Molecular cloning and characterization of Antheraea mylitta cytoplasmic polyhedrosis virus polyhedrin gene and its variant forms

International Nuclear Information System (INIS)

Sinha-Datta, Uma; Chavali, Venkata Ramana Murthy; Ghosh, Ananta K.

2005-01-01

The segments 10 (S10) of the 11 double stranded RNA genomes from Antheraea mylitta cytoplasmic polyhedrosis virus (AmCPV) encoding a novel polyhedrin polypeptide was converted to cDNA, cloned, and sequenced. Three cDNA clones consisting of 1502 (AmCPV10-1), 1120 (AmCPV10-2), and 1415 (AmCPV10-3) nucleotides encoding polyhedrin of 254, 339, and 319 amino acids with molecular masses of 29, 39, and 37 kDa, respectively, were obtained, and verified by Northern analysis. These clones showed 70-94% sequence identity among them but none with any sequences in databases. The expression of AmCPV10-1 cDNA encoded polyhedrin in Sf-9 cells was detected by immunoblot analysis and formation of polyhedra by electron microscopy, as observed in AmCPV-infected gut cells, but no expression of AmCPV10-2 or AmCPV10-3 cDNA was detected, indicating that during AmCPV replication, along with functional S10 RNA, some defective variant forms of S10 RNAs are packaged in virion particles

Suscetibilidade de Spodoptera Frugiperda a isolados geográficos de um vírus de poliedrose nuclear Spodoptera Frugiperda susceptibility to nuclear polyhedrosis virus geographical isolates

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Sérgio Arce Gomez

1999-09-01

Full Text Available O presente trabalho objetivou verificar a suscetibilidade de larvas de segundo ínstar de Spodoptera frugiperda (Smith, 1797 a sete isolados geográficos de um vírus de poliedrose nuclear (VPN, conduzindo-se sete bioensaios no Laboratório de Patologia de Insetos da Embrapa-Centro Nacional de Pesquisa de Soja, Londrina. Para cada isolado preparou-se dieta artificial contendo 0, 2x10³, 4x10³, 8x10³, 16x10³, 32x10³ e 64x10³ corpos poliédricos de inclusão (CPI/mL. Cada dose foi oferecida às larvas em copos de plástico de 50 mL, sob condições controladas (temperatura: 26±2ºC; umidade relativa: 60±10%; fotófase:14 horas. A análise (Probits realizada sobre o somatório de larvas mortas (contadas, diariamente, do quinto ao décimo quarto dia após a inoculação mostrou, com base na ausência de sobreposição das amplitudes dos intervalos de confiança das concentrações letais médias (CL50, que: o isolado de Sertaneja, PR (5.631 CPI/mL, foi o mais virulento; o da Guatemala (11.520 CPI/mL equivaleu aos de Ponta Grossa, PR (14.184 CPI/mL, Argentina (15.891 CPI/mL e Alabama, EUA (17.558 CPI/mL, mas foi superior aos isolados de Louisiana, EUA (19.325 CPI/mL e Sete Lagoas, MG (25.310 CPI/mL. A variação do tempo letal médio, de 8,3 a 10 dias, não foi significativa em relação aos isolados.In order to verify the Spodoptera frugiperda (Smith,1797 second instar larvae susceptibility to seven nuclear polyhedrosis virus (NPV geographical isolates, seven bioassays were carried out at Embrapa-Centro Nacional de Pesquisa de Soja, Insect Pathology Laboratory, Londrina, Paraná State, Brazil. Artificial diet containing 0 (control, 2x10³, 4x10³, 8x10³, 16x10³, 32x10³, and 64x10³ polyhedral inclusion bodies (PIB/mL was prepared for each virus isolate; each dose was offered, in 50 mL plastic cups to the larvae under controlled conditions (temperature 26±2ºC; relative humidity: 60±10% and photophase: 14 hours. The statistical analysis

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

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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.

Viral Small-RNA Analysis of Bombyx mori Larval Midgut during Persistent and Pathogenic Cytoplasmic Polyhedrosis Virus Infection.

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Zografidis, Aris; Van Nieuwerburgh, Filip; Kolliopoulou, Anna; Apostolou-Karampelis, Konstantinos; Head, Steven R; Deforce, Dieter; Smagghe, Guy; Swevers, Luc

2015-11-01

The lepidopteran innate immune response against RNA viruses remains poorly understood, while in other insects several studies have highlighted an essential role for the exo-RNAi pathway in combating viral infection. Here, by using deep-sequencing technology for viral small-RNA (vsRNA) assessment, we provide evidence that exo-RNAi is operative in the silkworm Bombyx mori against both persistent and pathogenic infection of B. mori cytoplasmic polyhedrosis virus (BmCPV) which is characterized by a segmented double-stranded RNA (dsRNA) genome. Further, we show that Dicer-2 predominantly targets viral dsRNA and produces 20-nucleotide (nt) vsRNAs, whereas an additional pathway is responsive to viral mRNA derived from segment 10. Importantly, vsRNA distributions, which define specific hot and cold spot profiles for each viral segment, to a considerable degree overlap between Dicer-2-related (19 to 21 nt) and Dicer-2-unrelated vsRNAs, suggesting a common origin for these profiles. We found a degenerate motif significantly enriched at the cut sites of vsRNAs of various lengths which link an unknown RNase to the origins of vsRNAs biogenesis and distribution. Accordingly, the indicated RNase activity may be an important early factor for the host's antiviral defense in Lepidoptera. This work contributes to the elucidation of the lepidopteran antiviral response against infection of segmented double-stranded RNA (dsRNA) virus (CPV; Reoviridae) and highlights the importance of viral small-RNA (vsRNA) analysis for getting insights into host-pathogen interactions. Three vsRNA pathways are implicated in antiviral defense. For dsRNA, two pathways are proposed, either based on Dicer-2 cleavage to generate 20-nucleotide vsRNAs or based on the activity of an uncharacterized endo-RNase that cleaves the viral RNA substrate at a degenerate motif. The analysis also indicates the existence of a degradation pathway that targets the positive strand of segment 10. Copyright © 2015, American

BACULOVIRUS REPLICATION ALTERS HORMONE-REGULATED HOST DEVELOPMENT.

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The baculovirus Lymantria dispar nuclear polyhedrosis virus interferes with insect larval development by altering the host's hormonal system. The level of haemolymph ecdysteroids, the insect moulting hormone, was found to be higher in virus-infected larvae than in uninfected cont...

Functional analysis of the baculovirus 10 kilodalton protein

NARCIS (Netherlands)

Oers, van M.M.

1994-01-01

Nuclear polyhedrosis viruses belong to the family Baculoviridae and cause fatal diseases in arthropods predominantly in insects of the order Lepidoptera . These viruses were first reported in silk worms where viral infections could have devastating

Granulosis viruses, with emphasis on the GV of the Indian meal moth, Plodia interpunctella.

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Consigli, R A; Tweeten, K A; Anderson, D K; Bulla, L A

1983-01-01

The granulosis viruses and nuclear polyhedrosis viruses are being considered for use as biological insecticides for control of their insect hosts. Many of these insect species, which include some of the most serious pests of agriculture and forests, have become difficult to control because they have developed resistance to chemical insecticides. Several laboratory and field studies have demonstrated that the baculoviruses (GV and NPV) are promising alternatives to chemicals for the control of economically important insects. These viruses are highly virulent, selective, and stable, and the impact on the environment following their application is minimal. A decision concerning the application of baculoviruses to stored grain and field crops must be based upon a prudent consideration of the benefits to be obtained and the potential risks of their use. Such decisions should be made only after consideration of the physical, chemical, and biological properties of these viruses. In addition, methods must be developed for the unequivocal identification of these viruses, and their effects on nontarget species at the cellular and molecular levels must be investigated. This can best be accomplished if a sufficient body of knowledge regarding the molecular properties of these viruses and their infection process is accumulated by an extensive quantitative approach. Much of this knowledge is lacking because, prior to their consideration for use as insecticides, the baculoviruses appeared to have little medical or economic importance. As a result, interest in studying them was limited. It has become obvious that the molecular properties of these viruses must be investigated if full advantage is to be taken of using them as insect control agents, and if present and future problems concerning their use as insecticides are to be handled properly. Fundamental research on the biochemical and biophysical properties of baculoviruses has concentrated mainly on a variety of nuclear

Disease-parasitoid relationships in natural populations of Lymantria dispar [Lep.: Lymantriidae] in the northeastern United States

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R.C. Reardon; J.D. Podgwaite

1976-01-01

Immature Lymantria dispar L. were collected from 6 geographically distinct populations over 2 years to determine correlations between parasitoid and disease incidences. Incidence of the nuclear polyhedrosis virus disease (NPV) was found to be positively correlated with incidences of the parasitoids Apanteles melanoscelus (Ratzeburg...

Characterization of Baculovirus Insecticides Expressing Tailored Bacillus thuringiensis CryIA(b) Crystal Proteins

NARCIS (Netherlands)

Martens, John W M; Knoester, Marga; Weijts, Franci; Groffen, Sander J A; Hu, Zhihong; Bosch, Dirk; Vlak, Just M.

1995-01-01

Full-length, truncated, and mature forms of the CryIA(b) insecticidal crystal protein gene of Bacillus thuringiensis were engineered into the p10 locus of Autographa californica nuclear polyhedrosis virus (AcNPV). A signal sequence of Heliothis virescens juvenile hormone esterase was introduced at

Molecular characterization of genome segments 1 and 3 encoding two capsid proteins of Antheraea mylitta cytoplasmic polyhedrosis virus

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Chakrabarti Mrinmay

2010-08-01

Full Text Available Abstract Background Antheraea mylitta cytoplasmic polyhedrosis virus (AmCPV, a cypovirus of Reoviridae family, infects Indian non-mulberry silkworm, Antheraea mylitta, and contains 11 segmented double stranded RNA (S1-S11 in its genome. Some of its genome segments (S2 and S6-S11 have been previously characterized but genome segments encoding viral capsid have not been characterized. Results In this study genome segments 1 (S1 and 3 (S3 of AmCPV were converted to cDNA, cloned and sequenced. S1 consisted of 3852 nucleotides, with one long ORF of 3735 nucleotides and could encode a protein of 1245 amino acids with molecular mass of ~141 kDa. Similarly, S3 consisted of 3784 nucleotides having a long ORF of 3630 nucleotides and could encode a protein of 1210 amino acids with molecular mass of ~137 kDa. BLAST analysis showed 20-22% homology of S1 and S3 sequence with spike and capsid proteins, respectively, of other closely related cypoviruses like Bombyx mori CPV (BmCPV, Lymantria dispar CPV (LdCPV, and Dendrolimus punctatus CPV (DpCPV. The ORFs of S1 and S3 were expressed as 141 kDa and 137 kDa insoluble His-tagged fusion proteins, respectively, in Escherichia coli M15 cells via pQE-30 vector, purified through Ni-NTA chromatography and polyclonal antibodies were raised. Immunoblot analysis of purified polyhedra, virion particles and virus infected mid-gut cells with the raised anti-p137 and anti-p141 antibodies showed specific immunoreactive bands and suggest that S1 and S3 may code for viral structural proteins. Expression of S1 and S3 ORFs in insect cells via baculovirus recombinants showed to produce viral like particles (VLPs by transmission electron microscopy. Immunogold staining showed that S3 encoded proteins self assembled to form viral outer capsid and VLPs maintained their stability at different pH in presence of S1 encoded protein. Conclusion Our results of cloning, sequencing and functional analysis of AmCPV S1 and S3 indicate that S3

Genetically-engineered baculovirus pesticides and their environmental safety

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H. Alan Wood; Yu Zailin

1991-01-01

Baculoviruses such as the Lymantria dispar nuclear polyhedrosis virus (LdMNPV) are ecologically attractive alternatives to chemical insect pesticides but have a slow rate of control. To overcome this we have developed and are field testing an environmentally acceptable strategy which can be used for the introduction and expression of pesticide-...

Inactivation of certain insect pathogens by ultraviolet radiation

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Krieg, A.; Groener, A.; Huber, J.; Zimmermann, G.

1981-01-01

The UV-sensitivity of two baculoviruses (granulosis virus, nuclear polyhedrosis virus) and two entomopathogenic microorganisms (Bacillus thuringiensis, Beauveria bassiana) was determined by radiation tests. In the far UV (254 nm) the stability, measured at an inactivation rate of 99%, was in declining order: nuclear polyhedra >= conidia of B. bassiana > granula > spores of B. thuringiensis >= vegetative cells of B. thuringiensis. In the near UV (285-380 nm) the following order could be found: conidia of B. bassiana >= nuclear polyhedra > spores of B. thuringiensis >= granula > vegetative cells of B. thuringiensis. Far UV had a much higher germicidal effect for all pathogens tested than near UV.

Microarray and RT-PCR screening for white spot syndrome virus immediate-early genes in cycloheximide-treated shrimp

International Nuclear Information System (INIS)

Liu Wangjing; Chang Yunshiang; Wang Chunghsiung; Kou, Guang-Hsiung; Lo Chufang

2005-01-01

Here, we report for the first time the successful use of cycloheximide (CHX) as an inhibitor to block de novo viral protein synthesis during WSSV (white spot syndrome virus) infection. Sixty candidate IE (immediate-early) genes were identified using a global analysis microarray technique. RT-PCR showed that the genes corresponding to ORF126, ORF242 and ORF418 in the Taiwan isolate were consistently CHX-insensitive, and these genes were designated ie1, ie2 and ie3, respectively. The sequences for these IE genes also appear in the two other WSSV isolates that have been sequenced. Three corresponding ORFs were identified in the China WSSV isolate, but only an ORF corresponding to ie1 was predicted in the Thailand isolate. In a promoter activity assay in Sf9 insect cells using EGFP (enhanced green fluorescence protein) as a reporter, ie1 showed very strong promoter activity, producing higher EGFP signals than the insect Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus (OpMNPV) ie2 promoter

Controle da lagarta-da-soja com aplicações de seu vírus de poliedrose nuclear por vias aérea e terrestre Control of the velvetbean caterpillar through air and land applications of its nuclear polyhedrosis virus

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SÉRGIO ARCE GOMEZ

2000-03-01

Full Text Available De 1983 a 1988 foram conduzidos, na região de Dourados, MS, seis experimentos e três campos-piloto, objetivando controlar a lagarta Anticarsia gemmatalis Hübner, 1818, com aplicações aérea e terrestre de seu vírus de poliedrose nuclear (VPN Ag. Cem lagartas equivalentes (LE de VPN Ag associadas a óleo de soja, melaço de cana-de-açúcar e água, foram aplicadas com avião agrícola equipado com Micronair. Os preparados oleosos (5,5 e 5 L ha-1 e com melaço (10 L ha-1 controlaram 75-89% e 79-96% das lagartas, respectivamente. A suspensão aquosa de 3 L ha-1 foi ineficaz, porém as de 15, 20 e 25 L ha-1 controlaram de 81% a 90% das lagartas. Cinqüenta LE, aplicadas com avião agrícola (3 L ha-1 ou atomizador (15 L ha-1, foram ineficientes. Aplicações da mesma dose com pulverizador de barra (134 e 150 L ha-1 proporcionaram controle de 87% e 90%, respectivamente, e com avião (15, 20 e 25 L ha-1, entre 93% e 98%. Aplicações aéreas de 50 LE com óleo de soja (5 L ha-1 ou melaço (10 L ha-1 foram eficientes (86-88% e 99%, respectivamente. Aplicações aéreas de suspensões aquosas e formulado oleoso, em campos-piloto, confirmaram os resultados experimentais.From 1983 to 1988 six experiments and three pilot fields were carried out at Dourados, Mato Grosso do Sul State, Brazil, aimed at controlling Anticarsia gemmatalis Hübner, 1818 (Lepidoptera: Noctuidae larvae through air and land applications of its nuclear polyhedrosis virus (Ag NPV. One hundred larval equivalents (LE of NPV were applied, with soybean oil, sugar cane molasses and water, with an Ipanema spraying plane equipped with Micronair nozzles. The oil (5.5 and 5 L ha-1 and molasses (10 L ha-1 preparations yielded 75-89% and 79-96% control, respectively. The use of aqueous formulation (3 L ha-1 didn't provide good control, but 15, 20 and 25 L ha-1 were effective (81-90%. Fifty LE applied by plane at 3 L ha-1 or by a tractor propelled atomizer (15 L ha-1 was inefficient. Fifty

Inactivation of certain insect pathogens by ultraviolet radiation

International Nuclear Information System (INIS)

Krieg, A.; Groener, A.; Huber, J.; Zimmermann, G.

1981-01-01

The UV-sensitivity of two baculoviruses (granulosis virus, nuclear polyhedrosis virus) and two entomopathogenic microorganisms (Bacillus thuringiensis, Beauveria bassiana) was determined by radiation tests. In the far UV (254 nm) the stability, measured at an inactivation rate of 99%, was in declining order: nuclear polyhedra >= conidia of B. bassiana > granula > spores of B. thuringiensis >= vegetative cells of B. thuringiensis. In the near UV (285-380 nm) the following order could be found: conidia of B. bassiana >= nuclear polyhedra > spores of B. thuringiensis >= granula > vegetative cells of B. thuringiensis. Far UV had a much higher germicidal effect for all pathogens tested than near UV. (orig.) [de

Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses

International Nuclear Information System (INIS)

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. - 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

Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses

Energy Technology Data Exchange (ETDEWEB)

Lloyd, Richard E., E-mail: rlloyd@bcm.edu

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.

Localization of influenza virus proteins to nuclear dot 10 structures in influenza virus-infected cells

International Nuclear Information System (INIS)

Sato, Yoshiko; Yoshioka, Kenichi; Suzuki, Chie; Awashima, Satoshi; Hosaka, Yasuhiro; Yewdell, Jonathan; Kuroda, Kazumichi

2003-01-01

We studied influenza virus M1 protein by generating HeLa and MDCK cell lines that express M1 genetically fused to green fluorescent protein (GFP). GFP-M1 was incorporated into virions produced by influenza virus infected MDCK cells expressing the fusion protein indicating that the fusion protein is at least partially functional. Following infection of either HeLa or MDCK cells with influenza A virus (but not influenza B virus), GFP-M1 redistributes from its cytosolic/nuclear location and accumulates in nuclear dots. Immunofluorescence revealed that the nuclear dots represent nuclear dot 10 (ND10) structures. The colocalization of authentic M1, as well as NS1 and NS2 protein, with ND10 was confirmed by immunofluorescence following in situ isolation of ND10. These findings demonstrate a previously unappreciated involvement of influenza virus with ND10, a structure involved in cellular responses to immune cytokines as well as the replication of a rapidly increasing list of viruses

Old foes, new understandings: nuclear entry of small non-enveloped DNA viruses.

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Fay, Nikta; Panté, Nelly

2015-06-01

The nuclear import of viral genomes is an important step of the infectious cycle for viruses that replicate in the nucleus of their host cells. Although most viruses use the cellular nuclear import machinery or some components of this machinery, others have developed sophisticated ways to reach the nucleus. Some of these have been known for some time; however, recent studies have changed our understanding of how some non-enveloped DNA viruses access the nucleus. For example, parvoviruses enter the nucleus through small disruptions of the nuclear membranes and nuclear lamina, and adenovirus tugs at the nuclear pore complex, using kinesin-1, to disassemble their capsids and deliver viral proteins and genomes into the nucleus. Here we review recent findings of the nuclear import strategies of three small non-enveloped DNA viruses, including adenovirus, parvovirus, and the polyomavirus simian virus 40. Copyright © 2015 Elsevier B.V. All rights reserved.

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

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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. Copyright © 2012 Elsevier B.V. All rights reserved.

Outer nuclear membrane fusion of adjacent nuclei in varicella-zoster virus-induced syncytia.

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Wang, Wei; Yang, Lianwei; Huang, Xiumin; Fu, Wenkun; Pan, Dequan; Cai, Linli; Ye, Jianghui; Liu, Jian; Xia, Ningshao; Cheng, Tong; Zhu, Hua

2017-12-01

Syncytia formation has been considered important for cell-to-cell spread and pathogenesis of many viruses. As a syncytium forms, individual nuclei often congregate together, allowing close contact of nuclear membranes and possibly fusion to occur. However, there is currently no reported evidence of nuclear membrane fusion between adjacent nuclei in wild-type virus-induced syncytia. Varicella-zoster virus (VZV) is one typical syncytia-inducing virus that causes chickenpox and shingles in humans. Here, we report, for the first time, an interesting observation of apparent fusion of the outer nuclear membranes from juxtaposed nuclei that comprise VZV syncytia both in ARPE-19 human epithelial cells in vitro and in human skin xenografts in the SCID-hu mouse model in vivo. This work reveals a novel aspect of VZV-related cytopathic effect in the context of multinucleated syncytia. Additionally, the information provided by this study could be helpful for future studies on interactions of viruses with host cell nuclei. Copyright © 2017 Elsevier Inc. All rights reserved.

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

International Nuclear Information System (INIS)

Lee, Changhee; Hodgins, Douglas; Calvert, Jay G.; Welch, Siao-Kun W.; Jolie, Rika; Yoo, Dongwan

2006-01-01

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

How baculovirus polyhedra fit square pegs into round holes to robustly package viruses.

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Ji, Xiaoyun; Sutton, Geoff; Evans, Gwyndaf; Axford, Danny; Owen, Robin; Stuart, David I

2010-01-20

Natural protein crystals (polyhedra) armour certain viruses, allowing them to survive for years under hostile conditions. We have determined the structure of polyhedra of the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV), revealing a highly symmetrical covalently cross-braced robust lattice, the subunits of which possess a flexible adaptor enabling this supra-molecular assembly to specifically entrap massive baculoviruses. Inter-subunit chemical switches modulate the controlled release of virus particles in the unusual high pH environment of the target insect's gut. Surprisingly, the polyhedrin subunits are more similar to picornavirus coat proteins than to the polyhedrin of cytoplasmic polyhedrosis virus (CPV). It is, therefore, remarkable that both AcMNPV and CPV polyhedra possess identical crystal lattices and crystal symmetry. This crystalline arrangement must be particularly well suited to the functional requirements of the polyhedra and has been either preserved or re-selected during evolution. The use of flexible adaptors to generate a powerful system for packaging irregular particles is characteristic of the AcMNPV polyhedrin and may provide a vehicle to sequester a wide range of objects such as biological nano-particles.

Radiation enhanced reactivation of nuclear replicating mammalian viruses

International Nuclear Information System (INIS)

Bockstahler, L.E.; Lytle, C.D.

1977-01-01

When CV-1 monkey kidney cells were UV-irradiated (0 to 18 J/m 2 ) or X-irradiated (0 to 10 krads) before infection with UV-irradiated simian adenovirus 7 (SA7) or simian virus 40 (SV40), increases in the infectivity of these nuclear replicating viruses as measured by plaque formation were observed. These radiation enhanced reactivations, UV enhanced reactivation (UVER) and X-ray enhanced reactivation (X-ray ER), occurred both when virus infection immediately followed irradiation of the cells (except for X-ray ER with SA7) and when virus infection was delayed until 3 to 5 days after cell irradiation. While there was little difference in the levels of reactivation of UV-irradiated SV40 between immediate and delayed infection, delayed infection resulted in higher levels of reactivation of SA7. X-ray enhanced reactivation of UV-irradiated Herpes simplex virus persisted for several days but did not increase. Thus, X-ray enhanced and UV enhanced reactivations of these mammalian viruses were relatively long-lived effects. Essentially no UVER or X-ray ER was found in CV-1 cells for either immediate or delayed infection with UV-irradiated vaccinia virus or poliovirus, both of which replicate in the cell cytoplasm. These results suggest UVER and X-ray ER in mammalian cells may be restricted to viruses which are replicated in the cell nucleus. (author)

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

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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. Copyright © 2014 Elsevier Inc. All rights reserved.

Natural occurrence of baculoviruses in populations of some Heliconiini (Lepidoptera; Nymphalidae with symptomatological notes

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G. F. S. Andrade

1983-01-01

Full Text Available Natural occurrence of nuclear polyhedrosis viruses were detected in populations of some Heliconiini in the field as well as in the laboratory. The epizootics appeared under field conditions in populations of Dione juno juno, D. moneta and Agraulis vanillae maculosa. In the laboratory, however, larvae of Heliconius numata mirus, H. hecale vetustus and H. erato phyllis in addition to two hybrids and Eueides Isabella dianasa, all suffered the same disease. The effect of several factors which might contribute to the occurrence of the disease are discussed. Symptoms, histopathology and description of viral particles and polyhedra are given.

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

Energy Technology Data Exchange (ETDEWEB)

Lalime, Erin N.; Pekosz, Andrew, E-mail: apekosz@jhsph.edu

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.

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

International Nuclear Information System (INIS)

Lalime, Erin N.; Pekosz, Andrew

2014-01-01

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

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

Energy Technology Data Exchange (ETDEWEB)

Maric, Martina; Haugo, Alison C. [Department of Microbiology, University of Iowa, Iowa City, IA 52242 (United States); Dauer, William [Department of Neurology, University of Michigan, Ann Arbor, MI 48109 (United States); Johnson, David [Department of Microbiology and Immunology, Oregon Health Sciences University, Portland, OR 97201 (United States); Roller, Richard J., E-mail: richard-roller@uiowa.edu [Department of Microbiology, University of Iowa, Iowa City, IA 52242 (United States)

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.

AcMNPV ac143 (odv-e18) is essential for mediating budded virus production and is the 30th baculovirus core gene

International Nuclear Information System (INIS)

McCarthy, Christina B.; Theilmann, David A.

