Ginger M.; Schindelin, Johannes; Dietz, Christian; Eliceiri, Kevin W.; Swanson, Chad M.; Malim, Michael H.; Ahlquist, Paul

    2014-01-01

    ABSTRACT Murine cells exhibit a profound block to HIV-1 virion production that was recently mapped to a species-specific structural attribute of the murine version of the chromosomal region maintenance 1 (mCRM1) nuclear export receptor and rescued by the expression of human CRM1 (hCRM1). In human cells, the HIV-1 Rev protein recruits hCRM1 to intron-containing viral mRNAs encoding the Rev response element (RRE), thereby facilitating viral late gene expression. Here we exploited murine 3T3 fibroblasts as a gain-of-function system to study hCRM1's species-specific role in regulating Rev's effector functions. We show that Rev is rapidly exported from the nucleus by mCRM1 despite only weak contributions to HIV-1's posttranscriptional stages. Indeed, Rev preferentially accumulates in the cytoplasm of murine 3T3 cells with or without hCRM1 expression, in contrast to human HeLa cells, where Rev exhibits striking en masse transitions between the nuclear and cytoplasmic compartments. Efforts to bias Rev's trafficking either into or out of the nucleus revealed that Rev encoding a second CRM1 binding domain (Rev-2xNES) or Rev-dependent viral gag-pol mRNAs bearing tandem RREs (GP-2xRRE), rescue virus particle production in murine cells even in the absence of hCRM1. Combined, these results suggest a model wherein Rev-associated nuclear export signals cooperate to regulate the number or quality of CRM1's interactions with viral Rev/RRE ribonucleoprotein complexes in the nucleus. This mechanism regulates CRM1-dependent viral gene expression and is a determinant of HIV-1's capacity to produce virions in nonhuman cell types. IMPORTANCE Cells derived from mice and other nonhuman species exhibit profound blocks to HIV-1 replication. Here we elucidate a block to HIV-1 gene expression attributable to the murine version of the CRM1 (mCRM1) nuclear export receptor. In human cells, hCRM1 regulates the nuclear export of viral intron-containing mRNAs through the activity of the viral Rev

  9. hnRNP A2/B1 interacts with influenza A viral protein NS1 and inhibits virus replication potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nuclear export

    SciTech Connect

    Wang, Yimeng; Zhou, Jianhong; Du, Yuchun

    2014-01-20

    The NS1 protein of influenza viruses is a major virulence factor and exerts its function through interacting with viral/cellular RNAs and proteins. In this study, we identified heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) as an interacting partner of NS1 proteins by a proteomic method. Knockdown of hnRNP A2/B1 by small interfering RNA (siRNA) resulted in higher levels of NS vRNA, NS1 mRNA, and NS1 protein in the virus-infected cells. In addition, we demonstrated that hnRNP A2/B1 proteins are associated with NS1 and NS2 mRNAs and that knockdown of hnRNP A2/B1 promotes transport of NS1 mRNA from the nucleus to the cytoplasm in the infected cells. Lastly, we showed that knockdown of hnRNP A2/B1 leads to enhanced virus replication. Our results suggest that hnRNP A2/B1 plays an inhibitory role in the replication of influenza A virus in host cells potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nucleocytoplasmic translocation. - Highlights: • Cellular protein hnRNP A2/B1 interacts with influenza viral protein NS1. • hnRNP A2/B1 suppresses the levels of NS1 protein, vRNA and mRNA in infected cells. • hnRNP A2/B1 protein is associated with NS1 and NS2 mRNAs. • hnRNP A2/B1 inhibits the nuclear export of NS1 mRNAs. • hnRNP A2/B1 inhibits influenza virus replication.

  10. Epstein-Barr virus nuclear protein 2 transactivation of the latent membrane protein 1 promoter is mediated by J kappa and PU.1.

    PubMed Central

    Johannsen, E; Koh, E; Mosialos, G; Tong, X; Kieff, E; Grossman, S R

    1995-01-01

    Expression of the Epstein-Barr virus (EBV) latent membrane protein 1 (LMP-1) oncogene is regulated by the EBV nuclear protein 2 (EBNA-2) transactivator. EBNA-2 is known to interact with the cellular DNA-binding protein J kappa and is recruited to promoters containing the GTGGGAA J kappa recognition sequence. The minimal EBNA-2-responsive LMP-1 promoter includes one J kappa-binding site, and we now show that mutation of that site, such that J kappa cannot bind, reduces EBNA-2 responsiveness by 60%. To identify other factors which interact with the LMP-1 EBNA-2 response element (E2RE), a -236/-145 minimal E2RE was used as a probe in an electrophoretic mobility shift assay. The previously characterized factors J kappa, PU.1, and AML1 bind to the LMP-1 E2RE, along with six other unidentified factors (LBF2 to LBF7). Binding sites were mapped for each factor. LBF4 is B- and T-cell specific and recognizes the PU.1 GGAA core sequence as shown by methylation interference. LBF4 has a molecular mass of 105 kDa and is probably unrelated to PU.1. LBF2 was found only in epithelial cell lines, whereas LBF3, LBF5, LBF6, and LBF7 were not cell type specific. Mutations of the AML1- or LBF4-binding sites had no effect on EBNA-2 transactivation, whereas mutation of the PU.1-binding site completely eliminated EBNA-2 responses. A gst-EBNA-2 fusion protein specifically depleted PU.1 from nuclear extracts and bound in vitro translated PU.1, providing biochemical evidence for a direct EBNA-2-PU.1 interaction. Thus, EBNA-2 transactivation of the LMP-1 promoter is dependent on interaction with at least two distinct sequence-specific DNA-binding proteins, J kappa and PU.1. LBF3, LBF5, LBF6, or LBF7 may also be involved, since their binding sites also contribute to EBNA-2 responsiveness. PMID:7983717

  11. Epstein-Barr Virus Nuclear Antigen 3C Facilitates G1-S Transition by Stabilizing and Enhancing the Function of Cyclin D1

    PubMed Central

    Saha, Abhik; Halder, Sabyasachi; Upadhyay, Santosh K.; Lu, Jie; Kumar, Pankaj; Murakami, Masanao; Cai, Qiliang; Robertson, Erle S.

    2011-01-01

    EBNA3C, one of the Epstein-Barr virus (EBV)-encoded latent antigens, is essential for primary B-cell transformation. Cyclin D1, a key regulator of G1 to S phase progression, is tightly associated and aberrantly expressed in numerous human cancers. Previously, EBNA3C was shown to bind to Cyclin D1 in vitro along with Cyclin A and Cyclin E. In the present study, we provide evidence which demonstrates that EBNA3C forms a complex with Cyclin D1 in human cells. Detailed mapping experiments show that a small N-terminal region which lies between amino acids 130–160 of EBNA3C binds to two different sites of Cyclin D1- the N-terminal pRb binding domain (residues 1–50), and C-terminal domain (residues 171–240), known to regulate Cyclin D1 stability. Cyclin D1 is short-lived and ubiquitin-mediated proteasomal degradation has been targeted as a means of therapeutic intervention. Here, we show that EBNA3C stabilizes Cyclin D1 through inhibition of its poly-ubiquitination, and also increases its nuclear localization by blocking GSK3β activity. We further show that EBNA3C enhances the kinase activity of Cyclin D1/CDK6 which enables subsequent ubiquitination and degradation of pRb. EBNA3C together with Cyclin D1-CDK6 complex also efficiently nullifies the inhibitory effect of pRb on cell growth. Moreover, an sh-RNA based strategy for knock-down of both cyclin D1 and EBNA3C genes in EBV transformed lymphoblastoid cell lines (LCLs) shows a significant reduction in cell-growth. Based on these results, we propose that EBNA3C can stabilize as well as enhance the functional activity of Cyclin D1 thereby facilitating the G1-S transition in EBV transformed lymphoblastoid cell lines. PMID:21347341

  12. Epstein-Barr virus nuclear antigen 3A partially coincides with EBNA3C genome-wide and is tethered to DNA through BATF complexes.

    PubMed

    Schmidt, Stefanie C S; Jiang, Sizun; Zhou, Hufeng; Willox, Bradford; Holthaus, Amy M; Kharchenko, Peter V; Johannsen, Eric C; Kieff, Elliott; Zhao, Bo

    2015-01-13

    Epstein-Barr Virus (EBV) conversion of B-lymphocytes to Lymphoblastoid Cell Lines (LCLs) requires four EBV nuclear antigen (EBNA) oncoproteins: EBNA2, EBNALP, EBNA3A, and EBNA3C. EBNA2 and EBNALP associate with EBV and cell enhancers, up-regulate the EBNA promoter, MYC, and EBV Latent infection Membrane Proteins (LMPs), which up-regulate BCL2 to protect EBV-infected B-cells from MYC proliferation-induced cell death. LCL proliferation induces p16(INK4A) and p14(ARF)-mediated cell senescence. EBNA3A and EBNA3C jointly suppress p16(INK4A) and p14(ARF), enabling continuous cell proliferation. Analyses of the EBNA3A human genome-wide ChIP-seq landscape revealed 37% of 10,000 EBNA3A sites to be at strong enhancers; 28% to be at weak enhancers; 4.4% to be at active promoters; and 6.9% to be at weak and poised promoters. EBNA3A colocalized with BATF-IRF4, ETS-IRF4, RUNX3, and other B-cell Transcription Factors (TFs). EBNA3A sites clustered into seven unique groups, with differing B-cell TFs and epigenetic marks. EBNA3A coincidence with BATF-IRF4 or RUNX3 was associated with stronger EBNA3A ChIP-Seq signals. EBNA3A was at MYC, CDKN2A/B, CCND2, CXCL9/10, and BCL2, together with RUNX3, BATF, IRF4, and SPI1. ChIP-re-ChIP revealed complexes of EBNA3A on DNA with BATF. These data strongly support a model in which EBNA3A is tethered to DNA through a BATF-containing protein complexes to enable continuous cell proliferation.

  13. SUMO Ligase Protein Inhibitor of Activated STAT1 (PIAS1) Is a Constituent Promyelocytic Leukemia Nuclear Body Protein That Contributes to the Intrinsic Antiviral Immune Response to Herpes Simplex Virus 1

    PubMed Central

    Brown, James R.; Conn, Kristen L.; Wasson, Peter; Charman, Matthew; Tong, Lily; Grant, Kyle; McFarlane, Steven

    2016-01-01

    ABSTRACT Aspects of intrinsic antiviral immunity are mediated by promyelocytic leukemia nuclear body (PML-NB) constituent proteins. During herpesvirus infection, these antiviral proteins are independently recruited to nuclear domains that contain infecting viral genomes to cooperatively promote viral genome silencing. Central to the execution of this particular antiviral response is the small ubiquitin-like modifier (SUMO) signaling pathway. However, the participating SUMOylation enzymes are not fully characterized. We identify the SUMO ligase protein inhibitor of activated STAT1 (PIAS1) as a constituent PML-NB protein. We show that PIAS1 localizes at PML-NBs in a SUMO interaction motif (SIM)-dependent manner that requires SUMOylated or SUMOylation-competent PML. Following infection with herpes simplex virus 1 (HSV-1), PIAS1 is recruited to nuclear sites associated with viral genome entry in a SIM-dependent manner, consistent with the SIM-dependent recruitment mechanisms of other well-characterized PML-NB proteins. In contrast to that of Daxx and Sp100, however, the recruitment of PIAS1 is enhanced by PML. PIAS1 promotes the stable accumulation of SUMO1 at nuclear sites associated with HSV-1 genome entry, whereas the accumulation of other evaluated PML-NB proteins occurs independently of PIAS1. We show that PIAS1 cooperatively contributes to HSV-1 restriction through mechanisms that are additive to those of PML and cooperative with those of PIAS4. The antiviral mechanisms of PIAS1 are counteracted by ICP0, the HSV-1 SUMO-targeted ubiquitin ligase, which disrupts the recruitment of PIAS1 to nuclear domains that contain infecting HSV-1 genomes through mechanisms that do not directly result in PIAS1 degradation. IMPORTANCE Adaptive, innate, and intrinsic immunity cooperatively and efficiently restrict the propagation of viral pathogens. Intrinsic immunity mediated by constitutively expressed cellular proteins represents the first line of intracellular defense against

  14. The N-terminal half of the influenza virus NS1 protein is sufficient for nuclear retention of mRNA and enhancement of viral mRNA translation.

    PubMed

    Marión, R M; Aragón, T; Beloso, A; Nieto, A; Ortín, J

    1997-11-01

    A collection of C-terminal deletion mutants of the influenza A virus NS1 gene has been used to define the regions of the NS1 protein involved in its functionality. Immunofluorescence analyses showed that the NS1 protein sequences downstream from position 81 are not required for nuclear transport. The capacity of these mutants to bind RNA was studied by in vitro binding tests using a model vRNA probe. These experiments showed that the N-terminal 81 amino acids of NS1 protein are sufficient for RNA binding activity. The collection of mutants also served to map the NS1 sequences required for nuclear retention of mRNA and for stimulation of viral mRNA translation, using the NP gene as reporter. The results obtained indicated that the N-terminal 113 amino acids of NS1 protein are sufficient for nuclear retention of mRNA and stimulation of viral mRNA translation. The possibility that this region of the protein may be sufficient for virus viability is discussed in relation to the sequences of NS1 genes of field isolates and to the phenotype of known viral mutants affected in the NS1 gene.

  15. Ceramide induces early and late apoptosis in human papilloma virus+ cervical cancer cells by inhibiting reactive oxygen species decay, diminishing the intracellular concentration of glutathione and increasing nuclear factor-kappaB translocation.

    PubMed

    Gutiérrez, Gisela; Mendoza, Criselda; Montaño, Luis F; López-Marure, Rebeca

    2007-02-01

    Ceramide is regarded as an important cellular signal for the induction of cell death. We have previously shown that ceramide induces the death of cervical tumor cells without biochemical and morphological markers of apoptosis. The mechanisms by which ceramide induces cell death are not understood, therefore we evaluated the effect of C6-ceramide, a synthetic cell-permeable analog of endogenous ceramides, in signaling pathways involved in the oxidative stress of three cervical human papilloma virus cancer cell lines. Reactive oxygen species production was determined by fluorescent 2,7-dichlorofluorescein, nitrite concentration by the Griess reaction (as an indirect measure of nitric oxide production), mitochondrial membrane potential by staining with Rh123, reduced-glutathione concentration by high-pressure liquid chromatography, nuclear factor-kappaB translocation by electrophoretic mobility shift assay, inhibitory protein of nuclear factor-kappaB expression by Western blot and cell death by a poly-caspases fluorochrome-labeled inhibitors of caspases apoptosis assay. C6-ceramide induced early and late apoptosis, which was associated with an increase in reactive oxygen species and nitric oxide production, a loss in mitochondrial membrane potential, an increase in nuclear factor-kappaB translocation, and a decrease in reduced glutathione concentration. C6-ceramide did not modify the expression of inhibitory protein of nuclear factor-kappaB and its antiproliferative effect was not abrogated by Bay 11-7082, an inhibitory protein of nuclear factor-kappaB kinase inhibitor. Our results suggest that oxidative stress might participate in the ceramide-induced damage to human papilloma virus cervical cancer cells.

  16. Inventing Viruses.

    PubMed

    Summers, William C

    2014-11-01

    In the nineteenth century, "virus" commonly meant an agent (usually unknown) that caused disease in inoculation experiments. By the 1890s, however, some disease-causing agents were found to pass through filters that retained the common bacteria. Such an agent was called "filterable virus," the best known being the virus that caused tobacco mosaic disease. By the 1920s there were many examples of filterable viruses, but no clear understanding of their nature. However, by the 1930s, the term "filterable virus" was being abandoned in favor of simply "virus," meaning an agent other than bacteria. Visualization of viruses by the electron microscope in the late 1930s finally settled their particulate nature. This article describes the ever-changing concept of "virus" and how virologists talked about viruses. These changes reflected their invention and reinvention of the concept of a virus as it was revised in light of new knowledge, new scientific values and interests, and new hegemonic technologies.

  17. Inventing Viruses.

    PubMed

    Summers, William C

    2014-11-01

    In the nineteenth century, "virus" commonly meant an agent (usually unknown) that caused disease in inoculation experiments. By the 1890s, however, some disease-causing agents were found to pass through filters that retained the common bacteria. Such an agent was called "filterable virus," the best known being the virus that caused tobacco mosaic disease. By the 1920s there were many examples of filterable viruses, but no clear understanding of their nature. However, by the 1930s, the term "filterable virus" was being abandoned in favor of simply "virus," meaning an agent other than bacteria. Visualization of viruses by the electron microscope in the late 1930s finally settled their particulate nature. This article describes the ever-changing concept of "virus" and how virologists talked about viruses. These changes reflected their invention and reinvention of the concept of a virus as it was revised in light of new knowledge, new scientific values and interests, and new hegemonic technologies. PMID:26958713

  18. MHC II tetramers visualize human CD4+ T cell responses to Epstein-Barr virus infection and demonstrate atypical kinetics of the nuclear antigen EBNA1 response.

    PubMed

    Long, Heather M; Chagoury, Odette L; Leese, Alison M; Ryan, Gordon B; James, Eddie; Morton, Laura T; Abbott, Rachel J M; Sabbah, Shereen; Kwok, William; Rickinson, Alan B

    2013-05-01

    Virus-specific CD4(+) T cells are key orchestrators of host responses to viral infection yet, compared with their CD8(+) T cell counterparts, remain poorly characterized at the single cell level. Here we use nine MHC II-epitope peptide tetramers to visualize human CD4(+) T cell responses to Epstein-Barr virus (EBV), the causative agent of infectious mononucleosis (IM), a disease associated with large virus-specific CD8(+) T cell responses. We find that, while not approaching virus-specific CD8(+) T cell expansions in magnitude, activated CD4(+) T cells specific for epitopes in the latent antigen EBNA2 and four lytic cycle antigens are detected at high frequencies in acute IM blood. They then fall rapidly to values typical of life-long virus carriage where most tetramer-positive cells display conventional memory markers but some, unexpectedly, revert to a naive-like phenotype. In contrast CD4(+) T cell responses to EBNA1 epitopes are greatly delayed in IM patients, in line with the well-known but hitherto unexplained delay in EBNA1 IgG antibody responses. We present evidence from an in vitro system that may explain these unusual kinetics. Unlike other EBNAs and lytic cycle proteins, EBNA1 is not naturally released from EBV-infected cells as a source of antigen for CD4(+) T cell priming. PMID:23569328

  19. Nuclear imprisonment: viral strategies to arrest host mRNA nuclear export.

    PubMed

    Kuss, Sharon K; Mata, Miguel A; Zhang, Liang; Fontoura, Beatriz M A

    2013-07-01

    Viruses possess many strategies to impair host cellular responses to infection. Nuclear export of host messenger RNAs (mRNA) that encode antiviral factors is critical for antiviral protein production and control of viral infections. Several viruses have evolved sophisticated strategies to inhibit nuclear export of host mRNAs, including targeting mRNA export factors and nucleoporins to compromise their roles in nucleo-cytoplasmic trafficking of cellular mRNA. Here, we present a review of research focused on suppression of host mRNA nuclear export by viruses, including influenza A virus and vesicular stomatitis virus, and the impact of this viral suppression on host antiviral responses.

  20. Nuclear Imprisonment: Viral Strategies to Arrest Host mRNA Nuclear Export

    PubMed Central

    Kuss, Sharon K.; Mata, Miguel A.; Zhang, Liang; Fontoura, Beatriz M. A.

    2013-01-01

    Viruses possess many strategies to impair host cellular responses to infection. Nuclear export of host messenger RNAs (mRNA) that encode antiviral factors is critical for antiviral protein production and control of viral infections. Several viruses have evolved sophisticated strategies to inhibit nuclear export of host mRNAs, including targeting mRNA export factors and nucleoporins to compromise their roles in nucleo-cytoplasmic trafficking of cellular mRNA. Here, we present a review of research focused on suppression of host mRNA nuclear export by viruses, including influenza A virus and vesicular stomatitis virus, and the impact of this viral suppression on host antiviral responses. PMID:23872491

  1. A nuclear localization signal in the matrix of spleen necrosis virus (SNV) does not allow efficient gene transfer into quiescent cells with SNV-derived vectors

    SciTech Connect

    Caron, Marie-Christine; Caruso, Manuel . E-mail: manuel.caruso@crhdq.ulaval.ca

    2005-08-01

    A major limitation in gene therapy for vectors derived from Moloney murine leukemia virus (MLV) is that they only deliver genes into dividing cells. In this study, a careful comparison of spleen necrosis virus (SNV)-derived vectors with MLV and human immunodeficiency virus (HIV)-1 retroviral vectors indicated that SNV vectors can deliver genes 4-fold more efficiently than MLV vectors into aphidicolin-arrested cells, although at a 25-fold lower efficiency than HIV-1-derived vectors. Furthermore, the addition of a NLS in the SNV matrix (MA) that mimics the one located in HIV-1 MA did not increase the ability of SNV vectors to transfer genes into arrested cells. Also, we found that the RD114 envelope was able to pseudotype SNV viral particles in a very efficient manner.

  2. Temperature-Sensitive Mutants in the Influenza A Virus RNA Polymerase: Alterations in the PA Linker Reduce Nuclear Targeting of the PB1-PA Dimer and Result in Viral Attenuation

    PubMed Central

    Da Costa, Bruno; Sausset, Alix; Munier, Sandie; Ghounaris, Alexandre; Naffakh, Nadia; Le Goffic, Ronan

    2015-01-01

    ABSTRACT The influenza virus RNA-dependent RNA polymerase catalyzes genome replication and transcription within the cell nucleus. Efficient nuclear import and assembly of the polymerase subunits PB1, PB2, and PA are critical steps in the virus life cycle. We investigated the structure and function of the PA linker (residues 197 to 256), located between its N-terminal endonuclease domain and its C-terminal structured domain that binds PB1, the polymerase core. Circular dichroism experiments revealed that the PA linker by itself is structurally disordered. A large series of PA linker mutants exhibited a temperature-sensitive (ts) phenotype (reduced viral growth at 39.5°C versus 37°C/33°C), suggesting an alteration of folding kinetic parameters. The ts phenotype was associated with a reduced efficiency of replication/transcription of a pseudoviral reporter RNA in a minireplicon assay. Using a fluorescent-tagged PB1, we observed that ts and lethal PA mutants did not efficiently recruit PB1 to reach the nucleus at 39.5°C. A protein complementation assay using PA mutants, PB1, and β-importin IPO5 tagged with fragments of the Gaussia princeps luciferase showed that increasing the temperature negatively modulated the PA-PB1 and the PA-PB1-IPO5 interactions or complex stability. The selection of revertant viruses allowed the identification of different types of compensatory mutations located in one or the other of the three polymerase subunits. Two ts mutants were shown to be attenuated and able to induce antibodies in mice. Taken together, our results identify a PA domain critical for PB1-PA nuclear import and that is a “hot spot” to engineer ts mutants that could be used to design novel attenuated vaccines. IMPORTANCE By targeting a discrete domain of the PA polymerase subunit of influenza virus, we were able to identify a series of 9 amino acid positions that are appropriate to engineer temperature-sensitive (ts) mutants. This is the first time that a large

  3. Identification and molecular characterization of nuclear Citrus leprosis virus, an unassigned Dichorhavirus genus member associated with citrus leprosis disease in Mexico

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Citrus leprosis is a difficult viral disease causing significant damage to citrus fruit in South America and Central America. The disease is marked by dramatic lesions on fruit, leaves and stems resulting in unmarketable product. Citrus leprosis virus cytoplasmic type (CiLV-C) was detected in states...

  4. Transcription of the Epstein-Barr virus nuclear antigen 1 (EBNA1) gene occurs before induction of the BCR2 (Cp) EBNA gene promoter during the initial stages of infection in B cells.

    PubMed

    Schlager, S; Speck, S H; Woisetschläger, M

    1996-06-01

    The purpose of this study was to gain insights into the regulation of Epstein-Barr virus (EBV) gene transcription during the establishment of viral latency in B cells. During the early stages of EBV infection in B lymphocytes, transcription of six viral nuclear antigens (EBNAs) is initiated from an early promoter (Wp). This is followed by a switch of promoter usage to an upstream promoter, Cp, whose activity is autoregulated by both EBNA1 and EBNA2. Previously it was demonstrated that infection of primary B cells with EBNA2-negative (EBNA2-) EBNA4-mutant (EBNA4mut) virus resulted only in the expression of mutant EBNA4 protein and failure to express the other EBNA gene products (C. Rooney H. G. Howe, S. H. Speck, and G. Miller, J. Virol. 63:1531-1539, 1989). We extended this research to demonstrate that Wp-to-Cp switching did not occur upon infection of primary B cells with an EBNA2- EBNA4mut virus (M. Woisetschlaeger, X. W. Jin, C. N. Yandara, L. A. Furmanski, J. L. Strominger, and S. H. Speck, Proc. Natl. Acad. Sci. USA 88:3942-3946, 1991). Further characterization of this phenomenon led to the identification of an EBNA2-dependent enhancer upstream of Cp. On the basis of these data, a model was proposed in which initial transcription from Wp gives rise to the expression of EBNA2 and EBNA4, and then transcription is upregulated from Cp via the EBNA2- dependent enhancer (Woisetschlaeger et al., as noted above). Implicit in this model is that transcription of the EBNA1 and EBNA3a to -3c genes is dependent on the switch from Wp to Cp, since primary cells infected with EBNA2- EBNA4mut virus fail to switch and also fail to express these viral antigens. Here we critically evaluate this model and demonstrate, in contrast to the predictions of the model, that transcription of both the EBNA1 and EBNA2 genes precedes activation of Cp. Furthermore, the level of EBNA1 gene transcription was strongly reduced in primary B cells infected with EBNA2- EBNA4mut virus compared with

  5. Zika Virus

    MedlinePlus

    Zika is a virus that is spread mostly by mosquitoes. A pregnant mother can pass it to ... through blood transfusions. There have been outbreaks of Zika virus in the United States, Africa, Southeast Asia, ...

  6. Zika Virus

    MedlinePlus

    ... Search The CDC Cancel Submit Search The CDC Zika Virus Note: Javascript is disabled or is not supported ... Areas with Zika Countries and territories with active Zika virus transmission... Mosquito Control Prevent mosquito bites, integrated mosquito ...

  7. Chikungunya Virus

    MedlinePlus

    ... traveling to countries with chikungunya virus, use insect repellent, wear long sleeves and pants, and stay in ... Chikungunya Prevention is key! Prevent Infection. Use mosquito repellent. Chikungunya Virus Distribution Chikungunya in the U.S. What's ...

  8. Residues R{sup 199}H{sup 200} of prototype foamy virus transactivator Bel1 contribute to its binding with LTR and IP promoters but not its nuclear localization

    SciTech Connect

    Ma, Qinglin; Tan, Juan; Cui, Xiaoxu; Luo, Di; Yu, Miao; Liang, Chen; Qiao, Wentao

    2014-01-20

    Prototype foamy virus encodes a transactivator called Bel1 that enhances viral gene transcription and is essential for PFV replication. Nuclear localization of Bel1 has been reported to rely on two proximal basic motifs R{sup 199}H{sup 200} and R{sup 221}R{sup 222}R{sup 223} that likely function together as a bipartite nuclear localization signal. In this study, we report that mutating R{sup 221}R{sup 222}R{sup 223}, but not R{sup 199}H{sup 200}, relocates Bel1 from the nucleus to the cytoplasm, suggesting an essential role for R{sup 221}R{sup 222}R{sup 223} in the nuclear localization of Bel1. Although not affecting the nuclear localization of Bel1, mutating R{sup 199}H{sup 200} disables Bel1 from transactivating PFV promoters. Results of EMSA reveal that the R{sup 199}H{sup 200} residues are vital for the binding of Bel1 to viral promoter DNA. Moreover, mutating R{sup 199}H{sup 200} in Bel1 impairs PFV replication to a much greater extent than mutating R{sup 221}R{sup 222}R{sup 223}. Collectively, our findings suggest that R{sup 199}H{sup 200} directly participate in Bel1 binding to viral promoter DNA and are indispensible for Bel1 transactivation activity. - Highlights: • The R{sup 221}R{sup 222}R{sup 223} residues are essential for the nuclear localization of Bel1. • Although not affecting the nuclear localization of Bel1, mutating R{sup 199}H{sup 200} disables Bel1 from transactivating PFV promoters. • The R{sup 199}H{sup 200} residues directly participate in Bel1 binding to viral promoter DNA. • Mutating R{sup 199}H{sup 200} in Bel1 impairs PFV replication to a much greater extent than mutating R{sup 221}R{sup 222}R{sup 223}.

  9. Virus Maturation

    PubMed Central

    Veesler, David; Johnson, John E.

    2013-01-01

    We examined virus maturation of selected non-enveloped and enveloped ssRNA viruses; retroviruses; bacteriophages and herpes virus. Processes associated with maturation in the RNA viruses range from subtle (noda and picornaviruses) to dramatic (tetraviruses and togaviruses). The elaborate assembly and maturation pathway of HIV is discussed in contrast to the less sophisticated but highly efficient processes associated with togaviruses. Bacteriophage assembly and maturation are discussed in general terms with specific examples chosen for emphasis. Finally the herpes viruses are compared with bacteriophages. The data support divergent evolution of noda, picorna and tetraviruses from a common ancestor and divergent evolution of alpha and flaviviruses from a common ancestor. Likewise, bacteriophages and herpes viruses almost certainly share a common ancestor in their evolution. Comparing all the viruses, we conclude that maturation is a convergent process that is required to solve conflicting requirements in biological dynamics and function. PMID:22404678

  10. The 5' flanking region of the gene for the Epstein-Barr virus-encoded nuclear antigen 2 contains a cell type specific cis-acting regulatory element that activates transcription in transfected B-cells.

    PubMed Central

    Ricksten, A; Olsson, A; Andersson, T; Rymo, L

    1988-01-01

    We have recently identified the promoter that positions the initiation (cap) site for RNA encoding the Epstein-Barr virus (EBV) determined nuclear antigen 2 (EBNA2) in transfected COS-1 cells. The cells were transfected with recombinant vectors that contained the BamHI WYH region of the EBV genome. In order to delineate regulatory DNA sequences required for the expression of EBNA2 the 5' flanking region of the gene was linked to reporter genes in expression vectors and transfected into EBV genome-negative lymphoid DG75 cells. We demonstrate that several cis-acting elements contribute to a transcriptional enhancer activity found in the region between nucleotides-553 and -86 relative to the cap site. The enhancer was active in lymphoid DG75 cells but not in HeLa cells and stimulated transcription also from the heterologous thymidine kinase (TK) and beta-globin promoters. Nuclear extracts of lymphoid cells contained protein factors that bound to the enhancer. The in vitro introduction of a mutation in the enhancer sequence that substantially reduced the transcription stimulatory activity concurrently blocked the binding of one of the factors. Images PMID:2843816

  11. Significance of host cell kinases in herpes simplex virus type 1 egress and lamin-associated protein disassembly from the nuclear lamina

    SciTech Connect

    Leach, Natalie R.; Roller, Richard J.