2008-01-01

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) ac143 (odv-e18) is a late gene that encodes for a predicted 9.6 kDa structural protein that locates to the occlusion derived viral envelope and viral induced intranuclear microvesicles [Braunagel, S.C., He, H., Ramamurthy, P., and Summers, M.D. (1996). Transcription, translation, and cellular localization of three Autographa californica nuclear polyhedrosis virus structural proteins: ODV-E18, ODV-E35, and ODV-EC27. Virology 222, 100-114.]. In this study we demonstrate that ac143 is actually a previously unrecognized core gene and that it is essential for mediating budded virus production. To examine the role of ac143 in the baculovirus life cycle, we used the AcMNPV bacmid system to generate an ac143 knockout (KO) virus (AcBAC ac142REP-ac143KO ). Fluorescence and light microscopy showed that infection by AcBAC ac142REP-ac143KO is limited to a single cell and titration assays confirmed that AcBAC ac142REP-ac143KO was unable to produce budded virus (BV). Progression to very late phases of the viral infection was evidenced by the development of occlusion bodies in the nuclei of transfected cells. This correlated with the fact that viral DNA replication was unaffected in AcBAC ac142REP-ac143KO transfected cells. The entire ac143 promoter, which includes three late promoter motifs, is contained within the ac142 open reading frame. Different deletion mutants of this region showed that the integrity of the ac142-ac143 core gene cluster was required for the bacmids to display wild-type patterns of viral replication, BV production and RNA transcription

Ubiquitin-regulated nuclear-cytoplasmic trafficking of the Nipah virus matrix protein is important for viral budding.

Directory of Open Access Journals (Sweden)

Yao E Wang

2010-11-01

Full Text Available Paramyxoviruses are known to replicate in the cytoplasm and bud from the plasma membrane. Matrix is the major structural protein in paramyxoviruses that mediates viral assembly and budding. Curiously, the matrix proteins of a few paramyxoviruses have been found in the nucleus, although the biological function associated with this nuclear localization remains obscure. We report here that the nuclear-cytoplasmic trafficking of the Nipah virus matrix (NiV-M protein and associated post-translational modification play a critical role in matrix-mediated virus budding. Nipah virus (NiV is a highly pathogenic emerging paramyxovirus that causes fatal encephalitis in humans, and is classified as a Biosafety Level 4 (BSL4 pathogen. During live NiV infection, NiV-M was first detected in the nucleus at early stages of infection before subsequent localization to the cytoplasm and the plasma membrane. Mutations in the putative bipartite nuclear localization signal (NLS and the leucine-rich nuclear export signal (NES found in NiV-M impaired its nuclear-cytoplasmic trafficking and also abolished NiV-M budding. A highly conserved lysine residue in the NLS served dual functions: its positive charge was important for mediating nuclear import, and it was also a potential site for monoubiquitination which regulates nuclear export of the protein. Concordantly, overexpression of ubiquitin enhanced NiV-M budding whereas depletion of free ubiquitin in the cell (via proteasome inhibitors resulted in nuclear retention of NiV-M and blocked viral budding. Live Nipah virus budding was exquisitely sensitive to proteasome inhibitors: bortezomib, an FDA-approved proteasome inhibitor for treating multiple myeloma, reduced viral titers with an IC(50 of 2.7 nM, which is 100-fold less than the peak plasma concentration that can be achieved in humans. This opens up the possibility of using an "off-the-shelf" therapeutic against acute NiV infection.

Radiation dose response of strand breaks in SINPV-DNA

International Nuclear Information System (INIS)

Zhang Chunxiang; Luo Daling; Li Mianfeng; Liu Xiaowei; Zeng Rong; Wang Xunzhang

1995-01-01

The Spodoplera litura Nuclear Polyhedrosis Viruses (SINPV) is a kind of insectile virus with a simple structure, in which a double helix DNA is encapsulated in a protein coat and there is no function of enzymatic repair. The SINPV samples in dry powdered form held in sealed plastic tube were irradiated by 1-100 kGy gamma rays. The single strand breaks (SSB) and double strand breaks (DSB) induced in SINPV after irradiation were measured by neutral and alkaline agarose gel electrophoresis. A dose-response function combining the responses of one-hit and two-hit events was used to describe the SSB and DSB dose-response curves. It is shown that the SSB are one-hit events and the DSB are the combination of both one-hit, and two-hit events, and two-hit events are predominant in the DSB process

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

International Nuclear Information System (INIS)

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

2013-01-01

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

Identification of a functional, CRM-1-dependent nuclear export signal in hepatitis C virus core protein.

Directory of Open Access Journals (Sweden)

Andrea Cerutti

Full Text Available Hepatitis C virus (HCV infection is a major cause of chronic liver disease worldwide. HCV core protein is involved in nucleocapsid formation, but it also interacts with multiple cytoplasmic and nuclear molecules and plays a crucial role in the development of liver disease and hepatocarcinogenesis. The core protein is found mostly in the cytoplasm during HCV infection, but also in the nucleus in patients with hepatocarcinoma and in core-transgenic mice. HCV core contains nuclear localization signals (NLS, but no nuclear export signal (NES has yet been identified.We show here that the aa(109-133 region directs the translocation of core from the nucleus to the cytoplasm by the CRM-1-mediated nuclear export pathway. Mutagenesis of the three hydrophobic residues (L119, I123 and L126 in the identified NES or in the sequence encoding the mature core aa(1-173 significantly enhanced the nuclear localisation of the corresponding proteins in transfected Huh7 cells. Both the NES and the adjacent hydrophobic sequence in domain II of core were required to maintain the core protein or its fragments in the cytoplasmic compartment. Electron microscopy studies of the JFH1 replication model demonstrated that core was translocated into the nucleus a few minutes after the virus entered the cell. The blockade of nucleocytoplasmic export by leptomycin B treatment early in infection led to the detection of core protein in the nucleus by confocal microscopy and coincided with a decrease in virus replication.Our data suggest that the functional NLS and NES direct HCV core protein shuttling between the cytoplasmic and nuclear compartments, with at least some core protein transported to the nucleus. These new properties of HCV core may be essential for virus multiplication and interaction with nuclear molecules, influence cell signaling and the pathogenesis of HCV infection.

Identification of a functional, CRM-1-dependent nuclear export signal in hepatitis C virus core protein.

Science.gov (United States)

Cerutti, Andrea; Maillard, Patrick; Minisini, Rosalba; Vidalain, Pierre-Olivier; Roohvand, Farzin; Pecheur, Eve-Isabelle; Pirisi, Mario; Budkowska, Agata

2011-01-01

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease worldwide. HCV core protein is involved in nucleocapsid formation, but it also interacts with multiple cytoplasmic and nuclear molecules and plays a crucial role in the development of liver disease and hepatocarcinogenesis. The core protein is found mostly in the cytoplasm during HCV infection, but also in the nucleus in patients with hepatocarcinoma and in core-transgenic mice. HCV core contains nuclear localization signals (NLS), but no nuclear export signal (NES) has yet been identified.We show here that the aa(109-133) region directs the translocation of core from the nucleus to the cytoplasm by the CRM-1-mediated nuclear export pathway. Mutagenesis of the three hydrophobic residues (L119, I123 and L126) in the identified NES or in the sequence encoding the mature core aa(1-173) significantly enhanced the nuclear localisation of the corresponding proteins in transfected Huh7 cells. Both the NES and the adjacent hydrophobic sequence in domain II of core were required to maintain the core protein or its fragments in the cytoplasmic compartment. Electron microscopy studies of the JFH1 replication model demonstrated that core was translocated into the nucleus a few minutes after the virus entered the cell. The blockade of nucleocytoplasmic export by leptomycin B treatment early in infection led to the detection of core protein in the nucleus by confocal microscopy and coincided with a decrease in virus replication.Our data suggest that the functional NLS and NES direct HCV core protein shuttling between the cytoplasmic and nuclear compartments, with at least some core protein transported to the nucleus. These new properties of HCV core may be essential for virus multiplication and interaction with nuclear molecules, influence cell signaling and the pathogenesis of HCV infection.

Serotype-specific Differences in Dengue Virus Non-structural Protein 5 Nuclear Localization*

Science.gov (United States)

Hannemann, Holger; Sung, Po-Yu; Chiu, Han-Chen; Yousuf, Amjad; Bird, Jim; Lim, Siew Pheng; Davidson, Andrew D.

2013-01-01

The four serotypes of dengue virus (DENV-1 to -4) cause the most important arthropod-borne viral disease of humans. DENV non-structural protein 5 (NS5) contains enzymatic activities required for capping and replication of the viral RNA genome that occurs in the host cytoplasm. However, previous studies have shown that DENV-2 NS5 accumulates in the nucleus during infection. In this study, we examined the nuclear localization of NS5 for all four DENV serotypes. We demonstrate for the first time that there are serotypic differences in NS5 nuclear localization. Whereas the DENV-2 and -3 proteins accumulate in the nucleus, DENV-1 and -4 NS5 are predominantly if not exclusively localized to the cytoplasm. Comparative studies on the DENV-2 and -4 NS5 proteins revealed that the difference in DENV-4 NS5 nuclear localization was not due to rapid nuclear export but rather the lack of a functional nuclear localization sequence. Interaction studies using DENV-2 and -4 NS5 and human importin-α isoforms failed to identify an interaction that supported the differential nuclear localization of NS5. siRNA knockdown of the human importin-α isoform KPNA2, corresponding to the murine importin-α isoform previously shown to bind to DENV-2 NS5, did not substantially affect DENV-2 NS5 nuclear localization, whereas knockdown of importin-β did. The serotypic differences in NS5 nuclear localization did not correlate with differences in IL-8 gene expression. The results show that NS5 nuclear localization is not strictly required for virus replication but is more likely to have an auxiliary function in the life cycle of specific DENV serotypes. PMID:23770669

Serotype-specific differences in dengue virus non-structural protein 5 nuclear localization.

Science.gov (United States)

Hannemann, Holger; Sung, Po-Yu; Chiu, Han-Chen; Yousuf, Amjad; Bird, Jim; Lim, Siew Pheng; Davidson, Andrew D

2013-08-02

The four serotypes of dengue virus (DENV-1 to -4) cause the most important arthropod-borne viral disease of humans. DENV non-structural protein 5 (NS5) contains enzymatic activities required for capping and replication of the viral RNA genome that occurs in the host cytoplasm. However, previous studies have shown that DENV-2 NS5 accumulates in the nucleus during infection. In this study, we examined the nuclear localization of NS5 for all four DENV serotypes. We demonstrate for the first time that there are serotypic differences in NS5 nuclear localization. Whereas the DENV-2 and -3 proteins accumulate in the nucleus, DENV-1 and -4 NS5 are predominantly if not exclusively localized to the cytoplasm. Comparative studies on the DENV-2 and -4 NS5 proteins revealed that the difference in DENV-4 NS5 nuclear localization was not due to rapid nuclear export but rather the lack of a functional nuclear localization sequence. Interaction studies using DENV-2 and -4 NS5 and human importin-α isoforms failed to identify an interaction that supported the differential nuclear localization of NS5. siRNA knockdown of the human importin-α isoform KPNA2, corresponding to the murine importin-α isoform previously shown to bind to DENV-2 NS5, did not substantially affect DENV-2 NS5 nuclear localization, whereas knockdown of importin-β did. The serotypic differences in NS5 nuclear localization did not correlate with differences in IL-8 gene expression. The results show that NS5 nuclear localization is not strictly required for virus replication but is more likely to have an auxiliary function in the life cycle of specific DENV serotypes.

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

International Nuclear Information System (INIS)

Wanitchang, Asawin; Narkpuk, Jaraspim; Jongkaewwattana, Anan

2013-01-01

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

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

Energy Technology Data Exchange (ETDEWEB)

Wanitchang, Asawin; Narkpuk, Jaraspim; Jongkaewwattana, Anan, E-mail: anan.jon@biotec.or.th

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.

Detection of antibodies against porcine parvovirus nonstructural protein NS1 may distinguish between vaccinated and infected pigs

DEFF Research Database (Denmark)

Madsen, Eva Smedegaard; Madsen, Knud Gert; Nielsen, Jens

1997-01-01

The humoral antibody response against the nonstructural protein NS1 and the structural protein VP2 of porcine parvovirus (PPV) was evaluated by immuno-peroxidase test (IPT) and enzyme linked immune sorbent assay (ELISA) using recombinant PPV antigens. The coding sequence for NS1 and VP2...... was inserted into the baculovirus Autographa californica nuclear polyhedrosis virus (AcNPV) genome resulting in two recombinant baculoviruses AcNPV-NS1 and AcNPV-VP2, respectively. Sf9 cells (Spodoptora frugidiperda) inoculated with AcNPV-NS1 producing recombinant nonstructural protein (rNS1) and AcNPV-VP2...... producing recombinant virion protein (rVP2) were used in IPT and ELISA to analyse serum antibodies. Pigs vaccinated with an inactivated whole virus vaccine and experimentally infected pigs were studied. Significant titers against rVP2 were obtained in both vaccinated and infected pigs. Specific antibodies...

Occurrence, pathology, and ultrastructure of iridovirus and cytoplasmic polyhedrosis viruses in daphnids from the Czech Republic

Czech Academy of Sciences Publication Activity Database

Vávra, Jiří; Bílý, Tomáš; Nebesářová, Jana; Federici, B. A.

2016-01-01

Roč. 140, Jul 19 (2016), s. 35-38 ISSN 0022-2011 R&D Projects: GA MŠk LM2015062; GA TA ČR(CZ) TE01020118 Institutional support: RVO:60077344 Keywords : iridovirus * cytoplasmic polhedrosis virus * Crustacean * Daphnid * geographical occurrence * Czech Republic * AF microscopy * electron tomography Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.379, year: 2016

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

International Nuclear Information System (INIS)

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

2013-01-01

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

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

Energy Technology Data Exchange (ETDEWEB)

Sanchez-Aparicio, Maria Teresa; Rosas, Maria Flora [Centro de Biología Molecular, “Severo Ochoa” (CSIC-UAM), Cantoblanco 28049, Madrid (Spain); Sobrino, Francisco, E-mail: fsobrino@cbm.uam.es [Centro de Biología Molecular, “Severo Ochoa” (CSIC-UAM), Cantoblanco 28049, Madrid (Spain); Centro de Investigación en Sanidad Animal, INIA, Valdeolmos, 28130 Madrid (Spain)

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.

CD151, a novel host factor of nuclear export signaling in influenza virus infection.

Science.gov (United States)

Qiao, Yongkang; Yan, Yan; Tan, Kai Sen; Tan, Sheryl S L; Seet, Ju Ee; Arumugam, Thiruma Valavan; Chow, Vincent T K; Wang, De Yun; Tran, Thai

2018-05-01

Despite advances in our understanding of the mechanisms of influenza A virus (IAV) infection, the crucial virus-host interactions during the viral replication cycle still remain incomplete. Tetraspanin CD151 is highly expressed in the human respiratory tract, but its pathological role in IAV infection is unknown. We sought to characterize the functional role and mechanisms of action of CD151 in IAV infection of the upper and lower respiratory tracts with H1N1 and H3N2 strains. We used CD151-null mice in an in vivo model of IAV infection and clinical donor samples of in vitro-differentiated human nasal epithelial cells cultured at air-liquid interface. As compared with wild-type infected mice, CD151-null infected mice exhibited a significant reduction in virus titer and improvement in survival that is associated with pronounced host antiviral response and inflammasome activation together with accelerated lung repair. Interestingly, we show that CD151 complexes newly synthesized viral proteins with host nuclear export proteins and stabilizes microtubule complexes, which are key processes necessary for the polarized trafficking of viral progeny to the host plasma membrane for assembly. Our results provide new mechanistic insights into our understanding of IAV infection. We show that CD151 is a critical novel host factor of nuclear export signaling whereby the IAV nuclear export uses it to complement its own nuclear export proteins (a site not targeted by current therapy), making this regulation unique, and holds promise for the development of novel alternative/complementary strategies to reduce IAV severity. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Mutation of a Conserved Nuclear Export Sequence in Chikungunya Virus Capsid Protein Disrupts Host Cell Nuclear Import.

Science.gov (United States)

Jacobs, Susan C; Taylor, Adam; Herrero, Lara J; Mahalingam, Suresh; Fazakerley, John K

2017-10-20

Transmitted by mosquitoes; chikungunya virus (CHIKV) is responsible for frequent outbreaks of arthritic disease in humans. CHIKV is an arthritogenic alphavirus of the Togaviridae family. Capsid protein, a structural protein encoded by the CHIKV RNA genome, is able to translocate to the host cell nucleus. In encephalitic alphaviruses nuclear translocation induces host cell shut off; however, the role of capsid protein nuclear localisation in arthritogenic alphaviruses remains unclear. Using replicon systems, we investigated a nuclear export sequence (NES) in the N-terminal region of capsid protein; analogous to that found in encephalitic alphavirus capsid but uncharacterised in CHIKV. The chromosomal maintenance 1 (CRM1) export adaptor protein mediated CHIKV capsid protein export from the nucleus and a region within the N-terminal part of CHIKV capsid protein was required for active nuclear targeting. In contrast to encephalitic alphaviruses, CHIKV capsid protein did not inhibit host nuclear import; however, mutating the NES of capsid protein (∆NES) blocked host protein access to the nucleus. Interactions between capsid protein and the nucleus warrant further investigation.

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

International Nuclear Information System (INIS)

Alves, Carolina; Freitas, Natalia; Cunha, Celso

2008-01-01

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

Nucleocytoplasmic trafficking of Nipah virus W protein involves multiple discrete interactions with the nuclear import and export machinery

International Nuclear Information System (INIS)

Audsley, Michelle D.; Jans, David A.; Moseley, Gregory W.

2016-01-01

Paramyxoviruses replicate in the cytoplasm with no obvious requirement to interact with the nucleus. Nevertheless, the W protein of the highly lethal bat-borne paramyxovirus Nipah virus (NiV) is known to undergo specific targeting to the nucleus, mediated by a single nuclear localisation signal (NLS) within the C-terminal domain. Here, we report for the first time that additional sites modulate nucleocytoplasmic localisation of W. We show that the N-terminal domain interacts with importin α1 and contributes to nuclear accumulation of W, indicative of a novel N-terminal NLS. We also find that W undergoes exportin-1 mediated nuclear export, dependent on a leucine at position 174. Together, these data enable significant revision of the generally accepted model of W trafficking, with implications for understanding of the mechanisms of NiV immune evasion. - Highlights: • A new model for Nipah virus W protein nucleocytoplasmic trafficking is proposed. • Nipah W protein is shown to undergo active nuclear export via exportin-1. • Nipah W nuclear import is mediated by multiple nuclear localisation signals.

Functional analysis of the interaction of the human immunodeficiency virus type 1 Rev nuclear export signal with its cofactors

International Nuclear Information System (INIS)

Kiss, A.; Li, L.; Gettemeier, T.; Venkatesh, L.K.

2003-01-01

Human immunodeficiency virus type 1 (HIV-1) Rev-mediated nuclear export of viral RNAs involves the interaction of its leucine-rich nuclear export sequence (NES) with nuclear cofactors. In yeast two-hybrid screens of a human lymph node derived cDNA expression library, we identified the human nucleoporin Nup98 as a highly specific and potent interactor of the Rev NES. Using an extensive panel of nuclear export positive and negative mutants of the functionally homologous NESs of the HIV-1 Rev, human T cell leukemia virus type 1 (HTLV-1) Rex, and equine infectious anemia virus (EIAV) Rev proteins, physiologically significant interaction of hNup98 with the various NESs was demonstrated. Missense mutations in the yeast nuclear export factor Crm1p that abrogated Rev NES interaction with the XXFG repeat-containing nucleoporin, Rab/hRIP, had minimal effects on the interaction of GLFG repeat-containing hNup98. Functional analysis of Nup98 domains required for nuclear localization demonstrated that the entire ORF was required for efficient incorporation into the nuclear envelope. A putative nuclear localization signal was identified downstream of the GLFG repeat region. Whereas overexpression of both full-length Nup98 and the amino-terminal GLFG repeat region, but not the unique carboxy-terminal region, induced significant suppression of HIV unspliced RNA export, lower levels of exogenous Nup98 expression resulted in a relatively modest increase in unspliced RNA export. These results suggest a physiological role for hNup98 in modulating Rev-dependent RNA export during HIV infection

Nuclear imprisonment of host cellular mRNA by nsp1β protein of porcine reproductive and respiratory syndrome virus

International Nuclear Information System (INIS)

Han, Mingyuan; Ke, Hanzhong; Zhang, Qingzhan; Yoo, Dongwan

2017-01-01

Positive-strand RNA genomes function as mRNA for viral protein synthesis which is fully reliant on host cell translation machinery. Competing with cellular protein translation apparatus needs to ensure the production of viral proteins, but this also stifles host innate defense. In the present study, we showed that porcine reproductive and respiratory syndrome virus (PRRSV), whose replication takes place in the cytoplasm, imprisoned host cell mRNA in the nucleus, which suggests a novel mechanism to enhance translation of PRRSV genome. PRRSV nonstructural protein (nsp) 1β was identified as the nuclear protein playing the role for host mRNA nuclear retention and subversion of host protein synthesis. A SAP (SAF-A/B, Acinus, and PIAS) motif was identified in nsp1β with the consensus sequence of 126 -LQxxLxxxGL- 135 . In situ hybridization unveiled that SAP mutants were unable to cause nuclear retention of host cell mRNAs and did not suppress host protein synthesis. In addition, these SAP mutants reverted PRRSV-nsp1β-mediated suppression of interferon (IFN) production, IFN signaling, and TNF-α production pathway. Using reverse genetics, a series of SAP mutant PRRS viruses, vK124A, vL126A, vG134A, and vL135A were generated. No mRNA nuclear retention was observed during vL126A and vL135A infections. Importantly, vL126A and vL135A did not suppress IFN production. For other arteriviruses, mRNA nuclear accumulation was also observed for LDV-nsp1β and SHFV-nsp1β. EAV-nsp1 was exceptional and did not block the host mRNA nuclear export. - Highlights: •PRRS virus blocks host mRNA nuclear export to the cytoplasm. •PRRSV nsp1β is the viral protein responsible for host mRNA nuclear retention. •SAP domain in nsp1β is essential for host mRNA nuclear retention and type I interferon suppression. •Mutation in the SAP domain of nsp1β causes the loss of function. •Host mRNA nuclear retention by nsp1β is common in the family Arteriviridae, except equine arteritis virus.

Nuclear imprisonment of host cellular mRNA by nsp1β protein of porcine reproductive and respiratory syndrome virus

Energy Technology Data Exchange (ETDEWEB)

Han, Mingyuan, E-mail: hanming@umich.edu; Ke, Hanzhong; Zhang, Qingzhan; Yoo, Dongwan, E-mail: dyoo@illinois.edu

2017-05-15

Positive-strand RNA genomes function as mRNA for viral protein synthesis which is fully reliant on host cell translation machinery. Competing with cellular protein translation apparatus needs to ensure the production of viral proteins, but this also stifles host innate defense. In the present study, we showed that porcine reproductive and respiratory syndrome virus (PRRSV), whose replication takes place in the cytoplasm, imprisoned host cell mRNA in the nucleus, which suggests a novel mechanism to enhance translation of PRRSV genome. PRRSV nonstructural protein (nsp) 1β was identified as the nuclear protein playing the role for host mRNA nuclear retention and subversion of host protein synthesis. A SAP (SAF-A/B, Acinus, and PIAS) motif was identified in nsp1β with the consensus sequence of {sub 126}-LQxxLxxxGL-{sub 135}. In situ hybridization unveiled that SAP mutants were unable to cause nuclear retention of host cell mRNAs and did not suppress host protein synthesis. In addition, these SAP mutants reverted PRRSV-nsp1β-mediated suppression of interferon (IFN) production, IFN signaling, and TNF-α production pathway. Using reverse genetics, a series of SAP mutant PRRS viruses, vK124A, vL126A, vG134A, and vL135A were generated. No mRNA nuclear retention was observed during vL126A and vL135A infections. Importantly, vL126A and vL135A did not suppress IFN production. For other arteriviruses, mRNA nuclear accumulation was also observed for LDV-nsp1β and SHFV-nsp1β. EAV-nsp1 was exceptional and did not block the host mRNA nuclear export. - Highlights: •PRRS virus blocks host mRNA nuclear export to the cytoplasm. •PRRSV nsp1β is the viral protein responsible for host mRNA nuclear retention. •SAP domain in nsp1β is essential for host mRNA nuclear retention and type I interferon suppression. •Mutation in the SAP domain of nsp1β causes the loss of function. •Host mRNA nuclear retention by nsp1β is common in the family Arteriviridae, except equine

The Epstein-Barr virus nuclear antigen-6 protein co-localizes with EBNA-3 and survival of motor neurons protein

International Nuclear Information System (INIS)

Krauer, Kenia G.; Buck, Marion; Belzer, Deanna K.; Flanagan, James; Chojnowski, Grace M.; Sculley, Tom B.

2004-01-01

The Epstein-Barr virus nuclear antigen (EBNA)-6 protein is essential for Epstein-Barr virus (EBV)-induced immortalization of primary human B-lymphocytes in vitro. In this study, fusion proteins of EBNA-6 with green fluorescent protein (GFP) have been used to characterize its nuclear localization and organization within the nucleus. EBNA-6 associates with nuclear structures and in immunofluorescence demonstrate a punctate staining pattern. Herein, we show that the association of EBNA-6 with these nuclear structures was maintained throughout the cell cycle and with the use of GFP-E6 deletion mutants, that the region amino acids 733-808 of EBNA-6 contains a domain that can influence the association of EBNA-6 with these nuclear structures. Co-immunofluorescence and confocal analyses demonstrated that EBNA-6 and EBNA-3 co-localize in the nucleus of cells. Expression of EBNA-6, but not EBNA-3, caused a redistribution of nuclear survival of motor neurons protein (SMN) to the EBNA-6 containing nuclear structures resulting in co-localization of SMN with EBNA-6

Influenza A Virus NS1 Protein Promotes Efficient Nuclear Export of Unspliced Viral M1 mRNA.

Science.gov (United States)

Pereira, Carina F; Read, Eliot K C; Wise, Helen M; Amorim, Maria J; Digard, Paul

2017-08-01

Influenza A virus mRNAs are transcribed by the viral RNA-dependent RNA polymerase in the cell nucleus before being exported to the cytoplasm for translation. Segment 7 produces two major transcripts: an unspliced mRNA that encodes the M1 matrix protein and a spliced transcript that encodes the M2 ion channel. Export of both mRNAs is dependent on the cellular NXF1/TAP pathway, but it is unclear how they are recruited to the export machinery or how the intron-containing but unspliced M1 mRNA bypasses the normal quality-control checkpoints. Using fluorescent in situ hybridization to monitor segment 7 mRNA localization, we found that cytoplasmic accumulation of unspliced M1 mRNA was inefficient in the absence of NS1, both in the context of segment 7 RNPs reconstituted by plasmid transfection and in mutant virus-infected cells. This effect was independent of any major effect on steady-state levels of segment 7 mRNA or splicing but corresponded to a ∼5-fold reduction in the accumulation of M1. A similar defect in intronless hemagglutinin (HA) mRNA nuclear export was seen with an NS1 mutant virus. Efficient export of M1 mRNA required both an intact NS1 RNA-binding domain and effector domain. Furthermore, while wild-type NS1 interacted with cellular NXF1 and also increased the interaction of segment 7 mRNA with NXF1, mutant NS1 polypeptides unable to promote mRNA export did neither. Thus, we propose that NS1 facilitates late viral gene expression by acting as an adaptor between viral mRNAs and the cellular nuclear export machinery to promote their nuclear export. IMPORTANCE Influenza A virus is a major pathogen of a wide variety of mammalian and avian species that threatens public health and food security. A fuller understanding of the virus life cycle is important to aid control strategies. The virus has a small genome that encodes relatively few proteins that are often multifunctional. Here, we characterize a new function for the NS1 protein, showing that, as well as

Mapping the nuclear localization signal in the matrix protein of potato yellow dwarf virus.

Science.gov (United States)

Anderson, Gavin; Jang, Chanyong; Wang, Renyuan; Goodin, Michael

2018-05-01

The ability of the matrix (M) protein of potato yellow dwarf virus (PYDV) to remodel nuclear membranes is controlled by a di-leucine motif located at residues 223 and 224 of its primary structure. This function can be uncoupled from that of its nuclear localization signal (NLS), which is controlled primarily by lysine and arginine residues immediately downstream of the LL motif. In planta localization of green fluorescent protein fusions, bimolecular fluorescence complementation assays with nuclear import receptor importin-α1 and yeast-based nuclear import assays provided three independent experimental approaches to validate the authenticity of the M-NLS. The carboxy terminus of M is predicted to contain a nuclear export signal, which is belived to be functional, given the ability of M to bind the Arabidopsis nuclear export receptor 1 (XPO1). The nuclear shuttle activity of M has implications for the cell-to-cell movement of PYDV nucleocapsids, based upon its interaction with the N and Y proteins.

Nuclear Imprisonment: Viral Strategies to Arrest Host mRNA Nuclear Export

Directory of Open Access Journals (Sweden)

Beatriz M. A. Fontoura

2013-07-01

Full Text Available 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.

Nuclear Imprisonment: Viral Strategies to Arrest Host mRNA Nuclear Export

Science.gov (United States)

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

Phosphorylation of the Epstein-Barr virus nuclear antigen 2

DEFF Research Database (Denmark)

Grässer, F A; Göttel, S; Haiss, P

1992-01-01

A major in vivo phosphorylation site of the Epstein-Barr virus nuclear antigen 2 (EBNA-2) was found to be localized at the C-terminus of the protein. In vitro phosphorylation studies using casein kinase 1 (CK-1) and casein kinase 2 (CK-2) revealed that EBNA-2 is a substrate for CK-2, but not for CK......-1. The CK-2 specific phosphorylation site was localized in the 140 C-terminal amino acids using a recombinant trpE-C-terminal fusion protein. In a similar experiment, the 58 N-terminal amino acids expressed as a recombinant trpE-fusion protein were not phosphorylated. Phosphorylation of a synthetic...

Comparative analysis of seven viral nuclear export signals (NESs reveals the crucial role of nuclear export mediated by the third NES consensus sequence of nucleoprotein (NP in influenza A virus replication.

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Nopporn Chutiwitoonchai

Full Text Available The assembly of influenza virus progeny virions requires machinery that exports viral genomic ribonucleoproteins from the cell nucleus. Currently, seven nuclear export signal (NES consensus sequences have been identified in different viral proteins, including NS1, NS2, M1, and NP. The present study examined the roles of viral NES consensus sequences and their significance in terms of viral replication and nuclear export. Mutation of the NP-NES3 consensus sequence resulted in a failure to rescue viruses using a reverse genetics approach, whereas mutation of the NS2-NES1 and NS2-NES2 sequences led to a strong reduction in viral replication kinetics compared with the wild-type sequence. While the viral replication kinetics for other NES mutant viruses were also lower than those of the wild-type, the difference was not so marked. Immunofluorescence analysis after transient expression of NP-NES3, NS2-NES1, or NS2-NES2 proteins in host cells showed that they accumulated in the cell nucleus. These results suggest that the NP-NES3 consensus sequence is mostly required for viral replication. Therefore, each of the hydrophobic (Φ residues within this NES consensus sequence (Φ1, Φ2, Φ3, or Φ4 was mutated, and its viral replication and nuclear export function were analyzed. No viruses harboring NP-NES3 Φ2 or Φ3 mutants could be rescued. Consistent with this, the NP-NES3 Φ2 and Φ3 mutants showed reduced binding affinity with CRM1 in a pull-down assay, and both accumulated in the cell nucleus. Indeed, a nuclear export assay revealed that these mutant proteins showed lower nuclear export activity than the wild-type protein. Moreover, the Φ2 and Φ3 residues (along with other Φ residues within the NP-NES3 consensus were highly conserved among different influenza A viruses, including human, avian, and swine. Taken together, these results suggest that the Φ2 and Φ3 residues within the NP-NES3 protein are important for its nuclear export function

Human immunodeficiency virus type 1 KK26-27 matrix mutants display impaired infectivity, circularization and integration but not nuclear import

International Nuclear Information System (INIS)

Mannioui, Abdelkrim; Nelson, Elisabeth; Schiffer, Cecile

2005-01-01

We analyzed the role of human immunodeficiency virus (HIV)-1 matrix protein (MA) during the virus replication afferent phase. Single-round infection of H9 T lymphocytes showed that the combined mutation of MA Lys residues 26-27 in MA reported nuclear localization signal (NLS)-1 [Haffar, O.K., Popov, S., Dubrovsky, L., Agostini, I., Tang, H., Pushkarsky, T., Nadler, S.G., Bukrinsky, M., 2000. Two nuclear localization signals in the HIV-1 matrix protein regulate nuclear import of the HIV-1 pre-integration complex. J. Mol. Biol. 299 (2): 359-368] impaired infectivity, abrogated 2-LTR-circle formation and significantly reduced integration. However, the mutation did not affect viral DNA docking to chromatin in either interphasic or mitotic cells, indicating that MA N-terminal basic domain should not represent a major determinant of HIV-1 nuclear import in T lymphocytes. These data point to a previously unreported role of MA in the late, post-chromatin-binding, afferent phase of HIV-1 replication cycle

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

International Nuclear Information System (INIS)

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

2010-01-01

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 RBPJκ (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 RBPJκ 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 RBPJκ.

Humoral markers of active Epstein-Barr virus infection associate with anti-extractable nuclear antigen autoantibodies and plasma galectin-3 binding protein in systemic lupus erythematosus.

Science.gov (United States)

Rasmussen, N S; Nielsen, C T; Houen, G; Jacobsen, S

2016-12-01

We investigated if signs of active Epstein-Barr virus and cytomegalovirus infections associate with certain autoantibodies and a marker of type I interferon activity in patients with systemic lupus erythematosus. IgM and IgG plasma levels against Epstein-Barr virus early antigen diffuse and cytomegalovirus pp52 were applied as humoral markers of ongoing/recently active Epstein-Barr virus and cytomegalovirus infections, respectively. Plasma galectin-3 binding protein served as a surrogate marker of type I interferon activity. The measurements were conducted in 57 systemic lupus erythematosus patients and 29 healthy controls using ELISAs. Regression analyses and univariate comparisons were performed for associative evaluation between virus serology, plasma galectin-3 binding protein and autoantibodies, along with other clinical and demographic parameters. Plasma galectin-3 binding protein concentrations were significantly higher in systemic lupus erythematosus patients (P = 0.009) and associated positively with Epstein-Barr virus early antigen diffuse-directed antibodies and the presence of autoantibodies against extractable nuclear antigens in adjusted linear regressions (B = 2.02 and 2.02, P = 0.02 and P = 0.002, respectively). Furthermore, systemic lupus erythematosus patients with anti-extractable nuclear antigens had significantly higher antibody levels against Epstein-Barr virus early antigen diffuse (P = 0.02). Our study supports a link between active Epstein-Barr virus infections, positivity for anti-extractable nuclear antigens and increased plasma galectin-3 binding protein concentrations/type I interferon activity in systemic lupus erythematosus patients. © The Author(s) 2016.

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

Science.gov (United States)

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.

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

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Toshie Sakuma

2014-03-01

Full Text Available 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.

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

Science.gov (United States)

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.

Importin α1 is required for nuclear import of herpes simplex virus proteins and capsid assembly in fibroblasts and neurons

Science.gov (United States)

Anderson, Fenja; Rother, Franziska; Rudolph, Kathrin; Prank, Ute; Binz, Anne; Hügel, Stefanie; Hartmann, Enno; Bader, Michael; Bauerfeind, Rudolf; Sodeik, Beate

2018-01-01

Herpesviruses are large DNA viruses which depend on many nuclear functions, and therefore on host transport factors to ensure specific nuclear import of viral and host components. While some import cargoes bind directly to certain transport factors, most recruit importin β1 via importin α. We identified importin α1 in a small targeted siRNA screen to be important for herpes simplex virus (HSV-1) gene expression. Production of infectious virions was delayed in the absence of importin α1, but not in cells lacking importin α3 or importin α4. While nuclear targeting of the incoming capsids, of the HSV-1 transcription activator VP16, and of the viral genomes were not affected, the nuclear import of the HSV-1 proteins ICP4 and ICP0, required for efficient viral transcription, and of ICP8 and pUL42, necessary for DNA replication, were reduced. Furthermore, quantitative electron microscopy showed that fibroblasts lacking importin α1 contained overall fewer nuclear capsids, but an increased proportion of mature nuclear capsids indicating that capsid formation and capsid egress into the cytoplasm were impaired. In neurons, importin α1 was also not required for nuclear targeting of incoming capsids, but for nuclear import of ICP4 and for the formation of nuclear capsid assembly compartments. Our data suggest that importin α1 is specifically required for the nuclear localization of several important HSV1 proteins, capsid assembly, and capsid egress into the cytoplasm, and may become rate limiting in situ upon infection at low multiplicity or in terminally differentiated cells such as neurons. PMID:29304174

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

Energy Technology Data Exchange (ETDEWEB)

Lebrun, Marielle [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium); Thelen, Nicolas; Thiry, Marc [University of Liege (ULg), GIGA-Neurosciences, Laboratory of Cellular and Tissular Biology, Liege (Belgium); Riva, Laura; Ote, Isabelle; Condé, Claude; Vandevenne, Patricia [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium); Di Valentin, Emmanuel [University of Liege (ULg), GIGA-Viral Vectors Platform, Liege (Belgium); Bontems, Sébastien [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium); Sadzot-Delvaux, Catherine, E-mail: csadzot@ulg.ac.be [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium)

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.

NuMA and nuclear lamins are cleaved during viral infection - inhibition of caspase activity prevents cleavage and rescues HeLa cells from measles virus-induced but not from rhinovirus 1B-induced cell death

International Nuclear Information System (INIS)

Taimen, Pekka; Berghaell, Heidi; Vainionpaeae, Raija; Kallajoki, Markku

2004-01-01

Nuclear matrix is a structural framework of important nuclear processes. We studied the effect of two different types of viral infections on nuclear matrix. HeLa cells were infected with human rhinovirus 1B (HRV 1B) or measles virus (MV), and Nuclear Mitotic Apparatus protein (NuMA) and lamins A/C and B were used as markers for internal nuclear matrix and peripheral nuclear lamina, respectively. We show that NuMA, lamins, and poly(ADP-ribose) polymerase-1 are cleaved during viral infection in a virus family-specific manner suggesting that these viruses activate different sets of proteases. Morphologically, NuMA was excluded from the condensed chromatin, lamins showed a folded distribution, and both proteins finally remained around the nuclear fragments. A general caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (z-VAD-FMK) prevented the nuclear disintegration and the cleavage of the proteins studied. Interestingly, z-VAD-FMK rescued MV-infected but not HRV 1B-infected cells from cell death. These results show for the first time that NuMA and lamins are specific target proteins during virus-induced programmed cell death

Rift Valley fever virus NSS gene expression correlates with a defect in nuclear mRNA export.

Science.gov (United States)

Copeland, Anna Maria; Van Deusen, Nicole M; Schmaljohn, Connie S

2015-12-01

We investigated the localization of host mRNA during Rift Valley fever virus (RVFV) infection. Fluorescence in situ hybridization revealed that infection with RVFV altered the localization of host mRNA. mRNA accumulated in the nuclei of RVFV-infected but not mock-infected cells. Further, overexpression of the NSS gene, but not the N, GN or NSM genes correlated with mRNA nuclear accumulation. Nuclear accumulation of host mRNA was not observed in cells infected with a strain of RVFV lacking the gene encoding NSS, confirming that expression of NSS is likely responsible for this phenomenon. Published by Elsevier Inc.

Epstein–Barr virus glycoprotein gM can interact with the cellular protein p32 and knockdown of p32 impairs virus

International Nuclear Information System (INIS)

Changotra, Harish; Turk, Susan M.; Artigues, Antonio; Thakur, Nagendra; Gore, Mindy; Muggeridge, Martin I.; Hutt-Fletcher, Lindsey M.

2016-01-01

The Epstein–Barr virus glycoprotein complex gMgN has been implicated in assembly and release of fully enveloped virus, although the precise role that it plays has not been elucidated. We report here that the long predicted cytoplasmic tail of gM is not required for complex formation and that it interacts with the cellular protein p32, which has been reported to be involved in nuclear egress of human cytomegalovirus and herpes simplex virus. Although redistribution of p32 and colocalization with gM was not observed in virus infected cells, knockdown of p32 expression by siRNA or lentivirus-delivered shRNA recapitulated the phenotype of a virus lacking expression of gNgM. A proportion of virus released from cells sedimented with characteristics of virus lacking an intact envelope and there was an increase in virus trapped in nuclear condensed chromatin. The observations suggest the possibility that p32 may also be involved in nuclear egress of Epstein–Barr virus. - Highlights: • The predicted cytoplasmic tail of gM is not required to complex with gN. • Cellular p32 can interact with the predicted cytoplasmic tail of EBV gM. • Knockdown of p32 recapitulates the phenotype of virus lacking the gNgM complex.

Epstein–Barr virus glycoprotein gM can interact with the cellular protein p32 and knockdown of p32 impairs virus

Energy Technology Data Exchange (ETDEWEB)

Changotra, Harish; Turk, Susan M. [Department of Microbiology and Immunology, Center for Molecular and Tumor Virology and Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, LA (United States); Artigues, Antonio [Department of Biochemistry, University of Kansas Medical Center, Kansas City, KS (United States); Thakur, Nagendra; Gore, Mindy; Muggeridge, Martin I. [Department of Microbiology and Immunology, Center for Molecular and Tumor Virology and Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, LA (United States); Hutt-Fletcher, Lindsey M., E-mail: lhuttf@lsuhsc.edu [Department of Microbiology and Immunology, Center for Molecular and Tumor Virology and Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, LA (United States)

2016-02-15

The Epstein–Barr virus glycoprotein complex gMgN has been implicated in assembly and release of fully enveloped virus, although the precise role that it plays has not been elucidated. We report here that the long predicted cytoplasmic tail of gM is not required for complex formation and that it interacts with the cellular protein p32, which has been reported to be involved in nuclear egress of human cytomegalovirus and herpes simplex virus. Although redistribution of p32 and colocalization with gM was not observed in virus infected cells, knockdown of p32 expression by siRNA or lentivirus-delivered shRNA recapitulated the phenotype of a virus lacking expression of gNgM. A proportion of virus released from cells sedimented with characteristics of virus lacking an intact envelope and there was an increase in virus trapped in nuclear condensed chromatin. The observations suggest the possibility that p32 may also be involved in nuclear egress of Epstein–Barr virus. - Highlights: • The predicted cytoplasmic tail of gM is not required to complex with gN. • Cellular p32 can interact with the predicted cytoplasmic tail of EBV gM. • Knockdown of p32 recapitulates the phenotype of virus lacking the gNgM complex.

Immunohistochemical detection of virus through its nuclear cytopathic effect in idiopathic interstitial pneumonia other than acute exacerbation

Directory of Open Access Journals (Sweden)

G.C. dos Santos

2013-11-01

Full Text Available Idiopathic interstitial pneumonias include complex diseases that have a strong interaction between genetic makeup and environmental factors. However, in many cases, no infectious agent can be demonstrated, and these clinical diseases rapidly progress to death. Theoretically, idiopathic interstitial pneumonias could be caused by the Epstein-Barr virus, cytomegalovirus, adenovirus, hepatitis C virus, respiratory syncytial virus, and herpesvirus, which may be present in such small amounts or such configuration that routine histopathological analysis or viral culture techniques cannot detect them. To test the hypothesis that immunohistochemistry provides more accurate results than the mere histological demonstration of viral inclusions, this method was applied to 37 open lung biopsies obtained from patients with idiopathic interstitial pneumonias. As a result, immunohistochemistry detected measles virus and cytomegalovirus in diffuse alveolar damage-related histological patterns of acute exacerbation of idiopathic pulmonary fibrosis and nonspecific interstitial pneumonia in 38 and 10% of the cases, respectively. Alveolar epithelium infection by cytomegalovirus was observed in 25% of organizing pneumonia patterns. These findings were coincident with nuclear cytopathic effects but without demonstration of cytomegalovirus inclusions. These data indicate that diffuse alveolar damage-related cytomegalovirus or measles virus infections enhance lung injury, and a direct involvement of these viruses in diffuse alveolar damage-related histological patterns is likely. Immunohistochemistry was more sensitive than the histological demonstration of cytomegalovirus or measles virus inclusions. We concluded that all patients with diffuse alveolar damage-related histological patterns should be investigated for cytomegalovirus and measles virus using sensitive immunohistochemistry in conjunction with routine procedures.

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

International Nuclear Information System (INIS)

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

2009-01-01

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.

Rift Valley fever virus NS{sub S} gene expression correlates with a defect in nuclear mRNA export

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Copeland, Anna Maria; Van Deusen, Nicole M.; Schmaljohn, Connie S., E-mail: Connie.s.schmaljohn.civ@mail.mil

2015-12-15

We investigated the localization of host mRNA during Rift Valley fever virus (RVFV) infection. Fluorescence in situ hybridization revealed that infection with RVFV altered the localization of host mRNA. mRNA accumulated in the nuclei of RVFV-infected but not mock-infected cells. Further, overexpression of the NS{sub S} gene, but not the N, G{sub N} or NS{sub M} genes correlated with mRNA nuclear accumulation. Nuclear accumulation of host mRNA was not observed in cells infected with a strain of RVFV lacking the gene encoding NS{sub S}, confirming that expression of NS{sub S} is likely responsible for this phenomenon. - Highlights: • Rift Valley fever virus (RVFV) infection alters the localization of host mRNA. • mRNA accumulates in the nuclei of RVFV-infected but not mock-infected cells. • NS{sub S} is likely responsible for mRNA relocalization to the nucleus.

Parvoviral nuclear import: bypassing the host nuclear-transport machinery.

Science.gov (United States)

Cohen, Sarah; Behzad, Ali R; Carroll, Jeffrey B; Panté, Nelly

2006-11-01

The parvovirus Minute virus of mice (MVM) is a small DNA virus that replicates in the nucleus of its host cells. However, very little is known about the mechanisms underlying parvovirus' nuclear import. Recently, it was found that microinjection of MVM into the cytoplasm of Xenopus oocytes causes damage to the nuclear envelope (NE), suggesting that the nuclear-import mechanism of MVM involves disruption of the NE and import through the resulting breaks. Here, fluorescence microscopy and electron microscopy were used to examine the effect of MVM on host-cell nuclear structure during infection of mouse fibroblast cells. It was found that MVM caused dramatic changes in nuclear shape and morphology, alterations of nuclear lamin immunostaining and breaks in the NE of infected cells. Thus, it seems that the unusual nuclear-import mechanism observed in Xenopus oocytes is in fact used by MVM during infection of host cells.

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

International Nuclear Information System (INIS)

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

2008-01-01

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

A DNA Binding Protein Is Required for Viral Replication and Transcription in Bombyx mori Nucleopolyhedrovirus.

Science.gov (United States)

Zhao, Cui; Zhang, Chen; Chen, Bin; Shi, Yanghui; Quan, Yanping; Nie, Zuoming; Zhang, Yaozhou; Yu, Wei

2016-01-01

A DNA-binding protein (DBP) [GenBank accession number: M63416] of Bombyx mori nuclear polyhedrosis virus (BmNPV) has been reported to be a regulatory factor in BmNPV, but its detailed functions remain unknown. In order to study the regulatory mechanism of DBP on viral proliferation, genome replication, and gene transcription, a BmNPV dbp gene knockout virus dbp-ko-Bacmid was generated by the means of Red recombination system. In addition, dbp-repaired virus dbp-re-Bacmid was constructed by the means of the Bac to Bac system. Then, the Bacmids were transfected into BmN cells. The results of this viral titer experiment revealed that the TCID50 of the dbp-ko-Bacmid was 0; however, the dbp-re-Bacmid was similar to the wtBacmid (p>0.05), indicating that the dbp-deficient would lead to failure in the assembly of virus particles. In the next step, Real-Time PCR was used to analyze the transcriptional phases of dbp gene in BmN cells, which had been infected with BmNPV. The results of the latter experiment revealed that the transcript of dbp gene was first detected at 3 h post-infection. Furthermore, the replication level of virus genome and the transcriptional level of virus early, late, and very late genes in BmN cells, which had been transfected with 3 kinds of Bacmids, were analyzed by Real-Time PCR. The demonstrating that the replication level of genome was lower than that of wtBacmid and dbp-re-Bacmid (plife cycle.

Inhibition of human immunodeficiency virus type 1 (HIV-1) nuclear import via Vpr-Importin α interactions as a novel HIV-1 therapy

International Nuclear Information System (INIS)

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

2009-01-01

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 α, 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 α 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 α 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.

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

Science.gov (United States)

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

Nuclear TRIM25 Specifically Targets Influenza Virus Ribonucleoproteins to Block the Onset of RNA Chain Elongation.

Science.gov (United States)

Meyerson, Nicholas R; Zhou, Ligang; Guo, Yusong R; Zhao, Chen; Tao, Yizhi J; Krug, Robert M; Sawyer, Sara L

2017-11-08

TRIM25 is an E3 ubiquitin ligase that activates RIG-I to promote the antiviral interferon response. The NS1 protein from all strains of influenza A virus binds TRIM25, although not all virus strains block the interferon response, suggesting alternative mechanisms for TRIM25 action. Here we present a nuclear role for TRIM25 in specifically restricting influenza A virus replication. TRIM25 inhibits viral RNA synthesis through a direct mechanism that is independent of its ubiquitin ligase activity and the interferon pathway. This activity can be inhibited by the viral NS1 protein. TRIM25 inhibition of viral RNA synthesis results from its binding to viral ribonucleoproteins (vRNPs), the structures containing individual viral RNA segments, the viral polymerase, and multiple viral nucleoproteins. TRIM25 binding does not inhibit initiation of capped-RNA-primed viral mRNA synthesis by the viral polymerase. Rather, the onset of RNA chain elongation is inhibited because TRIM25 prohibits the movement of RNA into the polymerase complex. Copyright © 2017 Elsevier Inc. All rights reserved.

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

International Nuclear Information System (INIS)

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

2008-01-01

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

Sunlight persistence and rainfastness of spray-dried formulations of baculovirus isolated from Anagrapha falcifera (Lepidoptera: Noctuidae).