    2010-10-10

    The nuclear lamina is thought to be a steric barrier to the herpesvirus capsid. Disruption of the lamina accompanied by phosphorylation of lamina proteins is a conserved feature of herpesvirus infection. In HSV-1-infected cells, protein kinase C (PKC) alpha and delta isoforms are recruited to the nuclear membrane and PKC delta has been implicated in phosphorylation of emerin and lamin B. We tested two critical hypotheses about the mechanism and significance of lamina disruption. First, we show that chemical inhibition of all PKC isoforms reduced viral growth five-fold and inhibited capsid egress from the nucleus. However, specific inhibition of either conventional PKCs or PKC delta does not inhibit viral growth. Second, we show hyperphosphorylation of emerin by viral and cellular kinases is required for its disassociation from the lamina. These data support hypothesis that phosphorylation of lamina components mediates lamina disruption during HSV nuclear egress.

  12. Cell lines that support replication of a novel herpes simplex virus 1 U{sub L}31 deletion mutant can properly target U{sub L}34 protein to the nuclear rim in the absence of U{sub L}31

    SciTech Connect

    Liang Li; Tanaka, Michiko; Kawaguchi, Yasushi; Baines, Joel D. . E-mail: jdb11@cornell.edu

    2004-11-10

    Previous results indicated that the herpes simplex virus 1 (HSV-1) U{sub L}31 gene is necessary and sufficient for localization of the U{sub L}34 protein exclusively to the nuclear membrane of infected Hep2 cells. In the current studies, a bacterial artificial chromosome containing the entire HSV-1 strain F genome was used to construct a recombinant viral genome in which a gene encoding kanamycin resistance was inserted in place of 262 codons of the 306 codon U{sub L}31 open reading frame. The deletion virus produced virus titers approximately 10- to 50-fold lower in rabbit skin cells, more than 2000-fold lower in Vero cells, and more than 1500-fold lower in CV1 cells, compared to a virus bearing a restored U{sub L}31 gene. The replication of the U{sub L}31 deletion virus was restored on U{sub L}31-complementing cell lines derived either from rabbit skin cells or CV1 cells. Confocal microscopy indicated that the majority of U{sub L}34 protein localized aberrantly in the cytoplasm and nucleoplasm of Vero cells and CV1 cells, whereas U{sub L}34 protein localized at the nuclear membrane in rabbit skin cells, and U{sub L}31 complementing CV1 cells infected with the U{sub L}31 deletion virus. We conclude that rabbit skin cells encode a function that allows proper localization of U{sub L}34 protein to the nuclear membrane. We speculate that this function partially complements that of U{sub L}31 and may explain why U{sub L}31 is less critical for replication in rabbit skin cells as opposed to Vero and CV1 cells.

  13. Herpes simplex virus 2 modulates apoptosis and stimulates NF-{kappa}B nuclear translocation during infection in human epithelial HEp-2 cells

    SciTech Connect

    Yedowitz, Jamie C.; Blaho, John A. . E-mail: john.blaho@mssm.edu

    2005-11-25

    Virus-mediated apoptosis is well documented in various systems, including herpes simplex virus 1 (HSV-1). HSV-2 is closely related to HSV-1 but its apoptotic potential during infection has not been extensively scrutinized. We report that (i) HEp-2 cells infected with HSV-2(G) triggered apoptosis, assessed by apoptotic cellular morphologies, oligosomal DNA laddering, chromatin condensation, and death factor processing when a translational inhibitor (CHX) was added at 3 hpi. Thus, HSV-2 induced apoptosis but was unable to prevent the process from killing cells. (ii) Results from a time course of CHX addition experiment indicated that infected cell protein produced between 3 and 5 hpi, termed the apoptosis prevention window, are required for blocking virus-induced apoptosis. This corresponds to the same prevention time frame as reported for HSV-1. (iii) Importantly, CHX addition prior to 3 hpi led to less apoptosis than that at 3 hpi. This suggests that proteins produced immediately upon infection are needed for efficient apoptosis induction by HSV-2. This finding is different from that observed previously with HSV-1. (iv) Infected cell factors produced during the HSV-2(G) prevention window inhibited apoptosis induced by external TNF{alpha} plus cycloheximide treatment. (v) NF-{kappa}B translocated to nuclei and its presence in nuclei correlated with apoptosis prevention during HSV-2(G) infection. (vi) Finally, clinical HSV-2 isolates induced and prevented apoptosis in HEp-2 cells in a manner similar to that of laboratory strains. Thus, while laboratory and clinical HSV-2 strains are capable of modulating apoptosis in human HEp-2 cells, the mechanism of HSV-2 induction of apoptosis differs from that of HSV-1.

  14. Zika virus.

    PubMed

    2016-02-10

    Essential facts Zika virus disease is caused by a virus that is transmitted by the Aedes mosquito. While it generally causes a mild illness, there is increasing concern that it is harmful in pregnancy and can cause congenital abnormalities in infants born to women infected with the virus. There is no antiviral treatment or vaccine currently available. The best form of prevention is protection against mosquito bites.

  15. Ebola virus.

    PubMed

    Streether, L A

    1999-01-01

    Ebola virus was first identified as a filovirus in 1976, following epidemics of severe haemorrhagic fever in sub-Saharan Africa. Further outbreaks have occurred since, but, despite extensive and continued investigations, the natural reservoir for the virus remains unknown. The mortality rate is high and there is no cure for Ebola virus infection. Molecular technology is proving useful in extending our knowledge of the virus. Identification of the host reservoir, control and prevention of further outbreaks, rapid diagnosis of infection, and vaccine development remain areas of continued interest in the fight against this biosafety level-four pathogen.

  16. Virus Crystallography

    NASA Astrophysics Data System (ADS)

    Fry, Elizabeth; Logan, Derek; Stuart, David

    Crystallography provides a means of visualizing intact virus particles as well as their isolated constituent proteins and enzymes (1-3) at near-atomic resolution, and is thus an extraordinarily powerful tool in the pursuit of a fuller understanding of the functioning of these simple biological systems. We have already expanded our knowledge of virus evolution, assembly, antigenic variation, and host-cell interactions; further studies will no doubt reveal much more. Although the rewards are enormous, an intact virus structure determination is not a trivial undertaking and entails a significant scaling up in terms of time and resources through all stages of data collection and processing compared to a traditional protein crystallographic structure determination. It is the methodology required for such studies that will be the focus of this chapter. The computational requirements were satisfied in the late 1970s, and when combined with the introduction of phase improvement techniques utilizing the virus symmetry (4,5), the application of crystallography to these massive macromolecular assemblies became feasible. This led to the determination of the first virus structure (the small RNA plant virus, tomato bushy stunt virus), by Harrison and coworkers in 1978 (6). The structures of two other plant viruses followed rapidly (7,8). In the 1980s, a major focus of attention was a family of animal RNA viruses; the Picornaviridae.

  17. Fusion of Epstein-Barr virus nuclear antigen-1-derived glycine-alanine repeat to trans-dominant HIV-1 Gag increases inhibitory activities and survival of transduced cells in vivo.

    PubMed

    Hammer, Diana; Wild, Jens; Ludwig, Christine; Asbach, Benedikt; Notka, Frank; Wagner, Ralf

    2008-06-01

    Trans-dominant human immunodeficiency virus type 1 (HIV-1) Gag derivatives have been shown to efficiently inhibit late steps of HIV-1 replication in vitro by interfering with Gag precursor assembly, thus ranking among the interesting candidates for gene therapy approaches. However, efficient antiviral activities of corresponding transgenes are likely to be counteracted in particular by cell-mediated host immune responses toward the transgene-expressing cells. To decrease this potential immunogenicity, a 24-amino acid Gly-Ala (GA) stretch derived from Epstein-Barr virus nuclear antigen-1 (EBNA1) and known to overcome proteasomal degradation was fused to a trans-dominant Gag variant (sgD1). To determine the capacity of this fusion polypeptide to repress viral replication, PM-1 cells were transduced with sgD1 and GAsgD1 transgenes, using retroviral gene transfer. Challenge of stably transfected permissive cell lines with various viral strains indicated that N-terminal GA fusion even enhanced the inhibitory properties of sgD1. Further studies revealed that the GA stretch increased protein stability by blocking proteasomal degradation of Gag proteins. Immunization of BALB/c mice with a DNA vaccine vector expressing sgD1 induced substantial Gag-specific immune responses that were, however, clearly diminished in the presence of GA. Furthermore, recognition of cells expressing the GA-fused transgene by CD8(+) T cells was drastically reduced, both in vitro and in vivo, resulting in prolonged survival of the transduced cells in recipient mice.

  18. Live Virus Smallpox Vaccine

    MedlinePlus

    ... Index SMALLPOX FACT SHEET The Live Virus Smallpox Vaccine The vaccinia virus is the "live virus" used ... cannot cause smallpox. What is a "live virus" vaccine? A "live virus" vaccine is a vaccine that ...

  19. Stable transfection of Epstein-Barr virus (EBV) nuclear antigen 2 in lymphoma cells containing the EBV P3HR1 genome induces expression of B-cell activation molecules CD21 and CD23.

    PubMed Central

    Cordier, M; Calender, A; Billaud, M; Zimber, U; Rousselet, G; Pavlish, O; Banchereau, J; Tursz, T; Bornkamm, G; Lenoir, G M

    1990-01-01

    A set of B-cell activation molecules, including the Epstein-Barr virus (EBV) receptor CR2 (CD21) and the B-cell activation antigen CD23 (Blast2/Fc epsilon RII), is turned on by infecting EBV-negative B-lymphoma cell lines with immortalizing strains of the viruslike B95-8 (BL/B95 cells). This up regulation may represent one of the mechanisms involved in EBV-mediated B-cell immortalization. The P3HR1 nonimmortalizing strain of the virus, which is deleted for the entire Epstein-Barr nuclear antigen 2 (EBNA2) protein open reading frame, is incapable of inducing the expression of CR2 and CD23, suggesting a crucial role for EBNA2 in the activation of these molecules. In addition, lymphoma cells containing the P3HR1 genome (BL/P3HR1 cells) do not express the viral latent membrane protein (LMP), which is regularly expressed in cells infected with immortalizing viral strains. Using electroporation, we have transfected the EBNA2 gene cloned in an episomal vector into BL/P3HR1 cells and have obtained cell clones that stably express the EBNA2 protein. In these clones, EBNA2 expression was associated with an increased amount of CR2 and CD23 steady-state RNAs. Of the three species of CD23 mRNAs described, the Fc epsilon RIIa species was preferentially expressed in these EBNA2-expressing clones. An increased cell surface expression of CR2 but not of CD23 was observed, and the soluble form of CD23 molecule (SCD23) was released. We were, however, not able to detect any expression of LMP in these cell clones. These data demonstrate that EBNA2 gene is able to complement P3HR1 virus latent functions to induce the activation of CR2 and CD23 expression, and they emphasize the role of EBNA2 protein in the modulation of cellular gene implicated in B-cell proliferation and hence in EBV-mediated B-cell immortalization. Nevertheless, EBNA2 expression in BL/P3HR1 cells is not able to restore the level of CR2 and CD23 expression observed in BL/B95 cells, suggesting that other cellular or viral

  20. The C-terminal repressor region of herpes simplex virus type 1 ICP27 is required for the redistribution of small nuclear ribonucleoprotein particles and splicing factor SC35; however, these alterations are not sufficient to inhibit host cell splicing.

    PubMed Central

    Sandri-Goldin, R M; Hibbard, M K; Hardwicke, M A

    1995-01-01

    Herpes simplex virus type 1 infection results in a reorganization of antigens associated with the small nuclear ribonucleoprotein particles (snRNPs), resulting in the formation of prominent clusters near the nuclear periphery. In this study, we show that the immediate-early protein ICP27, which is involved in the impairment of host cell splicing and in the changes in the distribution of snRNPs, is also required for reassorting the SR domain splicing factor SC35. Other viral processes, such as adsorption and penetration, shutoff of host protein synthesis, early and late gene expression, and DNA replication, do not appear to play a role in changing the staining pattern of splicing antigens. Furthermore, the C-terminal repressor region of ICP27, which is required for the inhibitory effects on splicing, also is involved in redistributing the snRNPs and SC35. During infection or transfection with five different repressor mutants, the speckled staining pattern characteristic of uninfected cells was seen and the level of a spliced target mRNA was not reduced. Infections in the presence of activator mutants showed a redistributed snRNP pattern and a decreased accumulation of spliced target mRNA. Moreover, two arginine-rich regions in the N-terminal half of ICP27 were not required for the redistribution of snRNPs or SC35. Substitution of these regions with a lysine-rich sequence from simian virus 40 large-T antigen resulted in a redistribution of splicing antigens. Unexpectedly, a repressor mutant with a ts phenotype showed a redistributed staining pattern like that seen with wild-type infected cells. During infections with this ts mutant, splicing was not inhibited, as shown in this and previous studies, confirming its repressor phenotype. Furthermore, both the mutant and the wild-type protein colocalized with snRNPs. Therefore, the redistribution of snRNPs and SC35 correlates with ICP27-mediated impairment of host cell splicing, but these alterations are not sufficient to

  1. Primary monolayer culture of adult mouse hepatocytes -- a model for the study of hepatotropic viruses.

    PubMed

    Arnheiter, H

    1980-01-01

    Primary monolayer cultures of adult mouse hepatocytes isolated by collagenase perfusion of the liver in situ were exposed to 2 hepatotropic viruses, an avian influenza A virus adapted to grow in mouse liver in vivo and a herpes simplex type I virus. Influenza virus infection led to lysis ofindividual hepatocytes and total monolayer destruction within 18 to 120 hours after infection according to the virus dose used. Virus replication was evidenced by assaying hepatocyte supernates for hemagglutinin and infectivity, by immunofluorescent staining and by electron microscopy. Herpes virus infection resulted in polykaryocyte formation followed by nuclear pycnosis and cell lysis. Virus replication was assayed by titration of supernate infectivity.

  2. Design stars: how small DNA viruses remodel the host nucleus.

    PubMed

    Jiang, Mengxi; Imperiale, Michael J

    2012-05-01

    Numerous host components are encountered by viruses during the infection process. While some of these host structures are left unchanged, others may go through dramatic remodeling processes. In this review, we summarize these host changes that occur during small DNA virus infections, with a focus on host nuclear components and pathways. Although these viruses differ significantly in their genome structures and infectious pathways, there are common nuclear targets that are altered by various viral factors. Accumulating evidence suggests that these nuclear remodeling processes are often essential for productive viral infections and/or viral-induced transformation. Understanding the complex interactions between viruses and these host structures and pathways will help to build a more integrated network of how the virus completes its life cycle and point toward the design of novel therapeutic regimens that either prevent harmful viral infections or employ viruses as nontraditional treatment options or molecular tools.

  3. Computer viruses

    NASA Technical Reports Server (NTRS)

    Denning, Peter J.

    1988-01-01

    The worm, Trojan horse, bacterium, and virus are destructive programs that attack information stored in a computer's memory. Virus programs, which propagate by incorporating copies of themselves into other programs, are a growing menace in the late-1980s world of unprotected, networked workstations and personal computers. Limited immunity is offered by memory protection hardware, digitally authenticated object programs,and antibody programs that kill specific viruses. Additional immunity can be gained from the practice of digital hygiene, primarily the refusal to use software from untrusted sources. Full immunity requires attention in a social dimension, the accountability of programmers.

  4. Hendra virus.

    PubMed

    Middleton, Deborah

    2014-12-01

    Hendra virus infection of horses occurred sporadically between 1994 and 2010 as a result of spill-over from the viral reservoir in Australian mainland flying-foxes, and occasional onward transmission to people also followed from exposure to affected horses. An unprecedented number of outbreaks were recorded in 2011 leading to heightened community concern. Release of an inactivated subunit vaccine for horses against Hendra virus represents the first commercially available product that is focused on mitigating the impact of a Biosafety Level 4 pathogen. Through preventing the development of acute Hendra virus disease in horses, vaccine use is also expected to reduce the risk of transmission of infection to people.

  5. Hendra virus.

    PubMed

    Middleton, Deborah

    2014-12-01

    Hendra virus infection of horses occurred sporadically between 1994 and 2010 as a result of spill-over from the viral reservoir in Australian mainland flying-foxes, and occasional onward transmission to people also followed from exposure to affected horses. An unprecedented number of outbreaks were recorded in 2011 leading to heightened community concern. Release of an inactivated subunit vaccine for horses against Hendra virus represents the first commercially available product that is focused on mitigating the impact of a Biosafety Level 4 pathogen. Through preventing the development of acute Hendra virus disease in horses, vaccine use is also expected to reduce the risk of transmission of infection to people. PMID:25281398

  6. Nuclear Medicine

    MedlinePlus

    ... Parents/Teachers Resource Links for Students Glossary Nuclear Medicine What is nuclear medicine? What are radioactive tracers? ... funded researchers advancing nuclear medicine? What is nuclear medicine? Nuclear medicine is a medical specialty that uses ...

  7. Zika Virus

    MedlinePlus

    ... be at risk for developing fetal complications. Blood, organ and tissue donor screening tests are also needed to assure the safety of transfusion and transplantation in areas of active mosquito-borne virus transmission. ...

  8. Chikungunya virus

    MedlinePlus

    ... first time in the Americas in the Caribbean Islands. In the Americas, local transmission of the disease ... in Florida, Puerto Rico, and the U.S. Virgin Islands. How Chikungunya can spread Mosquitoes spread the virus ...

  9. Zika Virus.

    PubMed

    Phillips, Jennan A; Neyland, Anavernyel

    2016-08-01

    Zika virus (ZIKV) infections are the latest global public health emergency. Occupational health nurses can protect society by educating workers, women of childbearing age, and others traveling in ZIKV-infected areas about prevention strategies.

  10. Dengue virus.

    PubMed

    Ross, Ted M

    2010-03-01

    Dengue is the most prevalent arthropod-borne virus affecting humans today. The virus group consists of 4 serotypes that manifest with similar symptoms. Dengue causes a spectrum of disease, ranging from a mild febrile illness to a life-threatening dengue hemorrhagic fever. Breeding sites for the mosquitoes that transmit dengue virus have proliferated, partly because of population growth and uncontrolled urbanization in tropical and subtropical countries. Successful vector control programs have also been eliminated, often because of lack of governmental funding. Dengue viruses have evolved rapidly as they have spread worldwide, and genotypes associated with increased virulence have spread across Asia and the Americas. This article describes the virology, epidemiology, clinical manifestations and outcomes, and treatments/vaccines associated with dengue infection.

  11. Zika Virus.

    PubMed

    Phillips, Jennan A; Neyland, Anavernyel

    2016-08-01

    Zika virus (ZIKV) infections are the latest global public health emergency. Occupational health nurses can protect society by educating workers, women of childbearing age, and others traveling in ZIKV-infected areas about prevention strategies. PMID:27411846

  12. Epstein-Barr virus nuclear antigen 3A promotes cellular proliferation by repression of the cyclin-dependent kinase inhibitor p21WAF1/CIP1.

    PubMed

    Tursiella, Melissa L; Bowman, Emily R; Wanzeck, Keith C; Throm, Robert E; Liao, Jason; Zhu, Junjia; Sample, Clare E

    2014-10-01

    Latent infection by Epstein-Barr virus (EBV) is highly associated with the endemic form of Burkitt lymphoma (eBL), which typically limits expression of EBV proteins to EBNA-1 (Latency I). Interestingly, a subset of eBLs maintain a variant program of EBV latency - Wp-restricted latency (Wp-R) - that includes expression of the EBNA-3 proteins (3A, 3B and 3C), in addition to EBNA-1. In xenograft assays, Wp-R BL cell lines were notably more tumorigenic than their counterparts that maintain Latency I, suggesting that the additional latency-associated proteins expressed in Wp-R influence cell proliferation and/or survival. Here, we evaluated the contribution of EBNA-3A. Consistent with the enhanced tumorigenic potential of Wp-R BLs, knockdown of EBNA-3A expression resulted in abrupt cell-cycle arrest in G0/G1 that was concomitant with conversion of retinoblastoma protein (Rb) to its hypophosphorylated state, followed by a loss of Rb protein. Comparable results were seen in EBV-immortalized B lymphoblastoid cell lines (LCLs), consistent with the previous observation that EBNA-3A is essential for sustained growth of these cells. In agreement with the known ability of EBNA-3A and EBNA-3C to cooperatively repress p14(ARF) and p16(INK4a) expression, knockdown of EBNA-3A in LCLs resulted in rapid elevation of p14(ARF) and p16I(NK4a). By contrast, p16(INK4a) was not detectably expressed in Wp-R BL and the low-level expression of p14(ARF) was unchanged by EBNA-3A knockdown. Amongst other G1/S regulatory proteins, only p21(WAF1/CIP1), a potent inducer of G1 arrest, was upregulated following knockdown of EBNA-3A in Wp-R BL Sal cells and LCLs, coincident with hypophosphorylation and destabilization of Rb and growth arrest. Furthermore, knockdown of p21(WAF1/CIP1) expression in Wp-R BL correlated with an increase in cellular proliferation. This novel function of EBNA-3A is distinct from the functions previously described that are shared with EBNA-3C, and likely contributes to the

  13. Detection of virus-specific antigen in the nuclei or nucleoli of cells infected with Zika or Langat virus.

    PubMed

    Buckley, A; Gould, E A

    1988-08-01

    Two monoclonal antibodies (MAbs) with molecular specificities for either the viral envelope glycoprotein (MAb 541) or the non-structural NS1 glycoprotein (MAb 109) were derived using West Nile and yellow fever (YF) viruses respectively. Their antigenic reactivity with a large number of flaviviruses was tested by indirect immunofluorescence microscopy. Both produced cytoplasmic fluorescent staining patterns with the homologous virus against which they were raised. Additionally, MAb 541 reacted with two substrains of YF virus whereas MAb 109 reacted with Bussuquara, YF and Ntaya viruses. These reactions were exclusively cytoplasmic. Two unexpected patterns of fluorescent labelling were observed when the antibodies were tested with Zika and Langat viruses. MAb 541 produced fluorescent staining of the nuclei, but not the cytoplasm, of cells infected with Zika virus and MAb 109 labelled only the nucleoli of cells infected with Langat virus. Double-labelling experiments showed that the nuclear fluorescent label was confined to virus-infected cells, and antibody absorption experiments with virus-infected cell packs confirmed the virus specificity of the nuclear antigen. The unexpected presence of virus-specific antigen in the nuclei or nucleoli of Zika or Langat virus-infected cells brings into question the role of the nucleus in flavivirus replication.

  14. Computer Viruses. Technology Update.

    ERIC Educational Resources Information Center

    Ponder, Tim, Comp.; Ropog, Marty, Comp.; Keating, Joseph, Comp.

    This document provides general information on computer viruses, how to help protect a computer network from them, measures to take if a computer becomes infected. Highlights include the origins of computer viruses; virus contraction; a description of some common virus types (File Virus, Boot Sector/Partition Table Viruses, Trojan Horses, and…

  15. Bromodomain and extraterminal (BET) protein inhibition suppresses human T cell leukemia virus 1 (HTLV-1) Tax protein-mediated tumorigenesis by inhibiting nuclear factor κB (NF-κB) signaling.

    PubMed

    Wu, Xuewei; Qi, Jun; Bradner, James E; Xiao, Gutian; Chen, Lin-Feng

    2013-12-13

    The etiology of human T cell leukemia virus 1 (HTLV-1)-mediated adult T cell leukemia is associated with the ability of viral oncoprotein Tax to induce sustained NF-κB activation and the expression of many NF-κB target genes. Acetylation of the RelA subunit of NF-κB and the subsequent recruitment of bromodomain-containing factor Brd4 are important for the expression of NF-κB target genes in response to various stimuli. However, their contributions to Tax-mediated NF-κB target gene expression and tumorigenesis remain unclear. Here we report that Tax induced the acetylation of lysine 310 of RelA and the binding of Brd4 to acetylated RelA to facilitate Tax-mediated transcriptional activation of NF-κB. Depletion of Brd4 down-regulated Tax-mediated NF-κB target gene expression and cell proliferation. Inhibiting the interaction of Brd4 and acetylated RelA with the bromodomain extraterminal protein inhibitor JQ1 suppressed the proliferation of Tax-expressing rat fibroblasts and Tax-positive HTLV-1-infected cells and Tax-mediated cell transformation and tumorigenesis. Moreover, JQ1 attenuated the Tax-mediated transcriptional activation of NF-κB, triggering the polyubiquitination and proteasome-mediated degradation of constitutively active nuclear RelA. Our results identify Brd4 as a key regulator for Tax-mediated NF-κB gene expression and suggest that targeting epigenetic regulators such as Brd4 with the bromodomain extraterminal protein inhibitor might be a potential therapeutic strategy for cancers and other diseases associated with HTLV-1 infection.

  16. Parainfluenza Viruses

    PubMed Central

    Henrickson, Kelly J.

    2003-01-01

    Human parainfluenza viruses (HPIV) were first discovered in the late 1950s. Over the last decade, considerable knowledge about their molecular structure and function has been accumulated. This has led to significant changes in both the nomenclature and taxonomic relationships of these viruses. HPIV is genetically and antigenically divided into types 1 to 4. Further major subtypes of HPIV-4 (A and B) and subgroups/genotypes of HPIV-1 and HPIV-3 have been described. HPIV-1 to HPIV-3 are major causes of lower respiratory infections in infants, young children, the immunocompromised, the chronically ill, and the elderly. Each subtype can cause somewhat unique clinical diseases in different hosts. HPIV are enveloped and of medium size (150 to 250 nm), and their RNA genome is in the negative sense. These viruses belong to the Paramyxoviridae family, one of the largest and most rapidly growing groups of viruses causing significant human and veterinary disease. HPIV are closely related to recently discovered megamyxoviruses (Hendra and Nipah viruses) and metapneumovirus. PMID:12692097

  17. Hendra virus

    PubMed Central

    Middleton, Deborah

    2014-01-01

    Synopsis Hendra virus infection of horses occurred sporadically between 1994 and 2010 as a result of spill-over from the viral reservoir in Australian mainland flying-foxes, and occasional onward transmission to people also followed from exposure to affected horses. For reasons that are not well understood an unprecedented number of outbreaks were recorded in 2011, including the first recorded field infection of a dog, leading to heightened community concern. Increasingly, pressure mounted to instigate measures for control of flying-fox numbers, and equine health care workers started to leave the industry on account of risk and liability concerns. Release of an inactivated subunit vaccine for horses against Hendra virus represents the first commercially available product that is focused on mitigating the impact of a Biosafety Level 4 pathogen. Through preventing the development of acute Hendra virus disease in horses, vaccine use is also expected to reduce the risk of transmission of infection to people. This approach to emerging infectious disease management focuses on the role of horses as the proximal cause of human Hendra virus disease, and may assist in redirecting community concerns away from the flying-fox reservoirs, keystone species for the ongoing health of Australia’s native forests. PMID:25281398

  18. [Influenza virus].

    PubMed

    Juozapaitis, Mindaugas; Antoniukas, Linas

    2007-01-01

    Every year, especially during the cold season, many people catch an acute respiratory disease, namely flu. It is easy to catch this disease; therefore, it spreads very rapidly and often becomes an epidemic or a global pandemic. Airway inflammation and other body ailments, which form in a very short period, torment the patient several weeks. After that, the symptoms of the disease usually disappear as quickly as they emerged. The great epidemics of flu have rather unique characteristics; therefore, it is possible to identify descriptions of such epidemics in historic sources. Already in the 4th century bc, Hippocrates himself wrote about one of them. It is known now that flu epidemics emerge rather frequently, but there are no regular intervals between those events. The epidemics can differ in their consequences, but usually they cause an increased mortality of elderly people. The great flu epidemics of the last century took millions of human lives. In 1918-19, during "The Spanish" pandemic of flu, there were around 40-50 millions of deaths all over the world; "Pandemic of Asia" in 1957 took up to one million lives, etc. Influenza virus can cause various disorders of the respiratory system: from mild inflammations of upper airways to acute pneumonia that finally results in the patient's death. Scientist Richard E. Shope, who investigated swine flu in 1920, had a suspicion that the cause of this disease might be a virus. Already in 1933, scientists from the National Institute for Medical Research in London - Wilson Smith, Sir Christopher Andrewes, and Sir Patrick Laidlaw - for the first time isolated the virus, which caused human flu. Then scientific community started the exhaustive research of influenza virus, and the great interest in this virus and its unique features is still active even today.

  19. Nuclear rights - nuclear wrongs

    SciTech Connect

    Paul, E.F.; Miller, F.D.; Paul, J.; Ahrens, J.

    1986-01-01

    This book contains 11 selections. The titles are: Three Ways to Kill Innocent Bystanders: Some Conundrums Concerning the Morality of War; The International Defense of Liberty; Two Concepts of Deterrence; Nuclear Deterrence and Arms Control; Ethical Issues for the 1980s; The Moral Status of Nuclear Deterrent Threats; Optimal Deterrence; Morality and Paradoxical Deterrence; Immoral Risks: A Deontological Critique of Nuclear Deterrence; No War Without Dictatorship, No Peace Without Democracy: Foreign Policy as Domestic Politics; Marxism-Leninism and its Strategic Implications for the United States; Tocqueveille War.

  20. The Geometry of Viruses.

    ERIC Educational Resources Information Center

    Case, Christine L.

    1991-01-01

    Presented is an activity in which students make models of viruses, which allows them to visualize the shape of these microorganisms. Included are some background on viruses, the biology and geometry of viruses, directions for building viruses, a comparison of cells and viruses, and questions for students. (KR)

  1. Epstein-Barr virus nuclear antigen 3C binds to BATF/IRF4 or SPI1/IRF4 composite sites and recruits Sin3A to repress CDKN2A.

    PubMed

    Jiang, Sizun; Willox, Bradford; Zhou, Hufeng; Holthaus, Amy M; Wang, Anqi; Shi, Tommy T; Maruo, Seiji; Kharchenko, Peter V; Johannsen, Eric C; Kieff, Elliott; Zhao, Bo

    2014-01-01

    Epstein-Barr virus nuclear antigen 3C (EBNA3C) repression of CDKN2A p14(ARF) and p16(INK4A) is essential for immortal human B-lymphoblastoid cell line (LCL) growth. EBNA3C ChIP-sequencing identified >13,000 EBNA3C sites in LCL DNA. Most EBNA3C sites were associated with active transcription; 64% were strong H3K4me1- and H3K27ac-marked enhancers and 16% were active promoters marked by H3K4me3 and H3K9ac. Using ENCODE LCL transcription factor ChIP-sequencing data, EBNA3C sites coincided (±250 bp) with RUNX3 (64%), BATF (55%), ATF2 (51%), IRF4 (41%), MEF2A (35%), PAX5 (34%), SPI1 (29%), BCL11a (28%), SP1 (26%), TCF12 (23%), NF-κB (23%), POU2F2 (23%), and RBPJ (16%). EBNA3C sites separated into five distinct clusters: (i) Sin3A, (ii) EBNA2/RBPJ, (iii) SPI1, and (iv) strong or (v) weak BATF/IRF4. EBNA3C signals were positively affected by RUNX3, BATF/IRF4 (AICE) and SPI1/IRF4 (EICE) cooccupancy. Gene set enrichment analyses correlated EBNA3C/Sin3A promoter sites with transcription down-regulation (P < 1.6 × 10(-4)). EBNA3C signals were strongest at BATF/IRF4 and SPI1/IRF4 composite sites. EBNA3C bound strongly to the p14(ARF) promoter through SPI1/IRF4/BATF/RUNX3, establishing RBPJ-, Sin3A-, and REST-mediated repression. EBNA3C immune precipitated with Sin3A and conditional EBNA3C inactivation significantly decreased Sin3A binding at the p14(ARF) promoter (P < 0.05). These data support a model in which EBNA3C binds strongly to BATF/IRF4/SPI1/RUNX3 sites to enhance transcription and recruits RBPJ/Sin3A- and REST/NRSF-repressive complexes to repress p14(ARF) and p16(INK4A) expression. PMID:24344258

  2. Mutations of amino acids in the DNA-recognition domain of Epstein-Barr virus ZEBRA protein alter its sub-nuclear localization and affect formation of replication compartments

    SciTech Connect

    Park, Richard; Heston, Lee; Shedd, Duane; Delecluse, Henri-Jacques; Miller, George

    2008-12-20

    ZEBRA, a transcription factor and DNA replication protein encoded by the Epstein-Barr virus (EBV) BZLF1 gene, plays indispensable roles in the EBV lytic cycle. We recently described the phenotypes of 46 single amino acid substitutions introduced into the DNA-recognition region of ZEBRA [Heston, L., El-Guindy, A., Countryman, J., Dela Cruz, C., Delecluse, H.J., and Miller, G. 2006]. The 27 DNA-binding-proficient mutants exhibited distinct defects in their ability to activate expression of the kinetic classes of viral genes. Four phenotypic variants could be discerned: wild-type, defective at activating Rta, defective at activating early genes, and defective at activating late genes. Here we analyze the distribution of ZEBRA within the nucleus and the localization of EA-D (the viral DNA polymerase processivity factor), an indicator of the development of replication compartments, in representatives of each phenotypic group. Plasmids encoding wild-type (WT) and mutant ZEBRA were transfected into 293 cells containing EBV-bacmids. WT ZEBRA protein was diffusely and smoothly distributed throughout the nucleus, sparing nucleoli, and partially recruited to globular replication compartments. EA-D induced by WT ZEBRA was present diffusely in some cells and concentrated in globular replication compartments in other cells. The distribution of ZEBRA and EA-D proteins was identical to WT following transfection of K188R, a mutant with a conservative change. The distribution of S186A mutant ZEBRA protein, defective for activation of Rta and EA-D, was identical to WT, except that the mutant ZEBRA was never found in globular compartments. Co-expression of Rta with S186A mutant rescued diffuse EA-D but not globular replication compartments. The most striking observation was that several mutant ZEBRA proteins defective in activating EA-D (R179A, K181A and A185V) and defective in activating lytic viral DNA replication and late genes (Y180E and K188A) were localized to numerous punctate

  3. Nuclear actin and lamins in viral infections.

    PubMed

    Cibulka, Jakub; Fraiberk, Martin; Forstova, Jitka

    2012-03-01

    Lamins are the best characterized cytoskeletal components of the cell nucleus that help to maintain the nuclear shape and participate in diverse nuclear processes including replication or transcription. Nuclear actin is now widely accepted to be another cytoskeletal protein present in the nucleus that fulfills important functions in the gene expression. Some viruses replicating in the nucleus evolved the ability to interact with and probably utilize nuclear actin for their replication, e.g., for the assembly and transport of capsids or mRNA export. On the other hand, lamins play a role in the propagation of other viruses since nuclear lamina may represent a barrier for virions entering or escaping the nucleus. This review will summarize the current knowledge about the roles of nuclear actin and lamins in viral infections.

  4. Plant Virus Metagenomics: Advances in Virus Discovery.

    PubMed

    Roossinck, Marilyn J; Martin, Darren P; Roumagnac, Philippe

    2015-06-01

    In recent years plant viruses have been detected from many environments, including domestic and wild plants and interfaces between these systems-aquatic sources, feces of various animals, and insects. A variety of methods have been employed to study plant virus biodiversity, including enrichment for virus-like particles or virus-specific RNA or DNA, or the extraction of total nucleic acids, followed by next-generation deep sequencing and bioinformatic analyses. All of the methods have some shortcomings, but taken together these studies reveal our surprising lack of knowledge about plant viruses and point to the need for more comprehensive studies. In addition, many new viruses have been discovered, with most virus infections in wild plants appearing asymptomatic, suggesting that virus disease may be a byproduct of domestication. For plant pathologists these studies are providing useful tools to detect viruses, and perhaps to predict future problems that could threaten cultivated plants.

  5. A single vertebrate DNA virus protein disarms invertebrate immunity to RNA virus infection

    PubMed Central

    Gammon, Don B; Duraffour, Sophie; Rozelle, Daniel K; Hehnly, Heidi; Sharma, Rita; Sparks, Michael E; West, Cara C; Chen, Ying; Moresco, James J; Andrei, Graciela; Connor, John H; Conte, Darryl; Gundersen-Rindal, Dawn E; Marshall, William L; Yates, John R; Silverman, Neal; Mello, Craig C

    2014-01-01

    Virus-host interactions drive a remarkable diversity of immune responses and countermeasures. We found that two RNA viruses with broad host ranges, vesicular stomatitis virus (VSV) and Sindbis virus (SINV), are completely restricted in their replication after entry into Lepidopteran cells. This restriction is overcome when cells are co-infected with vaccinia virus (VACV), a vertebrate DNA virus. Using RNAi screening, we show that Lepidopteran RNAi, Nuclear Factor-κB, and ubiquitin-proteasome pathways restrict RNA virus infection. Surprisingly, a highly conserved, uncharacterized VACV protein, A51R, can partially overcome this virus restriction. We show that A51R is also critical for VACV replication in vertebrate cells and for pathogenesis in mice. Interestingly, A51R colocalizes with, and stabilizes, host microtubules and also associates with ubiquitin. We show that A51R promotes viral protein stability, possibly by preventing ubiquitin-dependent targeting of viral proteins for destruction. Importantly, our studies reveal exciting new opportunities to study virus-host interactions in experimentally-tractable Lepidopteran systems. DOI: http://dx.doi.org/10.7554/eLife.02910.001 PMID:24966209

  6. Zika Virus.

    PubMed

    Musso, Didier; Gubler, Duane J

    2016-07-01

    Zika virus (ZIKV) is an arthropod-borne virus (arbovirus) in the genus Flavivirus and the family Flaviviridae. ZIKV was first isolated from a nonhuman primate in 1947 and from mosquitoes in 1948 in Africa, and ZIKV infections in humans were sporadic for half a century before emerging in the Pacific and the Americas. ZIKV is usually transmitted by the bite of infected mosquitoes. The clinical presentation of Zika fever is nonspecific and can be misdiagnosed as other infectious diseases, especially those due to arboviruses such as dengue and chikungunya. ZIKV infection was associated with only mild illness prior to the large French Polynesian outbreak in 2013 and 2014, when severe neurological complications were reported, and the emergence in Brazil of a dramatic increase in severe congenital malformations (microcephaly) suspected to be associated with ZIKV. Laboratory diagnosis of Zika fever relies on virus isolation or detection of ZIKV-specific RNA. Serological diagnosis is complicated by cross-reactivity among members of the Flavivirus genus. The adaptation of ZIKV to an urban cycle involving humans and domestic mosquito vectors in tropical areas where dengue is endemic suggests that the incidence of ZIKV infections may be underestimated. There is a high potential for ZIKV emergence in urban centers in the tropics that are infested with competent mosquito vectors such as Aedes aegypti and Aedes albopictus.

  7. Zika Virus and Pregnancy

    MedlinePlus

    ... Management Education & Events Advocacy For Patients About ACOG Zika Virus and Pregnancy Home For Patients Zika Virus ... Patient Education Pamphlets - Spanish Share: PEV002, September 2016 Zika Virus and Pregnancy There are risks to your ...

  8. SAMPLING VIRUSES FROM SOIL

    EPA Science Inventory

    This chapter describes in detail methods for detecting viruses of bacteria and humans in soil. Methods also are presented for the assay of these viruses. Reference sources are provided for information on viruses of plants.

  9. Hanta virus (image)

    MedlinePlus

    Hanta virus is a distant cousin of Ebola virus, but is found worldwide. The virus is spread by human contact with rodent waste. Dangerous respiratory illness develops. Effective treatment is not yet ...

  10. Ebola Virus Disease

    MedlinePlus

    ... 2014 Fact sheets Features Commentaries 2014 Multimedia Contacts Ebola virus disease Fact sheet Updated January 2016 Key ... for survivors of Ebola virus disease Symptoms of Ebola virus disease The incubation period, that is, the ...

  11. Computer Viruses: An Overview.

    ERIC Educational Resources Information Center

    Marmion, Dan

    1990-01-01

    Discusses the early history and current proliferation of computer viruses that occur on Macintosh and DOS personal computers, mentions virus detection programs, and offers suggestions for how libraries can protect themselves and their users from damage by computer viruses. (LRW)

  12. Virus Movement Maintains Local Virus Population Diversity

    SciTech Connect

    J. Snyder; B. Wiedenheft; M. Lavin; F. Roberto; J. Spuhler; A. Ortmann; T. Douglas; M. Young

    2007-11-01

    Viruses are the largest reservoir of genetic material on the planet, yet little is known about the population dynamics of any virus within its natural environment. Over a 2-year period, we monitored the diversity of two archaeal viruses found in hot springs within Yellowstone National Park (YNP). Both temporal phylogeny and neutral biodiversity models reveal that virus diversity in these local environments is not being maintained by mutation but rather by high rates of immigration from a globally distributed metacommunity. These results indicate that geographically isolated hot springs are readily able to exchange viruses. The importance of virus movement is supported by the detection of virus particles in air samples collected over YNP hot springs and by their detection in metacommunity sequencing projects conducted in the Sargasso Sea. Rapid rates of virus movement are not expected to be unique to these archaeal viruses but rather a common feature among virus metacommunities. The finding that virus immigration rather than mutation can dominate community structure has significant implications for understanding virus circulation and the role that viruses play in ecology and evolution by providing a reservoir of mobile genetic material.

  13. Nuclear ventriculography

    MedlinePlus

    ... ventriculography (RNV); Multiple gate acquisition scan (MUGA); Nuclear cardiology; Cardiomyopathy - nuclear ventriculography ... 56. Udelson JE, Dilsizian V, Bonow RO. Nuclear cardiology. In: Bonow RO, Mann DL, Zipes DP, Libby ...

  14. Nuclear Medicine.

    ERIC Educational Resources Information Center

    Badawi, Ramsey D.

    2001-01-01

    Describes the use of nuclear medicine techniques in diagnosis and therapy. Describes instrumentation in diagnostic nuclear medicine and predicts future trends in nuclear medicine imaging technology. (Author/MM)

  15. [BK virus nephropathy after kidney transplantation].

    PubMed

    Bröcker, V; Schwarz, A; Becker, J U

    2011-09-01

    JC and BK viruses are strains of the polyomavirus group with pathogenic potential in humans. BK is the most frequent pathogenic agent of polyomavirus nephropathy (BKVN) in kidney transplant patients, which is only exceptionally caused by JC virus. Asymptomatic BK virus infection is often acquired in childhood and the virus persists in urothelium and kidneys of healthy individuals, where it can be reactivated under immunosuppression. Up to 10% of transplanted kidneys are affected by BKVN, while the risk of transplant failure due to BKVN exceeds 50% in some publications. In kidney biopsies BKVN leads to tubulointerstitial nephritis, which may be difficult to distinguish from acute cellular rejection without additional use of immunohistochemistry for a polyomavirus antigen. Typical hallmarks of BKVN include cytopathic effects caused by the virus with cell lysis, denudation of tubular basement membranes and nuclear inclusion bodies. An early diagnosis is essential for transplant survival, making screening of blood and urine for BK virus after kidney transplantation part of the standard care of renal transplant patients today. In the case of significant viremia or biopsy-proven BKVN immunosuppression is reduced to allow clearing of the virus.

  16. Viruses and Virus Diseases of Rubus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rubus species are propagated vegetatively and are subject to infection by viruses during development, propagation and fruit production stages. Reports of initial detection and symptoms of more than 30 viruses, virus-like diseases and phytoplasmas affecting Rubus spp. have been reviewed more than 20 ...

  17. Crystallization of viruses and virus proteins

    NASA Astrophysics Data System (ADS)

    Sehnke, Paul C.; Harrington, Melissa; Hosur, M. V.; Li, Yunge; Usha, R.; Craig Tucker, R.; Bomu, Wu; Stauffacher, Cynthia V.; Johnson, John E.

    1988-07-01

    Methods for crystallizing six isometric plant and insect viruses are presented. Procedures developed for modifying, purifying and crystallizing coat protein subunits isolated from a virus forming asymmetric, spheroidal particles, stabilized almost exclusively by protein-RNA interactions, are also discussed. The tertiary and quaternary structures of small RNA viruses are compared.

  18. Nuclear weapons and nuclear war

    SciTech Connect

    Cassel, C.; McCally, M.; Abraham, H.

    1984-01-01

    This book examines the potential radiation hazards and environmental impacts of nuclear weapons. Topics considered include medical responsibility and thermonuclear war, the threat of nuclear war, nuclear weaponry, biological effects, radiation injury, decontamination, long-term effects, ecological effects, psychological aspects, the economic implications of nuclear weapons and war, ethics, civil defense, arms control, nuclear winter, and long-term biological consequences of nuclear war.

  19. Nuclear Theory - Nuclear Power

    NASA Astrophysics Data System (ADS)

    Svenne, J. P.; Canton, L.; Kozier, K. S.

    2008-01-01

    The results from modern nuclear theory are accurate and reliable enough to be used for practical applications, in particular for scattering that involves few-nucleon systems of importance to nuclear power. Using well-established nucleon-nucleon (NN) interactions that fit well the NN scattering data, and the AGS form of the three-body theory, we have performed precise calculations of low-energy neutron-deuteron (n+d) scattering. We show that three-nucleon force effects that have impact on the low-energy vector analyzing powers have no practical effects on the angular distribution of the n+d cross-section. There appear to be problems for this scattering in the evaluated nuclear data file (ENDF) libraries, at the incident neutron energies less than 3.2 MeV. Supporting experimental data in this energy region are rather old (>25 years), sparse and often inconsistent. Our three-body results at low energies, 50 keV to 10.0 MeV, are compared to the ENDF/B-VII.0 and JENDL (Japanese Evaluated Nuclear Data Library) -3.3 evaluated angular distributions. The impact of these results on the calculated reactivity for various critical systems involving heavy water is shown.

  20. Role Bending: Complex Relationships Between Viruses, Hosts, and Vectors Related to Citrus Leprosis, an Emerging Disease.

    PubMed

    Roy, Avijit; Hartung, John S; Schneider, William L; Shao, Jonathan; Leon, Guillermo; Melzer, Michael J; Beard, Jennifer J; Otero-Colina, Gabriel; Bauchan, Gary R; Ochoa, Ronald; Brlansky, Ronald H

    2015-07-01

    Citrus leprosis complex is an emerging disease in the Americas, associated with two unrelated taxa of viruses distributed in South, Central, and North America. The cytoplasmic viruses are Citrus leprosis virus C (CiLV-C), Citrus leprosis virus C2 (CiLV-C2), and Hibiscus green spot virus 2, and the nuclear viruses are Citrus leprosis virus N (CiLV-N) and Citrus necrotic spot virus. These viruses cause local lesion infections in all known hosts, with no natural systemic host identified to date. All leprosis viruses were believed to be transmitted by one species of mite, Brevipalpus phoenicis. However, mites collected from CiLV-C and CiLV-N infected citrus groves in Mexico were identified as B. yothersi and B. californicus sensu lato, respectively, and only B. yothersi was detected from CiLV-C2 and CiLV-N mixed infections in the Orinoco regions of Colombia. Phylogenetic analysis of the helicase, RNA-dependent RNA polymerase 2 domains and p24 gene amino acid sequences of cytoplasmic leprosis viruses showed a close relationship with recently deposited mosquito-borne negevirus sequences. Here, we present evidence that both cytoplasmic and nuclear viruses seem to replicate in viruliferous Brevipalpus species. The possible replication in the mite vector and the close relationship with mosquito borne negeviruses are consistent with the concept that members of the genus Cilevirus and Higrevirus originated in mites and citrus may play the role of mite virus vector. PMID:25775106

  1. Role Bending: Complex Relationships Between Viruses, Hosts, and Vectors Related to Citrus Leprosis, an Emerging Disease.

    PubMed

    Roy, Avijit; Hartung, John S; Schneider, William L; Shao, Jonathan; Leon, Guillermo; Melzer, Michael J; Beard, Jennifer J; Otero-Colina, Gabriel; Bauchan, Gary R; Ochoa, Ronald; Brlansky, Ronald H

    2015-07-01

    Citrus leprosis complex is an emerging disease in the Americas, associated with two unrelated taxa of viruses distributed in South, Central, and North America. The cytoplasmic viruses are Citrus leprosis virus C (CiLV-C), Citrus leprosis virus C2 (CiLV-C2), and Hibiscus green spot virus 2, and the nuclear viruses are Citrus leprosis virus N (CiLV-N) and Citrus necrotic spot virus. These viruses cause local lesion infections in all known hosts, with no natural systemic host identified to date. All leprosis viruses were believed to be transmitted by one species of mite, Brevipalpus phoenicis. However, mites collected from CiLV-C and CiLV-N infected citrus groves in Mexico were identified as B. yothersi and B. californicus sensu lato, respectively, and only B. yothersi was detected from CiLV-C2 and CiLV-N mixed infections in the Orinoco regions of Colombia. Phylogenetic analysis of the helicase, RNA-dependent RNA polymerase 2 domains and p24 gene amino acid sequences of cytoplasmic leprosis viruses showed a close relationship with recently deposited mosquito-borne negevirus sequences. Here, we present evidence that both cytoplasmic and nuclear viruses seem to replicate in viruliferous Brevipalpus species. The possible replication in the mite vector and the close relationship with mosquito borne negeviruses are consistent with the concept that members of the genus Cilevirus and Higrevirus originated in mites and citrus may play the role of mite virus vector.

  2. The Tobacco Mosaic Virus.

    ERIC Educational Resources Information Center

    Sulzinski, Michael A.

    1992-01-01

    Explains how the tobacco mosaic virus can be used to study virology. Presents facts about the virus, procedures to handle the virus in the laboratory, and four laboratory exercises involving the viruses' survival under inactivating conditions, dilution end point, filterability, and microscopy. (MDH)

  3. Nuclear choices

    SciTech Connect

    Wolfson, R.

    1991-01-01

    This book contains part of the series New Liberal Arts, which is intended to make science and technology more accessible to students of the liberal arts. Volume in hand provides a comprehensive, multifaceted examination of nuclear energy, in nontechnical terms. Wolfson explains the basics of nuclear energy and radiation, nuclear power..., and nuclear weapons..., and he invites readers to make their own judgments on controversial nuclear issues. Illustrated with photos and diagrams. Each chapter contains suggestions for additional reading and a glossary. For policy, science, and general collections in all libraries. (ES) Topics contained include Atoms and nuclei. Effects and uses of radiation. Energy and People. Reactor safety. Nuclear strategy. Defense in the nuclear age. Nuclear power, nuclear weapons, and nuclear futures.

  4. Viruses of potato.

    PubMed

    Loebenstein, Gad; Gaba, Victor

    2012-01-01

    Potatoes are an important crop in Mediterranean countries both for local consumption and for export to other countries, mainly during the winter. Many Mediterranean countries import certified seed potato in addition to their own seed production. The local seeds are mainly used for planting in the autumn and winter, while the imported seed are used for early and late spring plantings. Potato virus Y is the most important virus in Mediterranean countries, present mainly in the autumn plantings. The second important virus is Potato leafroll virus, though in recent years its importance seems to be decreasing. Potato virus X, Potato virus A, Potato virus S, Potato virus M, and the viroid, Potato spindle tuber viroid, were also recorded in several Mediterranean countries. For each virus the main strains, transmission, characterization of the virus particle, its genome organization, detection, and control methods including transgenic approaches will be discussed. PMID:22682169

  5. Understanding Ebola Virus Transmission

    PubMed Central

    Judson, Seth; Prescott, Joseph; Munster, Vincent

    2015-01-01

    An unprecedented number of Ebola virus infections among healthcare workers and patients have raised questions about our understanding of Ebola virus transmission. Here, we explore different routes of Ebola virus transmission between people, summarizing the known epidemiological and experimental data. From this data, we expose important gaps in Ebola virus research pertinent to outbreak situations. We further propose experiments and methods of data collection that will enable scientists to fill these voids in our knowledge about the transmission of Ebola virus. PMID:25654239

  6. Multiple overlapping homologies between two rheumatoid antigens and immunosuppressive viruses.

    PubMed Central

    Douvas, A; Sobelman, S

    1991-01-01

    Amino acid (aa) sequence homologies between viruses and autoimmune nuclear antigens are suggestive of viral involvement in disorders such as systemic lupus erythematosus (SLE) and scleroderma. We analyzed the frequency of exact homologies of greater than or equal to 5 aa between 61 viral proteins (19,827 aa), 8 nuclear antigens (3813 aa), and 41 control proteins (11,743 aa). Both pentamer and hexamer homologies between control proteins and viruses are unexpectedly abundant, with hexamer matches occurring in 1 of 3 control proteins (or once every 769 aa). However, 2 nuclear antigens, the SLE-associated 70-kDa antigen and the scleroderma-associated CENP-B protein, are highly unusual in containing multiple homologies to a group of synergizing immunosuppressive viruses. Two viruses, herpes simplex virus 1 (HSV-1) and human immunodeficiency virus 1 (HIV-1), contain sequences exactly duplicated at 15 sites in the 70-kDa antigen and at 10 sites in CENP-B protein. The immediate-early (IE) protein of HSV-1, which activates HIV-1 regulatory functions, contains three homologies to the 70-kDa antigen (two hexamers and a pentamer) and two to CENP-B (a hexamer and pentamer). There are four homologies (including a hexamer) common to the 70-kDa antigen and Epstein-Barr virus, and three homologies (including two hexamers) common to CENP-B and cytomegalovirus. The majority of homologies in both nuclear antigens are clustered in highly charged C-terminal domains containing epitopes for human autoantibodies. Furthermore, most homologies have a contiguous or overlapping distribution, thereby creating a high density of potential epitopes. In addition to the exact homologies tabulated, motifs of matching sequences are repeated frequently in these domains. Our analysis suggests that coexpression of heterologous viruses having common immunosuppressive functions may generate autoantibodies cross-reacting with certain nuclear proteins. PMID:1712488

  7. Virus-Vectored Influenza Virus Vaccines

    PubMed Central

    Tripp, Ralph A.; Tompkins, S. Mark

    2014-01-01

    Despite the availability of an inactivated vaccine that has been licensed for >50 years, the influenza virus continues to cause morbidity and mortality worldwide. Constant evolution of circulating influenza virus strains and the emergence of new strains diminishes the effectiveness of annual vaccines that rely on a match with circulating influenza strains. Thus, there is a continued need for new, efficacious vaccines conferring cross-clade protection to avoid the need for biannual reformulation of seasonal influenza vaccines. Recombinant virus-vectored vaccines are an appealing alternative to classical inactivated vaccines because virus vectors enable native expression of influenza antigens, even from virulent influenza viruses, while expressed in the context of the vector that can improve immunogenicity. In addition, a vectored vaccine often enables delivery of the vaccine to sites of inductive immunity such as the respiratory tract enabling protection from influenza virus infection. Moreover, the ability to readily manipulate virus vectors to produce novel influenza vaccines may provide the quickest path toward a universal vaccine protecting against all influenza viruses. This review will discuss experimental virus-vectored vaccines for use in humans, comparing them to licensed vaccines and the hurdles faced for licensure of these next-generation influenza virus vaccines. PMID:25105278

  8. Saikosaponin A inhibits influenza A virus replication and lung immunopathology

    PubMed Central

    Zhao, Yaqin; Ling, Fangfang; Xiao, Kun; Li, Qian; Li, Bin; Lu, Chunni; Qi, Wenbao; Zeng, Zhenling; Liao, Ming; Liu, Yahong; Chen, Weisan

    2015-01-01

    Fatal influenza outcomes result from a combination of rapid virus replication and collateral lung tissue damage caused by exaggerated pro-inflammatory host immune cell responses. There are few therapeutic agents that target both biological processes for the attenuation of influenza-induced lung pathology. We show that Saikosaponin A, a bioactive triterpene saponin with previouslyestablished anti-inflammatory effects, demonstrates both in vitro and in vivo anti-viral activity against influenza A virus infections. Saikosaponin A attenuated the replication of three different influenza A virus strains, including a highly pathogenic H5N1 strain, in human alveolar epithelial A549 cells. This anti-viral activity occurred through both downregulation of NF-κB signaling and caspase 3-dependent virus ribonucleoprotein nuclear export as demonstrated by NF-κB subunit p65 and influenza virus nucleoprotein nuclear translocation studies in influenza virus infected A549 cells. Critically, Saikosaponin A also attenuated viral replication, aberrant pro-inflammatory cytokine production and lung histopathology in the widely established H1N1 PR8 model of influenza A virus lethality in C57BL/6 mice. Flow cytometry studies of mouse bronchoalveolar lavage cells revealed that SSa exerted immunomodulatory effects through a selective attenuation of lung neutrophil and monocyte recruitment during the early peak of the innate immune response to PR8 infection. Altogether, our results indicate that Saikosaponin A possesses novel therapeutic potential for the treatment of pathological influenza virus infections. PMID:26637810

  9. Saikosaponin A inhibits influenza A virus replication and lung immunopathology.

    PubMed

    Chen, Jianxin; Duan, Mubing; Zhao, Yaqin; Ling, Fangfang; Xiao, Kun; Li, Qian; Li, Bin; Lu, Chunni; Qi, Wenbao; Zeng, Zhenling; Liao, Ming; Liu, Yahong; Chen, Weisan

    2015-12-15

    Fatal influenza outcomes result from a combination of rapid virus replication and collateral lung tissue damage caused by exaggerated pro-inflammatory host immune cell responses. There are few therapeutic agents that target both biological processes for the attenuation of influenza-induced lung pathology. We show that Saikosaponin A, a bioactive triterpene saponin with previouslyestablished anti-inflammatory effects, demonstrates both in vitro and in vivo anti-viral activity against influenza A virus infections. Saikosaponin A attenuated the replication of three different influenza A virus strains, including a highly pathogenic H5N1 strain, in human alveolar epithelial A549 cells. This anti-viral activity occurred through both downregulation of NF-κB signaling and caspase 3-dependent virus ribonucleoprotein nuclear export as demonstrated by NF-κB subunit p65 and influenza virus nucleoprotein nuclear translocation studies in influenza virus infected A549 cells. Critically, Saikosaponin A also attenuated viral replication, aberrant pro-inflammatory cytokine production and lung histopathology in the widely established H1N1 PR8 model of influenza A virus lethality in C57BL/6 mice. Flow cytometry studies of mouse bronchoalveolar lavage cells revealed that SSa exerted immunomodulatory effects through a selective attenuation of lung neutrophil and monocyte recruitment during the early peak of the innate immune response to PR8 infection. Altogether, our results indicate that Saikosaponin A possesses novel therapeutic potential for the treatment of pathological influenza virus infections.

  10. Nuclear Exodus: Herpesviruses Lead the Way

    PubMed Central

    Bigalke, Janna M.; Heldwein, Ekaterina E.

    2016-01-01

    Most DNA viruses replicate in the nucleus and exit it either by passing through the nuclear pores or by rupturing the nuclear envelope. Unusually, herpesviruses have evolved a complex mechanism of nuclear escape whereby nascent capsids bud at the inner nuclear membrane to form perinuclear virions that subsequently fuse with the outer nuclear membrane, releasing capsids into the cytosol. Although this general scheme is accepted in the field, the players and their roles are still debated. Recent studies illuminated critical mechanistic features of this enigmatic process and uncovered surprising parallels with a novel cellular nuclear export process. This review summarizes our current understanding of nuclear egress in herpesviruses, examines the experimental evidence and models, and outlines outstanding questions with the goal of stimulating new research in this area. PMID:27482898

  11. Viruses infecting reptiles.

    PubMed

    Marschang, Rachel E

    2011-11-01

    A large number of viruses have been described in many different reptiles. These viruses include arboviruses that primarily infect mammals or birds as well as viruses that are specific for reptiles. Interest in arboviruses infecting reptiles has mainly focused on the role reptiles may play in the epidemiology of these viruses, especially over winter. Interest in reptile specific viruses has concentrated on both their importance for reptile medicine as well as virus taxonomy and evolution. The impact of many viral infections on reptile health is not known. Koch's postulates have only been fulfilled for a limited number of reptilian viruses. As diagnostic testing becomes more sensitive, multiple infections with various viruses and other infectious agents are also being detected. In most cases the interactions between these different agents are not known. This review provides an update on viruses described in reptiles, the animal species in which they have been detected, and what is known about their taxonomic positions.

  12. Viruses Infecting Reptiles

    PubMed Central

    Marschang, Rachel E.

    2011-01-01

    A large number of viruses have been described in many different reptiles. These viruses include arboviruses that primarily infect mammals or birds as well as viruses that are specific for reptiles. Interest in arboviruses infecting reptiles has mainly focused on the role reptiles may play in the epidemiology of these viruses, especially over winter. Interest in reptile specific viruses has concentrated on both their importance for reptile medicine as well as virus taxonomy and evolution. The impact of many viral infections on reptile health is not known. Koch’s postulates have only been fulfilled for a limited number of reptilian viruses. As diagnostic testing becomes more sensitive, multiple infections with various viruses and other infectious agents are also being detected. In most cases the interactions between these different agents are not known. This review provides an update on viruses described in reptiles, the animal species in which they have been detected, and what is known about their taxonomic positions. PMID:22163336

  13. Nuclear Winter.

    ERIC Educational Resources Information Center

    Ehrlich, Anne

    1984-01-01

    "Nuclear Winter" was recently coined to describe the climatic and biological effects of a nuclear war. These effects are discussed based on models, simulations, scenarios, and projections. Effects on human populations are also considered. (JN)

  14. Nuclear Chemistry.

    ERIC Educational Resources Information Center

    Chemical and Engineering News, 1979

    1979-01-01

    Provides a brief review of the latest developments in nuclear chemistry. Nuclear research today is directed toward increased activity in radiopharmaceuticals and formation of new isotopes by high-energy, heavy-ion collisions. (Author/BB)

  15. Nuclear Scans

    MedlinePlus

    Nuclear scans use radioactive substances to see structures and functions inside your body. They use a special ... images. Most scans take 20 to 45 minutes. Nuclear scans can help doctors diagnose many conditions, including ...