Science.gov (United States)

Tamez-Guerra, P; McGuire, M R; Behle, R W; Hamm, J J; Sumner, H R; Shasha, B S

2000-04-01

Nuclear polyhedrosis viruses such as the one isolated from the celery looper, Anagrapha falcifera (Kirby) (AfMNPV), have the potential to be successful bioinsecticides if improved formulations can prevent rapid loss of insecticidal activity from environmental conditions such as sunlight and rainfall. We tested 16 spray-dried formulations of AfMNPV to determine the effect of different ingredients (e.g., lignin, corn flour, and so on) on insecticidal activity after simulated rain and simulated sunlight (at Peoria, IL) and natural sunlight exposures (at Tifton, GA). The most effective formulation contained pregelatinized corn flour and potassium lignate, which retained more than half of its original activity after 5 cm of simulated rain, and almost full activity after 8 h of simulated sunlight. In Georgia, formulations made with and without lignin were compared for persistence of insecticidal activity when exposed to natural sunlight. In addition, the effect of fluorescent brighteners as formulation components and spray tank additives was tested. Results showed that the formulations with lignin had more insecticidal activity remaining after sunlight exposure than formulations without lignin. The inclusion of brighteners in the formulation did not improve initial activity or virus persistence. However, a 1% tank mix significantly enhanced activity and improved persistence. Scanning electron micrographs revealed discreet particles, and transmission electron micrographs showed virus embedded within microgranules. Results demonstrated that formulations made with natural ingredients could improve persistence of virus-based biopesticides.

Differential association with cellular substructures of pseudorabies virus DNA during early and late phases of replication

International Nuclear Information System (INIS)

Ben-Porat, T.; Veach, R.A.; Blankenship, M.L.; Kaplan, A.S.

1984-01-01

Pseudorabies virus DNA synthesis can be divided into two phases, early and late, which can be distinguished from each other on the basis of the structures of the replicating DNA. The two types of replicating virus DNA can also be distinguished from each other on the basis of the cellular substructures with which each is associated. Analysis by electron microscopic autoradiography showed that during the first round of replication, nascent virus DNA was found in the vicinity of the nuclear membrane; during later rounds of replication the nascent virus DNA was located centrally within the nucleus. The degree of association of virus DNA synthesized at early and late phases with the nuclear matrix fractions also differed; a larger proportion of late than of early nascent virus DNA was associated with this fraction. While nascent cellular DNA only was associated in significant amounts with the nuclear matrix fraction, a large part (up to 40%) of all the virus DNA remained associated with this fraction. However, no retention of specific virus proteins in this fraction was observed. Except for two virus proteins, which were preferentially extracted from the nuclear matrix, approximately 20% of all virus proteins remained in the nuclear matrix fraction. The large proportion of virus DNA associated with the nuclear fraction indicated that virus DNA may be intimately associated with some proteins

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

Energy Technology Data Exchange (ETDEWEB)

Bejerman, Nicolás, E-mail: n.bejerman@uq.edu.au [Instituto de Patología Vegetal (IPAVE), Centro de Investigaciones Agropecuarias (CIAP), Instituto Nacional de Tecnología Agropecuaria INTA, Camino a 60 Cuadras k 5,5, Córdoba X5020ICA (Argentina); Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St Lucia, QLD 4072 (Australia); Giolitti, Fabián; Breuil, Soledad de; Trucco, Verónica; Nome, Claudia; Lenardon, Sergio [Instituto de Patología Vegetal (IPAVE), Centro de Investigaciones Agropecuarias (CIAP), Instituto Nacional de Tecnología Agropecuaria INTA, Camino a 60 Cuadras k 5,5, Córdoba X5020ICA (Argentina); Dietzgen, Ralf G. [Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St Lucia, QLD 4072 (Australia)

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.

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

International Nuclear Information System (INIS)

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

2015-01-01

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

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

International Nuclear Information System (INIS)

Mannioui, Abdelkrim; Schiffer, Cecile; Felix, Nathalie

2004-01-01

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

Identification of a novel NLS of herpes simplex virus type 1 (HSV-1) VP19C and its nuclear localization is required for efficient production of HSV-1.

Science.gov (United States)

Li, You; Zhao, Lei; Wang, Shuai; Xing, Junji; Zheng, Chunfu

2012-09-01

Herpes simplex virus type 1 (HSV-1) triplex is a complex of three protein subunits, consisting of two copies of VP23 and one copy of VP19C. Here, we identified a non-classical NLS of VP19C between aa 50 and 61, and the nuclear import of VP19C was mediated by RanGTP and importin β1-, but not importin α5-, dependent pathway. Additionally, recombinant virus harbouring this NLS mutation (NLSm) replicates less efficiently as wild-type. These data strongly suggested that the nuclear import of VP19C is required for efficient HSV-1 production.

Pharmacological inhibition of feline immunodeficiency virus (FIV).

Science.gov (United States)

Mohammadi, Hakimeh; Bienzle, Dorothee

2012-05-01

Feline immunodeficiency virus (FIV) is a member of the retroviridae family of viruses and causes an acquired immunodeficiency syndrome (AIDS) in domestic and non-domestic cats worldwide. Genome organization of FIV and clinical characteristics of the disease caused by the virus are similar to those of human immunodeficiency virus (HIV). Both viruses infect T lymphocytes, monocytes and macrophages, and their replication cycle in infected cells is analogous. Due to marked similarity in genomic organization, virus structure, virus replication and disease pathogenesis of FIV and HIV, infection of cats with FIV is a useful tool to study and develop novel drugs and vaccines for HIV. Anti-retroviral drugs studied extensively in HIV infection have targeted different steps of the virus replication cycle: (1) inhibition of virus entry into susceptible cells at the level of attachment to host cell surface receptors and co-receptors; (2) inhibition of fusion of the virus membrane with the cell membrane; (3) blockade of reverse transcription of viral genomic RNA; (4) interruption of nuclear translocation and viral DNA integration into host genomes; (5) prevention of viral transcript processing and nuclear export; and (6) inhibition of virion assembly and maturation. Despite much success of anti-retroviral therapy slowing disease progression in people, similar therapy has not been thoroughly investigated in cats. In this article we review current pharmacological approaches and novel targets for anti-lentiviral therapy, and critically assess potentially suitable applications against FIV infection in cats.

Disruption of Microtubules Post-Virus Entry Enhances Adeno-Associated Virus Vector Transduction

Science.gov (United States)

Xiao, Ping-Jie; Mitchell, Angela M.; Huang, Lu; Li, Chengwen; Samulski, R. Jude

2016-01-01

Perinuclear retention of viral particles is a poorly understood phenomenon observed during many virus infections. In this study, we investigated whether perinuclear accumulation acts as a barrier to limit recombinant adeno-associated virus (rAAV) transduction. After nocodazole treatment to disrupt microtubules at microtubule-organization center (MT-MTOC) after virus entry, we observed higher rAAV transduction. To elucidate the role of MT-MTOC in rAAV infection and study its underlying mechanisms, we demonstrated that rAAV's perinuclear localization was retained by MT-MTOC with fluorescent analysis, and enhanced rAAV transduction from MT-MTOC disruption was dependent on the rAAV capsid's nuclear import signals. Interestingly, after knocking down RhoA or inhibiting its downstream effectors (ROCK and Actin), MT-MTOC disruption failed to increase rAAV transduction or nuclear entry. These data suggest that enhancement of rAAV transduction is the result of increased trafficking to the nucleus via the RhoA-ROCK-Actin pathway. Ten-fold higher rAAV transduction was also observed by disrupting MT-MTOC in brain, liver, and tumor in vivo. In summary, this study indicates that virus perinuclear accumulation at MT-MTOC is a barrier-limiting parameter for effective rAAV transduction and defines a novel defense mechanism by which host cells restrain viral invasion. PMID:26942476

Fast track, dynein-dependent nuclear targeting of human immunodeficiency virus Vpr protein; impaired trafficking in a clinical isolate

Energy Technology Data Exchange (ETDEWEB)

Caly, Leon [Department of Biochemistry and Molecular Biology, Monash University, Clayton, Vic. 3800 (Australia); Kassouf, Vicki T. [Centre for Virus Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145 (Australia); Moseley, Gregory W. [Department of Biochemistry and Molecular Biology, Monash University, Clayton, Vic. 3800 (Australia); Diefenbach, Russell J.; Cunningham, Anthony L. [Centre for Virus Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW 2145 (Australia); Jans, David A., E-mail: david.jans@monash.edu [Department of Biochemistry and Molecular Biology, Monash University, Clayton, Vic. 3800 (Australia)

2016-02-12

Nuclear import of the accessory protein Vpr is central to infection by human immunodeficiency virus (HIV). We previously identified the Vpr F72L mutation in a HIV-infected, long-term non-progressor, showing that it resulted in reduced Vpr nuclear accumulation and altered cytoplasmic localisation. Here we demonstrate for the first time that the effects of nuclear accumulation of the F72L mutation are due to impairment of microtubule-dependent-enhancement of Vpr nuclear import. We use high resolution imaging approaches including fluorescence recovery after photobleaching and other approaches to document interaction between Vpr and the dynein light chain protein, DYNLT1, and impaired interaction of the F72L mutant with DYNLT1. The results implicate MTs/DYNLT1 as drivers of Vpr nuclear import and HIV infection, with important therapeutic implications. - Highlights: • HIV-1 Vpr utilizes the microtubule network to traffic towards the nucleus. • Mechanism relies on interaction between Vpr and dynein light chain protein DYNLT1. • Long-term non-progressor derived mutation (F72L) impairs this interaction. • Key residues in the vicinity of F72 contribute to interaction with DYNLT1.

Fast track, dynein-dependent nuclear targeting of human immunodeficiency virus Vpr protein; impaired trafficking in a clinical isolate

International Nuclear Information System (INIS)

Caly, Leon; Kassouf, Vicki T.; Moseley, Gregory W.; Diefenbach, Russell J.; Cunningham, Anthony L.; Jans, David A.

2016-01-01

Nuclear import of the accessory protein Vpr is central to infection by human immunodeficiency virus (HIV). We previously identified the Vpr F72L mutation in a HIV-infected, long-term non-progressor, showing that it resulted in reduced Vpr nuclear accumulation and altered cytoplasmic localisation. Here we demonstrate for the first time that the effects of nuclear accumulation of the F72L mutation are due to impairment of microtubule-dependent-enhancement of Vpr nuclear import. We use high resolution imaging approaches including fluorescence recovery after photobleaching and other approaches to document interaction between Vpr and the dynein light chain protein, DYNLT1, and impaired interaction of the F72L mutant with DYNLT1. The results implicate MTs/DYNLT1 as drivers of Vpr nuclear import and HIV infection, with important therapeutic implications. - Highlights: • HIV-1 Vpr utilizes the microtubule network to traffic towards the nucleus. • Mechanism relies on interaction between Vpr and dynein light chain protein DYNLT1. • Long-term non-progressor derived mutation (F72L) impairs this interaction. • Key residues in the vicinity of F72 contribute to interaction with DYNLT1.

Nucleocytoplasmic transport of nucleocapsid proteins of enveloped RNA viruses

Directory of Open Access Journals (Sweden)

Wahyu eWulan

2015-06-01

Full Text Available Most viruses with non-segmented single stranded RNA genomes complete their life cycle in the cytoplasm of infected cells. However, despite undergoing replication in the cytoplasm, the structural proteins of some of these RNA viruses localize to the nucleus at specific times in the virus life cycle, primarily early in infection. Limited evidence suggests that this enhances successful viral replication by interfering with or inhibiting the host antiviral response. Nucleocapsid proteins of RNA viruses have a well-established, essential cytoplasmic role in virus replication and assembly. Intriguingly, nucleocapsid proteins of some RNA viruses also localize to the nucleus/nucleolus of infected cells. Their nuclear function is less well understood although significant advances have been made in recent years. This review will focus on the nucleocapsid protein of cytoplasmic enveloped RNA viruses, including their localization to the nucleus/nucleolus and function therein. A greater understanding of the nuclear localization of nucleocapsid proteins has the potential to enhance therapeutic strategies as it can be a target for the development of live-attenuated vaccines or antiviral drugs.

A Human Nuclear Shuttling Protein That Interacts with Human Immunodeficiency Virus Type 1 Matrix Is Packaged into Virions

Science.gov (United States)

Gupta, Kalpana; Ott, David; Hope, Thomas J.; Siliciano, Robert F.; Boeke, Jef D.

2000-01-01

Active nuclear import of the human immunodeficiency virus type 1 (HIV-1) preintegration complex (PIC) is essential for the productive infection of nondividing cells. Nuclear import of the PIC is mediated by the HIV-1 matrix protein, which also plays several critical roles during viral entry and possibly during virion production facilitating the export of Pr55Gag and genomic RNA. Using a yeast two-hybrid screen, we identified a novel human virion-associated matrix-interacting protein (VAN) that is highly conserved in vertebrates and expressed in most human tissues. Its expression is upregulated upon activation of CD4+ T cells. VAN is efficiently incorporated into HIV-1 virions and, like matrix, shuttles between the nucleus and cytoplasm. Furthermore, overexpression of VAN significantly inhibits HIV-1 replication in tissue culture. We propose that VAN regulates matrix nuclear localization and, by extension, both nuclear import of the PIC and export of Pr55Gag and viral genomic RNA during virion production. Our data suggest that this regulatory mechanism reflects a more global process for regulation of nucleocytoplasmic transport. PMID:11090181

Humoral markers of active Epstein-Barr virus infection associate with anti-extractable nuclear antigen autoantibodies and plasma galectin-3 binding protein in systemic lupus erythematosus

DEFF Research Database (Denmark)

Rasmussen, N S; Nielsen, C T; Houen, G

2016-01-01

We investigated if signs of active Epstein-Barr virus and cytomegalovirus infections associate with certain autoantibodies and a marker of type I interferon activity in patients with systemic lupus erythematosus. IgM and IgG plasma levels against Epstein-Barr virus early antigen diffuse...... and cytomegalovirus pp52 were applied as humoral markers of ongoing/recently active Epstein-Barr virus and cytomegalovirus infections, respectively. Plasma galectin-3 binding protein served as a surrogate marker of type I interferon activity. The measurements were conducted in 57 systemic lupus erythematosus patients...... concentrations were significantly higher in systemic lupus erythematosus patients (P = 0.009) and associated positively with Epstein-Barr virus early antigen diffuse-directed antibodies and the presence of autoantibodies against extractable nuclear antigens in adjusted linear regressions (B = 2.02 and 2.02, P...

Pharmacological Inhibition of Feline Immunodeficiency Virus (FIV

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Dorothee Bienzle

2012-04-01

Full Text Available Feline immunodeficiency virus (FIV is a member of the retroviridae family of viruses and causes an acquired immunodeficiency syndrome (AIDS in domestic and non-domestic cats worldwide. Genome organization of FIV and clinical characteristics of the disease caused by the virus are similar to those of human immunodeficiency virus (HIV. Both viruses infect T lymphocytes, monocytes and macrophages, and their replication cycle in infected cells is analogous. Due to marked similarity in genomic organization, virus structure, virus replication and disease pathogenesis of FIV and HIV, infection of cats with FIV is a useful tool to study and develop novel drugs and vaccines for HIV. Anti-retroviral drugs studied extensively in HIV infection have targeted different steps of the virus replication cycle: (1 inhibition of virus entry into susceptible cells at the level of attachment to host cell surface receptors and co-receptors; (2 inhibition of fusion of the virus membrane with the cell membrane; (3 blockade of reverse transcription of viral genomic RNA; (4 interruption of nuclear translocation and viral DNA integration into host genomes; (5 prevention of viral transcript processing and nuclear export; and (6 inhibition of virion assembly and maturation. Despite much success of anti-retroviral therapy slowing disease progression in people, similar therapy has not been thoroughly investigated in cats. In this article we review current pharmacological approaches and novel targets for anti-lentiviral therapy, and critically assess potentially suitable applications against FIV infection in cats.

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

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Caitlin L Rowe

Full Text Available 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

Cell lines that support replication of a novel herpes simplex virus 1 UL31 deletion mutant can properly target UL34 protein to the nuclear rim in the absence of UL31

International Nuclear Information System (INIS)

Liang Li; Tanaka, Michiko; Kawaguchi, Yasushi; Baines, Joel D.

2004-01-01

Previous results indicated that the herpes simplex virus 1 (HSV-1) U L 31 gene is necessary and sufficient for localization of the U 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 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 L 31 gene. The replication of the U L 31 deletion virus was restored on U L 31-complementing cell lines derived either from rabbit skin cells or CV1 cells. Confocal microscopy indicated that the majority of U L 34 protein localized aberrantly in the cytoplasm and nucleoplasm of Vero cells and CV1 cells, whereas U L 34 protein localized at the nuclear membrane in rabbit skin cells, and U L 31 complementing CV1 cells infected with the U L 31 deletion virus. We conclude that rabbit skin cells encode a function that allows proper localization of U L 34 protein to the nuclear membrane. We speculate that this function partially complements that of U L 31 and may explain why U L 31 is less critical for replication in rabbit skin cells as opposed to Vero and CV1 cells

The nuclear export protein of H5N1 influenza A viruses recruits Matrix 1 (M1) protein to the viral ribonucleoprotein to mediate nuclear export.

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Brunotte, Linda; Flies, Joe; Bolte, Hardin; Reuther, Peter; Vreede, Frank; Schwemmle, Martin

2014-07-18

In influenza A virus-infected cells, replication and transcription of the viral genome occurs in the nucleus. To be packaged into viral particles at the plasma membrane, encapsidated viral genomes must be exported from the nucleus. Intriguingly, the nuclear export protein (NEP) is involved in both processes. Although NEP stimulates viral RNA synthesis by binding to the viral polymerase, its function during nuclear export implicates interaction with viral ribonucleoprotein (vRNP)-associated M1. The observation that both interactions are mediated by the C-terminal moiety of NEP raised the question whether these two features of NEP are linked functionally. Here we provide evidence that the interaction between M1 and the vRNP depends on the NEP C terminus and its polymerase activity-enhancing property for the nuclear export of vRNPs. This suggests that these features of NEP are linked functionally. Furthermore, our data suggest that the N-terminal domain of NEP interferes with the stability of the vRNP-M1-NEP nuclear export complex, probably mediated by its highly flexible intramolecular interaction with the NEP C terminus. On the basis of our data, we propose a new model for the assembly of the nuclear export complex of Influenza A vRNPs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Identification of three redundant segments responsible for herpes simplex virus 1 ICP0 to fuse with ND10 nuclear bodies.

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Zheng, Yi; Gu, Haidong

2015-04-01

Infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) is a key regulator in both lytic and latent infections. In lytic infection, an important early event is the colocalization of ICP0 to nuclear domain 10 (ND10), the discrete nuclear bodies that impose restrictions on viral expression. ICP0 contains an E3 ubiquitin ligase that degrades promyelocytic leukemia protein (PML) and Sp100, two major components of ND10, and disperses ND10 to alleviate repression. We previously reported that the association between ICP0 and ND10 is a dynamic process that includes three steps: adhesion, fusion, and retention. ICP0 residues 245 to 474, defined as ND10 entry signal (ND10-ES), is a region required for the fusion step. Without ND10-ES, ICP0 adheres at the ND10 surface but fails to enter. In the present study, we focus on characterizing ND10-ES. Here we report the following. (i) Fusion of ICP0 with ND10 relies on specific sequences located within ND10-ES. Replacement of ND10-ES by the corresponding region from ORF61 of varicella-zoster virus did not rescue ND10 fusion. (ii) Three tandem ND10 fusion segments (ND10-FS1, ND10-FS2, and ND10-FS3), encompassing 200 amino acids within ND10-ES, redundantly facilitate fusion. Each of the three segments is sufficient to independently drive the fusion process, but none of the segments by themselves are necessary for ND10 fusion. Only when all three segments are deleted is fusion blocked. (iii) The SUMO interaction motif located within ND10-FS2 is not required for ND10 fusion but is required for the complete degradation of PML, suggesting that PML degradation and ND10 fusion are regulated by different molecular mechanisms. ND10 nuclear bodies are part of the cell-intrinsic antiviral defenses that restrict viral gene expression upon virus infection. As a countermeasure, infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) localizes to ND10s, degrades the ND10 organizer, and disperses ND10 components in order to

Importin α5 negatively regulates importin β1-mediated nuclear import of Newcastle disease virus matrix protein and viral replication and pathogenicity in chicken fibroblasts.

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Duan, Zhiqiang; Xu, Haixu; Ji, Xinqin; Zhao, Jiafu; Xu, Houqiang; Hu, Yan; Deng, Shanshan; Hu, Shunlin; Liu, Xiufan

2018-12-31

The matrix (M) protein of Newcastle disease virus (NDV) is demonstrated to localize in the nucleus via intrinsic nuclear localization signal (NLS), but cellular proteins involved in the nuclear import of NDV M protein and the role of M's nuclear localization in the replication and pathogenicity of NDV remain unclear. In this study, importin β1 was screened to interact with NDV M protein by yeast two-hybrid screening. This interaction was subsequently confirmed by co-immunoprecipitation and pull-down assays. In vitro binding studies indicated that the NLS region of M protein and the amino acids 336-433 of importin β1 that belonged to the RanGTP binding region were important for binding. Importantly, a recombinant virus with M/NLS mutation resulted in a pathotype change of NDV and attenuated viral replication and pathogenicity in chicken fibroblasts and SPF chickens. In agreement with the binding data, nuclear import of NDV M protein in digitonin-permeabilized HeLa cells required both importin β1 and RanGTP. Interestingly, importin α5 was verified to interact with M protein through binding importin β1. However, importin β1 or importin α5 depletion by siRNA resulted in different results, which showed the obviously cytoplasmic or nuclear accumulation of M protein and the remarkably decreased or increased replication ability and pathogenicity of NDV in chicken fibroblasts, respectively. Our findings therefore demonstrate for the first time the nuclear import mechanism of NDV M protein and the negative regulation role of importin α5 in importin β1-mediated nuclear import of M protein and the replication and pathogenicity of a paramyxovirus.

Dual function of the nuclear export signal of the Borna disease virus nucleoprotein in nuclear export activity and binding to viral phosphoprotein.

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Yanai, Mako; Sakai, Madoka; Makino, Akiko; Tomonaga, Keizo

2017-07-11

Borna disease virus (BoDV), which has a negative-sense, single-stranded RNA genome, causes persistent infection in the cell nucleus. The nuclear export signal (NES) of the viral nucleoprotein (N) consisting of leucine at positions 128 and 131 and isoleucine at positions 133 and 136 overlaps with one of two predicted binding sites for the viral phosphoprotein (P). A previous study demonstrated that higher expression of BoDV-P inhibits nuclear export of N; however, the function of N NES in the interaction with P remains unclear. We examined the subcellular localization, viral polymerase activity, and P-binding ability of BoDV-N NES mutants. We also characterized a recombinant BoDV (rBoDV) harboring an NES mutation of N. BoDV-N with four alanine-substitutions in the leucine and isoleucine residues of the NES impaired its cytoplasmic localization and abolished polymerase activity and P-binding ability. Although an alanine-substitution at position 131 markedly enhanced viral polymerase activity as determined by a minigenome assay, rBoDV harboring this mutation showed expression of viral RNAs and proteins relative to that of wild-type rBoDV. Our results demonstrate that BoDV-N NES has a dual function in BoDV replication, i.e., nuclear export of N and an interaction with P, affecting viral polymerase activity in the nucleus.

The stress granule component G3BP is a novel interaction partner for the nuclear shuttle proteins of the nanovirus pea necrotic yellow dwarf virus and geminivirus abutilon mosaic virus.

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Krapp, Susanna; Greiner, Eva; Amin, Bushra; Sonnewald, Uwe; Krenz, Björn

2017-01-02

Stress granules (SGs) are structures within cells that regulate gene expression during stress response, e.g. viral infection. In mammalian cells assembly of SGs is dependent on the Ras-GAP SH3-domain-binding protein (G3BP). The C-terminal domain of the viral nonstructural protein 3 (nsP3) of Semliki Forest virus (SFV) forms a complex with mammalian G3BP and sequesters it into viral RNA replication complexes in a manner that inhibits the formation of SGs. The binding domain of nsP3 to HsG3BP was mapped to two tandem 'FGDF' repeat motifs close to the C-terminus of the viral proteins. It was speculated that plant viruses employ a similar strategy to inhibit SG function. This study identifies an Arabidopsis thaliana NTF2-RRM domain-containing protein as a G3BP-like protein (AtG3BP), which localizes to plant SGs. Moreover, the nuclear shuttle protein (NSP) of the begomovirus abutilon mosaic virus (AbMV), which harbors a 'FVSF'-motif at its C-terminal end, interacts with the AtG3BP-like protein, as does the 'FNGSF'-motif containing NSP of pea necrotic yellow dwarf virus (PNYDV), a member of the Nanoviridae family. We therefore propose that SG formation upon stress is conserved between mammalian and plant cells and that plant viruses may follow a similar strategy to inhibit plant SG function as it has been shown for their mammalian counterparts. Copyright © 2016 Elsevier B.V. All rights reserved.

Virus-Induced Chaperone-Enriched (VICE domains function as nuclear protein quality control centers during HSV-1 infection.

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Christine M Livingston

2009-10-01

Full Text Available Virus-Induced Chaperone-Enriched (VICE domains form adjacent to nuclear viral replication compartments (RC during the early stages of HSV-1 infection. Between 2 and 3 hours post infection at a MOI of 10, host protein quality control machinery such as molecular chaperones (e.g. Hsc70, the 20S proteasome and ubiquitin are reorganized from a diffuse nuclear distribution pattern to sequestration in VICE domains. The observation that VICE domains contain putative misfolded proteins suggests that they may be similar to nuclear inclusion bodies that form under conditions in which the protein quality control machinery is overwhelmed by the presence of misfolded proteins. The detection of Hsc70 in VICE domains, but not in nuclear inclusion bodies, indicates that Hsc70 is specifically reorganized by HSV-1 infection. We hypothesize that HSV-1 infection induces the formation of nuclear protein quality control centers to remodel or degrade aberrant nuclear proteins that would otherwise interfere with productive infection. Detection of proteolytic activity in VICE domains suggests that substrates may be degraded by the 20S proteasome in VICE domains. FRAP analysis reveals that GFP-Hsc70 is dynamically associated with VICE domains, suggesting a role for Hsc70 in scanning the infected nucleus for misfolded proteins. During 42 degrees C heat shock, Hsc70 is redistributed from VICE domains into RC perhaps to remodel viral replication and regulatory proteins that have become insoluble in these compartments. The experiments presented in this paper suggest that VICE domains are nuclear protein quality control centers that are modified by HSV-1 to promote productive infection.