  16. Nuclear Fuels.

    ERIC Educational Resources Information Center

    Nash, J. Thomas

    1983-01-01

    Trends in and factors related to the nuclear industry and nuclear fuel production are discussed. Topics addressed include nuclear reactors, survival of the U.S. uranium industry, production costs, budget cuts by the Department of Energy and U.S. Geological survey for resource studies, mining, and research/development activities. (JN)

  17. Morphogenesis of Bittner Virus

    PubMed Central

    Gay, Frederick W.; Clarke, John K.; Dermott, Evelyn

    1970-01-01

    The morphogenesis of Bittner virus (mouse mammary tumor virus) was studied in sectioned mammary tumor cells. Internal components of the virus (type A particles) were seen being assembled in virus factories close to the nucleus and were also seen forming at the plasma membrane. The particles in virus factories became enveloped by budding through the membrane of cytoplasmic vacuoles which were derived from dilated endoplasmic reticulum. Complete virus particles were liberated from these vacuoles by cell lysis. Particles budding at the plasma membrane were released into intercellular spaces. Maturation of enveloped virus occurred after release, but mature internal components were rarely seen in the cytoplasm before envelopment. Direct cell-to-cell transfer of virus by pinocytosis of budding particles by an adjacent cell was observed, and unusual forms of budding virus which participated in this process are illustrated and described. There was evidence that some virus particles contained cytoplasmic constituents, including ribosomes. Certain features of the structure of internal components are discussed in relation to a recently proposed model for the internal component of the mouse leukemia virus. Intracisternal virus-like particles were occasionally seen in tumor cells, but there was no evidence that these structures were developmentally related to Bittner virus. Images PMID:4193837

  18. Morphogenesis of Bittner virus.

    PubMed

    Gay, F W; Clarke, J K; Dermott, E

    1970-06-01

    The morphogenesis of Bittner virus (mouse mammary tumor virus) was studied in sectioned mammary tumor cells. Internal components of the virus (type A particles) were seen being assembled in virus factories close to the nucleus and were also seen forming at the plasma membrane. The particles in virus factories became enveloped by budding through the membrane of cytoplasmic vacuoles which were derived from dilated endoplasmic reticulum. Complete virus particles were liberated from these vacuoles by cell lysis. Particles budding at the plasma membrane were released into intercellular spaces. Maturation of enveloped virus occurred after release, but mature internal components were rarely seen in the cytoplasm before envelopment. Direct cell-to-cell transfer of virus by pinocytosis of budding particles by an adjacent cell was observed, and unusual forms of budding virus which participated in this process are illustrated and described. There was evidence that some virus particles contained cytoplasmic constituents, including ribosomes. Certain features of the structure of internal components are discussed in relation to a recently proposed model for the internal component of the mouse leukemia virus. Intracisternal virus-like particles were occasionally seen in tumor cells, but there was no evidence that these structures were developmentally related to Bittner virus. PMID:4193837

  19. Nuclear weapons, nuclear effects, nuclear war

    SciTech Connect

    Bing, G.F.

    1991-08-20

    This paper provides a brief and mostly non-technical description of the militarily important features of nuclear weapons, of the physical phenomena associated with individual explosions, and of the expected or possible results of the use of many weapons in a nuclear war. Most emphasis is on the effects of so-called ``strategic exchanges.``

  20. Respiratory Syncytial Virus Infections

    MedlinePlus

    Respiratory syncytial virus (RSV) causes mild, cold-like symptoms in adults and older healthy children. It can cause serious problems in ... tests can tell if your child has the virus. There is no specific treatment. You should give ...

  1. Viruses and human cancer

    SciTech Connect

    Gallo, R.C.; Haseltine, W.; Klein, G.; Zur Hausen, H.

    1987-01-01

    This book contains papers on the following topics: Immunology and Epidemiology, Biology and Pathogenesis, Models of Pathogenesis and Treatment, Simian and Bovine Retroviruses, Human Papilloma Viruses, EBV and Herpesvirus, and Hepatitis B Virus.

  2. Densonucleosis virus structural proteins.

    PubMed

    Kelly, D C; Moore, N F; Spilling, C R; Barwise, A H; Walker, I O

    1980-10-01

    The protein coats of two densonucleosis viruses (types 1 and 2) were examined by a variety of biophysical, biochemical, and serological techniques. The viruses were 24 nm in diameter, contained at least four polypeptides, were remarkably stable to extremes of pH and denaturing agents, and were serologically closely related. The two viruses could, however, be distinguished serologically and by differences in migration of their structural polypeptides. For each virus the "top component" (i.e., the protein coat minus DNA, found occurring naturally in infections) appeared to have a composition identical to that of the coat of the virus and was a more stable structure. Electrometric titration curves of the virus particles and top components demonstrated that the DNA phosphate in densonucleosis virus particles was neutralized by cations other than basic amino acid side chains of the protein coat. Circular dichroism studies showed that there was a conformational difference between the protein coats of top components and virus particles.

  3. Viruses and Breast Cancer

    PubMed Central

    Lawson, James S.; Heng, Benjamin

    2010-01-01

    Viruses are the accepted cause of many important cancers including cancers of the cervix and anogenital area, the liver, some lymphomas, head and neck cancers and indirectly human immunodeficiency virus associated cancers. For over 50 years, there have been serious attempts to identify viruses which may have a role in breast cancer. Despite these efforts, the establishment of conclusive evidence for such a role has been elusive. However, the development of extremely sophisticated new experimental techniques has allowed the recent development of evidence that human papilloma virus, Epstein-Barr virus, mouse mammary tumor virus and bovine leukemia virus may each have a role in the causation of human breast cancers. This is potentially good news as effective vaccines are already available to prevent infections from carcinogenic strains of human papilloma virus, which causes cancer of the uterine cervix. PMID:24281093

  4. Zika Virus Fact Sheet

    MedlinePlus

    ... 2014 Fact sheets Features Commentaries 2014 Multimedia Contacts Zika virus Fact sheet Updated 6 September 2016 Key facts ... and last for 2-7 days. Complications of Zika virus disease After a comprehensive review of evidence, there ...

  5. Human Parainfluenza Viruses

    MedlinePlus

    ... HPIVs Are Not the Same as Influenza (Flu) Viruses People usually get HPIV infections more often in ... hands, and touching objects or surfaces with the viruses on them then touching your mouth, nose, or ...

  6. Herpes Simplex Virus (HSV)

    MedlinePlus

    ... rashes clinical tools newsletter | contact Share | Herpes Simplex Virus (HSV) A parent's guide to condition and treatment ... skin or mouth sores with the herpes simplex virus (HSV) is called primary herpes. This may be ...

  7. Tumorigenic DNA viruses

    SciTech Connect

    Klein, G.

    1989-01-01

    The eighth volume of Advances in Viral Oncology focuses on the three major DNA virus groups with a postulated or proven tumorigenic potential: papillomaviruses, animal hepatitis viruses, and the Epstein-Bar virus. In the opening chapters, the contributors analyze the evidence that papillomaviruses and animal hepatitis viruses are involved in tumorigenesis and describe the mechanisms that trigger virus-host cell interactions. A detailed section on the Epstein-Barr virus (EBV) - comprising more than half the book - examines the transcription and mRNA processing patterns of the virus genome; the mechanisms by which EBV infects lymphoid and epithelial cells; the immunological aspects of the virus; the actions of EBV in hosts with Acquired Immune Deficiency Syndrome; and the involvement of EBV in the etiology of Burkitt's lymphoma.

  8. Advances in virus research

    SciTech Connect

    Maramorosch, K. ); Murphy, F.A. ); Shatkin, A.J. )

    1988-01-01

    This book contains eight chapters. Some of the titles are: Initiation of viral DNA replication; Vaccinia: virus, vector, vaccine; The pre-S region of hepadnavirus envelope proteins; and Archaebacterial viruses.

  9. West Nile virus

    MedlinePlus

    ... believe West Nile virus is spread when a mosquito bites an infected bird and then bites a person. ... avoid getting West Nile virus infection after a mosquito bite. People in good health generally do not develop ...

  10. Virus Assembly and Maturation

    NASA Astrophysics Data System (ADS)

    Johnson, John E.

    2004-03-01

    We use two techniques to look at three-dimensional virus structure: electron cryomicroscopy (cryoEM) and X-ray crystallography. Figure 1 is a gallery of virus particles whose structures Timothy Baker, one of my former colleagues at Purdue University, used cryoEM to determine. It illustrates the variety of sizes of icosahedral virus particles. The largest virus particle on this slide is the Herpes simplex virus, around 1200Å in diameter; the smallest we examined was around 250Å in diameter. Viruses bear their genomic information either as positive-sense DNA and RNA, double-strand DNA, double-strand RNA, or negative-strand RNA. Viruses utilize the various structure and function "tactics" seen throughout cell biology to replicate at high levels. Many of the biological principles that we consider general were in fact discovered in the context of viruses ...

  11. Avian influenza virus and Newcastle disease virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Avian influenza virus (AIV) and Newcastle disease virus (NDV) severely impact poultry egg production. Decreased egg yield and hatchability, as well as misshapen eggs, are often observed during infection with AIV and NDV, even with low-virulence strains or in vaccinated flocks. Data suggest that in...

  12. Computer Virus Protection

    ERIC Educational Resources Information Center

    Rajala, Judith B.

    2004-01-01

    A computer virus is a program--a piece of executable code--that has the unique ability to replicate. Like biological viruses, computer viruses can spread quickly and are often difficult to eradicate. They can attach themselves to just about any type of file, and are spread by replicating and being sent from one individual to another. Simply having…

  13. Avian influenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Avian influenza virus (AIV) is type A influenza, which is adapted to an avian host. Although avian influenza has been isolated from numerous avian species, the primary natural hosts for the virus are dabbling ducks, shorebirds, and gulls. The virus can be found world-wide in these species and in o...

  14. Respiratory Syncytial Virus

    MedlinePlus

    ... Palsy: Shannon's Story" 5 Things to Know About Zika & Pregnancy Respiratory Syncytial Virus KidsHealth > For Parents > Respiratory Syncytial Virus Print A ... often get it when older kids carry the virus home from school and pass it to ... often happen in epidemics that last from late fall through early spring. ...

  15. Zika virus - an overview.

    PubMed

    Zanluca, Camila; Dos Santos, Claudia Nunes Duarte

    2016-05-01

    Zika virus (ZIKV) is currently one of the most important emerging viruses in the world. Recently, it has caused outbreaks and epidemics, and has been associated with severe clinical manifestations and congenital malformations. However to date, little is known about the pathogenicity of the virus and the consequences of ZIKV infection. In this paper, we provide an overview of the current knowledge on ZIKV.

  16. The taxonomy of viruses should include viruses.

    PubMed

    Calisher, Charles H

    2016-05-01

    Having lost sight of its goal, the International Committee on Taxonomy of Viruses has redoubled its efforts. That goal is to arrive at a consensus regarding virus classification, i.e., proper placement of viruses in a hierarchical taxonomic scheme; not an easy task given the wide variety of recognized viruses. Rather than suggesting a continuation of the bureaucratic machinations of the past, this opinion piece is a call for insertion of common sense in sorting out the avalanche of information already, and soon-to-be, accrued data. In this way information about viruses ideally would be taxonomically correct as well as useful to working virologists and journal editors, rather than being lost, minimized, or ignored.

  17. The influenza virus nucleoprotein synthesized from cloned DNA in a simian virus 40 vector is detected in the nucleus.

    PubMed Central

    Lin, B C; Lai, C J

    1983-01-01

    We obtained DNA sequences coding for the nucleoprotein (NP) of an influenza A virus by reverse transcription of virion RNA with synthetic oligonucleotide primers. Terminal sequence analysis showed that the cloned gene contained a full-length copy of the virion RNA segment. The NP-specific DNA was inserted into the late region of a simian virus 40 vector, and the DNA recombinant was propagated in the presence of an early simian virus 40 temperature-sensitive mutant helper. Infection of African green monkey kidney cells with the recombinant produced a polypeptide immunoprecipitable with NP-specific antisera. The polypeptide product had a molecular weight of 56,000, identical to that of the nucleoprotein of influenza virus as estimated on polyacrylamide gels. The putative NP was detected in the nucleus of infected primate cells by an immunofluorescence assay. This nuclear localization of NP from recombinant DNA was similar to that seen during influenza virus infection. Images PMID:6296449

  18. Viruses of asparagus.

    PubMed

    Tomassoli, Laura; Tiberini, Antonio; Vetten, Heinrich-Josef

    2012-01-01

    The current knowledge on viruses infecting asparagus (Asparagus officinalis) is reviewed. Over half a century, nine virus species belonging to the genera Ilarvirus, Cucumovirus, Nepovirus, Tobamovirus, Potexvirus, and Potyvirus have been found in this crop. The potyvirus Asparagus virus 1 (AV1) and the ilarvirus Asparagus virus 2 (AV2) are widespread and negatively affect the economic life of asparagus crops reducing yield and increasing the susceptibility to biotic and abiotic stress. The main properties and epidemiology of AV1 and AV2 as well as diagnostic techniques for their detection and identification are described. Minor viruses and control are briefly outlined.

  19. Viruses of asparagus.

    PubMed

    Tomassoli, Laura; Tiberini, Antonio; Vetten, Heinrich-Josef

    2012-01-01

    The current knowledge on viruses infecting asparagus (Asparagus officinalis) is reviewed. Over half a century, nine virus species belonging to the genera Ilarvirus, Cucumovirus, Nepovirus, Tobamovirus, Potexvirus, and Potyvirus have been found in this crop. The potyvirus Asparagus virus 1 (AV1) and the ilarvirus Asparagus virus 2 (AV2) are widespread and negatively affect the economic life of asparagus crops reducing yield and increasing the susceptibility to biotic and abiotic stress. The main properties and epidemiology of AV1 and AV2 as well as diagnostic techniques for their detection and identification are described. Minor viruses and control are briefly outlined. PMID:22682173

  20. Double-Stranded RNA Is Detected by Immunofluorescence Analysis in RNA and DNA Virus Infections, Including Those by Negative-Stranded RNA Viruses

    PubMed Central

    Son, Kyung-No; Liang, Zhiguo

    2015-01-01

    of IFN-stimulated genes. The present study demonstrates that infections, including those by ssDNA viruses and positive- and negative-strand RNA viruses, produce dsRNAs detectable by standard immunofluorescence staining. While dsRNA staining was primarily observed in the cytoplasm, nuclear staining was also present in some RNA and DNA virus infections. The nucleus is unlikely to have pathogen-associated molecular pattern (PAMP) receptors for dsRNA because of the presence of host dsRNA molecules. Thus, it is likely that most animal virus infections produce dsRNA species detectable by immunofluorescence staining, which may prove useful in viral discovery as well. PMID:26136565

  1. Nuclear mortality

    SciTech Connect

    Krauthammer, C.

    1983-10-01

    The author notes that the anti-nuclear movement is shifting its focus from bodily harm to concern for the impact on our souls from building and threatening the use of nuclear weapons. Two aspects of nuclear deterrence receiving the most public attention are the freeze effort to halt weapons modernization and the no-first-use effort to take down the nuclear umbrella. Opponents attack both the countervalue and the counterforce approach, but the arguments of the Catholic bishops, Jonathan Schell, and others stop short of unilateral disarmament, which would be the greatest threat to our survival. Mr. Krauthammer observes that nuclear deterrence has worked, however, and will continue to be useful only if potential adversaries believe we have the will to use nuclear weapons. 2 references. (DCK)

  2. Serodiagnosis for Tumor Viruses

    PubMed Central

    Morrison, Brian J.; Labo, Nazzarena; Miley, Wendell J.; Whitby, Denise

    2015-01-01

    The known human tumor viruses include the DNA viruses Epstein-Barr virus, Kaposi sarcoma herpesvirus, Merkel cell polyomavirus, human papillomavirus, and hepatitis B virus. RNA tumor viruses include Human T-cell lymphotrophic virus type-1 and hepatitis C virus. The serological identification of antigens/antibodies in plasma serum is a rapidly progressing field with utility for both scientists and clinicians. Serology is useful for conducting seroepidemiology studies and to inform on the pathogenesis and host immune response to a particular viral agent. Clinically, serology is useful for diagnosing current or past infection and for aiding in clinical management decisions. Serology is useful for screening blood donations for infectious agents and for monitoring the outcome of vaccination against these viruses. Serodiagnosis of human tumor viruses has improved in recent years with increased specificity and sensitivity of the assays, as well as reductions in cost and the ability to assess multiple antibody/antigens in single assays. Serodiagnosis of tumor viruses plays an important role in our understanding of the prevalence and transmission of these viruses and ultimately in the ability to develop treatments/preventions for these globally important diseases. PMID:25843726

  3. Discovering novel zoonotic viruses.

    PubMed

    Wang, Lin-Fa

    2011-07-01

    From the emergence of Hendra virus and Menangle virus in Australia to the global pandemics of severe acute respiratory syndrome and influenza viruses (both H5N1 and H1N1), there has been a surge of zoonotic virus outbreaks in the last two decades. Although the drivers for virus emergence remain poorly understood, the rate of discovery of new viruses is accelerating. This is due to a combination of true emergence of new pathogens and the advance of new technologies making rapid detection and characterisation possible. While molecular approaches will continue to lead the way in virus discovery, other technological platforms are required to increase the chance of success. The lessons learnt in the last 20 years confirm that the One Health approach, involving inclusive collaborations between physicians, veterinarians and other health and environmental professionals, will be the key to combating future zoonotic disease outbreaks.

  4. Nuclear astrophysics

    SciTech Connect

    Haxton, W.C.

    1992-01-01

    The problem of core-collapse supernovae is used to illustrate the many connections between nuclear astrophysics and the problems nuclear physicists study in terrestrial laboratories. Efforts to better understand the collapse and mantle ejection are also motivated by a variety of interdisciplinary issues in nuclear, particle, and astrophysics, including galactic chemical evolution, neutrino masses and mixing, and stellar cooling by the emission of new particles. The current status of theory and observations is summarized.

  5. Nuclear astrophysics

    SciTech Connect

    Haxton, W.C.

    1992-12-31

    The problem of core-collapse supernovae is used to illustrate the many connections between nuclear astrophysics and the problems nuclear physicists study in terrestrial laboratories. Efforts to better understand the collapse and mantle ejection are also motivated by a variety of interdisciplinary issues in nuclear, particle, and astrophysics, including galactic chemical evolution, neutrino masses and mixing, and stellar cooling by the emission of new particles. The current status of theory and observations is summarized.

  6. The Polyomaviridae: Contributions of virus structure to our understanding of virus receptors and infectious entry

    SciTech Connect

    Neu, Ursula; Stehle, Thilo Atwood, Walter J.

    2009-02-20

    This review summarizes the field's major findings related to the characterization of polyomavirus structures and to the characterization of virus receptors and mechanisms of host cell invasion. The four members of the family that have received the most attention in this regard are the mouse polyomavirus (mPyV), the monkey polyomavirus SV40, and the two human polyomaviruses, JCV and BKV. The structures of both the mPyV and SV40 alone and in complex with receptor fragments have been solved to high resolution. The majority of polyomaviruses recognize terminal sialic acid in either an {alpha}2,3 linkage or an {alpha}2,6 linkage to the underlying galactose. Studies on virus structure, receptor utilization and mechanisms of entry have led to new insights into how these viruses interact in an active way with cells to ensure the nuclear delivery and expression of their genomes. Critical work on virus entry has led to the discovery of a pH neutral endocytic compartment that accepts cargo from caveolae and to novel roles for endoplasmic reticulum (ER) associated factors in virus uncoating and penetration of ER membranes. This review will summarize the major findings and compare and contrast the mechanisms used by these viruses to infect cells.

  7. Virus-Associated Lymphomagenesis

    PubMed Central

    Tarantul, V. Z.

    2006-01-01

    At least 2 billion people are affected by viral infections worldwide. The infections induce a lot of various human diseases and are one of the main causes of human mortality. In particular, they can lead to development of various human cancers. Up to 15-20% of human cancer incidence can be attributed to viruses. Although viral infections are very common in the general population, only few of them result in clinically relevant lesions. Certain associations between virus infections and malignancy are strong and irrefutable, the others are still speculative. The criteria most often used for determining the causality are the consistence of the association, either epidemiologic or at the molecular level, and oncogenicity of viruses or particular viral genes in animal models or cell cultures. Due to some ambiguity of such a determination, it is instructive to consider by specific cases what evidence is generally accepted as sufficient to establish a causal relation between virus and cancer. Lymphomas are one of the best studied cancer types closely associated with a small but definite range of viruses. Numerous data show a close interrelation between lymphomagenesis and infection by such viruses as Kaposi’s sarcoma herpesvirus (KSHV), Epstein-Barr virus (EBV), hepatitis C virus (HCV), human T-cell leukemia virus (HTLV), and human immunodeficiency virus (HIV). For instance, experiments on monkeys artificially infected with viruses and data on anti-cancer effect of specific antiviral preparations strongly suggest the involvement of viruses in lymphoma development. The present review is devoted to the association of different viruses with human lymphomas and to viral genes potentially involved in the neoplastic process. The recognition of virus involvement in lymphomagenesis may facilitate new strategies for cancer therapy, diagnosis and screening and can lead to a reduction in the number of individuals at risk of disease. PMID:23674972

  8. [The great virus comeback].

    PubMed

    Forterre, Patrick

    2013-01-01

    Viruses have been considered for a long time as by-products of biological evolution. This view is changing now as a result of several recent discoveries. Viral ecologists have shown that viral particles are the most abundant biological entities on our planet, whereas metagenomic analyses have revealed an unexpected abundance and diversity of viral genes in the biosphere. Comparative genomics have highlighted the uniqueness of viral sequences, in contradiction with the traditional view of viruses as pickpockets of cellular genes. On the contrary, cellular genomes, especially eukaryotic ones, turned out to be full of genes derived from viruses or related elements (plasmids, transposons, retroelements and so on). The discovery of unusual viruses infecting archaea has shown that the viral world is much more diverse than previously thought, ruining the traditional dichotomy between bacteriophages and viruses. Finally, the discovery of giant viruses has blurred the traditional image of viruses as small entities. Furthermore, essential clues on virus history have been obtained in the last ten years. In particular, structural analyses of capsid proteins have uncovered deeply rooted homologies between viruses infecting different cellular domains, suggesting that viruses originated before the last universal common ancestor (LUCA). These studies have shown that several lineages of viruses originated independently, i.e., viruses are polyphyletic. From the time of LUCA, viruses have coevolved with their hosts, and viral lineages can be viewed as lianas wrapping around the trunk, branches and leaves of the tree of life. Although viruses are very diverse, with genomes encoding from one to more than one thousand proteins, they can all be simply defined as organisms producing virions. Virions themselves can be defined as infectious particles made of at least one protein associated with the viral nucleic acid, endowed with the capability to protect the viral genome and ensure its

  9. Evidence for ubiquitin-regulated nuclear and subnuclear trafficking among Paramyxovirinae matrix proteins.

    PubMed

    Pentecost, Mickey; Vashisht, Ajay A; Lester, Talia; Voros, Tim; Beaty, Shannon M; Park, Arnold; Wang, Yao E; Yun, Tatyana E; Freiberg, Alexander N; Wohlschlegel, James A; Lee, Benhur

    2015-03-01

    The paramyxovirus matrix (M) protein is a molecular scaffold required for viral morphogenesis and budding at the plasma membrane. Transient nuclear residence of some M proteins hints at non-structural roles. However, little is known regarding the mechanisms that regulate the nuclear sojourn. Previously, we found that the nuclear-cytoplasmic trafficking of Nipah virus M (NiV-M) is a prerequisite for budding, and is regulated by a bipartite nuclear localization signal (NLSbp), a leucine-rich nuclear export signal (NES), and monoubiquitination of the K258 residue within the NLSbp itself (NLSbp-lysine). To define whether the sequence determinants of nuclear trafficking identified in NiV-M are common among other Paramyxovirinae M proteins, we generated the homologous NES and NLSbp-lysine mutations in M proteins from the five major Paramyxovirinae genera. Using quantitative 3D confocal microscopy, we determined that the NES and NLSbp-lysine are required for the efficient nuclear export of the M proteins of Nipah virus, Hendra virus, Sendai virus, and Mumps virus. Pharmacological depletion of free ubiquitin or mutation of the conserved NLSbp-lysine to an arginine, which inhibits M ubiquitination, also results in nuclear and nucleolar retention of these M proteins. Recombinant Sendai virus (rSeV-eGFP) bearing the NES or NLSbp-lysine M mutants rescued at similar efficiencies to wild type. However, foci of cells expressing the M mutants displayed marked fusogenicity in contrast to wild type, and infection did not spread. Recombinant Mumps virus (rMuV-eGFP) bearing the homologous mutations showed similar defects in viral morphogenesis. Finally, shotgun proteomics experiments indicated that the interactomes of Paramyxovirinae M proteins are significantly enriched for components of the nuclear pore complex, nuclear transport receptors, and nucleolar proteins. We then synthesize our functional and proteomics data to propose a working model for the ubiquitin-regulated nuclear

  10. Viruses of botrytis.

    PubMed

    Pearson, Michael N; Bailey, Andrew M

    2013-01-01

    Botrytis cinerea (gray mold) is one of the most widespread and destructive fungal diseases of horticultural crops. Propagation and dispersal is usually by asexual conidia but the sexual stage (Botryotinia fuckeliana (de Bary) Whetzel) also occurs in nature. DsRNAs, indicative of virus infection, are common in B. cinerea, but only four viruses (Botrytis virus F (BVF), Botrytis virus X (BVX), Botrytis cinerea mitovirus 1 (BcMV1), and Botrytis porri RNA virus) have been sequenced. BVF and BVX are unusual mycoviruses being ssRNA flexous rods and have been designated the type species of the genera Mycoflexivirus and Botrexvirus (family Betaflexivirdae), respectively. The reported effects of viruses on Botrytis range from negligible to severe, with Botrytis cinerea mitovirus 1 causing hypovirulence. Little is currently known about the effects of viruses on Botrytis metabolism but recent complete sequencing of the B. cinerea genome now provides an opportunity to investigate the host-pathogen interactions at the molecular level. There is interest in the possible use of mycoviruses as biological controls for Botrytis because of the common problem of fungicide resistance. Unfortunately, hyphal anastomosis is the only known mechanism of horizontal virus transmission and the large number of vegetative incompatibility groups in Botrytis is a potential constraint on the spread of an introduced virus. Although some Botrytis viruses, such as BVF and BVX, are known to have international distribution, there is a distinct lack of epidemiological data and the means of spread are unknown.

  11. RNA Viruses Infecting Pest Insects

    Technology Transfer Automated Retrieval System (TEKTRAN)

    RNA viruses are viruses whose genetic material is ribonucleic acid (RNA). RNA viruses may be double or single-stranded based on the type of RNA they contain. Single-stranded RNA viruses can be further grouped into negative sense or positive-sense viruses according to the polarity of their RNA. Fur...

  12. Postmortem stability of Ebola virus.

    PubMed

    Prescott, Joseph; Bushmaker, Trenton; Fischer, Robert; Miazgowicz, Kerri; Judson, Seth; Munster, Vincent J

    2015-05-01

    The ongoing Ebola virus outbreak in West Africa has highlighted questions regarding stability of the virus and detection of RNA from corpses. We used Ebola virus-infected macaques to model humans who died of Ebola virus disease. Viable virus was isolated <7 days posteuthanasia; viral RNA was detectable for 10 weeks.

  13. Nuclear Astrophysics

    NASA Astrophysics Data System (ADS)

    Drago, Alessandro

    2005-04-01

    The activity of the Italian nuclear physicists community in the field of Nuclear Astrophysics is reported. The researches here described have been performed within the project "Fisica teorica del nucleo e dei sistemi a multi corpi", supported by the Ministero dell'Istruzione, dell'Università e della Ricerca.

  14. Nuclear safety

    NASA Technical Reports Server (NTRS)

    Buden, D.

    1991-01-01

    Topics dealing with nuclear safety are addressed which include the following: general safety requirements; safety design requirements; terrestrial safety; SP-100 Flight System key safety requirements; potential mission accidents and hazards; key safety features; ground operations; launch operations; flight operations; disposal; safety concerns; licensing; the nuclear engine for rocket vehicle application (NERVA) design philosophy; the NERVA flight safety program; and the NERVA safety plan.

  15. Nuclear hostages

    SciTech Connect

    O'Keefe, B.J.

    1983-01-01

    Classical physics since Roentgen's discovery of X-rays led quickly to work on atomic structure and the Nuclear Age. The author traces the history of decisions to pursue nuclear fission, the organization of the Manhattan Project, the compromises of the 1963 test ban treaty, and the dilemma of nuclear weapons development and deployment that now hold mankind hostage. He reviews the rationale for limited nuclear war, first strike, massive retaliation, non-proliferation, and the Strategic Arms Limitation Talks (SALT) treaties. He argues that the concepts of mobile MX weapons, fratricide, and population dispersal for civil defense are unworkable, suggesting a program of unilaterally withdrawing tactical nuclear weapons from Europe and strengthening intelligence and law-enforcement powers to withstand terrorist activity. Economic cooperation and political reconciliation may take a generation to achieve, but should be our national goal.

  16. A Role for Nuclear F-Actin Induction in Human Cytomegalovirus Nuclear Egress

    PubMed Central

    Wilkie, Adrian R.; Lawler, Jessica L.

    2016-01-01

    ABSTRACT Herpesviruses, which include important pathogens, remodel the host cell nucleus to facilitate infection. This remodeling includes the formation of structures called replication compartments (RCs) in which herpesviruses replicate their DNA. During infection with the betaherpesvirus, human cytomegalovirus (HCMV), viral DNA synthesis occurs at the periphery of RCs within the nuclear interior, after which assembled capsids must reach the inner nuclear membrane (INM) for translocation to the cytoplasm (nuclear egress). The processes that facilitate movement of HCMV capsids to the INM during nuclear egress are unknown. Although an actin-based mechanism of alphaherpesvirus capsid trafficking to the INM has been proposed, it is controversial. Here, using a fluorescently-tagged, nucleus-localized actin-binding peptide, we show that HCMV, but not herpes simplex virus 1, strongly induced nuclear actin filaments (F-actin) in human fibroblasts. Based on studies using UV inactivation and inhibitors, this induction depended on viral gene expression. Interestingly, by 24 h postinfection, nuclear F-actin formed thicker structures that appeared by super-resolution microscopy to be bundles of filaments. Later in infection, nuclear F-actin primarily localized along the RC periphery and between the RC periphery and the nuclear rim. Importantly, a drug that depolymerized nuclear F-actin caused defects in production of infectious virus, capsid accumulation in the cytoplasm, and capsid localization near the nuclear rim, without decreasing capsid accumulation in the nucleus. Thus, our results suggest that for at least one herpesvirus, nuclear F-actin promotes capsid movement to the nuclear periphery and nuclear egress. We discuss our results in terms of competing models for these processes. PMID:27555312

  17. Crystal Structure of the Human Cytomegalovirus pUL50-pUL53 Core Nuclear Egress Complex Provides Insight into a Unique Assembly Scaffold for Virus-Host Protein Interactions.

    PubMed

    Walzer, Sascha A; Egerer-Sieber, Claudia; Sticht, Heinrich; Sevvana, Madhumati; Hohl, Katharina; Milbradt, Jens; Muller, Yves A; Marschall, Manfred

    2015-11-13

    Nuclear replication of cytomegalovirus relies on elaborate mechanisms of nucleocytoplasmic egress of viral particles. Thus, the role of two essential and conserved viral nuclear egress proteins, pUL50 and pUL53, is pivotal. pUL50 and pUL53 heterodimerize and form a core nuclear egress complex (NEC), which is anchored to the inner nuclear membrane and provides a scaffold for the assembly of a multimeric viral-cellular NEC. Here, we report the crystal structure of the pUL50-pUL53 heterodimer (amino acids 1-175 and 50-292, respectively) at 2.44 Å resolution. Both proteins adopt a globular fold with mixed α and β secondary structure elements. pUL53-specific features include a zinc-binding site and a hook-like N-terminal extension, the latter representing a hallmark element of the pUL50-pUL53 interaction. The hook-like extension (amino acids 59-87) embraces pUL50 and contributes 1510 Å(2) to the total interface area (1880 Å(2)). The pUL50 structure overall resembles the recently published NMR structure of the murine cytomegalovirus homolog pM50 but reveals a considerable repositioning of the very C-terminal α-helix of pUL50 upon pUL53 binding. pUL53 shows structural resemblance with the GHKL domain of bacterial sensory histidine kinases. A close examination of the crystal structure indicates partial assembly of pUL50-pUL53 heterodimers to hexameric ring-like structures possibly providing additional scaffolding opportunities for NEC. In combination, the structural information on pUL50-pUL53 considerably improves our understanding of the mechanism of HCMV nuclear egress. It may also accelerate the validation of the NEC as a unique target for developing a novel type of antiviral drug and improved options of broad-spectrum antiherpesviral therapy.

  18. Viruses of Entamoeba histolytica. I. Identification of transmissible virus-like agents.

    PubMed

    Diamond, L S; Mattern, C F; Bartgis, I L

    1972-02-01

    This and a companion report deal with the identification and morphogenesis of viruses in axenized cultures of Entamoeba histolytica. There are probably two different types of virus each producing a different pathological picture in different amoebal strains, or, less likely, there is one type of agent having widely different morphological and morphogenetical pictures in different strains of E. histolytica. Both types of agent produce a lytic response in axenized amoebae and have been serially passaged to an extent assuring their replicating nature. One appears to replicate in the nucleus as multiple clusters of fine filaments which ultimately lyse the nucleus, causing cell death. The second type of agent appears to be a typical polyhedral virus, seen only in the cytoplasm and also resulting in lysis of the cell. A particle morphologically indistinguishable from this second agent is also found in late passages of the agent producing the nuclear pathology. PMID:4335522

  19. Water system virus detection

    NASA Technical Reports Server (NTRS)

    Fraser, A. S.; Wells, A. F.; Tenoso, H. J. (Inventor)

    1978-01-01

    The performance of a waste water reclamation system is monitored by introducing a non-pathogenic marker virus, bacteriophage F2, into the waste-water prior to treatment and, thereafter, testing the reclaimed water for the presence of the marker virus. A test sample is first concentrated by absorbing any marker virus onto a cellulose acetate filter in the presence of a trivalent cation at low pH and then flushing the filter with a limited quantity of a glycine buffer solution to desorb any marker virus present on the filter. Photo-optical detection of indirect passive immune agglutination by polystyrene beads indicates the performance of the water reclamation system in removing the marker virus. A closed system provides for concentrating any marker virus, initiating and monitoring the passive immune agglutination reaction, and then flushing the system to prepare for another sample.

  20. Constructing computer virus phylogenies

    SciTech Connect

    Goldberg, L.A.; Goldberg, P.W.; Phillips, C.A.; Sorkin, G.B.

    1996-03-01

    There has been much recent algorithmic work on the problem of reconstructing the evolutionary history of biological species. Computer virus specialists are interested in finding the evolutionary history of computer viruses--a virus is often written using code fragments from one or more other viruses, which are its immediate ancestors. A phylogeny for a collection of computer viruses is a directed acyclic graph whose nodes are the viruses and whose edges map ancestors to descendants and satisfy the property that each code fragment is ``invented`` only once. To provide a simple explanation for the data, we consider the problem of constructing such a phylogeny with a minimal number of edges. In general, this optimization problem cannot be solved in quasi-polynomial time unless NQP=QP; we present positive and negative results for associated approximated problems. When tree solutions exist, they can be constructed and randomly sampled in polynomial time.

  1. Viruses within animal genomes.

    PubMed

    De Brognier, A; Willems, L

    2016-04-01

    Viruses and their hosts can co-evolve to reach a fragile equilibrium that allows the survival of both. An excess of pathogenicity in the absence of a reservoir would be detrimental to virus survival. A significant proportion of all animal genomes has been shaped by the insertion of viruses that subsequently became 'fossilised'. Most endogenous viruses have lost the capacity to replicate via an infectious cycle and now replicate passively. The insertion of endogenous viruses has contributed to the evolution of animal genomes, for example in the reproductive biology of mammals. However, spontaneous viral integration still occasionally occurs in a number of virus-host systems. This constitutes a potential risk to host survival but also provides an opportunity for diversification and evolution.

  2. Viruses of lower vertebrates.

    PubMed

    Essbauer, S; Ahne, W

    2001-08-01

    Viruses of lower vertebrates recently became a field of interest to the public due to increasing epizootics and economic losses of poikilothermic animals. These were reported worldwide from both wildlife and collections of aquatic poikilothermic animals. Several RNA and DNA viruses infecting fish, amphibians and reptiles have been studied intensively during the last 20 years. Many of these viruses induce diseases resulting in important economic losses of lower vertebrates, especially in fish aquaculture. In addition, some of the DNA viruses seem to be emerging pathogens involved in the worldwide decline in wildlife. Irido-, herpes- and polyomavirus infections may be involved in the reduction in the numbers of endangered amphibian and reptile species. In this context the knowledge of several important RNA viruses such as orthomyxo-, paramyxo-, rhabdo-, retro-, corona-, calici-, toga-, picorna-, noda-, reo- and birnaviruses, and DNA viruses such as parvo-, irido-, herpes-, adeno-, polyoma- and poxviruses, is described in this review. PMID:11550762

  3. Viruses in Antarctic lakes

    NASA Technical Reports Server (NTRS)

    Kepner, R. L. Jr; Wharton, R. A. Jr; Suttle, C. A.; Wharton RA, J. r. (Principal Investigator)

    1998-01-01

    Water samples collected from four perennially ice-covered Antarctic lakes during the austral summer of 1996-1997 contained high densities of extracellular viruses. Many of these viruses were found to be morphologically similar to double-stranded DNA viruses that are known to infect algae and protozoa. These constitute the first observations of viruses in perennially ice-covered polar lakes. The abundance of planktonic viruses and data suggesting substantial production potential (relative to bacteria] secondary and photosynthetic primary production) indicate that viral lysis may be a major factor in the regulation of microbial populations in these extreme environments. Furthermore, we suggest that Antarctic lakes may be a reservoir of previously undescribed viruses that possess novel biological and biochemical characteristics.

  4. Global Reprogramming of Host SUMOylation during Influenza Virus Infection

    PubMed Central

    Domingues, Patricia; Golebiowski, Filip; Tatham, Michael H.; Lopes, Antonio M.; Taggart, Aislynn; Hay, Ronald T.; Hale, Benjamin G.

    2015-01-01

    Summary Dynamic nuclear SUMO modifications play essential roles in orchestrating cellular responses to proteotoxic stress, DNA damage, and DNA virus infection. Here, we describe a non-canonical host SUMOylation response to the nuclear-replicating RNA pathogen, influenza virus, and identify viral RNA polymerase activity as a major contributor to SUMO proteome remodeling. Using quantitative proteomics to compare stress-induced SUMOylation responses, we reveal that influenza virus infection triggers unique re-targeting of SUMO to 63 host proteins involved in transcription, mRNA processing, RNA quality control, and DNA damage repair. This is paralleled by widespread host deSUMOylation. Depletion screening identified ten virus-induced SUMO targets as potential antiviral factors, including C18orf25 and the SMC5/6 and PAF1 complexes. Mechanistic studies further uncovered a role for SUMOylation of the PAF1 complex component, parafibromin (CDC73), in potentiating antiviral gene expression. Our global characterization of influenza virus-triggered SUMO redistribution provides a proteomic resource to understand host nuclear SUMOylation responses to infection. PMID:26549460

  5. Tobacco mosaic virus-directed reprogramming of auxin/indole acetic acid protein transcriptional responses enhances virus phloem loading.

    PubMed

    Collum, Tamara D; Padmanabhan, Meenu S; Hsieh, Yi-Cheng; Culver, James N

    2016-05-10

    Vascular phloem loading has long been recognized as an essential step in the establishment of a systemic virus infection. In this study, an interaction between the replication protein of tobacco mosaic virus (TMV) and phloem-specific auxin/indole acetic acid (Aux/IAA) transcriptional regulators was found to modulate virus phloem loading in an age-dependent manner. Promoter expression studies show that in mature tissues TMV 126/183-kDa-interacting Aux/IAAs predominantly express and accumulate within the nuclei of phloem companion cells (CCs). Furthermore, CC Aux/IAA nuclear localization is disrupted upon infection with an interacting virus. In situ analysis of virus spread shows that the inability to disrupt Aux/IAA CC nuclear localization correlates with a reduced ability to load into the vascular tissue. Subsequent systemic movement assays also demonstrate that a virus capable of disrupting Aux/IAA localization is significantly more competitive at moving out of older plant tissues than a noninteracting virus. Similarly, CC expression and overaccumulation of a degradation-resistant Aux/IAA-interacting protein was found to inhibit TMV accumulation and phloem loading selectively in flowering plants. Transcriptional expression studies demonstrate a role for Aux/IAA-interacting proteins in the regulation of salicylic and jasmonic acid host defense responses as well as virus-specific movement factors, including pectin methylesterase, that are involved in regulating plasmodesmata size-exclusion limits and promoting virus cell-to-cell movement. Combined, these findings indicate that TMV directs the reprogramming of auxin-regulated gene expression within the vascular phloem of mature tissues as a means to enhance phloem loading and systemic spread. PMID:27118842

  6. The human oncogenic viruses

    SciTech Connect

    Luderer, A.A.; Weetall, H.H

    1986-01-01

    This book contains eight selections. The titles are: Cytogenetics of the Leukemias and Lymphomas; Cytogenetics of Solid Tumors: Renal Cell Carcinoma, Malignant Melanoma, Retinoblastoma, and Wilms' Tumor; Elucidation of a Normal Function for a Human Proto-Oncogene; Detection of HSV-2 Genes and Gene Products in Cervical Neoplasia; Papillomaviruses in Anogennital Neoplasms; Human Epstein-Barr Virus and Cancer; Hepatitis B Virus and Hepatocellular Carcinoma; and Kaposi's Sarcoma: Acquired Immunodeficiency Syndrome (AIDS) and Associated Viruses.

  7. Water system virus detection

    NASA Technical Reports Server (NTRS)

    Fraser, A. S.; Wells, A. F.; Tenoso, H. J.

    1975-01-01

    A monitoring system developed to test the capability of a water recovery system to reject the passage of viruses into the recovered water is described. A nonpathogenic marker virus, bacteriophage F2, is fed into the process stream before the recovery unit and the reclaimed water is assayed for its presence. Detection of the marker virus consists of two major components, concentration and isolation of the marker virus, and detection of the marker virus. The concentration system involves adsorption of virus to cellulose acetate filters in the presence of trivalent cations and low pH with subsequent desorption of the virus using volumes of high pH buffer. The detection of the virus is performed by a passive immune agglutination test utilizing specially prepared polystyrene particles. An engineering preliminary design was performed as a parallel effort to the laboratory development of the marker virus test system. Engineering schematics and drawings of a fully functional laboratory prototype capable of zero-G operation are presented. The instrument consists of reagent pump/metering system, reagent storage containers, a filter concentrator, an incubation/detector system, and an electronic readout and control system.

  8. [Zika virus epidemic].

    PubMed

    Kronborg, Gitte; Fomsgaard, Anders

    2016-03-21

    Zika virus is endemic in several parts of the world. February 1, 2016 Zika virus was declared a public health emergency by the WHO. This declaration is mainly due to a convincing association between Zika virus infection during pregnancy and birth defects, like microcephaly, among some of the newborns. Imported cases of Zika virus infection to North America, Europe and Denmark have been described. The infection in itself is mild and self-limiting. The available diagnostic methods are under development, validation and evaluation. In Denmark, some promising diagnostics are available at Statens Serum Institut.

  9. Nuclear Astrophysics

    NASA Astrophysics Data System (ADS)

    Bombaci, Ignazio

    2003-04-01

    In this report I will try to illustrate some of the main research themes and "hot topics" in nuclear astrophysics. The particular aim of the present report is to briefly illustrate the research activities, in the field of nuclear astrophysics, performed by the Italian nuclear physicist community within the "Programma di Interesse Nazionale su Fisica Teorica del Nucleo e dei Sistemi a Molti Corpi" (National Research Program on Theoretical Physics of Nuclei and Many Body Systems) supported by the "Ministero dell'Istruzione dell'Università e della Ricerca".

  10. Nuclear Speckles

    PubMed Central

    Spector, David L.; Lamond, Angus I.

    2011-01-01

    Nuclear speckles, also known as interchromatin granule clusters, are nuclear domains enriched in pre-mRNA splicing factors, located in the interchromatin regions of the nucleoplasm of mammalian cells. When observed by immunofluorescence microscopy, they usually appear as 20–50 irregularly shaped structures that vary in size. Speckles are dynamic structures, and their constituents can exchange continuously with the nucleoplasm and other nuclear locations, including active transcription sites. Studies on the composition, structure, and dynamics of speckles have provided an important paradigm for understanding the functional organization of the nucleus and the dynamics of the gene expression machinery. PMID:20926517

  11. (Nuclear theory). [Research in nuclear physics

    SciTech Connect

    Haxton, W.

    1990-01-01

    This report discusses research in nuclear physics. Topics covered in this paper are: symmetry principles; nuclear astrophysics; nuclear structure; quark-gluon plasma; quantum chromodynamics; symmetry breaking; nuclear deformation; and cold fusion. (LSP)

  12. Hsp90 inhibitors reduce influenza virus replication in cell culture

    SciTech Connect

    Chase, Geoffrey; Deng, Tao; Fodor, Ervin; Leung, B.W.; Mayer, Daniel; Schwemmle, Martin Brownlee, George

    2008-08-01

    The viral RNA polymerase complex of influenza A virus consists of three subunits PB1, PB2 and PA. Recently, the cellular chaperone Hsp90 was shown to play a role in nuclear import and assembly of the trimeric polymerase complex by binding to PB1 and PB2. Here we show that Hsp90 inhibitors, geldanamycin or its derivative 17-AAG, delay the growth of influenza virus in cell culture resulting in a 1-2 log reduction in viral titre early in infection. We suggest that this is caused by the reduced half-life of PB1 and PB2 and inhibition of nuclear import of PB1 and PA which lead to reduction in viral RNP assembly. Hsp90 inhibitors may represent a new class of antiviral compounds against influenza viruses.

  13. Nuclear forces

    SciTech Connect

    Machleidt, R.

    2013-06-10

    These lectures present an introduction into the theory of nuclear forces. We focus mainly on the modern approach, in which the forces between nucleons emerge from low-energy QCD via chiral effective field theory.

  14. Nuclear Disarmament.

    ERIC Educational Resources Information Center

    Johnson, Christopher

    1982-01-01

    Material about nuclear disarmament and the arms race should be included in secondary school curricula. Teachers can present this technical, controversial, and frightening material in a balanced and comprehensible way. Resources for instructional materials are listed. (PP)

  15. Nuclear battlefields

    SciTech Connect

    Arkin, W.M.; Fieldhouse, R.W.

    1985-01-01

    This book provides complete data on the nuclear operations and research facilities in the U.S.A., the U.S.S.R., France, China and the U.K. It describes detailed estimates on the U.S.S.R.'s nuclear stockpile for over 500 locations. It shows how non-nuclear countries cooperate with the world-wide war machine. And it maps the U.S. nuclear facilities from Little America, WY, and Charleston, SC, to the battleships patroling the world's oceans and subs stalking under the sea. The data were gathered from unclassified sources through the Freedom of Information Act, from data supplied to military installations, and from weapons source books. It provides guidance for policymakers, government and corporate officials.

  16. NUCLEAR REACTOR

    DOEpatents

    Sherman, J.; Sharbaugh, J.E.; Fauth, W.L. Jr.; Palladino, N.J.; DeHuff, P.G.

    1962-10-23

    A nuclear reactor incorporating seed and blanket assemblies is designed. Means are provided for obtaining samples of the coolant from the blanket assemblies and for varying the flow of coolant through the blanket assemblies. (AEC)

  17. Nuclear Data

    SciTech Connect

    White, Morgan C.

    2014-01-23

    PowerPoint presentation targeted for educational use. Nuclear data comes from a variety of sources and in many flavors. Understanding where the data you use comes from and what flavor it is can be essential to understand and interpret your results. This talk will discuss the nuclear data pipeline with particular emphasis on providing links to additional resources that can be used to explore the issues you will encounter.

  18. Nuclear Nonproliferation

    SciTech Connect

    Atkins-Duffin, C E

    2008-12-10

    With an explosion equivalent of about 20kT of TNT, the Trinity test was the first demonstration of a nuclear weapon. Conducted on July 16, 1945 in Alamogordo, NM this site is now a Registered National Historic Landmark. The concept and applicability of nuclear power was demonstrated on December 20, 1951 with the Experimental Breeder Reactor Number One (EBR-1) lit four light bulbs. This reactor is now a Registered National Historic Landmark, located near Arco, ID. From that moment forward it had been clearly demonstrated that nuclear energy has both peaceful and military applications and that the civilian and military fuel cycles can overlap. For the more than fifty years since the Atoms for Peace program, a key objective of nuclear policy has been to enable the wider peaceful use of nuclear energy while preventing the spread of nuclear weapons. Volumes have been written on the impact of these two actions on the world by advocates and critics; pundits and practioners; politicians and technologists. The nations of the world have woven together a delicate balance of treaties, agreements, frameworks and handshakes that are representative of the timeframe in which they were constructed and how they have evolved in time. Collectively these vehicles attempt to keep political will, nuclear materials and technology in check. This paper captures only the briefest abstract of the more significant aspects on the Nonproliferation Regime. Of particular relevance to this discussion is the special nonproliferation sensitivity associated with the uranium isotope separation and spent fuel reprocessing aspects of the nuclear fuel cycle.

  19. Nuclear Structure

    NASA Astrophysics Data System (ADS)

    Gargano, Angela

    2003-04-01

    An account of recent studies in the field of theoretical nuclear structure is reported. These studies concern essentially research activities performed under the Italian project "Fisica Teorica del Nucleo e dei Sistemi a Molti Corpi". Special attention is addressed to results obtained during the last two years as regards the development of new many-body techniques as well as the interpretation of new experimental aspects of nuclear structure.

  20. Nuclear astrophysics

    NASA Astrophysics Data System (ADS)

    Arnould, M.; Takahashi, K.

    1999-03-01

    Nuclear astrophysics is that branch of astrophysics which helps understanding of the Universe, or at least some of its many faces, through the knowledge of the microcosm of the atomic nucleus. It attempts to find as many nuclear physics imprints as possible in the macrocosm, and to decipher what those messages are telling us about the varied constituent objects in the Universe at present and in the past. In the last decades much advance has been made in nuclear astrophysics thanks to the sometimes spectacular progress made in the modelling of the structure and evolution of the stars, in the quality and diversity of the astronomical observations, as well as in the experimental and theoretical understanding of the atomic nucleus and of its spontaneous or induced transformations. Developments in other subfields of physics and chemistry have also contributed to that advance. Notwithstanding the accomplishment, many long-standing problems remain to be solved, and the theoretical understanding of a large variety of observational facts needs to be put on safer grounds. In addition, new questions are continuously emerging, and new facts endangering old ideas. This review shows that astrophysics has been, and still is, highly demanding to nuclear physics in both its experimental and theoretical components. On top of the fact that large varieties of nuclei have to be dealt with, these nuclei are immersed in highly unusual environments which may have a significant impact on their static properties, the diversity of their transmutation modes, and on the probabilities of these modes. In order to have a chance of solving some of the problems nuclear astrophysics is facing, the astrophysicists and nuclear physicists are obviously bound to put their competence in common, and have sometimes to benefit from the help of other fields of physics, like particle physics, plasma physics or solid-state physics. Given the highly varied and complex aspects, we pick here some specific nuclear

  1. Nuclear telemedicine

    NASA Astrophysics Data System (ADS)

    Morrison, R. T.; Szasz, I. J.

    1990-06-01

    Diagnostic nuclear medicine patient images have been transniitted for 8 years from a regional conununity hospital to a university teaching hospital 700 kiloinetres away employing slow scan TV and telephone. Transruission and interpretation were done at the end of each working day or as circumstances required in cases of emergencies. Referring physicians received the nuclear medicine procedure report at the end of the completion day or within few minutes of completion in case of emergency procedures. To date more than 25 patient studies have been transmitted for interpretation. Blinded reinterpretation of the original hard copy data of 350 patient studies resulted in 100 agreement with the interpretation of transmitted data. This technique provides high quality diagnostic and therapeutic nuclear medicine services in remote hospitals where the services of an on-site nuclear physician is not available. 2. HISTORY Eight years ago when the nuclear medicine physician at Trail Regional Hospital left the Trail area and an other could not be recruited we examined the feasibility of image transmission by phone for interpretation since closing the department would have imposed unacceptable physical and financial hardship and medical constraints on the patient population the nearest nuclear medicine facility was at some 8 hours drive away. In hospital patients would have to be treated either based purely on physical findings or flown to Vancouver at considerable cost to the health care system (estimated cost $1500.

  2. Paramecium bursaria chlorella virus 1 proteome reveals novel architectural and regulatory features of a giant virus.

    PubMed

    Dunigan, David D; Cerny, Ronald L; Bauman, Andrew T; Roach, Jared C; Lane, Leslie C; Agarkova, Irina V; Wulser, Kurt; Yanai-Balser, Giane M; Gurnon, James R; Vitek, Jason C; Kronschnabel, Bernard J; Jeanniard, Adrien; Blanc, Guillaume; Upton, Chris; Duncan, Garry A; McClung, O William; Ma, Fangrui; Van Etten, James L

    2012-08-01

    The 331-kbp chlorovirus Paramecium bursaria chlorella virus 1 (PBCV-1) genome was resequenced and annotated to correct errors in the original 15-year-old sequence; 40 codons was considered the minimum protein size of an open reading frame. PBCV-1 has 416 predicted protein-encoding sequences and 11 tRNAs. A proteome analysis was also conducted on highly purified PBCV-1 virions using two mass spectrometry-based protocols. The mass spectrometry-derived data were compared to PBCV-1 and its host Chlorella variabilis NC64A predicted proteomes. Combined, these analyses revealed 148 unique virus-encoded proteins associated with the virion (about 35% of the coding capacity of the virus) and 1 host protein. Some of these proteins appear to be structural/architectural, whereas others have enzymatic, chromatin modification, and signal transduction functions. Most (106) of the proteins have no known function or homologs in the existing gene databases except as orthologs with proteins of other chloroviruses, phycodnaviruses, and nuclear-cytoplasmic large DNA viruses. The genes encoding these proteins are dispersed throughout the virus genome, and most are transcribed late or early-late in the infection cycle, which is consistent with virion morphogenesis.

  3. Cytoplasmic RNA viruses as potential vehicles for the delivery of therapeutic small RNAs

    PubMed Central

    2013-01-01

    Viral vectors have become the best option for the delivery of therapeutic genes in conventional and RNA interference-based gene therapies. The current viral vectors for the delivery of small regulatory RNAs are based on DNA viruses and retroviruses/lentiviruses. Cytoplasmic RNA viruses have been excluded as viral vectors for RNAi therapy because of the nuclear localization of the microprocessor complex and the potential degradation of the viral RNA genome during the excision of any virus-encoded pre-microRNAs. However, in the last few years, the presence of several species of small RNAs (e.g., virus-derived small interfering RNAs, virus-derived short RNAs, and unusually small RNAs) in animals and cell cultures that are infected with cytoplasmic RNA viruses has suggested the existence of a non-canonical mechanism of microRNA biogenesis. Several studies have been conducted on the tick-borne encephalitis virus and on the Sindbis virus in which microRNA precursors were artificially incorporated and demonstrated the production of mature microRNAs. The ability of these viruses to recruit Drosha to the cytoplasm during infection resulted in the efficient processing of virus-encoded microRNA without the viral genome entering the nucleus. In this review, we discuss the relevance of these findings with an emphasis on the potential use of cytoplasmic RNA viruses as vehicles for the efficient delivery of therapeutic small RNAs. PMID:23759022

  4. Equine Arteritis Virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    03. Nidovirales : 03.004. Arteriviridae : 03.004.0. {03.004.0. unknown} : 03.004.0.01. Arterivirus : 03.004.0.01.001. Equine arteritis virus will be published online. The article details the phenotypic and genotypic makeup of equine arteritis virus (EAV), and summarizes its biological properties....

  5. Virus separation using membranes.

    PubMed

    Grein, Tanja A; Michalsky, Ronald; Czermak, Peter

    2014-01-01

    Industrial manufacturing of cell culture-derived viruses or virus-like particles for gene therapy or vaccine production are complex multistep processes. In addition to the bioreactor, such processes require a multitude of downstream unit operations for product separation, concentration, or purification. Similarly, before a biopharmaceutical product can enter the market, removal or inactivation of potential viral contamination has to be demonstrated. Given the complexity of biological solutions and the high standards on composition and purity of biopharmaceuticals, downstream processing is the bottleneck in many biotechnological production trains. Membrane-based filtration can be an economically attractive and efficient technology for virus separation. Viral clearance, for instance, of up to seven orders of magnitude has been reported for state of the art polymeric membranes under best conditions.This chapter summarizes the fundamentals of virus ultrafiltration, diafiltration, or purification with adsorptive membranes. In lieu of an impractical universally applicable protocol for virus filtration, application of these principles is demonstrated with two examples. The chapter provides detailed methods for production, concentration, purification, and removal of a rod-shaped baculovirus (Autographa californica M nucleopolyhedrovirus, about 40 × 300 nm in size, a potential vector for gene therapy, and an industrially important protein expression system) or a spherical parvovirus (minute virus of mice, 22-26 nm in size, a model virus for virus clearance validation studies).

  6. Papaya Ringspot Virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The term papaya ringspot virus (PRSV) was coined by Jensen in 1949, to describe a papaya disease in Hawaii. Later work showed that diseases such as papaya mosaic and watermelon mosaic virus-1 were caused by PRSV. The primary host range of PRSV is papaya and cucurbits, with Chenopium amaranticolor ...

  7. Positive reinforcement for viruses

    PubMed Central

    Vigant, Frederic; Jung, Michael; Lee, Benhur

    2010-01-01

    Summary Virus-cell membrane fusion requires a critical transition from positive to negative membrane curvature. St. Vincent et al., in PNAS (St Vincent, et al., 2010), designed a class of antivirals that targets this transition. These Rigid Amphipathic Fusion Inhibitors are active against an array of enveloped viruses. PMID:21035726

  8. Positive reinforcement for viruses.

    PubMed

    Vigant, Frederic; Jung, Michael; Lee, Benhur

    2010-10-29

    Virus-cell membrane fusion requires a critical transition from positive to negative membrane curvature. St. Vincent et al. (2010), in PNAS, designed a class of antivirals that targets this transition. These rigid amphipathic fusion inhibitors are active against an array of enveloped viruses.

  9. Rift Valley Fever Virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rift Valley fever virus (RVFV) is a mosquito-transmitted virus or arbovirus that is endemic in sub-Saharan Africa. In the last decade, Rift Valley fever (RVF) outbreaks have resulted in loss of human and animal life, as well as had significant economic impact. The disease in livestock is primarily a...

  10. Papaya ringspot virus (Potyviridae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Papaya ringspot virus, a member of the family Potyviridae, is single stranded RNA plant virus with a monocistronic genome of about 10,326 nucleotides that is expressed via a large polyprotein subsequently cleaved into functional proteins. It causes severe damage on cucurbit crops such as squash and...

  11. Zika Virus Disease.

    PubMed

    Slenczka, Werner

    2016-06-01

    The history of Zika virus disease serves as a paradigm of a typical emerging viral infection. Zika virus disease, a mosquito-borne flavivirus, was first isolated in 1947 in the Zika forest of Uganda. The same virus was also isolated from jungle-dwelling mosquitoes (Aedes [Stegomyia] africanus). In many areas of Africa and South Asia human infections with Zika virus were detected by both serology and virus isolation. About 80% of infections are asymptomatic, and in 20% a mostly mild disease with fever, rash, arthralgia, and conjunctivitis may occur. Fetal infections with malformations were not recorded in Africa or Asia. Zika virus was imported to northern Brazil possibly during the world soccer championship that was hosted by Brazil in June through July 2014. A cluster of severe fetal malformations with microcephaly and ocular defects was noted in 2015 in the northeast of Brazil, and intrauterine infections with Zika virus were confirmed. The dramatic change in Zika virus pathogenicity upon its introduction to Brazil has remained an enigma.

  12. Zika Virus and Pregnancy.

    PubMed

    Stagg, Denise; Hurst, Helen M

    2016-01-01

    Recent outbreaks of Zika virus and reports linking infection in pregnant women with microcephaly in newborns have caused concern worldwide. Information has been evolving rapidly. Nurses and other clinicians, especially those who work with women of childbearing age, play a pivotal role in disseminating accurate information and identifying potential cases of Zika virus infection.

  13. Zika Virus and Pregnancy.

    PubMed

    Stagg, Denise; Hurst, Helen M

    2016-01-01

    Recent outbreaks of Zika virus and reports linking infection in pregnant women with microcephaly in newborns have caused concern worldwide. Information has been evolving rapidly. Nurses and other clinicians, especially those who work with women of childbearing age, play a pivotal role in disseminating accurate information and identifying potential cases of Zika virus infection. PMID:27287356

  14. Zika Virus Disease.

    PubMed

    Slenczka, Werner

    2016-06-01

    The history of Zika virus disease serves as a paradigm of a typical emerging viral infection. Zika virus disease, a mosquito-borne flavivirus, was first isolated in 1947 in the Zika forest of Uganda. The same virus was also isolated from jungle-dwelling mosquitoes (Aedes [Stegomyia] africanus). In many areas of Africa and South Asia human infections with Zika virus were detected by both serology and virus isolation. About 80% of infections are asymptomatic, and in 20% a mostly mild disease with fever, rash, arthralgia, and conjunctivitis may occur. Fetal infections with malformations were not recorded in Africa or Asia. Zika virus was imported to northern Brazil possibly during the world soccer championship that was hosted by Brazil in June through July 2014. A cluster of severe fetal malformations with microcephaly and ocular defects was noted in 2015 in the northeast of Brazil, and intrauterine infections with Zika virus were confirmed. The dramatic change in Zika virus pathogenicity upon its introduction to Brazil has remained an enigma. PMID:27337468

  15. Targeting Cell Division Cycle 25 Homolog B To Regulate Influenza Virus Replication

    PubMed Central

    Perwitasari, Olivia; Torrecilhas, Ana Claudia; Yan, Xiuzhen; Johnson, Scott; White, Caleb; Tompkins, S. Mark

    2013-01-01

    Influenza virus is a worldwide global health concern causing seasonal morbidity mortality and economic burden. Chemotherapeutics is available; however, rapid emergence of drug-resistant influenza virus strains has reduced its efficacy. Thus, there is a need to discover novel antiviral agents. In this study, RNA interference (RNAi) was used to screen host genes required for influenza virus replication. One pro-influenza virus host gene identified was dual-specificity phosphatase cell division cycle 25 B (CDC25B). RNAi screening of CDC25B resulted in reduced influenza A virus replication, and a CDC25B small-molecule inhibitor (NSC95397) inhibited influenza A virus replication in a dose-dependent fashion. Viral RNA synthesis was reduced by NSC95397 in favor of increased beta interferon (IFN-β) expression, and NSC95397 was found to interfere with nuclear localization and chromatin association of NS1, an influenza virus protein. As NS1 has been shown to be chromatin associated and to suppress host transcription, it is likely that CDC25B supports NS1 nuclear function to hijack host transcription machinery in favor of viral RNA synthesis, a process that is blocked by NSC95397. Importantly, NSC95397 treatment protects mice against lethal influenza virus challenge. The findings establish CDC25B as a pro-influenza A virus host factor that may be targeted as a novel influenza A therapeutic strategy. PMID:24109234

  16. Targeting cell division cycle 25 homolog B to regulate influenza virus replication.

    PubMed

    Perwitasari, Olivia; Torrecilhas, Ana Claudia; Yan, Xiuzhen; Johnson, Scott; White, Caleb; Tompkins, S Mark; Tripp, Ralph A

    2013-12-01

    Influenza virus is a worldwide global health concern causing seasonal morbidity mortality and economic burden. Chemotherapeutics is available; however, rapid emergence of drug-resistant influenza virus strains has reduced its efficacy. Thus, there is a need to discover novel antiviral agents. In this study, RNA interference (RNAi) was used to screen host genes required for influenza virus replication. One pro-influenza virus host gene identified was dual-specificity phosphatase cell division cycle 25 B (CDC25B). RNAi screening of CDC25B resulted in reduced influenza A virus replication, and a CDC25B small-molecule inhibitor (NSC95397) inhibited influenza A virus replication in a dose-dependent fashion. Viral RNA synthesis was reduced by NSC95397 in favor of increased beta interferon (IFN-β) expression, and NSC95397 was found to interfere with nuclear localization and chromatin association of NS1, an influenza virus protein. As NS1 has been shown to be chromatin associated and to suppress host transcription, it is likely that CDC25B supports NS1 nuclear function to hijack host transcription machinery in favor of viral RNA synthesis, a process that is blocked by NSC95397. Importantly, NSC95397 treatment protects mice against lethal influenza virus challenge. The findings establish CDC25B as a pro-influenza A virus host factor that may be targeted as a novel influenza A therapeutic strategy.