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

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Reimers, Kerstin [Klinik fuer Plastische, Hand-und Wiederherstellungschirurgie, Podbielskistrasse 380, D-30659 Hannover (Germany); Buchholz, Katja [Institut fuer Medizinische Mikrobiologie, Otto-von-Guericke-Universitaet Magdeburg, Leipzigerstrasse 44, D-39120 Magdeburg (Germany); Werchau, Hermann [Institut fuer Medizinische Mikrobiologie, Otto-von-Guericke-Universitaet Magdeburg, Leipzigerstrasse 44, D-39120 Magdeburg (Germany)

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.

A Novel Nuclear Trafficking Module Regulates the Nucleocytoplasmic Localization of the Rabies Virus Interferon Antagonist, P Protein*

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Oksayan, Sibil; Wiltzer, Linda; Rowe, Caitlin L.; Blondel, Danielle; Jans, David A.; Moseley, Gregory W.

2012-01-01

Regulated nucleocytoplasmic transport of proteins is central to cellular function and dysfunction during processes such as viral infection. Active protein trafficking into and out of the nucleus is dependent on the presence within cargo proteins of intrinsic specific modular signals for nuclear import (nuclear localization signals, NLSs) and export (nuclear export signals, NESs). Rabies virus (RabV) phospho (P) protein, which is largely responsible for antagonising the host anti-viral response, is expressed as five isoforms (P1–P5). The subcellular trafficking of these isoforms is thought to depend on a balance between the activities of a dominant N-terminal NES (N-NES) and a distinct C-terminal NLS (C-NLS). Specifically, the N-NES-containing isoforms P1 and P2 are cytoplasmic, whereas the shorter P3–P5 isoforms, which lack the N-NES, are believed to be nuclear through the activity of the C-NLS. Here, we show for the first time that RabV P contains an additional strong NLS in the N-terminal region (N-NLS), which, intriguingly, overlaps with the N-NES. This arrangement represents a novel nuclear trafficking module where the N-NLS is inactive in P1 but becomes activated in P3, concomitant with truncation of the N-NES, to become the principal targeting signal conferring nuclear accumulation. Understanding this unique switch arrangement of overlapping, co-regulated NES/NLS sequences is vital to delineating the critical role of RabV P protein in viral infection. PMID:22700958

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

International Nuclear Information System (INIS)

LeBlanc, Jason J.; Uddowla, Sabena; Abraham, Benjamin; Clatterbuck, Sarah; Beemon, Karen L.

2007-01-01

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

Cellular Promyelocytic Leukemia Protein Is an Important Dengue Virus Restriction Factor

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Giovannoni, Federico; Damonte, Elsa B.; Garc?a, Cybele C.

2015-01-01

The intrinsic antiviral defense is based on cellular restriction factors that are constitutively expressed and, thus, active even before a pathogen enters the cell. The promyelocytic leukemia (PML) nuclear bodies (NBs) are discrete nuclear foci that contain several cellular proteins involved in intrinsic antiviral responses against a number of viruses. Accumulating reports have shown the importance of PML as a DNA virus restriction factor and how these pathogens evade this antiviral activity....

Nuclear export and import of human hepatitis B virus capsid protein and particles.

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Hung-Cheng Li

Full Text Available It remains unclear what determines the subcellular localization of hepatitis B virus (HBV core protein (HBc and particles. To address this fundamental issue, we have identified four distinct HBc localization signals in the arginine rich domain (ARD of HBc, using immunofluorescence confocal microscopy and fractionation/Western blot analysis. ARD consists of four tight clustering arginine-rich subdomains. ARD-I and ARD-III are associated with two co-dependent nuclear localization signals (NLS, while ARD-II and ARD-IV behave like two independent nuclear export signals (NES. This conclusion is based on five independent lines of experimental evidence: i Using an HBV replication system in hepatoma cells, we demonstrated in a double-blind manner that only the HBc of mutant ARD-II+IV, among a total of 15 ARD mutants, can predominantly localize to the nucleus. ii These results were confirmed using a chimera reporter system by placing mutant or wild type HBc trafficking signals in the heterologous context of SV40 large T antigen (LT. iii By a heterokaryon or homokaryon analysis, the fusion protein of SV40 LT-HBc ARD appeared to transport from nuclei of transfected donor cells to nuclei of recipient cells, suggesting the existence of an NES in HBc ARD. This putative NES is leptomycin B resistant. iv We demonstrated by co-immunoprecipitation that HBc ARD can physically interact with a cellular factor TAP/NXF1 (Tip-associated protein/nuclear export factor-1, which is known to be important for nuclear export of mRNA and proteins. Treatment with a TAP-specific siRNA strikingly shifted cytoplasmic HBc to nucleus, and led to a near 7-fold reduction of viral replication, and a near 10-fold reduction in HBsAg secretion. v HBc of mutant ARD-II+IV was accumulated predominantly in the nucleus in a mouse model by hydrodynamic delivery. In addition to the revised map of NLS, our results suggest that HBc could shuttle rapidly between nucleus and cytoplasm via a novel

Inhibition of human T cell leukemia virus type 2 replication by the suppressive action of class II transactivator and nuclear factor Y.

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Tosi, Giovanna; Pilotti, Elisabetta; Mortara, Lorenzo; De Lerma Barbaro, Andrea; Casoli, Claudio; Accolla, Roberto S

2006-08-22

The master regulator of MHC-II gene transcription, class II transactivator (CIITA), acts as a potent inhibitor of human T cell leukemia virus type 2 (HTLV-2) replication by blocking the activity of the viral Tax-2 transactivator. Here, we show that this inhibitory effect takes place at the nuclear level and maps to the N-terminal 1-321 region of CIITA, where we identified a minimal domain, from positions 64-144, that is strictly required to suppress Tax-2 function. Furthermore, we show that Tax-2 specifically cooperates with cAMP response element binding protein-binding protein (CBP) and p300, but not with p300/CBP-associated factor, to enhance transcription from the viral promoter. This finding represents a unique difference with respect to Tax-1, which uses all three coactivators to transactivate the human T cell leukemia virus type 1 LTR. Direct sequestering of CBP or p300 is not the primary mechanism by which CIITA causes suppression of Tax-2. Interestingly, we found that the transcription factor nuclear factor Y, which interacts with CIITA to increase transcription of MHC-II genes, exerts a negative regulatory action on the Tax-2-mediated HTLV-2 LTR transactivation. Thus, CIITA may inhibit Tax-2 function, at least in part, through nuclear factor Y. These findings demonstrate the dual defensive role of CIITA against pathogens: it increases the antigen-presenting function for viral determinants and suppresses HTLV-2 replication in infected cells.

ANP32B is a nuclear target of henipavirus M proteins.

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Anja Bauer

Full Text Available Membrane envelopment and budding of negative strand RNA viruses (NSVs is mainly driven by viral matrix proteins (M. In addition, several M proteins are also known to be involved in host cell manipulation. Knowledge about the cellular targets and detailed molecular mechanisms, however, is poor for many M proteins. For instance, Nipah Virus (NiV M protein trafficking through the nucleus is essential for virus release, but nuclear targets of NiV M remain unknown. To identify cellular interactors of henipavirus M proteins, tagged Hendra Virus (HeV M proteins were expressed and M-containing protein complexes were isolated and analysed. Presence of acidic leucine-rich nuclear phosphoprotein 32 family member B (ANP32B in the complex suggested that this protein represents a direct or indirect interactor of the viral matrix protein. Over-expression of ANP32B led to specific nuclear accumulation of HeV M, providing a functional link between ANP32B and M protein. ANP32B-dependent nuclear accumulation was observed after plasmid-driven expression of HeV and NiV matrix proteins and also in NiV infected cells. The latter indicated that an interaction of henipavirus M protein with ANP32B also occurs in the context of virus replication. From these data we conclude that ANP32B is a nuclear target of henipavirus M that may contribute to virus replication. Potential effects of ANP32B on HeV nuclear shuttling and host cell manipulation by HeV M affecting ANP32B functions in host cell survival and gene expression regulation are discussed.

Autophagy in Measles Virus Infection

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Aurore Rozières

2017-11-01

Full Text Available Autophagy is a biological process that helps cells to recycle obsolete cellular components and which greatly contributes to maintaining cellular integrity in response to environmental stress factors. Autophagy is also among the first lines of cellular defense against invading microorganisms, including viruses. The autophagic destruction of invading pathogens, a process referred to as xenophagy, involves cytosolic autophagy receptors, such as p62/SQSTM1 (Sequestosome 1 or NDP52/CALCOCO2 (Nuclear Dot 52 KDa Protein/Calcium Binding And Coiled-Coil Domain 2, which bind to microbial components and target them towards growing autophagosomes for degradation. However, most, if not all, infectious viruses have evolved molecular tricks to escape from xenophagy. Many viruses even use autophagy, part of the autophagy pathway or some autophagy-associated proteins, to improve their infectious potential. In this regard, the measles virus, responsible for epidemic measles, has a unique interface with autophagy as the virus can induce multiple rounds of autophagy in the course of infection. These successive waves of autophagy result from distinct molecular pathways and seem associated with anti- and/or pro-measles virus consequences. In this review, we describe what the autophagy–measles virus interplay has taught us about both the biology of the virus and the mechanistic orchestration of autophagy.

Hepatitis B virus nuclear export elements: RNA stem-loop α and β, key parts of the HBV post-transcriptional regulatory element.

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Lim, Chun Shen; Brown, Chris M

2016-09-01

Many viruses contain RNA elements that modulate splicing and/or promote nuclear export of their RNAs. The RNAs of the major human pathogen, hepatitis B virus (HBV) contain a large (~600 bases) composite cis-acting 'post-transcriptional regulatory element' (PRE). This element promotes expression from these naturally intronless transcripts. Indeed, the related woodchuck hepadnavirus PRE (WPRE) is used to enhance expression in gene therapy and other expression vectors. These PRE are likely to act through a combination of mechanisms, including promotion of RNA nuclear export. Functional components of both the HBV PRE and WPRE are 2 conserved RNA cis-acting stem-loop (SL) structures, SLα and SLβ. They are within the coding regions of polymerase (P) gene, and both P and X genes, respectively. Based on previous studies using mutagenesis and/or nuclear magnetic resonance (NMR), here we propose 2 covariance models for SLα and SLβ. The model for the 30-nucleotide SLα contains a G-bulge and a CNGG(U) apical loop of which the first and the fourth loop residues form a CG pair and the fifth loop residue is bulged out, as observed in the NMR structure. The model for the 23-nucleotide SLβ contains a 7-base-pair stem and a 9-nucleotide loop. Comparison of the models with other RNA structural elements, as well as similarity searches of human transcriptome and viral genomes demonstrate that SLα and SLβ are specific to HBV transcripts. However, they are well conserved among the hepadnaviruses of non-human primates, the woodchuck and ground squirrel.

Induction of human interferon gene expression is associated with a nuclear factor that interacts with the site of the human immunodeficiency virus-enhancer

International Nuclear Information System (INIS)

Hiscott, J.; Alper, D.; Cohen, L.; Leblanc, J.F.; Sportza, L.; Wong, A.; Xanthoudakis, S.

1989-01-01

The relationship between transcription of alpha and beta interferon (IFN-α and IFN-β) genes and the interaction of IFN promoter-binding transcription factors has been examined in monoblastoid U937 cells following priming with recombinant IFN-α2 (rIFN-α2) and Sendai virus induction. Pretreatment of U937 cells with rIFN-α2 prior to Sendai virus infection increased the mRNA levels of IFN-α1, IFN-α2, and IFN-β as well as the final yield of biologically active IFN. Analysis of nuclear protein-IFN promoter DNA interactions by electrophoretic mobility-shift assays demonstrated increased factor binding to IFN-α1 and IFN-β regulatory domains, although no new induction-specific complexes were identified. On the basis of competition electrophoretic mobility-shift assay results, factors interacting with the IFN-α1 and IFN-β promoters appear to be distinct DNA-binding proteins. Hybrid promoter-chloramphenicol acetyltransferase fusion plasmids, containing either the IFN-β regulatory element or the human immunodeficiency virus enhancer element linked to the simian virus 40 promoter, were analyzed for virus and phorbol ester inducibility in epithelial and lymphoid cells, respectively. These experiments suggest that induction of IFN gene expression may be controlled in part by transcription regulatory proteins binding to an NF-κB-like site within the IFN-β promoter

Hepatitis B virus X promotes hepatocellular carcinoma development via nuclear protein 1 pathway

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Bak, Yesol; Shin, Hye-jun; Bak, In seon [Disease Model Research Laboratory, Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon (Korea, Republic of); Yoon, Do-young [Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul (Korea, Republic of); Yu, Dae-Yeul, E-mail: dyyu10@kribb.re.kr [Disease Model Research Laboratory, Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon (Korea, Republic of)

2015-10-30

Hepatocellular carcinoma (HCC) is one of the most common malignancies and chronic hepatitis B virus (HBV) infection is a major risk factor for HCC. Hepatitis B virus X (HBx) protein relates to trigger oncogenesis. HBx has oncogenic properties with a hyperproliferative response to HCC. Nuclear protein 1 (NUPR1) is a stress-response protein, frequently upregulated in several cancers. Recent data revealed that NUPR1 is involved in tumor progression, but its function in HCC is not revealed yet. Here we report HBx can induce NUPR1 in patients, mice, and HCC cell lines. In an HBx transgenic mouse model, we found that HBx overexpression upregulates NUPR1 expression consistently with tumor progression. Further, in cultured HBV positive cells, HBx knockdown induces downregulation of NUPR1. Smad4 is a representative transcription factor, regulated by HBx, and we showed that HBx upregulates NUPR1 by Smad4 dependent way. We found that NUPR1 can inhibit cell death and induce vasculogenic mimicry in HCC cell lines. Moreover, NUPR1 silencing in HepG2-HBx showed reduced cell motility. These results suggest that HBx can modulate NUPR1 expression through the Smad4 pathway and NUPR1 has a role in hepatocellular carcinoma progression. - Highlights: • NUPR1 is overexpressed in HBx transgenic mouse and HCC patients. • NUPR1 inactivation hampers the HBx induced growth, VM formation, and migration of HepG2 cells in vitro. • NUPR1 has a role for survival of HCC and mechanistically NUPR1 is activated by HBx-Smad4 axis.

Expression of the human multidrug transporter in insect cells by a recombinant baculovirus

International Nuclear Information System (INIS)

Germann, U.A.; Willingham, M.C.; Pastan, I.; Gottesman, M.M.

1990-01-01

The plasma membrane associated human multidrug resistance (MDR1) gene product, known as the 170-kDa P-glycoprotein or the multidrug transporter, acts as an ATP-dependent efflux pump for various cytotoxic agents. The authors expressed recombinant human multidrug transporter in a baculovirus expression system to obtain large quantities and further investigate its structure and mechanism of action. MDR1 cDNA was inserted into the genome of the Autographa californica nuclear polyhedrosis virus under the control of the polyhedrin promoter. Spodoptera frugiperda insect cells synthesized high levels of recombinant multidrug transporter 2-3 days after infection. The transporter was localized by immunocytochemical methods on the external surface of the plasma membranes, in the Golgi apparatus, and within the nuclear envelope. The human multidrug transporter expressed in insect cells is not susceptible to endoglycosidase F treatment and has a lower apparent molecular weight of 140,000, corresponding to the nonglycosylated precursor of its authentic counterpart expressed in multidrug-resistant cells. Labeling experiments showed that the recombinant multidrug transporter is phosphorylated and can be photoaffinity labeled by [ 3 H]azidopine, presumably at the same two sites as the native protein. Various drugs and reversing agents compete with the [ 3 H]azidopine binding reaction when added in excess, indicating that the recombinant human multidrug transporter expressed in insect cells is functionally similar to its authentic counterpart

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

Science.gov (United States)

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.

Characterization of a nuclear export signal within the human T cell leukemia virus type I transactivator protein Tax.

Science.gov (United States)

Alefantis, Timothy; Barmak, Kate; Harhaj, Edward W; Grant, Christian; Wigdahl, Brian

2003-06-13

Human T cell leukemia virus type I (HTLV-I) is the etiologic agent of adult T cell leukemia and HTLV-I-associated myelopathy/tropical spastic paraparesis. The HTLV-I transactivator protein Tax plays an integral role in the etiology of adult T cell leukemia, as expression of Tax in T lymphocytes has been shown to result in immortalization. In addition, Tax is known to interface with numerous transcription factor families, including activating transcription factor/cAMP response element-binding protein and nuclear factor-kappaB, requiring Tax to localize to both the nucleus and cytoplasm. In this report, the nucleocytoplasmic localization of Tax was examined in Jurkat, HeLa, and U-87 MG cells. The results reported herein indicate that Tax contains a leucine-rich nuclear export signal (NES) that, when fused to green fluorescent protein (GFP), can direct nuclear export via the CRM-1 pathway, as determined by leptomycin B inhibition of nuclear export. However, cytoplasmic localization of full-length Tax was not altered by treatment with leptomycin B, suggesting that native Tax utilizes another nuclear export pathway. Additional support for the presence of a functional NES has also been shown because the NES mutant Tax(L200A)-GFP localized to the nuclear membrane in the majority of U-87 MG cells. Evidence has also been provided suggesting that the Tax NES likely exists as a conditionally masked signal because the truncation mutant TaxDelta214-GFP localized constitutively to the cytoplasm. These results suggest that Tax localization may be directed by specific changes in Tax conformation or by specific interactions with cellular proteins leading to changes in the availability of the Tax NES and nuclear localization signal.

Frequent presence of subtype A virus in Epstein-Barr virus-associated malignancies

NARCIS (Netherlands)

Peh, SC; Kim, LH; Poppema, S

2002-01-01

Aims: Epstein-Barr virus (EBV) is associated with many human malignancies. It is implicated in a pathogenetic role in some of these tumours. Two subtypes, type A and B have been identified on the basis of DNA sequence divergence in the nuclear protein genes (EBNA) 2, 3, 4 and 6. They differ in their

Hsp90 inhibitors reduce influenza virus replication in cell culture

International Nuclear Information System (INIS)

Chase, Geoffrey; Deng, Tao; Fodor, Ervin; Leung, B.W.; Mayer, Daniel; Schwemmle, Martin; Brownlee, George

2008-01-01

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

Single-Domain Antibodies as Tools to Perturb and Study RNA Viruses

NARCIS (Netherlands)

Hanke, Leo

2017-01-01

In this thesis, I describe the generation and characterization of alpaca-derived, antiviral, single-domain antibody fragments (VHHs). The antiviral targets of the described VHHs are the nuclear proteins of influenza A virus (IAV) and vesicular stomatitis virus (VSV). The described VHHs protect cells

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

Science.gov (United States)

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.

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

International Nuclear Information System (INIS)

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

2008-01-01

The transfection of human cells with siRNA against adapter-related protein complex 2 alpha 1 subunit (AP2α) 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 AP2α. In contrast, viral nuclear translocation as well as the integration process was significantly up-regulated in cells transfected with siRNA against AP2α. Confocal fluorescence microscopy revealed that a subpopulation of AP2α was not only localized in the cytoplasm but was also partly co-localized with lamin B, importin β and Nup153, implying that AP2α negatively regulates HIV-1 replication in the process of nuclear translocation of viral DNA in the cytoplasm or the perinuclear region. We propose that AP2α may be a novel target for disrupting HIV-1 replication in the early stage of the viral life cycle

Epstein-Barr Virus Nuclear Antigen Leader Protein Coactivates EP300.

Science.gov (United States)

Wang, Chong; Zhou, Hufeng; Xue, Yong; Liang, Jun; Narita, Yohei; Gerdt, Catherine; Zheng, Amy Y; Jiang, Runsheng; Trudeau, Stephen; Peng, Chih-Wen; Gewurz, Benjamin E; Zhao, Bo

2018-05-01

Epstein-Barr virus nuclear antigen (EBNA) leader protein (EBNALP) is one of the first viral genes expressed upon B-cell infection. EBNALP is essential for EBV-mediated B-cell immortalization. EBNALP is thought to function primarily by coactivating EBNA2-mediated transcription. Chromatin immune precipitation followed by deep sequencing (ChIP-seq) studies highlight that EBNALP frequently cooccupies DNA sites with host cell transcription factors (TFs), in particular, EP300, implicating a broader role in transcription regulation. In this study, we investigated the mechanisms of EBNALP transcription coactivation through EP300. EBNALP greatly enhanced EP300 transcription activation when EP300 was tethered to a promoter. EBNALP coimmunoprecipitated endogenous EP300 from lymphoblastoid cell lines (LCLs). EBNALP W repeat serine residues 34, 36, and 63 were required for EP300 association and coactivation. Deletion of the EP300 histone acetyltransferase (HAT) domain greatly reduced EBNALP coactivation and abolished the EBNALP association. An EP300 bromodomain inhibitor also abolished EBNALP coactivation and blocked the EP300 association with EBNALP. EBNALP sites cooccupied by EP300 had significantly higher ChIP-seq signals for sequence-specific TFs, including SPI1, RelA, EBF1, IRF4, BATF, and PAX5. EBNALP- and EP300-cooccurring sites also had much higher H3K4me1 and H3K27ac signals, indicative of activated enhancers. EBNALP-only sites had much higher signals for DNA looping factors, including CTCF and RAD21. EBNALP coactivated reporters under the control of NF-κB or SPI1. EP300 inhibition abolished EBNALP coactivation of these reporters. Clustered regularly interspaced short palindromic repeat interference targeting of EBNALP enhancer sites significantly reduced target gene expression, including that of EP300 itself. These data suggest a previously unrecognized mechanism by which EBNALP coactivates transcription through subverting of EP300 and thus affects the expression of

Characterization of the N-terminal segment used by the barley yellow dwarf virus movement protein to promote interaction with the nuclear membrane of host plant cells.

Science.gov (United States)

Dennison, Sarah Rachel; Harris, Frederick; Brandenburg, Klaus; Phoenix, David Andrew

2007-11-01

The barley yellow dwarf virus movement protein (BYDV-MP) requires its N-terminal sequence to promote the transport of viral RNA into the nuclear compartment of host plant cells. Here, graphical analysis predicts that this sequence would form a membrane interactive amphiphilic alpha-helix. Confirming this prediction, NT1, a peptide homologue of the BYDV-MP N-terminal sequence, was found to be alpha-helical (65%) in the presence of vesicles mimics of the nuclear membrane. The peptide increased the fluidity of these nuclear membrane mimics (rise in wavenumber of circa 0.5-1.0 cm(-1)) and induced surface pressure changes of 2 mN m(-1) in lipid monolayers with corresponding compositions. Taken with isotherm analysis these results suggest that BYDV-MP forms an N-terminal amphiphilic alpha-helix, which partitions into the nuclear membrane primarily through thermodynamically stable associations with the membrane lipid headgroup region. We speculate that these associations may play a role in targeting of the nuclear membrane by BYDM-MP.

Single HIV-1 Imaging Reveals Progression of Infection through CA-Dependent Steps of Docking at the Nuclear Pore, Uncoating, and Nuclear Transport.

Science.gov (United States)

Francis, Ashwanth C; Melikyan, Gregory B

2018-04-11

The HIV-1 core consists of capsid proteins (CA) surrounding viral genomic RNA. After virus-cell fusion, the core enters the cytoplasm and the capsid shell is lost through uncoating. CA loss precedes nuclear import and HIV integration into the host genome, but the timing and location of uncoating remain unclear. By visualizing single HIV-1 infection, we find that CA is required for core docking at the nuclear envelope (NE), whereas early uncoating in the cytoplasm promotes proteasomal degradation of viral complexes. Only docked cores exhibiting accelerated loss of CA at the NE enter the nucleus. Interestingly, a CA mutation (N74D) altering virus engagement of host factors involved in nuclear transport does not alter the uncoating site at the NE but reduces the nuclear penetration depth. Thus, CA protects HIV-1 complexes from degradation, mediates docking at the nuclear pore before uncoating, and determines the depth of nuclear penetration en route to integration. Copyright © 2018 Elsevier Inc. All rights reserved.

The Mammalian Cell Cycle Regulates Parvovirus Nuclear Capsid Assembly

Science.gov (United States)

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

Investigation of radiation enhanced reactivation of cytoplasmic replicating human virus

International Nuclear Information System (INIS)

Bockstahler, L.E.; Haynes, K.F.; Stafford, J.E.

1976-01-01

When monolayers of CV-1 monkey kidney cells were exposed to ultraviolet (uv) radiation (0 to 200 erg/nm 2 ) or x rays (0 to 10 krads) before infection with uv-irradiated herpes simplex virus, an increase in the infectivity of this nuclear replicating virus occurred as measured by plaque formation. These phenomena are known as uv (Weigle) reactivation (WR) and x-ray reactivation (x-ray R). In this study the presence of WR and x-ray R was examined in CV-1 cells infected with uv-irradiated vaccinia virus or poliovirus, both cytoplasmic replicating viruses. Little or no WR or x-ray R was observed for either of these viruses. These results suggest that WR and x-ray R in mammalian cells may be restricted to viruses which are synthesized in the cell nucleus

Morbillivirus nucleoprotein possesses a novel nuclear localization signal and a CRM1-independent nuclear export signal

International Nuclear Information System (INIS)

Sato, Hiroki; Masuda, Munemitsu; Miura, Ryuichi; Yoneda, Misako; Kai, Chieko

2006-01-01

Morbilliviruses, which belong to the Mononegavirales, replicate its RNA genome in the cytoplasm of the host cell. However, they also form characteristic intranuclear inclusion bodies, consisting of nucleoprotein (N), in infected cells. To analyze the mechanisms of nucleocytoplasmic transport of N protein, we characterized the nuclear localization (NLS) and nuclear export (NES) signals of canine distemper virus (CDV) N protein by deletion mutation and alanine substitution of the protein. The NLS has a novel leucine/isoleucine-rich motif (TGILISIL) at positions 70-77, whereas the NES is composed of a leucine-rich motif (LLRSLTLF) at positions 4-11. The NLS and NES of the N proteins of other morbilliviruses, that is, measles virus (MV) and rinderpest virus (RPV), were also analyzed. The NLS of CDV-N protein is conserved at the same position in MV-N protein, whereas the NES of MV-N protein is located in the C-terminal region. The NES of RPV-N protein is also located at the same position as CDV-N protein, whereas the NLS motif is present not only at the same locus as CDV-N protein but also at other sites. Interestingly, the nuclear export of all these N proteins appears to proceed via a CRM1-independent pathway

Pathogenic characteristics of a novel triple-reasserted H1N2 swine influenza virus.

Science.gov (United States)

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. Copyright © 2016 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

Analysis of nuclear accumulation of influenza NP antigen in von Magnus virus-infected cells.

Science.gov (United States)

Maeno, K; Aoki, H; Hamaguchi, M; Iinuma, M; Nagai, Y; Matsumoto, T; Takeura, S; Shibata, M

1981-01-01

When 1-5C-4 cells were infected with von Magnus virus derived from influenza A/RI/5+ virus by successive undiluted passages in chick embryos, virus-specific proteins were synthesized but production of infectious virus was inhibited. In these cells the synthesis of viral RNA was suppressed and the nucleoprotein (NP) antigen was found predominantly in the nucleus in contrast to standard virus-infected cells in which the antigen was distributed throughout the whole cell. The intracellular location and migration of NP were determined by isotope labeling and sucrose gradient centrifugation of subcellular fractions. In standard virus-infected cell NP polypeptide was present predominantly in the cytoplasm in the form of viral ribonucleoprotein (RNP) and intranuclear RNP was detected in reduced amounts. In contrast, in von Magnus virus-infected cells NP polypeptide was present predominantly in the nucleus in a nonassembled, soluble from and the amount of cytoplasmic RNP was considerably reduced. After short-pulse labeling NP was detected exclusively in the cytoplasm in a soluble form and after a chase a large proportion of such soluble NP was seen in the nucleus. It is suggested that a large proportion of the NP synthesized in von Magnus virus-infected cells in not assembled into cytoplasmic RNP because of the lack of available RNA and the NP migrated into the nucleus and remained there.

Radiobiological inactivation of Epstein-Barr virus

International Nuclear Information System (INIS)

Henderson, E.; Heston, L.; Grogan, E.; Miller, G.

1978-01-01

Lymphocyte transforming properties of B95-8 strain Epstein-Barr virus (EBV) are very sensitive to inactivation by either uv or x irradiation. No dose of irradiation increases the transforming capacity of EBV. The x-ray dose needed for inactivation of EBV transformation (dose that results in 37% survival, 60,000 rads) is similar to the dose required for inactivation of plaque formation by herpes simplex virus type 1 (Fischer strain). Although herpes simplex virus is more sensitive than EBV to uv irradiation, this difference is most likely due to differences in the kinetics or mechanisms of repair of uv damage to the two viruses. The results lead to the hypothesis that a large part, or perhaps all, of the EBV genome is in some way needed to initiate transformation. The abilities of EBV to stimulate host cell DNA synthesis, to induce nuclear antigen, and to immortalize are inactivated in parallel. All clones of marmoset cells transformed by irradiated virus produce extracellular transforming virus. These findings suggest that the abilities of the virus to transform and to replicate complete progeny are inactivated together. The amounts of uv and x irradiation that inactivate transformation by B95-8 virus are less than the dose needed to inactivate early antigen induction by the nontransforming P 3 HR-1 strain of EBV. Based on radiobiological inactivation, 10 to 50% of the genome is needed for early antigen induction

Radiation-induced Epstein-Barr virus reactivation in gastric cancer cells with latent EBV infection.

Science.gov (United States)

Nandakumar, Athira; Uwatoko, Futoshi; Yamamoto, Megumi; Tomita, Kazuo; Majima, Hideyuki J; Akiba, Suminori; Koriyama, Chihaya

2017-07-01

Epstein-Barr virus, a ubiquitous human herpes virus with oncogenic activity, can be found in 6%-16% of gastric carcinomas worldwide. In Epstein-Barr virus-associated gastric carcinoma, only a few latent genes of the virus are expressed. Ionizing irradiation was shown to induce lytic Epstein-Barr virus infection in lymphoblastoid cell lines with latent Epstein-Barr virus infection. In this study, we examined the effect of ionizing radiation on the Epstein-Barr virus reactivation in a gastric epithelial cancer cell line (SNU-719, an Epstein-Barr virus-associated gastric carcinoma cell line). Irradiation with X-ray (dose = 5 and 10 Gy; dose rate = 0.5398 Gy/min) killed approximately 25% and 50% of cultured SNU-719 cells, respectively, in 48 h. Ionizing radiation increased the messenger RNA expression of immediate early Epstein-Barr virus lytic genes (BZLF1 and BRLF1), determined by real-time reverse transcription polymerase chain reaction, in a dose-dependent manner at 48 h and, to a slightly lesser extent, at 72 h after irradiation. Similar findings were observed for other Epstein-Barr virus lytic genes (BMRF1, BLLF1, and BcLF1). After radiation, the expression of transforming growth factor beta 1 messenger RNA increased and reached a peak in 12-24 h, and the high-level expression of the Epstein-Barr virus immediate early genes can convert latent Epstein-Barr virus infection into the lytic form and result in the release of infectious Epstein-Barr virus. To conclude, Ionizing radiation activates lytic Epstein-Barr virus gene expression in the SNU-719 cell line mainly through nuclear factor kappaB activation. We made a brief review of literature to explore underlying mechanism involved in transforming growth factor beta-induced Epstein-Barr virus reactivation. A possible involvement of nuclear factor kappaB was hypothesized.

Specific interaction of capsid protein and importin-α/β influences West Nile virus production

International Nuclear Information System (INIS)

Bhuvanakantham, Raghavan; Chong, Mun-Keat; Ng, Mah-Lee

2009-01-01

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-α. 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-α/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-α/C protein interaction in the context of flavivirus life-cycle.

Specific interaction of capsid protein and importin-{alpha}/{beta} influences West Nile virus production

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Bhuvanakantham, Raghavan; Chong, Mun-Keat [Flavivirology Laboratory, Department of Microbiology, 5 Science Drive 2, National University of Singapore, Singapore 117597 (Singapore); Ng, Mah-Lee, E-mail: micngml@nus.edu.sg [Flavivirology Laboratory, Department of Microbiology, 5 Science Drive 2, National University of Singapore, Singapore 117597 (Singapore)

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.

Epstein-Barr virus nuclear antigen 1 (EBNA1) induced cytotoxicity in epithelial cells is associated with EBNA1 degradation and processing

International Nuclear Information System (INIS)

Jones, Richard J.; Smith, Laura J.; Dawson, Christopher W.; Haigh, Tracy; Blake, Neil W.; Young, Lawrence S.

2003-01-01

Epstein-Barr virus nuclear antigen 1 (EBNA1) has a central role in the maintenance and segregation of the Epstein-Barr virus (EBV) episome and by virtue of a glycine-alanine repeat domain is prevented from being endogenously processed for recognition by HLA class I restricted cytotoxic T lymphocytes (CTLs). We found that EBNA1 expression resulted in growth inhibition and a G2/M arrest in human squamous epithelial cell lines (SCC12F, SVK) but not epithelial cell lines of glandular origin (Hela, Ad/AH). The cytotoxicity of EBNA1 was associated with EBNA1 degradation and both these effects were blocked in SCC12F cells expressing either the anti-apoptotic bcl-2 protein or the EBV homolog of bcl-2, BHRF1. The endogenous degradation of EBNA1 in SVK epithelial cells was associated with specific CTL recognition, an effect not evident in EBNA1-expressing Hela cells. Consistent with the inability of SVK cells to tolerate EBNA1 expression, studies with a recombinant EBV demonstrated that SVK cells are unable to maintain stable virus infection, whereas Hela cells are able to efficiently establish latent EBV infection. These data have important implications for both the cellular requirements necessary to sustain a stable EBV infection and for the possible role of CTL responses in controlling EBV infection of epithelial cells

Epstein-Barr virus nuclear antigen 3C stabilizes Gemin3 to block p53-mediated apoptosis.

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Qiliang Cai

2011-12-01

Full Text Available The Epstein-Barr nuclear antigen 3C (EBNA3C, one of the essential latent antigens for Epstein-Barr virus (EBV-induced immortalization of primary human B lymphocytes in vitro, has been implicated in regulating cell proliferation and anti-apoptosis via interaction with several cellular and viral factors. Gemin3 (also named DDX20 or DP103 is a member of DEAD RNA helicase family which exhibits diverse cellular functions including DNA transcription, recombination and repair, and RNA metabolism. Gemin3 was initially identified as a binding partner to EBNA2 and EBNA3C. However, the mechanism by which EBNA3C regulates Gemin3 function remains unclear. Here, we report that EBNA3C directly interacts with Gemin3 through its C-terminal domains. This interaction results in increased stability of Gemin3 and its accumulation in both B lymphoma cells and EBV transformed lymphoblastoid cell lines (LCLs. Moreover, EBNA3C promotes formation of a complex with p53 and Gemin3 which blocks the DNA-binding affinity of p53. Small hairpin RNA based knockdown of Gemin3 in B lymphoma or LCL cells remarkably attenuates the ability of EBNA3C to inhibit the transcription activity of p53 on its downstream genes p21 and Bax, as well as apoptosis. These findings provide the first evidence that Gemin3 may be a common target of oncogenic viruses for driving cell proliferation and anti-apoptotic activities.

Pathogenesis and role of nuclear medicine

International Nuclear Information System (INIS)

Freedman, P. N; Korowlay, N. A

2002-01-01

The means by which replication of viruses takes place is explained, as it helps in the understanding of how viruses spread in the blood and how antiretroviral drugs work. The most important viruses, from a health care workers point of view, are hepatitis B and C and human immunodefiency virus (HIV). Whether nuclear medicine has a role to play in the diagnosis of these viruses, and the oportunistic infections that go with them, is debatable. Several radiopharmaceuticals are extremely sensitive for infection and tumor imaging but lack specificity. Patients' treatment is often not based on the outcome of the investigation but rather on preset protocols. AIDS patients are put on prophylactic antibiotic treatment as protection against infections such as toxoplasmosis and pneumocystis carinii pneumonia and there is a poor prognosis for AIDS patients with tumors (Au)

Early warning: Avian flu and nuclear science

International Nuclear Information System (INIS)

Belak, S.

2006-01-01

Avian flu has spread to 51 countries, 36 this year alone, many of which are densely populated and deprived. The joint FAO/IAEA programme is working on the rapid detection of emerging diseases, including bird flu, and using nuclear and radiation techniques in the process. The problems are serious and challenging, but nuclear technologies may offer a solution. For most developing countries, TAD (transboundary animal diseases) detection is still vital. The bottleneck is their inability to rapidly detect the virus and to determine early enough whether it is H5N1 or another subtype, so that authorities can take appropriate control measures. Serious efforts are focused on the early detection of the agents. Timely recognition of such viral infections would prevent the spread of the diseases to large animal populations in huge geographic areas. Thus, the development of novel, powerful diagnostic nuclear and nuclear-related assays is a crucial issue today in veterinary research and animal health care. Molecular virology offers a range of new methods, which are able to accelerate and improve the diagnosis of infectious diseases in animals and in man. The molecular detection assays, like the polymerase chain reaction (PCR) technologies, provide possibilities for a very rapid diagnosis. The detection of viruses can be completed within hours or hopefully even within minutes with a sensitivity level of less than one pathogenic organism. Molecular approaches have contributed significantly to the rapid detection of well-established, as well as newly emerging, infectious agents such as Nipah and Hendra viruses or corona viruses in the SARS scenario and the detection and molecular characterisation of the highly pathogenic avian influenza H5N1 subtype that threatens the world today. The nucleic acid amplification assays, although they were at first expensive and cumbersome, have become relatively cheap and user-friendly tools in the diagnostic laboratories

Coilin, the signature protein of Cajal bodies, differentially modulates the interactions of plants with viruses in widely different taxa.

Science.gov (United States)

Shaw, Jane; Love, Andrew J; Makarova, Svetlana S; Kalinina, Natalia O; Harrison, Bryan D; Taliansky, Michael E

2014-01-01

Cajal bodies (CBs) are distinct nuclear bodies physically and functionally associated with the nucleolus. In addition to their traditional function in coordinating maturation of certain nuclear RNAs, CBs participate in cell cycle regulation, development, and regulation of stress responses. A key "signature" component of CBs is coilin, the scaffolding protein essential for CB formation and function. Using an RNA silencing (loss-of-function) approach, we describe here new phenomena whereby coilin also affects, directly or indirectly, a variety of interactions between host plants and viruses that have RNA or DNA genomes. Moreover, the effects of coilin on these interactions are manifested differently: coilin contributes to plant defense against tobacco rattle virus (tobravirus), tomato black ring virus (nepovirus), barley stripe mosaic virus (hordeivirus), and tomato golden mosaic virus (begomovirus). In contrast, with potato virus Y (potyvirus) and turnip vein clearing virus (tobamovirus), coilin serves to increase virus pathogenicity. These findings show that interactions with coilin (or CBs) may involve diverse mechanisms with different viruses and that these mechanisms act at different phases of virus infection. Thus, coilin (CBs) has novel, unexpected natural functions that may be recruited or subverted by plant viruses for their own needs or, in contrast, are involved in plant defense mechanisms that suppress host susceptibility to the viruses.

Host transcription factor Speckled 110 kDa (Sp110), a nuclear body protein, is hijacked by hepatitis B virus protein X for viral persistence.

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Sengupta, Isha; Das, Dipanwita; Singh, Shivaram Prasad; Chakravarty, Runu; Das, Chandrima

2017-12-15

Promyelocytic leukemia nuclear bodies (PML-NB) are sub-nuclear organelles that are the hub of numerous proteins. DNA/RNA viruses often hijack the cellular factors resident in PML-NBs to promote their proliferation in host cells. Hepatitis B virus (HBV), belonging to Hepadnaviridae family, remains undetected in early infection as it does not induce the innate immune response and is known to be the cause of several hepatic diseases leading to cirrhosis and hepatocellular carcinoma. The association of PML-NB proteins and HBV is being addressed in a number of recent studies. Here, we report that the PML-NB protein Speckled 110 kDa (Sp110) is SUMO1-modified and undergoes a deSUMOylation-driven release from the PML-NB in the presence of HBV. Intriguingly, Sp110 knockdown significantly reduced viral DNA load in the culture supernatant by activation of the type I interferon-response pathway. Furthermore, we found that Sp110 differentially regulates several direct target genes of hepatitis B virus protein X (HBx), a viral co-factor. Subsequently, we identified Sp110 as a novel interactor of HBx and found this association to be essential for the exit of Sp110 from the PML-NB during HBV infection and HBx recruitment on the promoter of these genes. HBx, in turn, modulates the recruitment of its associated transcription cofactors p300/HDAC1 to these co-regulated genes, thereby altering the host gene expression program in favor of viral persistence. Thus, we report a mechanism by which HBV can evade host immune response by hijacking the PML-NB protein Sp110, and therefore, we propose it to be a novel target for antiviral therapy. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Characterization of Elements Regulating the Nuclear-to-Cytoplasmic Translocation of ICP0 in Late Herpes Simplex Virus 1 Infection.

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Samrat, Subodh Kumar; Ha, Binh L; Zheng, Yi; Gu, Haidong

2018-01-15

Infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) is an immediate early protein containing a RING-type E3 ubiquitin ligase. It targets several host factors for proteasomal degradation and subsequently activates viral expression. ICP0 has a nuclear localization sequence and functions in the nucleus early during infection. However, later in infection, ICP0 is found solely in the cytoplasm. The molecular mechanism and biological function of the ICP0 nuclear-to-cytoplasmic translocation are not well understood. In this study, we sought to characterize elements important for this translocation. We found that (i) in human embryonic lung fibroblast (HEL) cells, ICP0 C-terminal residues 741 to 775 were necessary but not sufficient for the nuclear-to-cytoplasmic translocation; (ii) the loss of ICP0 E3 ubiquitin ligase activity, which led to defective viral replication in nonpermissive cells, also caused mutant ICP0 to be retained in the nucleus of HEL cells; (iii) in permissive U2OS cells, however, ICP0 lacking E3 ligase activity was translocated to the cytoplasm at a pace faster than that of wild-type ICP0, suggesting that nuclear retention of ICP0 occurs in an ICP0 E3 ligase-dependent manner; and (iv) the ICP0 C terminus and late viral proteins cooperate in order to overcome nuclear retention and stimulate ICP0 cytoplasmic translocation. Taken together, less ICP0 nuclear retention may contribute to the permissiveness of U2OS cells to HSV-1 in the absence of functional ICP0. IMPORTANCE A distinct characteristic for eukaryotes is the compartmentalization of cell metabolic pathways, which allows greater efficiency and specificity of cellular functions. ICP0 of HSV-1 is a multifunctional viral protein that travels through different compartments as infection progresses. Its main regulatory functions are carried out in the nucleus, but it is translocated to the cytoplasm late during HSV-1 infection. To understand the biological significance of cytoplasmic ICP0 in

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.

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

2016-07-01

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. 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 infection. PML

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

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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.

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

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Alene Kast

2015-05-01

Full Text Available 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.

The role of nuclear localization signal in parvovirus life cycle.

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Liu, Peng; Chen, Shun; Wang, Mingshu; Cheng, Anchun

2017-04-14

Parvoviruses are small, non-enveloped viruses with an approximately 5.0 kb, single-stranded DNA genome. Usually, the parvovirus capsid gene contains one or more nuclear localization signals (NLSs), which are required for guiding the virus particle into the nucleus through the nuclear pore. However, several classical NLSs (cNLSs) and non-classical NLSs (ncNLSs) have been identified in non-structural genes, and the ncNLSs can also target non-structural proteins into the nucleus. In this review, we have summarized recent research findings on parvovirus NLSs. The capsid protein of the adeno-associated virus has four potential nuclear localization sequences, named basic region 1 (BR), BR2, BR3 and BR4. BR3 was identified as an NLS by fusing it with green fluorescent protein. Moreover, BR3 and BR4 are required for infectivity and virion assembly. In Protoparvovirus, the canine parvovirus has a common cNLS located in the VP1 unique region, similar to parvovirus minute virus of mice (MVM) and porcine parvovirus. Moreover, an ncNLS is found in the C-terminal region of MVM VP1/2. Parvovirus B19 also contains an ncNLS in the C-terminal region of VP1/2, which is essential for the nuclear transport of VP1/VP2. Approximately 1 or 2 cNLSs and 1 ncNLS have been reported in the non-structural protein of bocaviruses. Understanding the role of the NLS in the process of parvovirus infection and its mechanism of nuclear transport will contribute to the development of therapeutic vaccines and novel antiviral medicines.

Continuous Lymphoid Cell Lines with Characteristics of B Cells (Bone-Marrow-Derived), Lacking the Epstein-Barr Virus Genome and Derived from Three Human Lymphomas

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Klein, George; Lindahl, Tomas; Jondal, Mikael; Leibold, Wolfgang; Menézes, José; Nilsson, Kenneth; Sundström, Christer

1974-01-01

Three exceptional cell lines have been tested for the presence of the Epstein-Barr virus genome by nucleic acid hybridization (complementary RNA·DNA) and Epstein-Barr virus-determined nuclear antigen tests. Two lines were derived from Swedish lymphoma cases and one from an African Burkitt-like lymphoma biopsy that was negative for Epstein-Barr virus DNA and the virus-determined nuclear antigen. All three lines apparently lacked the viral genome. Two of the three lines clearly had characteristics of B-cells (bone-marrow-derived). PMID:4369887

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

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Wang, Yimeng; Zhou, Jianhong; Du, Yuchun, E-mail: ydu@uark.edu

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.

Evasion of interferon responses by Ebola and Marburg viruses.

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Basler, Christopher F; Amarasinghe, Gaya K

2009-09-01

The filoviruses, Ebola virus (EBOV) and Marburg virus (MARV), cause frequently lethal viral hemorrhagic fever. These infections induce potent cytokine production, yet these host responses fail to prevent systemic virus replication. Consistent with this, filoviruses have been found to encode proteins VP35 and VP24 that block host interferon (IFN)-alpha/beta production and inhibit signaling downstream of the IFN-alpha/beta and the IFN-gamma receptors, respectively. VP35, which is a component of the viral nucleocapsid complex and plays an essential role in viral RNA synthesis, acts as a pseudosubstrate for the cellular kinases IKK-epsilon and TBK-1, which phosphorylate and activate interferon regulatory factor 3 (IRF-3) and interferon regulatory factor 7 (IRF-7). VP35 also promotes SUMOylation of IRF-7, repressing IFN gene transcription. In addition, VP35 is a dsRNA-binding protein, and mutations that disrupt dsRNA binding impair VP35 IFN-antagonist activity while leaving its RNA replication functions intact. The phenotypes of recombinant EBOV bearing mutant VP35s unable to inhibit IFN-alpha/beta demonstrate that VP35 IFN-antagonist activity is critical for full virulence of these lethal pathogens. The structure of the VP35 dsRNA-binding domain, which has recently become available, is expected to provide insight into how VP35 IFN-antagonist and dsRNA-binding functions are related. The EBOV VP24 protein inhibits IFN signaling through an interaction with select host cell karyopherin-alpha proteins, preventing the nuclear import of otherwise activated STAT1. It remains to be determined to what extent VP24 may also modulate the nuclear import of other host cell factors and to what extent this may influence the outcome of infection. Notably, the Marburg virus VP24 protein does not detectably block STAT1 nuclear import, and, unlike EBOV, MARV infection inhibits STAT1 and STAT2 phosphorylation. Thus, despite their similarities, there are fundamental differences by which

Contribution of the C-terminal tri-lysine regions of human immunodeficiency virus type 1 integrase for efficient reverse transcription and viral DNA nuclear import

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Fowke Keith R

2005-10-01

Full Text Available Abstract Background In addition to mediating the integration process, HIV-1 integrase (IN has also been implicated in different steps during viral life cycle including reverse transcription and viral DNA nuclear import. Although the karyophilic property of HIV-1 IN has been well demonstrated using a variety of experimental approaches, the definition of domain(s and/or motif(s within the protein that mediate viral DNA nuclear import and its mechanism are still disputed and controversial. In this study, we performed mutagenic analyses to investigate the contribution of different regions in the C-terminal domain of HIV-1 IN to protein nuclear localization as well as their effects on virus infection. Results Our analysis showed that replacing lysine residues in two highly conserved tri-lysine regions, which are located within previously described Region C (235WKGPAKLLWKGEGAVV and sequence Q (211KELQKQITK in the C-terminal domain of HIV-1 IN, impaired protein nuclear accumulation, while mutations for RK263,4 had no significant effect. Analysis of their effects on viral infection in a VSV-G pseudotyped RT/IN trans-complemented HIV-1 single cycle replication system revealed that all three C-terminal mutant viruses (KK215,9AA, KK240,4AE and RK263,4AA exhibited more severe defect of induction of β-Gal positive cells and luciferase activity than an IN class 1 mutant D64E in HeLa-CD4-CCR5-β-Gal cells, and in dividing as well as non-dividing C8166 T cells, suggesting that some viral defects are occurring prior to viral integration. Furthermore, by analyzing viral DNA synthesis and the nucleus-associated viral DNA level, the results clearly showed that, although all three C-terminal mutants inhibited viral reverse transcription to different extents, the KK240,4AE mutant exhibited most profound effect on this step, whereas KK215,9AA significantly impaired viral DNA nuclear import. In addition, our analysis could not detect viral DNA integration in each C

Human nucleoporins promote HIV-1 docking at the nuclear pore, nuclear import and integration.

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Francesca Di Nunzio

Full Text Available The nuclear pore complex (NPC mediates nucleo-cytoplasmic transport of macromolecules and is an obligatory point of passage and functional bottleneck in the replication of some viruses. The Human Immunodeficiency Virus (HIV has evolved the required mechanisms for active nuclear import of its genome through the NPC. However the mechanisms by which the NPC allows or even assists HIV translocation are still unknown. We investigated the involvement of four key nucleoporins in HIV-1 docking, translocation, and integration: Nup358/RanBP2, Nup214/CAN, Nup98 and Nup153. Although all induce defects in infectivity when depleted, only Nup153 actually showed any evidence of participating in HIV-1 translocation through the nuclear pore. We show that Nup358/RanBP2 mediates docking of HIV-1 cores on NPC cytoplasmic filaments by interacting with the cores and that the C-terminus of Nup358/RanBP2 comprising a cyclophilin-homology domain contributes to binding. We also show that Nup214/CAN and Nup98 play no role in HIV-1 nuclear import per se: Nup214/CAN plays an indirect role in infectivity read-outs through its effect on mRNA export, while the reduction of expression of Nup98 shows a slight reduction in proviral integration. Our work shows the involvement of nucleoporins in diverse and functionally separable steps of HIV infection and nuclear import.