  17. Pathogenic characteristics of a novel triple-reasserted H1N2 swine influenza virus.

    PubMed

    Liu, Huili; Tao, Jie; Zhang, Pengchao; Yin, Xiuchen; Ha, Zhuo; Zhang, Chunling

    2016-07-01

    A novel triple reasserted H1N2 virus A/swine/Shanghai/1/2007 (SH07) was isolated from nasal swabs of weaned pig showing clinical symptoms of coughing and sneezing. To explore the virus characteristics, mice, chickens and pigs were selected for pathogenicity study. Pigs inoculated intranasally with 10(6) TCID50 SH07 showed clinical symptoms with coughing and sneezing, but no death. The virus nuclear acid was detected in many tissues using real-time PCR, which was mainly distributed in respiratory system particularly in the lungs. The virus was low-pathogenic to chickens with 10(6) TCID50 dose inoculation either via intramuscular or intranasal routes. However virus nuclear acid detection and virus isolation confirmed that the virus can also be found in nasal and rectum. When virus was inoculated into mice by intramuscular or intranasal routes we observed 100% and 80% lethality respectively. The third generation of samples passaged on MDCK cell were SIV positive in indirect immunofluorescence assay (IFA) using antiserum against H1N2 SIV. Furthermore, the lungs of mice showed obvious lesion with interstitial pneumonia. Data in our study suggest that SH07 is preferentially pathogenic to mammals rather than birds although it is a reasserting virus with the fragments from swine, human and avian origin. PMID:27230301

  18. Influenza A virus reassortment.

    PubMed

    Steel, John; Lowen, Anice C

    2014-01-01

    Reassortment is the process by which influenza viruses swap gene segments. This genetic exchange is possible due to the segmented nature of the viral genome and occurs when two differing influenza viruses co-infect a cell. The viral diversity generated through reassortment is vast and plays an important role in the evolution of influenza viruses. Herein we review recent insights into the contribution of reassortment to the natural history and epidemiology of influenza A viruses, gained through population scale phylogenic analyses. We describe methods currently used to study reassortment in the laboratory, and we summarize recent progress made using these experimental approaches to further our understanding of influenza virus reassortment and the contexts in which it occurs.

  19. Respiratory viruses and children.

    PubMed

    Heikkinen, Terho

    2016-07-01

    Respiratory viruses place a great disease burden especially on the youngest children in terms of high rates of infection, bacterial complications and hospitalizations. In developing countries, some viral infections are even associated with substantial mortality in children. The interaction between viruses and bacteria is probably much more common and clinically significant than previously understood. Respiratory viruses frequently initiate the cascade of events that ultimately leads to bacterial infection. Effective antiviral agents can substantially shorten the duration of the viral illness and prevent the development of bacterial complications. Viral vaccines have the potential to not only prevent the viral infection but also decrease the incidence of bacterial complications. At present, antivirals and vaccines are only available against influenza viruses, but new vaccines and antivirals against other viruses, especially for RSV, are being developed. PMID:27177731

  20. Respiratory viruses and children.

    PubMed

    Heikkinen, Terho

    2016-07-01

    Respiratory viruses place a great disease burden especially on the youngest children in terms of high rates of infection, bacterial complications and hospitalizations. In developing countries, some viral infections are even associated with substantial mortality in children. The interaction between viruses and bacteria is probably much more common and clinically significant than previously understood. Respiratory viruses frequently initiate the cascade of events that ultimately leads to bacterial infection. Effective antiviral agents can substantially shorten the duration of the viral illness and prevent the development of bacterial complications. Viral vaccines have the potential to not only prevent the viral infection but also decrease the incidence of bacterial complications. At present, antivirals and vaccines are only available against influenza viruses, but new vaccines and antivirals against other viruses, especially for RSV, are being developed.

  1. Akabane virus infection.

    PubMed

    Kirkland, P D

    2015-08-01

    Akabane virus is a Culicoides-borne orthobunyavirus that is teratogenic to the fetus of cattle and small ruminant species. Depending upon the stage of gestation atwhich infection occurs, and the length of gestation of the mammalian host, a range of congenital defects may be observed. The developing central nervous system is usually the most severely affected, with hydranencephaly and arthrogryposis most frequently observed. Less commonly, some strains of Akabane virus can cause encephalitis in the neonate or, rarely, adult cattle. Akabane viruses are known to be widespread in temperate and tropical regions of Australia, Southeast Asia, the Middle East and some African countries. Disease is infrequently observed in regions where this virus is endemic and the presence of the virus remains unrecognised in the absence of serological surveillance. In some Asian countries, vaccines are used to minimise the occurrence of disease. PMID:26601444

  2. Infectious Viral Quantification of Chikungunya Virus-Virus Plaque Assay.

    PubMed

    Kaur, Parveen; Lee, Regina Ching Hua; Chu, Justin Jang Hann

    2016-01-01

    The plaque assay is an essential method for quantification of infectious virus titer. Cells infected with virus particles are overlaid with a viscous substrate. A suitable incubation period results in the formation of plaques, which can be fixed and stained for visualization. Here, we describe a method for measuring Chikungunya virus (CHIKV) titers via virus plaque assays.

  3. Brevipalpus-transmitted plant virus and virus-like diseases: cytopathology and some recent cases.

    PubMed

    Kitajima, E W; Chagas, C M; Rodrigues, J C V

    2003-01-01

    An increasing number of diseases transmitted by Brevipalpus mite species (Acari: Tenuipalpidae) is being identified that affect economically important plants such as citrus, coffee, passion fruit, orchids, and several ornamentals. All of these diseases are characterized by localized lesions (chlorotic, green spots, or ringspots) on leaves, stems, and fruits. Virus or virus-like agents are considered to be the causal agents, possibly transmitted in a circulative-propagative manner by Brevipalpus mites. The virus or virus-like particles are short, rod-like, or bacilliform, that induce two characteristic types of cell alteration: (1) 'Nuclear type'--nuclei of parenchyma and epidermal cells in the lesions often contain a large electron lucent inclusion. Short, naked, rod-like (40-50 nm x 100-110 nm) particles may be seen in the viroplasm or nucleoplasm and in the cytoplasm. These particles are commonly arranged perpendicularly on the membranes of the nuclear envelope or endoplasmic reticulum (ER). In a very few instances, they were found to be membrane-bound, within the ER cavities. (2) 'Cytoplasmic type'--short bacilliform particles (60-70 nm x 110-120 nm) are present within the cisternae of the ER and often have electron dense viroplasm of varied shapes present in the cytoplasm. Bacilliform particles may be seen budding into the ER lumen near the viroplasm. These particles resemble those of members of the Rhabdoviridae, but are shorter. The only sequenced virus of this group, orchid fleck virus (OFV), has a negative sense (bipartite) type ssRNA genome, but its organization is similar to known rhabdoviruses, which are monopartite. Both types of cytopathological effects have been found associated with citrus leprosis. In orchids, OFV has a 'nuclear type' of cytopathology, but in some species the 'cytoplasmic type' has been found associated with ringspot symptoms. In Hibiscus and Clerodendron, green spot symptoms have been associated with the cytoplasmic type of cell

  4. Virus discovery and recent insights into virus diversity in arthropods.

    PubMed

    Junglen, Sandra; Drosten, Christian

    2013-08-01

    Recent studies on virus discovery have focused mainly on mammalian and avian viruses. Arbovirology with its long tradition of ecologically oriented investigation is now catching up, with important novel insights into the diversity of arthropod-associated viruses. Recent discoveries include taxonomically outlying viruses within the families Flaviviridae, Togaviridae, and Bunyaviridae, and even novel virus families within the order Nidovirales. However, the current focusing of studies on blood-feeding arthropods has restricted the range of arthropod hosts analyzed for viruses so far. Future investigations should include species from other arthropod taxa than Ixodita, Culicidae and Phlebotominae in order to shed light on the true diversity of arthropod viruses.

  5. Nipah virus matrix protein: expert hacker of cellular machines.

    PubMed

    Watkinson, Ruth E; Lee, Benhur

    2016-08-01

    Nipah virus (NiV, Henipavirus) is a highly lethal emergent zoonotic paramyxovirus responsible for repeated human outbreaks of encephalitis in South East Asia. There are no approved vaccines or treatments, thus improved understanding of NiV biology is imperative. NiV matrix protein recruits a plethora of cellular machinery to scaffold and coordinate virion budding. Intriguingly, matrix also hijacks cellular trafficking and ubiquitination pathways to facilitate transient nuclear localization. While the biological significance of matrix nuclear localization for an otherwise cytoplasmic virus remains enigmatic, the molecular details have begun to be characterized, and are conserved among matrix proteins from divergent paramyxoviruses. Matrix protein appropriation of cellular machinery will be discussed in terms of its early nuclear targeting and later role in virion assembly. PMID:27350027

  6. Nipah virus matrix protein: expert hacker of cellular machines.

    PubMed

    Watkinson, Ruth E; Lee, Benhur

    2016-08-01

    Nipah virus (NiV, Henipavirus) is a highly lethal emergent zoonotic paramyxovirus responsible for repeated human outbreaks of encephalitis in South East Asia. There are no approved vaccines or treatments, thus improved understanding of NiV biology is imperative. NiV matrix protein recruits a plethora of cellular machinery to scaffold and coordinate virion budding. Intriguingly, matrix also hijacks cellular trafficking and ubiquitination pathways to facilitate transient nuclear localization. While the biological significance of matrix nuclear localization for an otherwise cytoplasmic virus remains enigmatic, the molecular details have begun to be characterized, and are conserved among matrix proteins from divergent paramyxoviruses. Matrix protein appropriation of cellular machinery will be discussed in terms of its early nuclear targeting and later role in virion assembly.

  7. RIG-I Signaling Is Essential for Influenza B Virus-Induced Rapid Interferon Gene Expression

    PubMed Central

    Österlund, Pamela; Westenius, Veera; Latvala, Sinikka; Diamond, Michael S.; Gale, Michael; Julkunen, Ilkka

    2015-01-01

    ABSTRACT Influenza B virus causes annual epidemics and, along with influenza A virus, accounts for substantial disease and economic burden throughout the world. Influenza B virus infects only humans and some marine mammals and is not responsible for pandemics, possibly due to a very low frequency of reassortment and a lower evolutionary rate than that of influenza A virus. Influenza B virus has been less studied than influenza A virus, and thus, a comparison of influenza A and B virus infection mechanisms may provide new insight into virus-host interactions. Here we analyzed the early events in influenza B virus infection and interferon (IFN) gene expression in human monocyte-derived macrophages and dendritic cells. We show that influenza B virus induces IFN regulatory factor 3 (IRF3) activation and IFN-λ1 gene expression with faster kinetics than does influenza A virus, without a requirement for viral protein synthesis or replication. Influenza B virus-induced activation of IRF3 required the fusion of viral and endosomal membranes, and nuclear accumulation of IRF3 and viral NP occurred concurrently. In comparison, immediate early IRF3 activation was not observed in influenza A virus-infected macrophages. Experiments with RIG-I-, MDA5-, and RIG-I/MDA5-deficient mouse fibroblasts showed that RIG-I is the critical pattern recognition receptor needed for the influenza B virus-induced activation of IRF3. Our results show that innate immune mechanisms are activated immediately after influenza B virus entry through the endocytic pathway, whereas influenza A virus avoids early IRF3 activation and IFN gene induction. IMPORTANCE Recently, a great deal of interest has been paid to identifying the ligands for RIG-I under conditions of natural infection, as many previous studies have been based on transfection of cells with different types of viral or synthetic RNA structures. We shed light on this question by analyzing the earliest step in innate immune recognition of

  8. Realms of the Viruses Online

    ERIC Educational Resources Information Center

    Liu, Dennis

    2007-01-01

    Viruses have evolved strategies for infecting all taxa, but most viruses are highly specific about their cellular host. In humans, viruses cause diverse diseases, from chronic but benign warts, to acute and deadly hemorrhagic fever. Viruses have entertaining names like Zucchini Yellow Mosaic, Semliki Forest, Coxsackie, and the original terminator,…

  9. Tembusu Virus in Ducks, China

    PubMed Central

    Cao, Zhenzhen; Zhang, Cun; Liu, Yuehuan; Ye, Weicheng; Han, Jingwen; Ma, Guoming; Zhang, Dongdong; Xu, Feng; Gao, Xuhui; Tang, Yi; Shi, Shaohua; Wan, Chunhe; Zhang, Chen; He, Bin; Yang, Mengjie; Lu, Xinhao; Huang, Yu; Diao, Youxiang; Ma, Xuejun

    2011-01-01

    In China in 2010, a disease outbreak in egg-laying ducks was associated with a flavivirus. The virus was isolated and partially sequenced. The isolate exhibited 87%–91% identity with strains of Tembusu virus, a mosquito-borne flavivirus of the Ntaya virus group. These findings demonstrate emergence of Tembusu virus in ducks. PMID:22000358

  10. Postmortem Stability of Ebola Virus

    PubMed Central

    Prescott, Joseph; Bushmaker, Trenton; Fischer, Robert; Miazgowicz, Kerri; Judson, Seth

    2015-01-01

    The ongoing Ebola virus outbreak in West Africa has highlighted questions regarding stability of the virus and detection of RNA from corpses. We used Ebola virus–infected macaques to model humans who died of Ebola virus disease. Viable virus was isolated <7 days posteuthanasia; viral RNA was detectable for 10 weeks. PMID:25897646

  11. Virus-PEDOT Biocomposite Films

    PubMed Central

    Donavan, Keith C.; Arter, Jessica A.

    2012-01-01

    Virus-poly(3,4-ethylenedioxythiophene) (virus-PEDOT) biocomposite films are prepared by electropolymerizing 3,4-ethylenedioxythiophene (EDOT) in aqueous electrolytes containing 12 mM LiClO4 and the bacteriophage M13. The concentration of virus in these solutions, [virus]soln, is varied from 3 nM to 15 nM. A quartz crystal microbalance is used to directly measure the total mass of the biocomposite film during its electrodeposition. In combination with a measurement of the electrodeposition charge, the mass of the virus incorporated into the film is calculated. These data show that concentration of the M13 within the electropolymerized film, [virus]film, increases linearly with [virus]soln. The incorporation of virus particles into the PEDOT film from solution is efficient, resulting in a concentration ratio: [virus]film:[virus]soln ≈450. Virus incorporation into the PEDOT causes roughening of the film topography that is observed using scanning electron microscopy and atomic force microscopy (AFM). The electrical conductivity of the virus-PEDOT film, measured perpendicular to the plane of the film using conductive tip AFM, decreases linearly with virus loading, from 270 μS/cm for pure PE-DOT films to 50 μS/cm for films containing 100 μM virus. The presence on the virus surface of displayed affinity peptides did not significantly influence the efficiency of incorporation into virus-PEDOT biocomposite films. PMID:22856875

  12. A Virus in Turbo Pascal.

    ERIC Educational Resources Information Center

    Teleky, Heidi Ann; And Others

    1993-01-01

    Addresses why the authors feel it is not inappropriate to teach about viruses in the how-to, hands-on fashion. Identifies the special features of Turbo Pascal that have to be used for the creation of an effective virus. Defines virus, derives its structure, and from this structure is derived the implemented virus. (PR)

  13. Computer Viruses: Pathology and Detection.

    ERIC Educational Resources Information Center

    Maxwell, John R.; Lamon, William E.

    1992-01-01

    Explains how computer viruses were originally created, how a computer can become infected by a virus, how viruses operate, symptoms that indicate a computer is infected, how to detect and remove viruses, and how to prevent a reinfection. A sidebar lists eight antivirus resources. (four references) (LRW)

  14. Nuclear waste

    SciTech Connect

    Not Available

    1988-05-01

    This paper discusses how, as part of the Department of Energy's implementation of the Nuclear Waste Policy Act of 1982, DOE is required to investigate a site at Yucca Mountain, Nevada and, if it determines that the site is suitable, recommend to the President its selection for a nuclear waste repository. The Nuclear Regulatory Commission, in considering development of the plan, issued five objections, one of which is DOE's failure to recognize the range of alternative conceptual models of the Yucca Mountain site that can be supported by the limited existing technical data. At the end of the quarter DOE directed its project offices in Washington and Texas to begin orderly phase-out of all site-specific repository activities. Costs for this phase-out are $53 million for the Deaf Smith site and $85 million for the Hanford site.

  15. Nuclear scales

    SciTech Connect

    Friar, J.L.

    1998-12-01

    Nuclear scales are discussed from the nuclear physics viewpoint. The conventional nuclear potential is characterized as a black box that interpolates nucleon-nucleon (NN) data, while being constrained by the best possible theoretical input. The latter consists of the longer-range parts of the NN force (e.g., OPEP, TPEP, the {pi}-{gamma} force), which can be calculated using chiral perturbation theory and gauged using modern phase-shift analyses. The shorter-range parts of the force are effectively parameterized by moments of the interaction that are independent of the details of the force model, in analogy to chiral perturbation theory. Results of GFMC calculations in light nuclei are interpreted in terms of fundamental scales, which are in good agreement with expectations from chiral effective field theories. Problems with spin-orbit-type observables are noted.

  16. Viruses of Chelonia.

    PubMed

    Ahne, W

    1993-02-01

    Viruses occurring in turtles and tortoises are hetergeneous but according to ecologic characteristics and pathogenic properties they can be divided in two major groups: 1. Arboviruses (toga-, flavi-, rhabdo- and bunyaviruses) transmitted by arthropods cause severe diseases in homoiothermic vertebrates. The viruses are of great epidemiological interest in human and veterinary medicine. Chelonia and other reptiles infected by bites of vectors e.g. Aedes, Anopheles, Culex develop cyclic viremia without injury. The ectothermic animals maintain inapparent arbovirus infections during hibernation and they play role as reservoirs for these viruses. 2. Viruses of Chelonia origin (papova-, herpes-, irido- and paramyxoviruses) associated with diseases of infected turtles and tortoises have been described frequently during the last 20 years. Several viruses or virus-like particles could be demonstrated in affected reptiles mainly by electron microscopy. Especially herpesviruses seem to attack Chelonia and epizootics due to infections with these viruses were reported in several reptiles in collections. However, the etiological role of the agents detected is not well documented yet. PMID:8456570

  17. Ocular Tropism of Respiratory Viruses

    PubMed Central

    Rota, Paul A.; Tumpey, Terrence M.

    2013-01-01

    SUMMARY Respiratory viruses (including adenovirus, influenza virus, respiratory syncytial virus, coronavirus, and rhinovirus) cause a broad spectrum of disease in humans, ranging from mild influenza-like symptoms to acute respiratory failure. While species D adenoviruses and subtype H7 influenza viruses are known to possess an ocular tropism, documented human ocular disease has been reported following infection with all principal respiratory viruses. In this review, we describe the anatomical proximity and cellular receptor distribution between ocular and respiratory tissues. All major respiratory viruses and their association with human ocular disease are discussed. Research utilizing in vitro and in vivo models to study the ability of respiratory viruses to use the eye as a portal of entry as well as a primary site of virus replication is highlighted. Identification of shared receptor-binding preferences, host responses, and laboratory modeling protocols among these viruses provides a needed bridge between clinical and laboratory studies of virus tropism. PMID:23471620

  18. Nuclear pursuits

    SciTech Connect

    Not Available

    1993-05-01

    This table lists quantities of warheads (in stockpile, peak number per year, total number built, number of known test explosions), weapon development milestones (developers of the atomic bomb and hydrogen bomb, date of first operational ICBM, first nuclear-powered naval SSN in service, first MIRVed missile deployed), and testing milestones (first fission test, type of boosted fission weapon, multistage thermonuclear test, number of months from fission bomb to multistage thermonuclear bomb, etc.), and nuclear infrastructure (assembly plants, plutonium production reactors, uranium enrichment plants, etc.). Countries included in the tally are the United States, Soviet Union, Britain, France, and China.

  19. Nuclear power: Fourth edition

    SciTech Connect

    Deutsch, R.W.

    1986-01-01

    This book describes the basics of nuclear power generation, explaining both the benefits and the real and imagined risks of nuclear power. It includes a discussion of the Three Mile Island accident and its effects. Nuclear Power has been used in the public information programs of more than 100 utilities. The contents discussed are: Nuclear Power and People; Why Nuclear Power. Electricity produced by coal; Electricity produced by nuclear fuel; Nuclear plant sites in the United States; Short History of Commercial Nuclear Power; U.S. nuclear submarines, Regulation of Nuclear Power Plants; Licensing process, Nuclear Power Plant Operator Training; Nuclear power plant simulator, Are Nuclear Plants Safe.; Containment structure, Nuclear Power Plant Insurance; Is Radiation Dangerous.; Man-made radiation, What is Nuclear Fuel.; Fuel cycle for commercial nuclear power plants; Warm Water Discharge; Cooling tower; Protection of Radioactive Materials; Plutonium and Proliferation; Disposal of Radioactive Wastes; Are Alternate Energy Sources Available.; Nuclear Opposition; and Nuclear Power in the Future.

  20. Herpes simplex virus 1 induces de novo phospholipid synthesis

    SciTech Connect

    Sutter, Esther; Oliveira, Anna Paula de; Tobler, Kurt; Schraner, Elisabeth M.; Sonda, Sabrina; Kaech, Andres; Lucas, Miriam S.; Ackermann, Mathias; Wild, Peter

    2012-08-01

    Herpes simplex virus type 1 capsids bud at nuclear membranes and Golgi membranes acquiring an envelope composed of phospholipids. Hence, we measured incorporation of phospholipid precursors into these membranes, and quantified changes in size of cellular compartments by morphometric analysis. Incorporation of [{sup 3}H]-choline into both nuclear and cytoplasmic membranes was significantly enhanced upon infection. [{sup 3}H]-choline was also part of isolated virions even grown in the presence of brefeldin A. Nuclei expanded early in infection. The Golgi complex and vacuoles increased substantially whereas the endoplasmic reticulum enlarged only temporarily. The data suggest that HSV-1 stimulates phospholipid synthesis, and that de novo synthesized phospholipids are inserted into nuclear and cytoplasmic membranes to i) maintain membrane integrity in the course of nuclear and cellular expansion, ii) to supply membrane constituents for envelopment of capsids by budding at nuclear membranes and Golgi membranes, and iii) to provide membranes for formation of transport vacuoles.

  1. Genome of horsepox virus.

    PubMed

    Tulman, E R; Delhon, G; Afonso, C L; Lu, Z; Zsak, L; Sandybaev, N T; Kerembekova, U Z; Zaitsev, V L; Kutish, G F; Rock, D L

    2006-09-01

    Here we present the genomic sequence of horsepox virus (HSPV) isolate MNR-76, an orthopoxvirus (OPV) isolated in 1976 from diseased Mongolian horses. The 212-kbp genome contained 7.5-kbp inverted terminal repeats and lacked extensive terminal tandem repetition. HSPV contained 236 open reading frames (ORFs) with similarity to those in other OPVs, with those in the central 100-kbp region most conserved relative to other OPVs. Phylogenetic analysis of the conserved region indicated that HSPV is closely related to sequenced isolates of vaccinia virus (VACV) and rabbitpox virus, clearly grouping together these VACV-like viruses. Fifty-four HSPV ORFs likely represented fragments of 25 orthologous OPV genes, including in the central region the only known fragmented form of an OPV ribonucleotide reductase large subunit gene. In terminal genomic regions, HSPV lacked full-length homologues of genes variably fragmented in other VACV-like viruses but was unique in fragmentation of the homologue of VACV strain Copenhagen B6R, a gene intact in other known VACV-like viruses. Notably, HSPV contained in terminal genomic regions 17 kbp of OPV-like sequence absent in known VACV-like viruses, including fragments of genes intact in other OPVs and approximately 1.4 kb of sequence present only in cowpox virus (CPXV). HSPV also contained seven full-length genes fragmented or missing in other VACV-like viruses, including intact homologues of the CPXV strain GRI-90 D2L/I4R CrmB and D13L CD30-like tumor necrosis factor receptors, D3L/I3R and C1L ankyrin repeat proteins, B19R kelch-like protein, D7L BTB/POZ domain protein, and B22R variola virus B22R-like protein. These results indicated that HSPV contains unique genomic features likely contributing to a unique virulence/host range phenotype. They also indicated that while closely related to known VACV-like viruses, HSPV contains additional, potentially ancestral sequences absent in other VACV-like viruses.

  2. Viruses in reptiles

    PubMed Central

    2011-01-01

    The etiology of reptilian viral diseases can be attributed to a wide range of viruses occurring across different genera and families. Thirty to forty years ago, studies of viruses in reptiles focused mainly on the zoonotic potential of arboviruses in reptiles and much effort went into surveys and challenge trials of a range of reptiles with eastern and western equine encephalitis as well as Japanese encephalitis viruses. In the past decade, outbreaks of infection with West Nile virus in human populations and in farmed alligators in the USA has seen the research emphasis placed on the issue of reptiles, particularly crocodiles and alligators, being susceptible to, and reservoirs for, this serious zoonotic disease. Although there are many recognised reptilian viruses, the evidence for those being primary pathogens is relatively limited. Transmission studies establishing pathogenicity and cofactors are likewise scarce, possibly due to the relatively low commercial importance of reptiles, difficulties with the availability of animals and permits for statistically sound experiments, difficulties with housing of reptiles in an experimental setting or the inability to propagate some viruses in cell culture to sufficient titres for transmission studies. Viruses as causes of direct loss of threatened species, such as the chelonid fibropapilloma associated herpesvirus and ranaviruses in farmed and wild tortoises and turtles, have re-focused attention back to the characterisation of the viruses as well as diagnosis and pathogenesis in the host itself. 1. Introduction 2. Methods for working with reptilian viruses 3. Reptilian viruses described by virus families 3.1. Herpesviridae 3.2. Iridoviridae 3.2.1 Ranavirus 3.2.2 Erythrocytic virus 3.2.3 Iridovirus 3.3. Poxviridae 3.4. Adenoviridae 3.5. Papillomaviridae 3.6. Parvoviridae 3.7. Reoviridae 3.8. Retroviridae and inclusion body disease of Boid snakes 3.9. Arboviruses 3.9.1. Flaviviridae 3.9.2. Togaviridae 3.10. Caliciviridae

  3. [Ebola virus disease].

    PubMed

    Nazimek, Katarzyna; Bociaga-Jasik, Monika; Bryniarski, Krzysztof; Gałas, Aleksander; Garlicki, Aleksander; Gawda, Anna; Gawlik, Grzegorz; Gil, Krzysztof; Kosz-Vnenchak, Magdalena; Mrozek-Budzyn, Dorota; Olszanecki, Rafał; Piatek, Anna; Zawilińska, Barbara; Marcinkiewicz, Janusz

    2014-01-01

    Ebola is one of the most virulent zoonotic RNA viruses causing in humans haemorrhagic fever with fatality ratio reaching 90%. During the outbreak of 2014 the number of deaths exceeded 8.000. The "imported" cases reported in Western Europe and USA highlighted the extreme risk of Ebola virus spreading outside the African countries. Thus, haemorrhagic fever outbreak is an international epidemiological problem, also due to the lack of approved prevention and therapeutic strategies. The editorial review article briefly summarizes current knowledge on Ebola virus disease epidemiology, etiology, pathogenesis, clinical presentation, diagnosis as well as possible prevention and treatment.

  4. Epstein-Barr virus nuclear protein 3C binds to the N-terminal (NTD) and beta trefoil domains (BTD) of RBP/CSL; Only the NTD interaction is essential for lymphoblastoid cell growth

    SciTech Connect

    Calderwood, Michael A.; Lee, Sungwook; Holthaus, Amy M.; Blacklow, Stephen C.; Kieff, Elliott; Johannsen, Eric

    2011-05-25

    Association of EBV nuclear proteins EBNA2, EBNA3A and EBNA3C with RBP/CSL, is essential for lymphoblastoid cell line (LCL) proliferation. Conserved residues in the EBNA3 homology domain, required for RBP/CSL interaction, lack the W{Phi}P motif that mediates EBNA2 and Notch binding to the RBP/CSL beta-trefoil domain (BTD). We map RBP/CSL interacting residues within EBNA3A(aa128-204) and EBNA3C(aa211-233). The EBNA3A results are consistent with an earlier report (aa125-222), but the EBNA3C domain is unexpectedly small and includes a 'WTP' sequence. This EBNA3C WTP motif confers RBP/CSL binding in vitro, in yeast, and in mammalian cells. Further, an EBNA3C WTP {yields} STP(W227S) mutation impaired BTD binding whereas EBNA3 homology domain mutations disrupted RBP/CSL N-terminal domain (NTD) binding. WTP was not essential for EBNA3C repression of EBNA2 in reporter assays or for maintenance of LCL growth. Our results indicate that EBNA3 proteins interact with multiple RBP/CSL domains, but only NTD interactions are required for LCL growth.

  5. Nuclear medicine

    SciTech Connect

    Wagner, H.N. Jr.

    1986-10-17

    In 1985 and 1986 nuclear medicine became more and more oriented toward in vov chemistry, chiefly as a result of advances in positron emission tomography (PET). The most important trend was the extension of PET technology into the care of patients with brain tumors, epilepsy, and heart disease. A second trend was the increasing use of single-photon emission computed tomography (SPECT).

  6. NUCLEAR REACTOR

    DOEpatents

    Treshow, M.

    1961-09-01

    A boiling-water nuclear reactor is described wherein control is effected by varying the moderator-to-fuel ratio in the reactor core. This is accomplished by providing control tubes containing a liquid control moderator in the reactor core and providing means for varying the amount of control moderatcr within the control tubes.

  7. Nuclear Science.

    ERIC Educational Resources Information Center

    Pennsylvania State Dept. of Education, Harrisburg. Bureau of Curriculum Services.

    This document is a report on a course in nuclear science for the high school curriculum. The course is designed to provide a basic but comprehensive understanding of the atom in the light of modern knowledge, and to show how people attempt to harness the tremendous energy liberated through fission and fusion reactions. The course crosses what are…

  8. Nuclear Misinformation

    ERIC Educational Resources Information Center

    Ford, Daniel F.; Kendall, Henry W.

    1975-01-01

    Many scientists feel that research into nuclear safety has been diverted or distorted, and the results of the research concealed or inaccurately reported on a large number of occasions. Of particular concern have been the emergency cooling systems which have not, as yet, been adequately tested. (Author/MA)

  9. Nuclear explosions

    SciTech Connect

    Broyles, A.A.

    1982-07-01

    A summary of the physics of a nuclear bomb explosion and its effects on human beings is presented at the level of a sophomore general physics course without calculus. It is designed to supplement a standard text for such a course and problems are included.

  10. Nuclear energy.

    PubMed

    Wilson, Peter D

    2010-01-01

    The technical principles and practices of the civil nuclear industry are described with particular reference to fission and its products, natural and artificial radioactivity elements principally concerned and their relationships, main types of reactor, safety issues, the fuel cycle, waste management, issues related to weapon proliferation, environmental considerations and possible future developments.

  11. Nuclear Terrorism.

    SciTech Connect

    Hecker, Siegfried S.

    2001-01-01

    As pointed out by several speakers, the level of violence and destruction in terrorist attacks has increased significantly during the past decade. Fortunately, few have involved weapons of mass destruction, and none have achieved mass casualties. The Aum Shinrikyo release of lethal nerve agent, sarin, in the Tokyo subway on March 20, 1995 clearly broke new ground by crossing the threshold in attempting mass casualties with chemical weapons. However, of all weapons of mass destruction, nuclear weapons still represent the most frightening threat to humankind. Nuclear weapons possess an enormous destructive force. The immediacy and scale of destruction are unmatched. In addition to destruction, terrorism also aims to create fear among the public and governments. Here also, nuclear weapons are unmatched. The public's fear of nuclear weapons or, for that matter, of all radioactivity is intense. To some extent, this fear arises from a sense of unlimited vulnerability. That is, radioactivity is seen as unbounded in three dimensions - distance, it is viewed as having unlimited reach; quantity, it is viewed as having deadly consequences in the smallest doses (the public is often told - incorrectly, of course - that one atom of plutonium will kill); and time, if it does not kill you immediately, then it will cause cancer decades hence.

  12. Hepatitis B virus (image)

    MedlinePlus

    Hepatitis B is also known as serum hepatitis and is spread through blood and sexual contact. It is ... population. This photograph is an electronmicroscopic image of hepatitis B virus particles. (Image courtesy of the Centers for ...

  13. [Zika, a neurotropic virus?].

    PubMed

    Del Carpio-Orantes, Luis

    2016-01-01

    In this paper, the neurotropism potential Zika virus is discussed, by comparison with viruses both RNA and DNA are neurotropic known, also it is said that compared with the new viruses that have affected the Americas, as the chikungunya, Zika has shown great affinity by brain tissue, manifested by a high incidence of acute neurological conditions, such as Guillain-Barré syndrome, among others, as well as the reported incidence of microcephaly that is abnormally high compared with the previous incidence, which, in a stillborn subject necropsied significant alterations demonstrated in brain tissue, identifying viral material and live virus in the fetoplacental complex, and demonstrating the impact both white matter and gray matter as well as basal ganglia, corpus callosum, ventricles and spinal cord, which could explain the microcephaly that concerns him. Although not a direct cause-effect relationship is demonstrated, however current evidence supports that relationship, hoping to be supported scientifically.

  14. The dengue viruses.

    PubMed Central

    Henchal, E A; Putnak, J R

    1990-01-01

    Dengue, a major public health problem throughout subtropical and tropical regions, is an acute infectious disease characterized by biphasic fever, headache, pain in various parts of the body, prostration, rash, lymphadenopathy, and leukopenia. In more severe or complicated dengue, patients present with a severe febrile illness characterized by abnormalities of hemostasis and increased vascular permeability, which in some instances results in a hypovolemic shock. Four distinct serotypes of the dengue virus (dengue-1, dengue-2, dengue-3, and dengue-4) exist, with numerous virus strains found worldwide. Molecular cloning methods have led to a greater understanding of the structure of the RNA genome and definition of virus-specific structural and nonstructural proteins. Progress towards producing safe, effective dengue virus vaccines, a goal for over 45 years, has been made. Images PMID:2224837

  15. VIRUS instrument enclosures

    NASA Astrophysics Data System (ADS)

    Prochaska, T.; Allen, R.; Mondrik, N.; Rheault, J. P.; Sauseda, M.; Boster, E.; James, M.; Rodriguez-Patino, M.; Torres, G.; Ham, J.; Cook, E.; Baker, D.; DePoy, Darren L.; Marshall, Jennifer L.; Hill, G. J.; Perry, D.; Savage, R. D.; Good, J. M.; Vattiat, Brian L.

    2014-08-01

    The Visible Integral-Field Replicable Unit Spectrograph (VIRUS) instrument will be installed at the Hobby-Eberly Telescope† in the near future. The instrument will be housed in two enclosures that are mounted adjacent to the telescope, via the VIRUS Support Structure (VSS). We have designed the enclosures to support and protect the instrument, to enable servicing of the instrument, and to cool the instrument appropriately while not adversely affecting the dome environment. The system uses simple HVAC air handling techniques in conjunction with thermoelectric and standard glycol heat exchangers to provide efficient heat removal. The enclosures also provide power and data transfer to and from each VIRUS unit, liquid nitrogen cooling to the detectors, and environmental monitoring of the instrument and dome environments. In this paper, we describe the design and fabrication of the VIRUS enclosures and their subsystems.

  16. Respiratory syncytial virus (RSV)

    MedlinePlus

    ... RSV often spreads quickly in crowded households and day care centers. The virus can live for a half ... The following increase the risk for RSV: Attending day care Being near tobacco smoke Having school-aged brothers ...

  17. West Nile Virus

    MedlinePlus

    ... to human beings through their bites. Credit: CDC Biology, Genetics, & Clinical Research NIAID conducts and funds basic and clinical research on WNV biology and viral structure, ways the virus causes human ...

  18. What's West Nile Virus?

    MedlinePlus

    ... is caused by a bite from an infected mosquito that's already carrying the virus, but it's important ... the risk of being bitten by an infected mosquito is greatest from July to early September. But ...

  19. Avoiding Computer Viruses.

    ERIC Educational Resources Information Center

    Rowe, Joyce; And Others

    1989-01-01

    The threat of computer sabotage is a real concern to business teachers and others responsible for academic computer facilities. Teachers can minimize the possibility. Eight suggestions for avoiding computer viruses are given. (JOW)

  20. West Nile Virus

    MedlinePlus

    ... appeared in the United States in 1999. Infected mosquitoes spread the virus that causes it. People who ... barrels Stay indoors between dusk and dawn, when mosquitoes are most active Use screens on windows to ...

  1. The dengue viruses.

    PubMed

    Henchal, E A; Putnak, J R

    1990-10-01

    Dengue, a major public health problem throughout subtropical and tropical regions, is an acute infectious disease characterized by biphasic fever, headache, pain in various parts of the body, prostration, rash, lymphadenopathy, and leukopenia. In more severe or complicated dengue, patients present with a severe febrile illness characterized by abnormalities of hemostasis and increased vascular permeability, which in some instances results in a hypovolemic shock. Four distinct serotypes of the dengue virus (dengue-1, dengue-2, dengue-3, and dengue-4) exist, with numerous virus strains found worldwide. Molecular cloning methods have led to a greater understanding of the structure of the RNA genome and definition of virus-specific structural and nonstructural proteins. Progress towards producing safe, effective dengue virus vaccines, a goal for over 45 years, has been made. PMID:2224837

  2. Nucleolus: the fascinating nuclear body

    PubMed Central

    Sirri, Valentina; Urcuqui-Inchima, Silvio; Roussel, Pascal

    2007-01-01

    Nucleoli are the prominent contrasted structures of the cell nucleus. In the nucleolus, ribosomal RNAs are synthesized, processed and assembled with ribosomal proteins. RNA polymerase I synthesizes the ribosomal RNAs and this activity is cell cycle regulated. The nucleolus reveals the functional organization of the nucleus in which the compartmentation of the different steps of ribosome biogenesis is observed whereas the nucleolar machineries are in permanent exchange with the nucleoplasm and other nuclear bodies. After mitosis, nucleolar assembly is a time and space regulated process controlled by the cell cycle. In addition, by generating a large volume in the nucleus with apparently no RNA polymerase II activity, the nucleolus creates a domain of retention/sequestration of molecules normally active outside the nucleolus. Viruses interact with the nucleolus and recruit nucleolar proteins to facilitate virus replication. The nucleolus is also a sensor of stress due to the redistribution of the ribosomal proteins in the nucleoplasm by nucleolus disruption. The nucleolus plays several crucial functions in the nucleus: in addition to its function as ribosome factory of the cells it is a multifunctional nuclear domain, and nucleolar activity is linked with several pathologies. Perspectives on the evolution of this research area are proposed. PMID:18046571

  3. Vaccinia virus protein C4 inhibits NF-κB activation and promotes virus virulence.

    PubMed

    Ember, Stuart W J; Ren, Hongwei; Ferguson, Brian J; Smith, Geoffrey L

    2012-10-01

    Vaccinia virus (VACV) strain Western Reserve protein C4 has been characterized and its function and contribution to virus virulence assessed. Bioinformatic analysis showed that C4 is conserved in six orthopoxvirus species and shares 43 % amino acid identity with VACV protein C16, a known virulence factor. A recombinant VACV expressing a C-terminally tagged version of C4 showed that, like C16, this 37 kDa protein is expressed early during infection and localizes to both the cytoplasm and the nucleus. Functional assays using a firefly luciferase reporter plasmid under the control of a nuclear factor kappa B (NF-κB)-dependent promoter demonstrated that C4 inhibits NF-κB activation at, or downstream of, the inhibitor of kappa kinase (IKK) complex. Consistent with this, C4 inhibited interleukin-1β-induced translocation of p65 into the nucleus. A VACV lacking the C4L gene (vΔC4) showed no significant differences from wild-type virus in growth kinetics or spread in cell culture, but had reduced virulence in a murine intranasal model of infection. vΔC4-infected mice exhibited fewer symptoms, lost less weight and recovered 7 days earlier than animals infected with control viruses expressing C4. Furthermore, bronchoalveolar lavage fluid from vΔC4-infected mice had increased cell numbers at day 5 post-infection, which correlated with reduced lung virus titres from this time onward. C4 represents the ninth VACV protein to inhibit NF-κB activation and remarkably, in every case examined, loss of each protein individually caused an alteration in virus virulence, despite the presence of other NF-κB inhibitors. PMID:22791606

  4. Vaccinia virus protein C4 inhibits NF-κB activation and promotes virus virulence.

    PubMed

    Ember, Stuart W J; Ren, Hongwei; Ferguson, Brian J; Smith, Geoffrey L

    2012-10-01

    Vaccinia virus (VACV) strain Western Reserve protein C4 has been characterized and its function and contribution to virus virulence assessed. Bioinformatic analysis showed that C4 is conserved in six orthopoxvirus species and shares 43 % amino acid identity with VACV protein C16, a known virulence factor. A recombinant VACV expressing a C-terminally tagged version of C4 showed that, like C16, this 37 kDa protein is expressed early during infection and localizes to both the cytoplasm and the nucleus. Functional assays using a firefly luciferase reporter plasmid under the control of a nuclear factor kappa B (NF-κB)-dependent promoter demonstrated that C4 inhibits NF-κB activation at, or downstream of, the inhibitor of kappa kinase (IKK) complex. Consistent with this, C4 inhibited interleukin-1β-induced translocation of p65 into the nucleus. A VACV lacking the C4L gene (vΔC4) showed no significant differences from wild-type virus in growth kinetics or spread in cell culture, but had reduced virulence in a murine intranasal model of infection. vΔC4-infected mice exhibited fewer symptoms, lost less weight and recovered 7 days earlier than animals infected with control viruses expressing C4. Furthermore, bronchoalveolar lavage fluid from vΔC4-infected mice had increased cell numbers at day 5 post-infection, which correlated with reduced lung virus titres from this time onward. C4 represents the ninth VACV protein to inhibit NF-κB activation and remarkably, in every case examined, loss of each protein individually caused an alteration in virus virulence, despite the presence of other NF-κB inhibitors.

  5. Nuclear IRS-1 and Cancer

    PubMed Central

    Reiss, Krzysztof; Valle, Luis Del; Lassak, Adam; Trojanek, Joanna

    2011-01-01

    The family of insulin receptor substrates (IRS) consists of four proteins (IRS-1 - IRS-4), which were initially characterized as typical cytosolic adaptor proteins involved in insulin receptor (IR) and insulin-like growth factor I receptor (IGF-IR) signaling. The first cloned and characterized member of the IRS family, IRS-1, has predicted molecular weight of 132 kDa, however, as a result of its extensive serine phosphorylation it separates on a SDS gel as a band of approximately 160–185 kDa. In addition to its metabolic and growth-promoting functions, IRS-1 is also suspected to play a role in malignant transformation. The mechanism by which IRS-1 supports tumor growth is not fully understood, and the argument that IRS-1 merely amplifies the signal from the IGF-1R and/or IR requires further investigation. Almost a decade ago, we reported the presence of nuclear IRS-1 in medulloblastoma clinical samples, which express viral oncoprotein, large T-antigen of human polyomavirus JC (JCV T-antigen). This first demonstration of nuclear IRS-1 was confirmed in several other laboratories. The nuclear IRS-1 was also detected by cells expressing the SV40 T-antigen, v-Src, in immortalized fibroblasts stimulated with IGF-I, in hepatocytes, 32D cells, and in an osteosarcoma cell line. More recently, nuclear IRS-1 was detected in breast cancer cells in association with estrogen receptor alpha (ERα), and in JC virus negative medulloblastoma cells expressing ERβ, further implicating nuclear IRS-1 in cellular transformation. Here, we discuss how nuclear IRS-1 acting on DNA repair fidelity, transcriptional activity, and cell growth can support tumor development and progression. PMID:22454254

  6. Viruses and Multiple Sclerosis

    PubMed Central

    Virtanen, Jussi Oskari; Jacobson, Steve

    2016-01-01

    Multiple sclerosis (MS) is a heterogeneous disease that develops as an interplay between the immune system and environmental stimuli in genetically susceptible individuals. There is increasing evidence that viruses may play a role in MS pathogenesis acting as these environmental triggers. However, it is not known if any single virus is causal, or rather several viruses can act as triggers in disease development. Here, we review the association of different viruses to MS with an emphasis on two herpesviruses, Epstein-Barr virus (EBV) and human herpesvirus 6 (HHV-6). These two agents have generated the most impact during recent years as possible co-factors in MS disease development. The strongest argument for association of EBV with MS comes from the link between symptomatic infectious mononucleosis and MS and from seroepidemiological studies. In contrast to EBV, HHV-6 has been found significantly more often in MS plaques than in MS normal appearing white matter or non-MS brains and HHV-6 re-activation has been reported during MS clinical relapses. In this review we also suggest new strategies, including the development of new infectious animal models of MS and antiviral MS clinical trials, to elucidate roles of different viruses in the pathogenesis of this disease. Furthermore, we introduce the idea of using unbiased sequence-independent pathogen discovery methodologies, such as next generation sequencing, to study MS brain tissue or body fluids for detection of known viral sequences or potential novel viral agents. PMID:22583435

  7. Human Influenza Virus Infections.

    PubMed

    Peteranderl, Christin; Herold, Susanne; Schmoldt, Carole

    2016-08-01

    Seasonal and pandemic influenza are the two faces of respiratory infections caused by influenza viruses in humans. As seasonal influenza occurs on an annual basis, the circulating virus strains are closely monitored and a yearly updated vaccination is provided, especially to identified risk populations. Nonetheless, influenza virus infection may result in pneumonia and acute respiratory failure, frequently complicated by bacterial coinfection. Pandemics are, in contrary, unexpected rare events related to the emergence of a reassorted human-pathogenic influenza A virus (IAV) strains that often causes increased morbidity and spreads extremely rapidly in the immunologically naive human population, with huge clinical and economic impact. Accordingly, particular efforts are made to advance our knowledge on the disease biology and pathology and recent studies have brought new insights into IAV adaptation mechanisms to the human host, as well as into the key players in disease pathogenesis on the host side. Current antiviral strategies are only efficient at the early stages of the disease and are challenged by the genomic instability of the virus, highlighting the need for novel antiviral therapies targeting the pulmonary host response to improve viral clearance, reduce the risk of bacterial coinfection, and prevent or attenuate acute lung injury. This review article summarizes our current knowledge on the molecular basis of influenza infection and disease progression, the key players in pathogenesis driving severe disease and progression to lung failure, as well as available and envisioned prevention and treatment strategies against influenza virus infection. PMID:27486731

  8. Virus templated metallic nanoparticles

    NASA Astrophysics Data System (ADS)

    Aljabali, Alaa A. A.; Barclay, J. Elaine; Lomonossoff, George P.; Evans, David J.

    2010-12-01

    Plant viruses are considered as nanobuilding blocks that can be used as synthons or templates for novel materials. Cowpea mosaic virus (CPMV) particles have been shown to template the fabrication of metallic nanoparticles by an electroless deposition metallization process. Palladium ions were electrostatically bound to the virus capsid and, when reduced, acted as nucleation sites for the subsequent metal deposition from solution. The method, although simple, produced highly monodisperse metallic nanoparticles with a diameter of ca. <=35 nm. CPMV-templated particles were prepared with cobalt, nickel, iron, platinum, cobalt-platinum and nickel-iron.Plant viruses are considered as nanobuilding blocks that can be used as synthons or templates for novel materials. Cowpea mosaic virus (CPMV) particles have been shown to template the fabrication of metallic nanoparticles by an electroless deposition metallization process. Palladium ions were electrostatically bound to the virus capsid and, when reduced, acted as nucleation sites for the subsequent metal deposition from solution. The method, although simple, produced highly monodisperse metallic nanoparticles with a diameter of ca. <=35 nm. CPMV-templated particles were prepared with cobalt, nickel, iron, platinum, cobalt-platinum and nickel-iron. Electronic supplementary information (ESI) available: Additional experimental detail, agarose gel electrophoresis results, energy dispersive X-ray spectra, ζ-potential measurements, dynamic light scattering data, nanoparticle tracking analysis and an atomic force microscopy image of Ni-CPMV. See DOI: 10.1039/c0nr00525h

  9. Human Influenza Virus Infections.

    PubMed

    Peteranderl, Christin; Herold, Susanne; Schmoldt, Carole

    2016-08-01

    Seasonal and pandemic influenza are the two faces of respiratory infections caused by influenza viruses in humans. As seasonal influenza occurs on an annual basis, the circulating virus strains are closely monitored and a yearly updated vaccination is provided, especially to identified risk populations. Nonetheless, influenza virus infection may result in pneumonia and acute respiratory failure, frequently complicated by bacterial coinfection. Pandemics are, in contrary, unexpected rare events related to the emergence of a reassorted human-pathogenic influenza A virus (IAV) strains that often causes increased morbidity and spreads extremely rapidly in the immunologically naive human population, with huge clinical and economic impact. Accordingly, particular efforts are made to advance our knowledge on the disease biology and pathology and recent studies have brought new insights into IAV adaptation mechanisms to the human host, as well as into the key players in disease pathogenesis on the host side. Current antiviral strategies are only efficient at the early stages of the disease and are challenged by the genomic instability of the virus, highlighting the need for novel antiviral therapies targeting the pulmonary host response to improve viral clearance, reduce the risk of bacterial coinfection, and prevent or attenuate acute lung injury. This review article summarizes our current knowledge on the molecular basis of influenza infection and disease progression, the key players in pathogenesis driving severe disease and progression to lung failure, as well as available and envisioned prevention and treatment strategies against influenza virus infection.

  10. Transmission of influenza A viruses.

    PubMed

    Neumann, Gabriele; Kawaoka, Yoshihiro

    2015-05-01

    Influenza A viruses cause respiratory infections that range from asymptomatic to deadly in humans. Widespread outbreaks (pandemics) are attributable to 'novel' viruses that possess a viral hemagglutinin (HA) gene to which humans lack immunity. After a pandemic, these novel viruses form stable virus lineages in humans and circulate until they are replaced by other novel viruses. The factors and mechanisms that facilitate virus transmission among hosts and the establishment of novel lineages are not completely understood, but the HA and basic polymerase 2 (PB2) proteins are thought to play essential roles in these processes by enabling avian influenza viruses to infect mammals and replicate efficiently in their new host. Here, we summarize our current knowledge of the contributions of HA, PB2, and other viral components to virus transmission and the formation of new virus lineages. PMID:25812763

  11. Transmission of Influenza A Viruses

    PubMed Central

    Neumann, Gabriele; Kawaoka, Yoshihiro

    2015-01-01

    Influenza A viruses cause respiratory infections that range from asymptomatic to deadly in humans. Widespread outbreaks (pandemics) are attributable to ‘novel’ viruses that possess a viral hemagglutinin (HA) gene to which humans lack immunity. After a pandemic, these novel viruses form stable virus lineages in humans and circulate until they are replaced by other novel viruses. The factors and mechanisms that facilitate virus transmission among hosts and the establishment of novel lineages are not completely understood, but the HA and basic polymerase 2 (PB2) proteins are thought to play essential roles in these processes by enabling avian influenza viruses to infect mammals and replicate efficiently in their new host. Here, we summarize our current knowledge of the contributions of HA, PB2, and other viral components to virus transmission and the formation of new virus lineages. PMID:25812763

  12. Smaller Fleas: Viruses of Microorganisms

    PubMed Central

    Hyman, Paul; Abedon, Stephen T.

    2012-01-01

    Life forms can be roughly differentiated into those that are microscopic versus those that are not as well as those that are multicellular and those that, instead, are unicellular. Cellular organisms seem generally able to host viruses, and this propensity carries over to those that are both microscopic and less than truly multicellular. These viruses of microorganisms, or VoMs, in fact exist as the world's most abundant somewhat autonomous genetic entities and include the viruses of domain Bacteria (bacteriophages), the viruses of domain Archaea (archaeal viruses), the viruses of protists, the viruses of microscopic fungi such as yeasts (mycoviruses), and even the viruses of other viruses (satellite viruses). In this paper we provide an introduction to the concept of viruses of microorganisms, a.k.a., viruses of microbes. We provide broad discussion particularly of VoM diversity. VoM diversity currently spans, in total, at least three-dozen virus families. This is roughly ten families per category—bacterial, archaeal, fungal, and protist—with some virus families infecting more than one of these microorganism major taxa. Such estimations, however, will vary with further discovery and taxon assignment and also are dependent upon what forms of life one includes among microorganisms. PMID:24278736

  13. Smaller fleas: viruses of microorganisms.

    PubMed

    Hyman, Paul; Abedon, Stephen T

    2012-01-01

    Life forms can be roughly differentiated into those that are microscopic versus those that are not as well as those that are multicellular and those that, instead, are unicellular. Cellular organisms seem generally able to host viruses, and this propensity carries over to those that are both microscopic and less than truly multicellular. These viruses of microorganisms, or VoMs, in fact exist as the world's most abundant somewhat autonomous genetic entities and include the viruses of domain Bacteria (bacteriophages), the viruses of domain Archaea (archaeal viruses), the viruses of protists, the viruses of microscopic fungi such as yeasts (mycoviruses), and even the viruses of other viruses (satellite viruses). In this paper we provide an introduction to the concept of viruses of microorganisms, a.k.a., viruses of microbes. We provide broad discussion particularly of VoM diversity. VoM diversity currently spans, in total, at least three-dozen virus families. This is roughly ten families per category-bacterial, archaeal, fungal, and protist-with some virus families infecting more than one of these microorganism major taxa. Such estimations, however, will vary with further discovery and taxon assignment and also are dependent upon what forms of life one includes among microorganisms.

  14. Nuclear politics

    NASA Astrophysics Data System (ADS)

    Ranson, John

    2009-04-01

    The sentiments expressed by Sidney Drell in his forum article "The nuclear threat: a new start" (February pp16-17) are laudable, but it was disappointing to find this almost entirely political story in isolation. The article, which outlined the prospects for reducing weapons stockpiles under the new US administration, would have been more pertinent as an introduction to a series describing the technology used in detecting nuclear-testing activity. It would have been interesting to discuss the specific equipment and methods used, together with the analysis and correlation techniques - along with an indication of how sensitive and reliable they are (if the information is not classified). It is far easier to detect an explosive event than it is to detect and quantify weapons stores, which is a key factor for any negotiated solution. Apart from deductions based on actual inspection and satellite surveillance, are there other techniques that can be applied to this issue?

  15. NUCLEAR REACTORS

    DOEpatents

    Long, E.; Ashby, J.W.

    1958-09-16

    ABS>A graphite moderator structure is presented for a nuclear reactor compriscd of an assembly of similarly orientated prismatic graphite blocks arranged on spaced longitudinal axes lying in common planes wherein the planes of the walls of the blocks are positioned so as to be twisted reintive to the planes of said axes so thatthe unlmpeded dtrect paths in direction wholly across the walls of the blocks are limited to the width of the blocks plus spacing between the blocks.

  16. NUCLEAR REACTOR

    DOEpatents

    Anderson, C.R.

    1962-07-24

    A fluidized bed nuclear reactor and a method of operating such a reactor are described. In the design means are provided for flowing a liquid moderator upwardly through the center of a bed of pellets of a nentron-fissionable material at such a rate as to obtain particulate fluidization while constraining the lower pontion of the bed into a conical shape. A smooth circulation of particles rising in the center and falling at the outside of the bed is thereby established. (AEC)

  17. Nuclear Chirality

    SciTech Connect

    Starosta, Krzysztof

    2005-04-05

    Nuclear chirality is a novel manifestation of spontaneous symmetry breaking resulting from an orthogonal coupling of angular momentum vectors in triaxial nuclei. Three perpendicular angular momenta can form two systems of opposite handedness; the time reversal operator, which reverses orientation of each of the angular momentum components, relates these two systems. The status of current experimental searches for chiral doubling of states, as well as recent progress on the theoretical side is reviewed.

  18. [Zika virus infection during pregnancy].

    PubMed

    Picone, O; Vauloup-Fellous, C; D'Ortenzio, E; Huissoud, C; Carles, G; Benachi, A; Faye, A; Luton, D; Paty, M-C; Ayoubi, J-M; Yazdanpanah, Y; Mandelbrot, L; Matheron, S

    2016-05-01

    A Zika virus epidemic is currently ongoing in the Americas. This virus is linked to congenital infections with potential severe neurodevelopmental dysfunction. However, incidence of fetal infection and whether this virus is responsible of other fetal complications are still unknown. National and international public health authorities recommend caution and several prevention measures. Declaration of Zika virus infection is now mandatory in France. Given the available knowledge on Zika virus, we suggest here a review of the current recommendations for management of pregnancy in case of suspicious or infection by Zika virus in a pregnant woman. PMID:27079865

  19. [Zika virus infection during pregnancy].

    PubMed

    Picone, O; Vauloup-Fellous, C; D'Ortenzio, E; Huissoud, C; Carles, G; Benachi, A; Faye, A; Luton, D; Paty, M-C; Ayoubi, J-M; Yazdanpanah, Y; Mandelbrot, L; Matheron, S

    2016-05-01

    A Zika virus epidemic is currently ongoing in the Americas. This virus is linked to congenital infections with potential severe neurodevelopmental dysfunction. However, incidence of fetal infection and whether this virus is responsible of other fetal complications are still unknown. National and international public health authorities recommend caution and several prevention measures. Declaration of Zika virus infection is now mandatory in France. Given the available knowledge on Zika virus, we suggest here a review of the current recommendations for management of pregnancy in case of suspicious or infection by Zika virus in a pregnant woman.

  20. Nuclear waste

    SciTech Connect

    Not Available

    1991-09-01

    Radioactive waste is mounting at U.S. nuclear power plants at a rate of more than 2,000 metric tons a year. Pursuant to statute and anticipating that a geologic repository would be available in 1998, the Department of Energy (DOE) entered into disposal contracts with nuclear utilities. Now, however, DOE does not expect the repository to be ready before 2010. For this reason, DOE does not want to develop a facility for monitored retrievable storage (MRS) by 1998. This book is concerned about how best to store the waste until a repository is available, congressional requesters asked GAO to review the alternatives of continued storage at utilities' reactor sites or transferring waste to an MRS facility, GAO assessed the likelihood of an MRSA facility operating by 1998, legal implications if DOE is not able to take delivery of wastes in 1998, propriety of using the Nuclear Waste Fund-from which DOE's waste program costs are paid-to pay utilities for on-site storage capacity added after 1998, ability of utilities to store their waste on-site until a repository is operating, and relative costs and safety of the two storage alternatives.

  1. Nuclear terrorism.

    PubMed

    Hogan, David E; Kellison, Ted

    2002-06-01

    Recent events have heightened awareness of the potential for terrorist attacks employing nonconventional weaponry such as biological agents and radiation. Historically, the philosophy of nuclear risk has focused on global or strategic nuclear exchanges and the resulting damage from large-scale releases. Currently, nuclear accidents or terrorist attacks involving low-level or regional release of radiation are considered the most likely events. Thus far, there have been several regional radiation incidents exposing hundreds of thousands of people to radiation, but there have been only a limited number of significant contaminations resulting in death. There are several different types of radioactive particles that differ in mass, extent of radiation emitted, and the degree to which tissue penetration occurs. Radiation affects its toxicity on biological systems by ionization, which creates tissue damage by the generation of free radicals, disruption of chemical bonds, and directly damaging cellular DNA and enzymes. The extent of damage depends on the type of radioisotope and the radiation dose. Radiation doses exceeding 2 to 10 Gy are considered lethal. Optimal management of radiation casualties requires knowledge of the type and dose of radiation received, a recognition of the manifestations of radiation sickness, and the use of standard medical care, decontamination, and decorporation techniques. PMID:12074488

  2. NUCLEAR REACTOR

    DOEpatents

    Grebe, J.J.

    1959-07-14

    High temperature reactors which are uniquely adapted to serve as the heat source for nuclear pcwered rockets are described. The reactor is comprised essentially of an outer tubular heat resistant casing which provides the main coolant passageway to and away from the reactor core within the casing and in which the working fluid is preferably hydrogen or helium gas which is permitted to vaporize from a liquid storage tank. The reactor core has a generally spherical shape formed entirely of an active material comprised of fissile material and a moderator material which serves as a diluent. The active material is fabricated as a gas permeable porous material and is interlaced in a random manner with very small inter-connecting bores or capillary tubes through which the coolant gas may flow. The entire reactor is divided into successive sections along the direction of the temperature gradient or coolant flow, each section utilizing materials of construction which are most advantageous from a nuclear standpoint and which at the same time can withstand the operating temperature of that particular zone. This design results in a nuclear reactor characterized simultaneously by a minimum critiral size and mass and by the ability to heat a working fluid to an extremely high temperature.