Non-coding RNAs and heme oxygenase-1 in vaccinia virus infection

International Nuclear Information System (INIS)

Meseda, Clement A.; Srinivasan, Kumar; Wise, Jasen; Catalano, Jennifer; Yamada, Kenneth M.; Dhawan, Subhash

2014-01-01

Highlights: • Heme oxygenase-1 (HO-1) induction inhibited vaccinia virus infection of macrophages. • Reduced infectivity inversely correlated with increased expression of non-coding RNAs. • The regulation of HO-1 and ncRNAs suggests a novel host defense response against vaccinia virus infection. - Abstract: Small nuclear RNAs (snRNAs) are <200 nucleotide non-coding uridylate-rich RNAs. Although the functions of many snRNAs remain undetermined, a population of snRNAs is produced during the early phase of infection of cells by vaccinia virus. In the present study, we demonstrate a direct correlation between expression of the cytoprotective enzyme heme oxygenase-1 (HO-1), suppression of selective snRNA expression, and inhibition of vaccinia virus infection of macrophages. Hemin induced HO-1 expression, completely reversed virus-induced host snRNA expression, and suppressed vaccinia virus infection. This involvement of specific virus-induced snRNAs and associated gene clusters suggests a novel HO-1-dependent host-defense pathway in poxvirus infection

Annulate lamellae in phloem cells of virus-infected Sonchus plants.

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Steinkamp, M P; Hoefert, L L

1977-07-01

The occurrence of annulate lamellae (AL) in differentiating phloem of Sonchus oleraceus (Compositae) singly infected with sowthistle yellow vein virus (SYVV) and doubly infected with a combination of SYVV and beet yellow stunt virus is documented by electron microscopy. Cell types in which AL were found were immature sieve elements and phloem parenchyma cells. AL were found only in cells that also contained SYVV particles although a direct association between the virus and AL was not apparent. The substructure of the AL and the relationships between the AL and the nuclear envelope and endoplasmic reticulum are similar to those reported in other descriptions of this organelle in the literature. This report appears to be the first one concerning the association of AL with a plant virus disease.

Autoantibodies from patients with systemic lupus erythematosus bind a shared sequence of SmD and Epstein-Barr virus-encoded nuclear antigen EBNA I.

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Sabbatini, A; Bombardieri, S; Migliorini, P

1993-05-01

SmD is one of the small nuclear ribonucleoproteins frequently targeted by autoantibodies in systemic lupus erythematosus. We isolated and characterized the antibodies present in lupus sera that are specific for the C-terminal region of SmD (sequence 95-119). This region is highly homologous to sequence 35-58 of the EBNA I antigen, one of the nuclear antigens induced by infection with Epstein-Barr virus. Antibodies affinity purified over a peptide 95-119 column were able to recognize this sequence in the context of the whole SmD molecule, as they reacted with blotted recombinant SmD. Anti-SmD 95-119 antibodies bound also the EBNA I 35-58 peptide and detected the EBNA I molecule in a total cell extract from Epstein-Barr virus-infected lines. A population of anti-SmD antibodies is, therefore, able to bind an epitope shared by the autoantigen and the viral antigen EBNA I. To investigate the involvement of this shared epitope in the generation of anti-SmD antibodies, we immunized mice with the EBNA I 35-58 peptide. Sera from immunized animals displayed the same pattern of reactivity of spontaneously produced anti-SmD antibodies. They reacted in fact with the EBNA peptide as well as with SmD 95-119 and recombinant SmD. These data suggest that molecular mimicry may play a role in the induction of anti-SmD autoantibodies.

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

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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...

Temporal expression of HIV-1 envelope proteins in baculovirus-infected insect cells: Implications for glycosylation and CD4 binding

International Nuclear Information System (INIS)

Murphy, C.I.; Lennick, M.; Lehar, S.M.; Beltz, G.A.; Young, E.

1990-01-01

Three different human immunodeficiency virus type I (HIV-1) envelope derived recombinant proteins and the full length human CD4 polypeptide were expressed in Spodoptera frugiperda (Sf9) cells. DNA constructs encoding CD4, gp120, gp160, and gp160 delta were cloned into the baculovirus expression vector pVL941 or a derivative and used to generate recombinant viruses in a cotransfection with DNA from Autographa californica nuclear polyhedrosis virus (AcMNPV). Western blotting of cell extracts of the recombinant HIV-1 proteins showed that for each construct two major bands specifically reacted with anti-HIV-1 envelope antiserum. These bands corresponded to glycosylated and nonglycosylated versions of the HIV proteins as determined by 3H-mannose labeling and tunicamycin treatment of infected cells. A time course of HIV envelope expression revealed that at early times post-infection (24 hours) the proteins were fully glycosylated and soluble in nonionic detergents. However, at later times postinfection (48 hours), expression levels of recombinant protein reached a maximum but most of the increase was due to a rise in the level of the nonglycosylated species, which was largely insoluble in nonionic detergents. Thus, it appears that Sf9 cells cannot process large amounts of glycosylated recombinant proteins efficiently. As a measure of biological activity, the CD4 binding ability of both glycosylated and nonglycosylated recombinant HIV envelope proteins was tested in a coimmunoprecipitation assay. The results showed that CD4 and the glycosylated versions of recombinant gp120 or gp160 delta specifically associated with one another in this analysis. Nonglycosylated gp120 or gp160 delta proteins from tunicamycin-treated cultures did immunoprecipitate with anti-HIV-1 antiserum but did not interact with CD4

Amino Acids of Epstein-Barr Virus Nuclear Antigen 3A Essential for Repression of Jκ-Mediated Transcription and Their Evolutionary Conservation

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Dalbiès-Tran, Rozenn; Stigger-Rosser, Evelyn; Dotson, Travis; Sample, Clare E.

2001-01-01

Epstein-Barr virus (EBV) nuclear antigen 3A (EBNA-3A) is essential for virus-mediated immortalization of B lymphocytes in vitro and is believed to regulate transcription of cellular and/or viral genes. One known mechanism of regulation is through its interaction with the cellular transcription factor Jκ. This interaction downregulates transcription mediated by EBNA-2 and Jκ. To identify the amino acids that play a role in this interaction, we have generated mutant EBNA-3A proteins. A mutant EBNA-3A protein in which alanine residues were substituted for amino acids 199, 200, and 202 no longer downregulated transcription. Surprisingly, this mutant protein remained able to coimmunoprecipitate with Jκ. Using a reporter gene assay based on the recruitment of Jκ by various regions spanning EBNA-3A, we have shown that this mutation abolished binding of Jκ to the N-proximal region (amino acids 125 to 222) and that no other region of EBNA-3A alone was sufficient to mediate an association with Jκ. To determine the biological significance of the interaction of EBNA-3A with Jκ, we have studied its conservation in the simian lymphocryptovirus herpesvirus papio (HVP) by cloning HVP-3A, the homolog of EBNA-3A encoded by this virus. This 903-amino-acid protein exhibited 37% identity with its EBV counterpart, mainly within the amino-terminal half. HVP-3A also interacted with Jκ through a region located between amino acids 127 and 223 and also repressed transcription mediated through EBNA-2 and Jκ. The evolutionary conservation of this function, in proteins that have otherwise significantly diverged, argues strongly for an important biological role in virus-mediated immortalization of B lymphocytes. PMID:11119577

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

International Nuclear Information System (INIS)

Lee, Boo-Ja; Kwon, Sun Jae; Kim, Sung-Kyu; Kim, Ki-Jeong; Park, Chang-Jin; Kim, Young-Jin; Park, Ohkmae K.; Paek, Kyung-Hee

2006-01-01

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 0 interaction, but not during compatible TMV-P 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

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

International Nuclear Information System (INIS)

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

2011-01-01

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.

Nuclear Exodus: Herpesviruses Lead the Way

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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

Immune responses to epstein-barr virus in atomic bomb survivors: Study of precursor frequency of cytotoxic lymphocytes and titer levels of anti-Epstein-Barr virus-related antibodies

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Kusunoki, Yoichiro; Kyoizumi, Seishi; Saito, Mayumi; Ozaki, Kyoko; Hirai, Yuko; Akiyama, Mitoshi (Radiation Effects Research Foundation, Hiroshima (Japan)); Fukuda, Yasuko (Radiation Effects Research Foundation, Hiroshima (Japan) Children' s Hospital Medical Center of Northern California, Oakland, CA (United States)); Huang, Hua (Radiation Effects Research Foundation, Hiroshima (Japan) Univ. of Wisconsin, Madison, WI (United States))

1994-04-01

Precursor frequencies of cytotoxic lymphocytes to autologous Epstein-Barr virus-transformed B cells and serum titers of anti-Epstein-Barr virus-related antibodies were measured in 68 atomic bomb survivors to clarify the immune mechanism controlling Epstein-Barr virus infection. The precursor frequency was negatively correlated with the titer of anti-early antigen lgG, which is probably produced at the stage of viral reactivation. A positive correlation between the precursor frequency and titer of anti-Epstein-Barr virus-associated nuclear antigen antibody was also observed, indicating that the precursor frequency reflects the degree of in vivo destruction by T cells of the virus-infected cells. These results suggest that T-cell memory specific to Epstein-Barr virus keeps the virus under control and that the precursor frequency assay is useful for the evaluation of immune responses to Epstein-Barr virus. However, no significant effect of atomic bomb radiation on the precursor frequency was observed in the present study, probably due to the limited number of participants. 24 refs., 4 figs., 2 tabs.

Immune responses to epstein-barr virus in atomic bomb survivors: Study of precursor frequency of cytotoxic lymphocytes and titer levels of anti-Epstein-Barr virus-related antibodies

International Nuclear Information System (INIS)

Kusunoki, Yoichiro; Kyoizumi, Seishi; Saito, Mayumi; Ozaki, Kyoko; Hirai, Yuko; Akiyama, Mitoshi; Fukuda, Yasuko; Huang, Hua

1994-01-01

Precursor frequencies of cytotoxic lymphocytes to autologous Epstein-Barr virus-transformed B cells and serum titers of anti-Epstein-Barr virus-related antibodies were measured in 68 atomic bomb survivors to clarify the immune mechanism controlling Epstein-Barr virus infection. The precursor frequency was negatively correlated with the titer of anti-early antigen lgG, which is probably produced at the stage of viral reactivation. A positive correlation between the precursor frequency and titer of anti-Epstein-Barr virus-associated nuclear antigen antibody was also observed, indicating that the precursor frequency reflects the degree of in vivo destruction by T cells of the virus-infected cells. These results suggest that T-cell memory specific to Epstein-Barr virus keeps the virus under control and that the precursor frequency assay is useful for the evaluation of immune responses to Epstein-Barr virus. However, no significant effect of atomic bomb radiation on the precursor frequency was observed in the present study, probably due to the limited number of participants. 24 refs., 4 figs., 2 tabs

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

International Nuclear Information System (INIS)

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

2011-01-01

Highlights: → LNA-modified oligonucleotides can pass through the plasma membrane of cultured cells even without using transfection machinery. → LNA-modified oligonucleotides passed efficiently across the cell membrane, and lipid-coating facilitated translocation from the cytoplasm to the nucleus. → 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.

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

Energy Technology Data Exchange (ETDEWEB)

Sun, Zhen [The State Key Laboratory of Genetic Engineering and The MOE Key Laboratory of Contemporary Anthropology, School of Life Science, Fudan University, Shanghai 200433 (China); Department of Biochemistry and Molecular Biology, Program in Molecular Cell Biology, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310058 (China); Xiang, Wenqing; Guo, Yajuan [Department of Biochemistry and Molecular Biology, Program in Molecular Cell Biology, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310058 (China); Chen, Zhi [The State Key Laboratory for Infectious Disease, Institute of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310003 (China); Liu, Wei, E-mail: liuwei666@zju.edu.cn [Department of Biochemistry and Molecular Biology, Program in Molecular Cell Biology, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310058 (China); Lu, Daru, E-mail: drlu@fudan.edu.cn [The State Key Laboratory of Genetic Engineering and The MOE Key Laboratory of Contemporary Anthropology, School of Life Science, Fudan University, Shanghai 200433 (China)

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.

Viral and Host Factors Required for Avian H5N1 Influenza A Virus Replication in Mammalian Cells

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Hong Zhang

2013-06-01

Full Text Available Following the initial and sporadic emergence into humans of highly pathogenic avian H5N1 influenza A viruses in Hong Kong in 1997, we have come to realize the potential for avian influenza A viruses to be transmitted directly from birds to humans. Understanding the basic viral and cellular mechanisms that contribute to infection of mammalian species with avian influenza viruses is essential for developing prevention and control measures against possible future human pandemics. Multiple physical and functional cellular barriers can restrict influenza A virus infection in a new host species, including the cell membrane, the nuclear envelope, the nuclear environment, and innate antiviral responses. In this review, we summarize current knowledge on viral and host factors required for avian H5N1 influenza A viruses to successfully establish infections in mammalian cells. We focus on the molecular mechanisms underpinning mammalian host restrictions, as well as the adaptive mutations that are necessary for an avian influenza virus to overcome them. It is likely that many more viral and host determinants remain to be discovered, and future research in this area should provide novel and translational insights into the biology of influenza virus-host interactions.

Tubular forms of papova viruses in human laryngeal papilloma.

Science.gov (United States)

Arnold, W

1979-01-01

In two cases of recurrent laryngeal papillomatosis tubular forms of papova viruses could be observed. The same material revealed the close relation between nuclear chromatine and the release of particles, as well as a capsomere like substructure of the virions.

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

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Christina Funk

2015-06-01

Full Text Available 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

Subcellular localization of low-abundance human immunodeficiency virus nucleic acid sequences visualized by fluorescence in situ hybridization

International Nuclear Information System (INIS)

Lawrence, J.B.; Marselle, L.M.; Byron, K.S.; Johnson, C.V.; Sullivan, J.L.; Singer, R.H.

1990-01-01

Detection and subcellular localization of human immunodeficiency virus (HIV) were investigated using sensitive high-resolution in situ hybridization methodology. Lymphocytes infected with HIV in vitro or in vivo were detected by fluorescence after hybridization with either biotin or digoxigenin-labeled probes. At 12 hr after infection in vitro, a single intense signal appeared in the nuclei of individual cells. Later in infection, when cytoplasmic fluorescence became intense, multiple nuclear foci frequently appeared. The nuclear focus consisted of newly synthesized HIV RNA as shown by hybridization in the absence of denaturation and by susceptibility to RNase and actinomycin D. Virus was detected in patient lymphocytes and it was shown that a singular nuclear focus also characterizes cells infected in vivo. The cell line 8E5/LAV containing one defective integrated provirus revealed a similar focus of nuclear RNA, and the single integrated HIV genome was unequivocally visualized on a D-group chromosome. This demonstrates an extremely sensitive single-cell assay for the presence of a single site of HIV transcription in vitro and in vivo and suggests that it derives from one (or very few) viral genomes per cell. In contrast, productive Epstein-Barr virus infection exhibited many foci of nuclear RNA per cell

A DNA Binding Protein Is Required for Viral Replication and Transcription in Bombyx mori Nucleopolyhedrovirus.

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Cui Zhao

Full Text Available A DNA-binding protein (DBP [GenBank accession number: M63416] of Bombyx mori nuclear polyhedrosis virus (BmNPV has been reported to be a regulatory factor in BmNPV, but its detailed functions remain unknown. In order to study the regulatory mechanism of DBP on viral proliferation, genome replication, and gene transcription, a BmNPV dbp gene knockout virus dbp-ko-Bacmid was generated by the means of Red recombination system. In addition, dbp-repaired virus dbp-re-Bacmid was constructed by the means of the Bac to Bac system. Then, the Bacmids were transfected into BmN cells. The results of this viral titer experiment revealed that the TCID50 of the dbp-ko-Bacmid was 0; however, the dbp-re-Bacmid was similar to the wtBacmid (p>0.05, indicating that the dbp-deficient would lead to failure in the assembly of virus particles. In the next step, Real-Time PCR was used to analyze the transcriptional phases of dbp gene in BmN cells, which had been infected with BmNPV. The results of the latter experiment revealed that the transcript of dbp gene was first detected at 3 h post-infection. Furthermore, the replication level of virus genome and the transcriptional level of virus early, late, and very late genes in BmN cells, which had been transfected with 3 kinds of Bacmids, were analyzed by Real-Time PCR. The demonstrating that the replication level of genome was lower than that of wtBacmid and dbp-re-Bacmid (p<0.01. The transcriptional level of dbp-ko-Bacmid early gene lef-3, ie-1, dnapol, late gene vp39 and very late gene p10 were statistically significantly lower than dbp-re-Bacmid and wtBacmid (p<0.01. The results presented are based on Western blot analysis, which indicated that the lack of dbp gene would lead to low expressions of lef3, vp39, and p10. In conclusion, dbp was not only essential for early viral replication, but also a viral gene that has a significant impact on transcription and expression during all periods of baculovirus life cycle.

Ebola Virus and Marburg Virus

Science.gov (United States)

Ebola virus and Marburg virus Overview Ebola virus and Marburg virus are related viruses that cause hemorrhagic fevers — illnesses marked by severe bleeding (hemorrhage), organ failure and, in many ...

Chlorotic spots on Clerodendrum, a disease caused by a nuclear type of Brevipalpus (Acari:Tenuipalpidae transmitted virus Mancha clorótica do Clerodendrum, uma enfermidade causada por um vírus do tipo nuclear, transmitido pelo ácaro Brevipalpus phoenicis (Acari:Tenuipalpidae

Directory of Open Access Journals (Sweden)

Elliot Watanabe Kitajima

2008-02-01

Full Text Available Chlorotic spots have been observed in plants of Clerodendrum x speciosum growing in residential gardens and parks in Piracicaba, SP, Brazil. Thin sections of diseased tissues revealed characteristic cytopathic effects of the nuclear type of the Brevipalpus (Acari: Tenuipalpidae mite-transmitted viruses (BTrV. Brevipalpus mites, identified as B. phoenicis, infesting symptomatic C. x speciosum plants transmitted the pathogen to healthy C. x speciosum and to C. thomsonae, Gomphrena globosa, Hibiscus cannabinus, H. coccineus, H. schizopetalus, Salvia leucantha, Spathiphyllum wallasi and Tetragonia expansa causing chlorotic spots on their leaves. Mechanical inoculation using leaf extracts from infected C. x speciosum resulted in chlorotic spots on inoculated C. x speciosum, Chenopodium quinoa, C. amaranticolor, G. globosa, H. cannabinus, H. coccineus and T. expansa leaves. C. amaranticolor and C. quinoa kept at 28 - 30°C became systemically infected. The same cytopathic effects caused by the nuclear type of BTrV were seen in tissues from all infected test plants by electron microscopy. The virus was purified from systemically infected leaves of C. amaranticolor and C. quinoa. A polyclonal antiserum obtained from an immunized rabbit presented a strong reaction with the homologous antigen in ELISA tests. The results suggest that this chlorotic spot disease of C. x speciosum is caused by a new species of the nuclear type of BTrV, tentatively named Clerodendrum chlorotic spot virus (ClCSV.Manchas cloróticas e necróticas foram observadas em folhas de várias plantas de coração-sangrento (Clerodendrum x speciosum cultivadas em parques e jardins em Piracicaba, SP, associadas à infestação pelo ácaro tenuipalpídeo Brevipalpus phoenicis. Exames preliminares de secções de tecido das manchas cloróticas ao microscópio eletrônico revelaram a ocorrência de efeitos citopáticos característicos dos induzidos pelos vírus do tipo nuclear, transmitido

Alterations in the nuclear proteome of HIV-1 infected T-cells

International Nuclear Information System (INIS)

DeBoer, Jason; Jagadish, Teena; Haverland, Nicole A.; Madson, Christian J.; Ciborowski, Pawel; Belshan, Michael

2014-01-01

Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines

Alterations in the nuclear proteome of HIV-1 infected T-cells

Energy Technology Data Exchange (ETDEWEB)

DeBoer, Jason [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Jagadish, Teena; Haverland, Nicole A. [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Ciborowski, Pawel [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States)

2014-11-15

Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines.

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

Energy Technology Data Exchange (ETDEWEB)

Sahu, Sushil Kumar; Mohanty, Suchitra; Kumar, Amit [Division of Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Chandrasekharpur, Bhubaneswar 751023 (India); Kundu, Chanakya N. [School of Biotechnology, KIIT University, Bhubaneswar (India); Verma, Subhash C. [Department of Microbiology and Immunology, University of Nevada, School of Medicine, Reno, NV 89557 (United States); Choudhuri, Tathagata, E-mail: tatha@ils.res.in [Division of Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Chandrasekharpur, Bhubaneswar 751023 (India); Department of Biotechnology, Siksha Bhavana, Visva Bharati, Santiniketan, Bolpur (India)

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.

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

International Nuclear Information System (INIS)

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

2014-01-01

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

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

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Myeong Kyu Choi

Full Text Available 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 (32EGDL(35 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 (32EGDL(35 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 (32EGDL(35 responsible for nuclear localization are important for the inhibitory activity of NV.

Nuclear relocalization of polyadenylate binding protein during rift valley fever virus infection involves expression of the NSs gene.

Science.gov (United States)

Copeland, Anna Maria; Altamura, Louis A; Van Deusen, Nicole M; Schmaljohn, Connie S

2013-11-01

Rift Valley fever virus (RVFV), an ambisense member of the family Bunyaviridae, genus Phlebovirus, is the causative agent of Rift Valley fever, an important zoonotic infection in Africa and the Middle East. Phlebovirus proteins are translated from virally transcribed mRNAs that, like host mRNA, are capped but, unlike host mRNAs, are not polyadenylated. Here, we investigated the role of PABP1 during RVFV infection of HeLa cells. Immunofluorescence studies of infected cells demonstrated a gross relocalization of PABP1 to the nucleus late in infection. Immunofluorescence microscopy studies of nuclear proteins revealed costaining between PABP1 and markers of nuclear speckles. PABP1 relocalization was sharply decreased in cells infected with a strain of RVFV lacking the gene encoding the RVFV nonstructural protein S (NSs). To determine whether PABP1 was required for RVFV infection, we measured the production of nucleocapsid protein (N) in cells transfected with small interfering RNAs (siRNAs) targeting PABP1. We found that the overall percentage of RVFV N-positive cells was not changed by siRNA treatment, indicating that PABP1 was not required for RVFV infection. However, when we analyzed populations of cells producing high versus low levels of PABP1, we found that the percentage of RVFV N-positive cells was decreased in cell populations producing physiologic levels of PABP1 and increased in cells with reduced levels of PABP1. Together, these results suggest that production of the NSs protein during RVFV infection leads to sequestration of PABP1 in the nuclear speckles, creating a state within the cell that favors viral protein production.

The host-dependent interaction of alpha-importins with influenza PB2 polymerase subunit is required for virus RNA replication.

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Patricia Resa-Infante

Full Text Available The influenza virus polymerase is formed by the PB1, PB2 and PA subunits and is required for virus transcription and replication in the nucleus of infected cells. As PB2 is a relevant host-range determinant we expressed a TAP-tagged PB2 in human cells and isolated intracellular complexes. Alpha-importin was identified as a PB2-associated factor by proteomic analyses. To study the relevance of this interaction for virus replication we mutated the PB2 NLS and analysed the phenotype of mutant subunits, polymerase complexes and RNPs. While mutant PB2 proteins showed reduced nuclear accumulation, they formed polymerase complexes normally when co expressed with PB1 and PA. However, mutant RNPs generated with a viral CAT replicon showed up to hundred-fold reduced CAT accumulation. Rescue of nuclear localisation of mutant PB2 by insertion of an additional SV40 TAg-derived NLS did not revert the mutant phenotype of RNPs. Furthermore, determination of recombinant RNP accumulation in vivo indicated that PB2 NLS mutations drastically reduced virus RNA replication. These results indicate that, above and beyond its role in nuclear accumulation, PB2 interaction with alpha-importins is required for virus RNA replication. To ascertain whether PB2-alpha-importin binding could contribute to the adaptation of H5N1 avian viruses to man, their association in vivo was determined. Human alpha importin isoforms associated efficiently to PB2 protein of an H3N2 human virus but bound to diminished and variable extents to PB2 from H5N1 avian or human strains, suggesting that the function of alpha importin during RNA replication is important for the adaptation of avian viruses to the human host.