  3. The Mammalian Cell Cycle Regulates Parvovirus Nuclear Capsid Assembly

    PubMed Central

    Riolobos, Laura; Domínguez, Carlos; Kann, Michael; Almendral, José M.

    2015-01-01

    It is unknown whether the mammalian cell cycle could impact the assembly of viruses maturing in the nucleus. We addressed this question using MVM, a reference member of the icosahedral ssDNA nuclear parvoviruses, which requires cell proliferation to infect by mechanisms partly understood. Constitutively expressed MVM capsid subunits (VPs) accumulated in the cytoplasm of mouse and human fibroblasts synchronized at G0, G1, and G1/S transition. Upon arrest release, VPs translocated to the nucleus as cells entered S phase, at efficiencies relying on cell origin and arrest method, and immediately assembled into capsids. In synchronously infected cells, the consecutive virus life cycle steps (gene expression, proteins nuclear translocation, capsid assembly, genome replication and encapsidation) proceeded tightly coupled to cell cycle progression from G0/G1 through S into G2 phase. However, a DNA synthesis stress caused by thymidine irreversibly disrupted virus life cycle, as VPs became increasingly retained in the cytoplasm hours post-stress, forming empty capsids in mouse fibroblasts, thereby impairing encapsidation of the nuclear viral DNA replicative intermediates. Synchronously infected cells subjected to density-arrest signals while traversing early S phase also blocked VPs transport, resulting in a similar misplaced cytoplasmic capsid assembly in mouse fibroblasts. In contrast, thymidine and density arrest signals deregulating virus assembly neither perturbed nuclear translocation of the NS1 protein nor viral genome replication occurring under S/G2 cycle arrest. An underlying mechanism of cell cycle control was identified in the nuclear translocation of phosphorylated VPs trimeric assembly intermediates, which accessed a non-conserved route distinct from the importin α2/β1 and transportin pathways. The exquisite cell cycle-dependence of parvovirus nuclear capsid assembly conforms a novel paradigm of time and functional coupling between cellular and virus life

  4. Nuclear photonics

    SciTech Connect

    Habs, D.; Guenther, M. M.; Jentschel, M.; Thirolf, P. G.

    2012-07-09

    With the planned new {gamma}-beam facilities like MEGa-ray at LLNL (USA) or ELI-NP at Bucharest (Romania) with 10{sup 13}{gamma}/s and a band width of {Delta}E{gamma}/E{gamma} Almost-Equal-To 10{sup -3}, a new era of {gamma} beams with energies up to 20MeV comes into operation, compared to the present world-leading HI{gamma}S facility at Duke University (USA) with 10{sup 8}{gamma}/s and {Delta}E{gamma}/E{gamma} Almost-Equal-To 3 Dot-Operator 10{sup -2}. In the long run even a seeded quantum FEL for {gamma} beams may become possible, with much higher brilliance and spectral flux. At the same time new exciting possibilities open up for focused {gamma} beams. Here we describe a new experiment at the {gamma} beam of the ILL reactor (Grenoble, France), where we observed for the first time that the index of refraction for {gamma} beams is determined by virtual pair creation. Using a combination of refractive and reflective optics, efficient monochromators for {gamma} beams are being developed. Thus, we have to optimize the total system: the {gamma}-beam facility, the {gamma}-beam optics and {gamma} detectors. We can trade {gamma} intensity for band width, going down to {Delta}E{gamma}/E{gamma} Almost-Equal-To 10{sup -6} and address individual nuclear levels. The term 'nuclear photonics' stresses the importance of nuclear applications. We can address with {gamma}-beams individual nuclear isotopes and not just elements like with X-ray beams. Compared to X rays, {gamma} beams can penetrate much deeper into big samples like radioactive waste barrels, motors or batteries. We can perform tomography and microscopy studies by focusing down to {mu}m resolution using Nuclear Resonance Fluorescence (NRF) for detection with eV resolution and high spatial resolution at the same time. We discuss the dominating M1 and E1 excitations like the scissors mode, two-phonon quadrupole octupole excitations, pygmy dipole excitations or giant dipole excitations under the new facet of

  5. Nuclear photonics

    NASA Astrophysics Data System (ADS)

    Habs, D.; Günther, M. M.; Jentschel, M.; Thirolf, P. G.

    2012-07-01

    With the planned new γ-beam facilities like MEGa-ray at LLNL (USA) or ELI-NP at Bucharest (Romania) with 1013 γ/s and a band width of ΔEγ/Eγ≈10-3, a new era of γ beams with energies up to 20MeV comes into operation, compared to the present world-leading HIγS facility at Duke University (USA) with 108 γ/s and ΔEγ/Eγ≈3ṡ10-2. In the long run even a seeded quantum FEL for γ beams may become possible, with much higher brilliance and spectral flux. At the same time new exciting possibilities open up for focused γ beams. Here we describe a new experiment at the γ beam of the ILL reactor (Grenoble, France), where we observed for the first time that the index of refraction for γ beams is determined by virtual pair creation. Using a combination of refractive and reflective optics, efficient monochromators for γ beams are being developed. Thus, we have to optimize the total system: the γ-beam facility, the γ-beam optics and γ detectors. We can trade γ intensity for band width, going down to ΔEγ/Eγ≈10-6 and address individual nuclear levels. The term "nuclear photonics" stresses the importance of nuclear applications. We can address with γ-beams individual nuclear isotopes and not just elements like with X-ray beams. Compared to X rays, γ beams can penetrate much deeper into big samples like radioactive waste barrels, motors or batteries. We can perform tomography and microscopy studies by focusing down to μm resolution using Nuclear Resonance Fluorescence (NRF) for detection with eV resolution and high spatial resolution at the same time. We discuss the dominating M1 and E1 excitations like the scissors mode, two-phonon quadrupole octupole excitations, pygmy dipole excitations or giant dipole excitations under the new facet of applications. We find many new applications in biomedicine, green energy, radioactive waste management or homeland security. Also more brilliant secondary beams of neutrons and positrons can be produced.

  6. Viruses and the nucleolus: the fatal attraction.

    PubMed

    Salvetti, Anna; Greco, Anna

    2014-06-01

    Viruses are small obligatory parasites and as a consequence, they have developed sophisticated strategies to exploit the host cell's functions to create an environment that favors their own replication. A common feature of most - if not all - families of human and non-human viruses concerns their interaction with the nucleolus. The nucleolus is a multifunctional nuclear domain, which, in addition to its well-known role in ribosome biogenesis, plays several crucial other functions. Viral infection induces important nucleolar alterations. Indeed, during viral infection numerous viral components localize in nucleoli, while various host nucleolar proteins are redistributed in other cell compartments or are modified, and non-nucleolar cellular proteins reach the nucleolus. This review highlights the interactions reported between the nucleolus and some human or animal viral families able to establish a latent or productive infection, selected on the basis of their known interactions with the nucleolus and the nucleolar activities, and their links with virus replication and/or pathogenesis. This article is part of a Special Issue entitled: Role of the Nucleolus in Human Disease.

  7. Recombinant Vaccinia Virus: Immunization against Multiple Pathogens

    NASA Astrophysics Data System (ADS)

    Perkus, Marion E.; Piccini, Antonia; Lipinskas, Bernard R.; Paoletti, Enzo

    1985-09-01

    The coding sequences for the hepatitis B virus surface antigen, the herpes simplex virus glycoprotein D, and the influenza virus hemagglutinin were inserted into a single vaccinia virus genome. Rabbits inoculated intravenously or intradermally with this polyvalent vaccinia virus recombinant produced antibodies reactive to all three authentic foreign antigens. In addition, the feasibility of multiple rounds of vaccination with recombinant vaccinia virus was demonstrated.

  8. Two African viruses serologically and morphologically related to rabies virus.

    PubMed

    Shope, R E; Murphy, F A; Harrison, A K; Causey, O R; Kemp, G E; Simpson, D I; Moore, D L

    1970-11-01

    Lagos bat virus and an isolate from shrews (IbAn 27377), both from Nigeria, were found to be bullet-shaped and to mature intracytoplasmically in association with a distinct matrix. They were related to, but readily distinguishable from, rabies virus and each other by complement fixation and neutralization tests. The three viruses, including rabies, form a subgrouping within the rhabdoviruses. PMID:5530013

  9. Strategies on the nuclear-targeted delivery of genes

    PubMed Central

    Yao, Jing; Fan, Ying; Li, Yuanke; Huang, Leaf

    2016-01-01

    To improve the nuclear-targeted delivery of non-viral vectors, extensive effort has been carried out on the development of smart vectors which could overcome multiple barriers. The nuclear envelope presents a major barrier to transgene delivery. Viruses are capable of crossing the nuclear envelope to efficiently deliver their genome into the nucleus through the specialized protein components. However, non-viral vectors are preferred over viral ones because of the safety concerns associated with the latter. Non-viral delivery systems have been designed to include various types of components to enable nuclear translocation at the periphery of the nucleus. This review summarizes the progress of research regarding nuclear transport mechanisms. “Smart” non-viral vectors that have been modified by peptides and other small molecules are able to facilitate the nuclear translocation and enhance the efficacy of gene expression. The resulting technology may also enhance delivery of other macromolecules to the nucleus. PMID:23964565

  10. The Nuclear Power and Nuclear Weapons Connection.

    ERIC Educational Resources Information Center

    Leventhal, Paul

    1990-01-01

    Explains problems enforcing the Nuclear Non-Proliferation Treaty (NPT) of 1968. Provides factual charts and details concerning the production of nuclear energy and arms, the processing and disposal of waste products, and outlines the nuclear fuel cycle. Discusses safeguards, the risk of nuclear terrorism, and ways to deal with these problems. (NL)

  11. The nuclear arsenals and nuclear disarmament.

    PubMed

    Barnaby, F

    1998-01-01

    Current world stockpiles of nuclear weapons and the status of treaties for nuclear disarmament and the ultimate elimination of nuclear weapons are summarised. The need for including stockpiles of civil plutonium in a programme for ending production and disposing of fissile materials is emphasized, and the ultimate difficulty of disposing of the last few nuclear weapons discussed.

  12. The Nuclear Power/Nuclear Weapons Connection.

    ERIC Educational Resources Information Center

    Totten, Sam; Totten, Martha Wescoat

    1985-01-01

    Once they have nuclear power, most countries will divert nuclear materials from commercial to military programs. In excerpts from the book "Facing the Danger" (by Totten, S. and M. W., Crossing Press, 1984), five anti-nuclear activists explain how and why they have been addressing the nuclear connection. (RM)

  13. [Viruses and civilization].

    PubMed

    Chastel, C

    1999-01-01

    A few million years ago, when primates moved from the east African forest to the savannah, they were already infected with endogenous viruses and occultly transmitted them to the prime Homo species. However it was much later with the building of the first large cities in Mesopotamia that interhuman viral transmission began in earnest. Spreading was further enhanced with the organization of the Egyptian, Greek, Roman, and Arab empires around the Mediterranean. Discovery of the New World in 1492 led to an unprecedented clash of civilizations and the destruction of pre-Columbian Indian civilizations. It also led to a rapid spread of viruses across the Atlantic Ocean with the emergence of yellow fever and appearance of smallpox and measles throughout the world. However the greatest opportunities for worldwide viral development have been created by our present, modern civilization. This fact is illustrated by epidemic outbreaks of human immunodeficiency virus, Venezuela hemorrhagic fever, Rift valley fever virus, and monkey pox virus. Close analysis underscores the major role of human intervention in producing these events.

  14. Viruses and Multiple Sclerosis

    PubMed Central

    Owens, Gregory P.; Gilden, Don; Burgoon, Mark P.; Yu, Xiaoli; Bennett, Jeffrey L.

    2012-01-01

    Multiple sclerosis (MS) is a chronic demyelinating disorder of unknown etiology, possibly caused by a virus or virus-triggered immunopathology. The virus might reactivate after years of latency and lyse oligodendrocytes, as in progressive multifocal leukoencephalopathy, or initiate immunopathological demyelination, as in animals infected with Theiler’s murine encephalomyelitis virus or coronaviruses. The argument for a viral cause of MS is supported by epidemiological analyses and studies of MS in identical twins, indicating that disease is acquired. However, the most important evidence is the presence of bands of oligoclonal IgG (OCBs) in MS brain and CSF that persist throughout the lifetime of the patient. OCBs are found almost exclusively in infectious CNS disorders, and antigenic targets of OCBs represent the agent that causes disease. Here, the authors review past attempts to identify an infectious agent in MS brain cells and discuss the promise of using recombinant antibodies generated from clonally expanded plasma cells in brain and CSF to identify disease-relevant antigens. They show how this strategy has been used successfully to analyze antigen specificity in subacute sclerosing panencephalitis, a chronic encephalitis caused by measles virus, and in neuromyelitis optica, a chronic autoimmune demyelinating disease produced by antibodies directed against the aquaporin-4 water channel. PMID:22130640

  15. Viruses and multiple sclerosis.

    PubMed

    Owens, Gregory P; Gilden, Don; Burgoon, Mark P; Yu, Xiaoli; Bennett, Jeffrey L

    2011-12-01

    Multiple sclerosis (MS) is a chronic demyelinating disorder of unknown etiology, possibly caused by a virus or virus-triggered immunopathology. The virus might reactivate after years of latency and lyse oligodendrocytes, as in progressive multifocal leukoencephalopathy, or initiate immunopathological demyelination, as in animals infected with Theiler's murine encephalomyelitis virus or coronaviruses. The argument for a viral cause of MS is supported by epidemiological analyses and studies of MS in identical twins, indicating that disease is acquired. However, the most important evidence is the presence of bands of oligoclonal IgG (OCBs) in MS brain and CSF that persist throughout the lifetime of the patient. OCBs are found almost exclusively in infectious CNS disorders, and antigenic targets of OCBs represent the agent that causes disease. Here, the authors review past attempts to identify an infectious agent in MS brain cells and discuss the promise of using recombinant antibodies generated from clonally expanded plasma cells in brain and CSF to identify disease-relevant antigens. They show how this strategy has been used successfully to analyze antigen specificity in subacute sclerosing panencephalitis, a chronic encephalitis caused by measles virus, and in neuromyelitis optica, a chronic autoimmune demyelinating disease produced by antibodies directed against the aquaporin-4 water channel. PMID:22130640

  16. Attenuation of Vaccinia Virus.

    PubMed

    Yakubitskiy, S N; Kolosova, I V; Maksyutov, R A; Shchelkunov, S N

    2015-01-01

    Since 1980, in the post-smallpox vaccination era the human population has become increasingly susceptible compared to a generation ago to not only the variola (smallpox) virus, but also other zoonotic orthopoxviruses. The need for safer vaccines against orthopoxviruses is even greater now. The Lister vaccine strain (LIVP) of vaccinia virus was used as a parental virus for generating a recombinant 1421ABJCN clone defective in five virulence genes encoding hemagglutinin (A56R), the IFN-γ-binding protein (B8R), thymidine kinase (J2R), the complement-binding protein (C3L), and the Bcl-2-like inhibitor of apoptosis (N1L). We found that disruption of these loci does not affect replication in mammalian cell cultures. The isogenic recombinant strain 1421ABJCN exhibits a reduced inflammatory response and attenuated neurovirulence relative to LIVP. Virus titers of 1421ABJCN were 3 lg lower versus the parent VACV LIVP when administered by the intracerebral route in new-born mice. In a subcutaneous mouse model, 1421ABJCN displayed levels of VACV-neutralizing antibodies comparable to those of LIVP and conferred protective immunity against lethal challenge by the ectromelia virus. The VACV mutant holds promise as a safe live vaccine strain for preventing smallpox and other orthopoxvirus infections. PMID:26798498

  17. [Markers of hepatitis virus].

    PubMed

    Suzuki, Fumitaka

    2008-11-01

    Hepatitis B virus (HBV) and hepatitis C virus (HCV) are the major viruses known to cause viral hepatitis. Serological markers are commonly used as diagnostic and/or prognostic indicators of acute or chronic HBV or HCV infection. The ability to detect HBV DNA in serum has been reported to have prognostic value for the outcome of chronic HBV infection. A rapid and sustained drop in HBV DNA or HCV RNA levels in patients under therapy has been shown to be a predictive factor for a favourable treatment outcome. Various techniques for detecting HBV DNA or HCV RNA have already been described; however, there are various problems with the sensitivity or detection range of those methods. New virus measuring methods have recently been reported and used. The Cobas Taq Man HCV Test is a new method to detect HBV DNA and HCV RNA with higher sensitivity and a broader range of quantitation than conventional methods. Some reports have shown that these methods improve therapy monitoring and the management of HBV or HCV infection. Moreover, hepatitis E virus (HEV) infection has been reported in Japan. The clinical features and viral markers of HEV have also been described. PMID:19086457

  18. Herpes simplex virus 1 induces egress channels through marginalized host chromatin.

    PubMed

    Myllys, Markko; Ruokolainen, Visa; Aho, Vesa; Smith, Elizabeth A; Hakanen, Satu; Peri, Piritta; Salvetti, Anna; Timonen, Jussi; Hukkanen, Veijo; Larabell, Carolyn A; Vihinen-Ranta, Maija

    2016-01-01

    Lytic infection with herpes simplex virus type 1 (HSV-1) induces profound modification of the cell nucleus including formation of a viral replication compartment and chromatin marginalization into the nuclear periphery. We used three-dimensional soft X-ray tomography, combined with cryogenic fluorescence, confocal and electron microscopy, to analyse the transformation of peripheral chromatin during HSV-1 infection. Our data showed an increased presence of low-density gaps in the marginalized chromatin at late infection. Advanced data analysis indicated the formation of virus-nucleocapsid-sized (or wider) channels extending through the compacted chromatin of the host. Importantly, confocal and electron microscopy analysis showed that these gaps frequently contained viral nucleocapsids. These results demonstrated that HSV-1 infection induces the formation of channels penetrating the compacted layer of cellular chromatin and allowing for the passage of progeny viruses to the nuclear envelope, their site of nuclear egress. PMID:27349677

  19. Specific interaction of capsid protein and importin-{alpha}/{beta} influences West Nile virus production

    SciTech Connect

    Bhuvanakantham, Raghavan; Chong, Mun-Keat; Ng, Mah-Lee

    2009-11-06

    West Nile virus (WNV) capsid (C) protein has been shown to enter the nucleus of infected cells. However, the mechanism by which C protein enters the nucleus is unknown. In this study, we have unveiled for the first time that nuclear transport of WNV and Dengue virus C protein is mediated by their direct association with importin-{alpha}. This interplay is mediated by the consensus sequences of bipartite nuclear localization signal located between amino acid residues 85-101 together with amino acid residues 42 and 43 of C protein. Elucidation of biological significance of importin-{alpha}/C protein interaction demonstrated that the binding efficiency of this association influenced the nuclear entry of C protein and virus production. Collectively, this study illustrated the molecular mechanism by which the C protein of arthropod-borne flavivirus enters the nucleus and showed the importance of importin-{alpha}/C protein interaction in the context of flavivirus life-cycle.

  20. Special Issue: Honey Bee Viruses.

    PubMed

    Gisder, Sebastian; Genersch, Elke

    2015-10-01

    Pollination of flowering plants is an important ecosystem service provided by wild insect pollinators and managed honey bees. Hence, losses and declines of pollinating insect species threaten human food security and are of major concern not only for apiculture or agriculture but for human society in general. Honey bee colony losses and bumblebee declines have attracted intensive research interest over the last decade and although the problem is far from being solved we now know that viruses are among the key players of many of these bee losses and bumblebee declines. With this special issue on bee viruses we, therefore, aimed to collect high quality original papers reflecting the current state of bee virus research. To this end, we focused on newly discovered viruses (Lake Sinai viruses, bee macula-like virus), or a so far neglected virus species (Apis mellifera filamentous virus), and cutting edge technologies (mass spectrometry, RNAi approach) applied in the field.

  1. Chlorella viruses isolated in China

    SciTech Connect

    Zhang, Y.; Burbank, D.E.; Van Etten, J.L. )

    1988-09-01

    Plaque-forming viruses of the unicellular, eukaryotic, exsymbiotic, Chlorella-like green algae strain NC64A, which are common in the United States, were also present in fresh water collected in the People's Republic of China. Seven of the Chinese viruses were examined in detail and compared with the Chlorella viruses previously isolated in the United States. Like the American viruses, the Chinese viruses were large polyhedra and sensitive to chloroform. They contained numerous structural proteins and large double-stranded DNA genomes of at least 300 kilobase pairs. Each of the DNAs from the Chinese viruses contained 5-methyldeoxycytosine, which varied from 12.6 to 46.7% of the deoxycytosine, and N{sup 6}-methyldeoxyadenosine, which varied from 2.2 to 28.3% of the deoxyadenosine. Four of the Chinese virus DNAs hybridized extensively with {sup 32}P-labeled DNA from the American virus PBCV-1, and three hybridized poorly.

  2. Special Issue: Honey Bee Viruses

    PubMed Central

    Gisder, Sebastian; Genersch, Elke

    2015-01-01

    Pollination of flowering plants is an important ecosystem service provided by wild insect pollinators and managed honey bees. Hence, losses and declines of pollinating insect species threaten human food security and are of major concern not only for apiculture or agriculture but for human society in general. Honey bee colony losses and bumblebee declines have attracted intensive research interest over the last decade and although the problem is far from being solved we now know that viruses are among the key players of many of these bee losses and bumblebee declines. With this special issue on bee viruses we, therefore, aimed to collect high quality original papers reflecting the current state of bee virus research. To this end, we focused on newly discovered viruses (Lake Sinai viruses, bee macula-like virus), or a so far neglected virus species (Apis mellifera filamentous virus), and cutting edge technologies (mass spectrometry, RNAi approach) applied in the field. PMID:26702462

  3. Testing for Human Immunodeficiency Virus

    MedlinePlus

    ... incisions made in the mother’s abdomen and uterus. Human Immunodeficiency Virus (HIV): A virus that attacks certain cells of the body’s immune system and causes acquired immunodeficiency syndrome (AIDS). Immune System: ...

  4. Emerging issues in virus taxonomy.

    PubMed

    van Regenmortel, Marc H V; Mahy, Brian W J

    2004-01-01

    Viruses occupy a unique position in biology. Although they possess some of the properties of living systems such as having a genome, they are actually nonliving infectious entities and should not be considered microorganisms. A clear distinction should be drawn between the terms virus, virion, and virus species. Species is the most fundamental taxonomic category used in all biological classification. In 1991, the International Committee on Taxonomy of Viruses (ICTV) decided that the category of virus species should be used in virus classification together with the categories of genus and family. More than 50 ICTV study groups were given the task of demarcating the 1,550 viral species that were recognized in the 7th ICTV report, which was published in 2000. We briefly describe the changes in virus classification that were introduced in that report. We also discuss recent proposals to introduce a nonlatinized binomial nomenclature for virus species. PMID:15078590

  5. Special Issue: Honey Bee Viruses.

    PubMed

    Gisder, Sebastian; Genersch, Elke

    2015-10-01

    Pollination of flowering plants is an important ecosystem service provided by wild insect pollinators and managed honey bees. Hence, losses and declines of pollinating insect species threaten human food security and are of major concern not only for apiculture or agriculture but for human society in general. Honey bee colony losses and bumblebee declines have attracted intensive research interest over the last decade and although the problem is far from being solved we now know that viruses are among the key players of many of these bee losses and bumblebee declines. With this special issue on bee viruses we, therefore, aimed to collect high quality original papers reflecting the current state of bee virus research. To this end, we focused on newly discovered viruses (Lake Sinai viruses, bee macula-like virus), or a so far neglected virus species (Apis mellifera filamentous virus), and cutting edge technologies (mass spectrometry, RNAi approach) applied in the field. PMID:26702462

  6. Ebola (Ebola Virus Disease): Prevention

    MedlinePlus

    ... Search The CDC Cancel Submit Search The CDC Ebola (Ebola Virus Disease) Note: Javascript is disabled or is ... message, please visit this page: About CDC.gov . Ebola (Ebola Virus Disease) About Ebola Questions & Answers 2014 ...

  7. Ebola (Ebola Virus Disease): Transmission

    MedlinePlus

    ... Search The CDC Cancel Submit Search The CDC Ebola (Ebola Virus Disease) Note: Javascript is disabled or is ... message, please visit this page: About CDC.gov . Ebola (Ebola Virus Disease) About Ebola Questions & Answers 2014 ...

  8. Ebola (Ebola Virus Disease): Treatment

    MedlinePlus

    ... Search The CDC Cancel Submit Search The CDC Ebola (Ebola Virus Disease) Note: Javascript is disabled or is ... message, please visit this page: About CDC.gov . Ebola (Ebola Virus Disease) About Ebola Questions & Answers 2014 ...

  9. Ebola (Ebola Virus Disease): Diagnosis

    MedlinePlus

    ... Search The CDC Cancel Submit Search The CDC Ebola (Ebola Virus Disease) Note: Javascript is disabled or is ... message, please visit this page: About CDC.gov . Ebola (Ebola Virus Disease) About Ebola Questions & Answers 2014 ...

  10. Production of virus resistant plants

    DOEpatents

    Dougherty, W.G.; Lindbo, J.A.

    1996-12-10

    A method of suppressing virus gene expression in plants using untranslatable plus sense RNA is disclosed. The method is useful for the production of plants that are resistant to virus infection. 9 figs.

  11. Zika Virus Infection and Microcephaly.

    PubMed

    Millichap, J Gordon

    2016-01-01

    A Task Force established by the Brazil Ministry of Health investigated the possible association of microcephaly with Zika virus infection during pregnancy and a registry for microcephaly cases among women suspected to have had Zika virus infection during pregnancy.

  12. Chlorella viruses isolated in China.

    PubMed Central

    Zhang, Y P; Burbank, D E; Van Etten, J L

    1988-01-01

    Plaque-forming viruses of the unicellular, eucaryotic, exsymbiotic, Chlorella-like green algae strain NC64A, which are common in the United States, were also present in fresh water collected in the People's Republic of China. Seven of the Chinese viruses were examined in detail and compared with the Chlorella viruses previously isolated in the United States. Like the American viruses, the Chinese viruses were large polyhedra and sensitive to chloroform. They contained numerous structural proteins and large double-stranded DNA genomes of at least 300 kilobase pairs. Each of the DNAs from the Chinese viruses contained 5-methyldeoxycytosine, which varied from 12.6 to 46.7% of the deoxycytosine, and N6-methyldeoxyadenosine, which varied from 2.2 to 28.3% of the deoxyadenosine. Four of the Chinese virus DNAs hybridized extensively with DNA from the American virus PBCV-1, and three hybridized poorly. Images PMID:2847652

  13. 9 CFR 113.215 - Bovine Virus Diarrhea Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Bovine Virus Diarrhea Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.215 Bovine Virus Diarrhea Vaccine, Killed Virus. Bovine Virus Diarrhea Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  14. 9 CFR 113.215 - Bovine Virus Diarrhea Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Bovine Virus Diarrhea Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.215 Bovine Virus Diarrhea Vaccine, Killed Virus. Bovine Virus Diarrhea Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  15. 9 CFR 113.215 - Bovine Virus Diarrhea Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Bovine Virus Diarrhea Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.215 Bovine Virus Diarrhea Vaccine, Killed Virus. Bovine Virus Diarrhea Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  16. 9 CFR 113.215 - Bovine Virus Diarrhea Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Bovine Virus Diarrhea Vaccine, Killed... REQUIREMENTS Killed Virus Vaccines § 113.215 Bovine Virus Diarrhea Vaccine, Killed Virus. Bovine Virus Diarrhea Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed...

  17. Epstein-Barr virus infection and replication in a human epithelial cell system.

    PubMed

    Li, Q X; Young, L S; Niedobitek, G; Dawson, C W; Birkenbach, M; Wang, F; Rickinson, A B

    1992-03-26

    Epstein-Barr virus, a human herpesvirus with oncogenic potential, infects two target tissues in vivo: B lymphocytes, where the infection is largely non-productive, and stratified squamous epithelium in which virus replication occurs. The interaction with B cells, initiated through virus binding to the B-cell surface molecule CR2 (ref. 4), has been studied in vitro and the virus 'latent' genes associated with B-cell growth transformation defined. By comparison, viral infection of epithelium remains poorly understood, reflecting the lack of an appropriate cell-culture model. Here we describe the development of such a model using as targets CR2-expressing transfected cells of two independent human epithelial lines. A high proportion of these cells bind virus and become actively infected, expressing the small EBER RNAs (small non-polyadenylated virus-coded RNAs) and the Epstein-Barr nuclear antigen 1 but not other latent proteins; thereafter, under conditions favouring epithelial differentiation, up to 30% of the cells can be induced to enter virus productive cycle with some progressing to full virus replication. We find significant differences between laboratory virus strains in their ability to infect epithelium that do not correlate with their B-cell growth-transforming activity. PMID:1312681

  18. RNA-Seq reveals virus-virus and virus-plant interactions in nature.

    PubMed

    Kamitani, Mari; Nagano, Atsushi J; Honjo, Mie N; Kudoh, Hiroshi

    2016-11-01

    As research on plant viruses has focused mainly on crop diseases, little is known about these viruses in natural environments. To understand the ecology of viruses in natural systems, comprehensive information on virus-virus and virus-host interactions is required. We applied RNA-Seq to plants from a natural population of Arabidopsis halleri subsp. gemmifera to simultaneously determine the presence/absence of all sequence-reported viruses, identify novel viruses and quantify the host transcriptome. By introducing the criteria of read number and genome coverage, we detected infections by Turnip mosaic virus (TuMV), Cucumber mosaic virus and Brassica yellows virus Active TuMV replication was observed by ultramicroscopy. De novo assembly further identified a novel partitivirus, Arabidopsis halleri partitivirus 1 Interestingly, virus reads reached a maximum level that was equivalent to that of the host's total mRNA, although asymptomatic infection was common. AhgAGO2, a key gene in host defence systems, was upregulated in TuMV-infected plants. Multiple infection was frequent in TuMV-infected leaves, suggesting that TuMV facilitates multiple infection, probably by suppressing host RNA silencing. Revealing hidden plant-virus interactions in nature can enhance our understanding of biological interactions and may have agricultural applications. PMID:27549115

  19. Nuclear security

    SciTech Connect

    Dingell, J.D.

    1991-02-01

    The Department of Energy's (DOE) Lawrence Livermore National Laboratory, located in Livermore, California, generates and controls large numbers of classified documents associated with the research and testing of nuclear weapons. Concern has been raised about the potential for espionage at the laboratory and the national security implications of classified documents being stolen. This paper determines the extent of missing classified documents at the laboratory and assesses the adequacy of accountability over classified documents in the laboratory's custody. Audit coverage was limited to the approximately 600,000 secret documents in the laboratory's custody. The adequacy of DOE's oversight of the laboratory's secret document control program was also assessed.

  20. NUCLEAR REACTOR

    DOEpatents

    Young, G.

    1963-01-01

    This patent covers a power-producing nuclear reactor in which fuel rods of slightly enriched U are moderated by heavy water and cooled by liquid metal. The fuel rods arranged parallel to one another in a circle are contained in a large outer closed-end conduit that extends into a tank containing the heavy water. Liquid metal is introduced into the large conduit by a small inner conduit that extends within the circle of fuel rods to a point near the lower closed end of the outer conduit. (AEC) Production Reactors