Identification of pyrrolo[3,2-c]pyridin-4-amine compounds as a new class of entry inhibitors against influenza viruses in vitro

International Nuclear Information System (INIS)

Chang, So Young; Cruz, Deu John M.; Ko, Yoonae; Min, Ji-Young

2016-01-01

Various influenza virus entry inhibitors are being developed as therapeutic antiviral agents in ongoing preparation for emerging influenza viruses, particularly those that may possess drug resistance to the current FDA-approved neuraminidase inhibitors. In this study, small molecules having the pyrrolopyridinamine (PPA), aminothiadiazole (ATD), dihydrofuropyridine carboxamide (HPC), or imidazopyridinamine (IPA) moiety were selected from a target-focused chemical library for their inhibitory activity against influenza A virus by high-throughput screening using the PR8GFP assay. Activity was evaluated by measuring changes the proportion of GFP-expressing cells as a reflection of influenza virus infection. Among them, PPA showed broad-spectrum activity against multiple influenza A viruses and influenza B virus. PPA was found to block the early stages of influenza virus infection using a time-of-addition assay. Using additional phenotypic assays that dissect the virus entry process, it appears that the antiviral activity of PPA against influenza virus can be attributed to interference of the post-fusion process: namely, virus uncoating and nuclear import of viral nucleoprotein complexes. Based on these results, PPA is an attractive chemical moiety that can be used to develop new antiviral drug candidates against influenza viruses. - Highlights: • Four chemical classes are identified from target-focused chemical library by HTS. • PPA inhibits the infection of various influenza A viruses and influenza B virus. • PPA is identified to inhibit the early stages of influenza virus infection. • PPA compound disrupts virus uncoating and nuclear import of viral ribonucleoprotein.

Epstein-Barr virus nuclear antigen EBNA-LP is essential for transforming naïve B cells, and facilitates recruitment of transcription factors to the viral genome.

Science.gov (United States)

Szymula, Agnieszka; Palermo, Richard D; Bayoumy, Amr; Groves, Ian J; Ba Abdullah, Mohammed; Holder, Beth; White, Robert E

2018-02-01

The Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is the first viral latency-associated protein produced after EBV infection of resting B cells. Its role in B cell transformation is poorly defined, but it has been reported to enhance gene activation by the EBV protein EBNA2 in vitro. We generated EBNA-LP knockout (LPKO) EBVs containing a STOP codon within each repeat unit of internal repeat 1 (IR1). EBNA-LP-mutant EBVs established lymphoblastoid cell lines (LCLs) from adult B cells at reduced efficiency, but not from umbilical cord B cells, which died approximately two weeks after infection. Adult B cells only established EBNA-LP-null LCLs with a memory (CD27+) phenotype. Quantitative PCR analysis of virus gene expression after infection identified both an altered ratio of the EBNA genes, and a dramatic reduction in transcript levels of both EBNA2-regulated virus genes (LMP1 and LMP2) and the EBNA2-independent EBER genes in the first 2 weeks. By 30 days post infection, LPKO transcription was the same as wild-type EBV. In contrast, EBNA2-regulated cellular genes were induced efficiently by LPKO viruses. Chromatin immunoprecipitation revealed that EBNA2 and the host transcription factors EBF1 and RBPJ were delayed in their recruitment to all viral latency promoters tested, whereas these same factors were recruited efficiently to several host genes, which exhibited increased EBNA2 recruitment. We conclude that EBNA-LP does not simply co-operate with EBNA2 in activating gene transcription, but rather facilitates the recruitment of several transcription factors to the viral genome, to enable transcription of virus latency genes. Additionally, our findings suggest that EBNA-LP is essential for the survival of EBV-infected naïve B cells.

Epstein-Barr virus nuclear antigen EBNA-LP is essential for transforming naïve B cells, and facilitates recruitment of transcription factors to the viral genome

Science.gov (United States)

Szymula, Agnieszka; Palermo, Richard D.; Bayoumy, Amr; Groves, Ian J.

2018-01-01

The Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is the first viral latency-associated protein produced after EBV infection of resting B cells. Its role in B cell transformation is poorly defined, but it has been reported to enhance gene activation by the EBV protein EBNA2 in vitro. We generated EBNA-LP knockout (LPKO) EBVs containing a STOP codon within each repeat unit of internal repeat 1 (IR1). EBNA-LP-mutant EBVs established lymphoblastoid cell lines (LCLs) from adult B cells at reduced efficiency, but not from umbilical cord B cells, which died approximately two weeks after infection. Adult B cells only established EBNA-LP-null LCLs with a memory (CD27+) phenotype. Quantitative PCR analysis of virus gene expression after infection identified both an altered ratio of the EBNA genes, and a dramatic reduction in transcript levels of both EBNA2-regulated virus genes (LMP1 and LMP2) and the EBNA2-independent EBER genes in the first 2 weeks. By 30 days post infection, LPKO transcription was the same as wild-type EBV. In contrast, EBNA2-regulated cellular genes were induced efficiently by LPKO viruses. Chromatin immunoprecipitation revealed that EBNA2 and the host transcription factors EBF1 and RBPJ were delayed in their recruitment to all viral latency promoters tested, whereas these same factors were recruited efficiently to several host genes, which exhibited increased EBNA2 recruitment. We conclude that EBNA-LP does not simply co-operate with EBNA2 in activating gene transcription, but rather facilitates the recruitment of several transcription factors to the viral genome, to enable transcription of virus latency genes. Additionally, our findings suggest that EBNA-LP is essential for the survival of EBV-infected naïve B cells. PMID:29462212

Nuclear Cyber Security Case Study and Analysis

Energy Technology Data Exchange (ETDEWEB)

Park, Sunae [ChungNam National Univ., Daejeon (Korea, Republic of); Kim, Kyung doo [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

2016-10-15

Due to the new trend in cyber attacks, there is an increased security threat towards every country's infrastructure. So, security measures are required now than ever before. Previous cyber attacks normal process consists of paralyzing a server function, data extraction, or data control into the IT system for trespassing. However, nowadays control systems and infrastructures are also targeted and attacking methods have changed a lot. These days, the virus is becoming increasingly serious and hacker attacks are also becoming more frequent. This virus is a computer virus produced for the purpose of destroying the infrastructure, such as power plants, airports, railways June 2010, and it was first discovered in Belarus. Israel, the US, and other countries are believed culprits behind Stuxnet attacks on other nations such as Iran. Recent malware distribution, such as website hacking threat is growing. In surveys today one of the most long-term posing security threats is from North Korea. In particular, North Korea has been caught launching ongoing cyber-attacks after their latest nuclear test. South Korea has identified national trends regarding North Korean nuclear tests and analyzed them in order to catch disclosed confidential information. Especially, many nuclear power plants in the world are found to be vulnerable to cyber-attacks. Industrial facilities should be more wary of the risk of a serious cyber attack in the middle is going to increase the reliance on universal and commercial digital systems (off the shelf) software, civilian nuclear infrastructure. Senior executives’ current risk rate levels are increasing. Digitalization of the perception of risk is lacking in nuclear power plants and workers are creating prevention methods to make them fully aware of the risks of cyber-attacks. It is suggested that it may be inappropriate to assume we are prepared for potential attacks. Due to advances in technology, a warning that the growing sense of crisis

Nuclear Cyber Security Case Study and Analysis

International Nuclear Information System (INIS)

Park, Sunae; Kim, Kyung doo

2016-01-01

Due to the new trend in cyber attacks, there is an increased security threat towards every country's infrastructure. So, security measures are required now than ever before. Previous cyber attacks normal process consists of paralyzing a server function, data extraction, or data control into the IT system for trespassing. However, nowadays control systems and infrastructures are also targeted and attacking methods have changed a lot. These days, the virus is becoming increasingly serious and hacker attacks are also becoming more frequent. This virus is a computer virus produced for the purpose of destroying the infrastructure, such as power plants, airports, railways June 2010, and it was first discovered in Belarus. Israel, the US, and other countries are believed culprits behind Stuxnet attacks on other nations such as Iran. Recent malware distribution, such as website hacking threat is growing. In surveys today one of the most long-term posing security threats is from North Korea. In particular, North Korea has been caught launching ongoing cyber-attacks after their latest nuclear test. South Korea has identified national trends regarding North Korean nuclear tests and analyzed them in order to catch disclosed confidential information. Especially, many nuclear power plants in the world are found to be vulnerable to cyber-attacks. Industrial facilities should be more wary of the risk of a serious cyber attack in the middle is going to increase the reliance on universal and commercial digital systems (off the shelf) software, civilian nuclear infrastructure. Senior executives’ current risk rate levels are increasing. Digitalization of the perception of risk is lacking in nuclear power plants and workers are creating prevention methods to make them fully aware of the risks of cyber-attacks. It is suggested that it may be inappropriate to assume we are prepared for potential attacks. Due to advances in technology, a warning that the growing sense of crisis about

Endoplasmic reticulum-to-Golgi transitions upon herpes virus infection [version 2; referees: 1 approved, 3 approved with reservations

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Peter Wild

2018-02-01

Full Text Available Background: Herpesvirus capsids are assembled in the nucleus, translocated to the perinuclear space by budding, acquiring tegument and envelope, or released to the cytoplasm via impaired nuclear envelope. One model proposes that envelopment, “de-envelopment” and “re-envelopment” is essential for production of infectious virus. Glycoproteins gB/gH were reported to be essential for de-envelopment, by fusion of the “primary” envelope with the outer nuclear membrane. Yet, a high proportion of enveloped virions generated from genomes with deleted gB/gH were found in the cytoplasm and extracellular space, suggesting the existence of alternative exit routes. Methods: We investigated the relatedness between the nuclear envelope and membranes of the endoplasmic reticulum and Golgi complex, in cells infected with either herpes simplex virus 1 (HSV-1 or a Us3 deletion mutant thereof, or with bovine herpesvirus 1 (BoHV-1 by transmission and scanning electron microscopy, employing freezing technique protocols. Results:  The Golgi complex is a compact entity in a juxtanuclear position covered by a membrane on the cis face. Golgi membranes merge with membranes of the endoplasmic reticulum forming an entity with the perinuclear space. All compartments contained enveloped virions. After treatment with brefeldin A, HSV-1 virions aggregated in the perinuclear space and endoplasmic reticulum, while infectious progeny virus was still produced. Conclusions: The data suggest that virions derived by budding at nuclear membranes are intraluminally transported from the perinuclear space via Golgi -endoplasmic reticulum transitions into Golgi cisternae for packaging. Virions derived by budding at nuclear membranes are infective like Us3 deletion mutants, which  accumulate in the perinuclear space. Therefore, i de-envelopment followed by re-envelopment is not essential for production of infective progeny virus, ii the process taking place at the outer nuclear

Baculovirus potential for agricultural pests management in Cuba

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Jorge Luis Ayala Sifontes

2017-10-01

Full Text Available Cuba has an international reputation for implementing widespread biological control of pests, and microbial biocontrol is an integral component of most pest management programs. One class of microbial pesticides however, has not been developed in Cuba, bio-insecticides based on the Baculoviridae. This class of safe and environmentally protective microbial pesticides is used ever more commonly worldwide as an alternative to chemical pesticides. The characteristics of the viruses of this family, particularly their high host specificity, safety to non-target organisms, capacity to persist in nature and create epizootics, and the economy with which they can be produced "in vivo", all make them attractive for incorporation into pest management programs along with other pesticides developed in Cuba. The mass production technology is well understood in Cuba and biofactories already exist for a number of microbial biocontrol products. In the province of Sancti Spíritus, the Plant Protection Laboratory of the Ministry of Agriculture, with the cooperation of the Institute for Sustainable Horticulture, Kwantlen Polytechnic University, are resuming the work which began in the 90´s to develop baculovirus products in support of sustainable agriculture in Cuba. This work is being carried out with the participation of young Canadian and Cuban students and professionals. The program includes research with the multicapsid nuclear polyhedrosis viruses of Spodoptera frugiperda (SfMNPV and S. exigua (SeMNPV and the search for native isolates of Baculovirus in Plutella xylostella, three priority pests in Cuba. In other jurisdictions they are well controlled by baculoviruses, and the expectation is that this same result is possible in Cuba.

Generation of covalently closed circular DNA of hepatitis B viruses via intracellular recycling is regulated in a virus specific manner.

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Josef Köck

Full Text Available Persistence of hepatitis B virus (HBV infection requires covalently closed circular (cccDNA formation and amplification, which can occur via intracellular recycling of the viral polymerase-linked relaxed circular (rc DNA genomes present in virions. Here we reveal a fundamental difference between HBV and the related duck hepatitis B virus (DHBV in the recycling mechanism. Direct comparison of HBV and DHBV cccDNA amplification in cross-species transfection experiments showed that, in the same human cell background, DHBV but not HBV rcDNA converts efficiently into cccDNA. By characterizing the distinct forms of HBV and DHBV rcDNA accumulating in the cells we find that nuclear import, complete versus partial release from the capsid and complete versus partial removal of the covalently bound polymerase contribute to limiting HBV cccDNA formation; particularly, we identify genome region-selectively opened nuclear capsids as a putative novel HBV uncoating intermediate. However, the presence in the nucleus of around 40% of completely uncoated rcDNA that lacks most if not all of the covalently bound protein strongly suggests a major block further downstream that operates in the HBV but not DHBV recycling pathway. In summary, our results uncover an unexpected contribution of the virus to cccDNA formation that might help to better understand the persistence of HBV infection. Moreover, efficient DHBV cccDNA formation in human hepatoma cells should greatly facilitate experimental identification, and possibly inhibition, of the human cell factors involved in the process.

Virus-Vectored Influenza Virus Vaccines

Science.gov (United States)

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

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

International Nuclear Information System (INIS)

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

2007-01-01

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

Comparative morpho-anatomical studies of the lesions caused by citrus leprosis virus on sweet orange

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João P.R. Marques

2010-06-01

Full Text Available The leprosis disease shows a viral etiology and the citrus leprosis virus is considered its etiologic agent. The disease may show two types of cytopatologic symptom caused by two virus: nuclear (CiLV-N and cytoplasmic (CiLV-C types. The aim of this study was to compare the morpho-anatomical differences in the lesions caused by leprosis virus-cytoplasmic and nuclear types in Citrus sinensis (L. Osbeck 'Pêra'. Leaf and fruit lesions were collected in Piracicaba/São Paulo (cytoplasmic type and Monte Alegre do Sul/São Paulo and Amparo/São Paulo (nuclear type. The lesions were photographed and then fixed in Karnovsky solution, dehydrated in a graded ethylic series, embedded in hydroxy-ethyl methacrylate resin (Leica Historesin, sectioned (5 μm thick, stained and mounted in synthetic resin. The digital images were acquired in a microscope with digital video camera. Leaf and fruit lesions caused by the two viruses were morphologically distinct. Only the lesion caused by CiLV-N virus presented three well-defined regions. In both lesions there was the accumulation of lipidic substances in necrotic areas that were surrounded by cells with amorphous or droplets protein. Only leaf and fruit lesions caused by CiLV-N virus exhibited traumatic gum ducts in the vascular bundles.A doença leprose dos citros tem etiologia viral sendo o citrus leprosis virus seu agente etiológico. Demonstrou-se que há dois vírus distintos que causam sintomas de leprose em ci-tros: citoplasmático (CiLV-C e o nuclear (CiLV-N. O objetivo desse estudo foi comparar as diferenças morfo-anatômicas nas lesões causadas por CiLV-C e por CiLV-N em laranjeira doce (Citrus sinensis (L. Osbeck 'Pêra'. As lesões foliares e dos frutos foram coletadas em Piracicaba/SP (tipo citoplas-mático e em Monte Alegre do Sul/SP e Amparo/SP (tipo nuclear. As lesões foram fotografadas e em seguida fixadas em solução Karnovsky, desidratadas em série etílica, incluídas em historesina e

Pharmacological Induction of Heme Oxygenase-1 Impairs Nuclear Accumulation of Herpes Simplex Virus Capsids upon Infection

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Francisco J. Ibáñez

2017-10-01

Full Text Available Heme oxygenase-1 (HO-1 is an inducible enzyme that is expressed in response to physical and chemical stresses, such as ultraviolet radiation, hyperthermia, hypoxia, reactive oxygen species (ROS, as well as cytokines, among others. Its activity can be positively modulated by cobalt protoporphyrin (CoPP and negatively by tin protoporphirin (SnPP. Once induced, HO-1 degrades iron-containing heme into ferrous iron (Fe2+, carbon monoxide (CO and biliverdin. Importantly, numerous products of HO-1 are cytoprotective with anti-apoptotic, anti-oxidant, anti-inflammatory, and anti-cancer effects. The products of HO-1 also display antiviral properties against several viruses, such as the human immunodeficiency virus (HIV, influenza, hepatitis B, hepatitis C, and Ebola virus. Here, we sought to assess the effect of modulating HO-1 activity over herpes simplex virus type 2 (HSV-2 infection in epithelial cells and neurons. There are no vaccines against HSV-2 and treatment options are scarce in the immunosuppressed, in which drug-resistant variants emerge. By using HSV strains that encode structural and non-structural forms of the green fluorescent protein (GFP, we found that pharmacological induction of HO-1 activity with CoPP significantly decreases virus plaque formation and the expression of virus-encoded genes in epithelial cells as determined by flow cytometry and western blot assays. CoPP treatment did not affect virus binding to the cell surface or entry into the cytoplasm, but rather downstream events in the virus infection cycle. Furthermore, we observed that treating cells with a CO-releasing molecule (CORM-2 recapitulated some of the anti-HSV effects elicited by CoPP. Taken together, these findings indicate that HO-1 activity interferes with the replication cycle of HSV and that its antiviral effects can be recapitulated by CO.

Lack of Connection Between Midgut Cell Autophagy Gene Expression and BmCPV Infection in the Midgut of Bombyx mori.

Science.gov (United States)

Yang, Xiaobing; Wu, Suli; Wu, Yongpeng; Liu, Yang; Qian, Yonghua; Jiao, Feng

2015-01-01

Autophagy is associated with multiple biological processes and has protective and defensive functions with respect to immunity, inflammation, and resistance to microbial infection. In this experiment, we wished to investigate whether autophagy is a factor in the midgut cell response of Bombyx mori to infection by the B. mori cytoplasmic polyhedrosis virus (BmCPV). Our results indicated that the expression of three autophagy-related genes (BmAtg8, BmAtg5, and BmAtg7) in the midgut did not change greatly after BmCPV infection in B. mori. Basal ATG8/ATG8PE protein expression was detected in different B. mori tissues by using western blot analysis. Immunohistochemistry showed that the ATG8/ATG8PE proteins were located mainly in the cytoplasm. ATG8/ATG8PE protein levels decreased at 12 and 16 h after BmCPV infection. Our results indicate that autophagy responded slightly to BmCPV infection, but could not prevent the invasion and replication of the virus. © The Author 2015. Published by Oxford University Press on behalf of the Entomological Society of America.

Late Maturation Steps Preceding Selective Nuclear Export and Egress of Progeny Parvovirus.

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Wolfisberg, Raphael; Kempf, Christoph; Ros, Carlos

2016-06-01

Although the mechanism is not well understood, growing evidence indicates that the nonenveloped parvovirus minute virus of mice (MVM) may actively egress before passive release through cell lysis. We have dissected the late maturation steps of the intranuclear progeny with the aims of confirming the existence of active prelytic egress and identifying critical capsid rearrangements required to initiate the process. By performing anion-exchange chromatography (AEX), we separated intranuclear progeny particles by their net surface charges. Apart from empty capsids (EC), two distinct populations of full capsids (FC) arose in the nuclei of infected cells. The earliest population of FC to appear was infectious but, like EC, could not be actively exported from the nucleus. Further maturation of this early population, involving the phosphorylation of surface residues, gave rise to a second, late population with nuclear export potential. While capsid surface phosphorylation was strictly associated with nuclear export capacity, mutational analysis revealed that the phosphoserine-rich N terminus of VP2 (N-VP2) was dispensable, although it contributed to passive release. The reverse situation was observed for the incoming particles, which were dephosphorylated in the endosomes. Our results confirm the existence of active prelytic egress and reveal a late phosphorylation event occurring in the nucleus as a selective factor for initiating the process. In general, the process of egress of enveloped viruses is active and involves host cell membranes. However, the release of nonenveloped viruses seems to rely more on cell lysis. At least for some nonenveloped viruses, an active process before passive release by cell lysis has been reported, although the underlying mechanism remains poorly understood. By using the nonenveloped model parvovirus minute virus of mice, we could confirm the existence of an active process of nuclear export and further characterize the associated capsid

The Role of Nuclear Medicine in the Staging and Management of Human Immune Deficiency Virus Infection and Associated Diseases.

Science.gov (United States)

Ankrah, Alfred O; Glaudemans, Andor W J M; Klein, Hans C; Dierckx, Rudi A J O; Sathekge, Mike

2017-06-01

Human immune deficiency virus (HIV) is a leading cause of death. It attacks the immune system, thereby rendering the infected host susceptible to many HIV-associated infections, malignancies and neurocognitive disorders. The altered immune system affects the way the human host responds to disease, resulting in atypical presentation of these disorders. This presents a diagnostic challenge and the clinician must use all diagnostic avenues available to diagnose and manage these conditions. The advent of highly active antiretroviral therapy (HAART) has markedly reduced the mortality associated with HIV infection but has also brought in its wake problems associated with adverse effects or drug interaction and may even modulate some of the HIV-associated disorders to the detriment of the infected human host. Nuclear medicine techniques allow non-invasive visualisation of tissues in the body. By using this principle, pathophysiology in the body can be targeted and the treatment of diseases can be monitored. Being a functional imaging modality, it is able to detect diseases at the molecular level, and thus it has increased our understanding of the immunological changes in the infected host at different stages of the HIV infection. It also detects pathological changes much earlier than conventional imaging based on anatomical changes. This is important in the immunocompromised host as in some of the associated disorders a delay in diagnosis may have dire consequences. Nuclear medicine has played a huge role in the management of many HIV-associated disorders in the past and continues to help in the diagnosis, prognosis, staging, monitoring and assessing the response to treatment of many HIV-associated disorders. As our understanding of the molecular basis of disease increases nuclear medicine is poised to play an even greater role. In this review we highlight the functional basis of the clinicopathological correlation of HIV from a metabolic view and discuss how the use of

The role of nuclear medicine in the staging and management of human immune deficiency virus infection and associated diseases

International Nuclear Information System (INIS)

Ankrah, Alfred O.; Sathekge, Mike; Glaudemans, Andor W. J. M.; Klein, Hans; Dierckx, Rudi A. J. O.

2017-01-01

Human immune deficiency virus (HIV) is a leading cause of death. It attacks the immune system, thereby rendering the infected host susceptible to many HIV-associated infections, malignancies and neurocognitive disorders. The altered immune system affects the way the human host responds to disease, resulting in atypical presentation of these disorders. This presents a diagnostic challenge and the clinician must use all diagnostic avenues available to diagnose and manage these conditions. The advent of highly active antiretroviral therapy (HAART) has markedly reduced the mortality associated with HIV infection but has also brought in its wake problems associated with adverse effects or drug interaction and may even modulate some of the HIV-associated disorders to the detriment of the infected human host. Nuclear medicine techniques allow non-invasive visualisation of tissues in the body. By using this principle, pathophysiology in the body can be targeted and the treatment of diseases can be monitored. Being a functional imaging modality, it is able to detect diseases at the molecular level, and thus it has increased our understanding of the immunological changes in the infected host at different stages of the HIV infection. It also detects pathological changes much earlier than conventional imaging based on anatomical changes. This is important in the immunocompromised host as in some of the associated disorders a delay in diagnosis may have dire consequences. Nuclear medicine has played a huge role in the management of many HIV-associated disorders in the past and continues to help in the diagnosis, prognosis, staging, monitoring and assessing the response to treatment of many HIV-associated disorders. As our understanding of the molecular basis of disease increases nuclear medicine is poised to play an even greater role. In this review we highlight the functional basis of the clinicopathological correlation of HIV from a metabolic view and discuss how the use of

The role of nuclear medicine in the staging and management of human immune deficiency virus infection and associated diseases

Energy Technology Data Exchange (ETDEWEB)

Ankrah, Alfred O.; Sathekge, Mike [Dept. of Nuclear Medicine, University of Pretoria and Steve Biko Academic Hospital, Pretoria (South Africa); Glaudemans, Andor W. J. M.; Klein, Hans; Dierckx, Rudi A. J. O. [University of Groningen, University Medical Centre Groningen, Groningen (Netherlands)

2017-06-15

Human immune deficiency virus (HIV) is a leading cause of death. It attacks the immune system, thereby rendering the infected host susceptible to many HIV-associated infections, malignancies and neurocognitive disorders. The altered immune system affects the way the human host responds to disease, resulting in atypical presentation of these disorders. This presents a diagnostic challenge and the clinician must use all diagnostic avenues available to diagnose and manage these conditions. The advent of highly active antiretroviral therapy (HAART) has markedly reduced the mortality associated with HIV infection but has also brought in its wake problems associated with adverse effects or drug interaction and may even modulate some of the HIV-associated disorders to the detriment of the infected human host. Nuclear medicine techniques allow non-invasive visualisation of tissues in the body. By using this principle, pathophysiology in the body can be targeted and the treatment of diseases can be monitored. Being a functional imaging modality, it is able to detect diseases at the molecular level, and thus it has increased our understanding of the immunological changes in the infected host at different stages of the HIV infection. It also detects pathological changes much earlier than conventional imaging based on anatomical changes. This is important in the immunocompromised host as in some of the associated disorders a delay in diagnosis may have dire consequences. Nuclear medicine has played a huge role in the management of many HIV-associated disorders in the past and continues to help in the diagnosis, prognosis, staging, monitoring and assessing the response to treatment of many HIV-associated disorders. As our understanding of the molecular basis of disease increases nuclear medicine is poised to play an even greater role. In this review we highlight the functional basis of the clinicopathological correlation of HIV from a metabolic view and discuss how the use of

Actin-myosin network is required for proper assembly of influenza virus particles

Energy Technology Data Exchange (ETDEWEB)

Kumakura, Michiko; Kawaguchi, Atsushi, E-mail: ats-kawaguchi@md.tsukuba.ac.jp; Nagata, Kyosuke, E-mail: knagata@md.tsukuba.ac.jp

2015-02-15

Actin filaments are known to play a central role in cellular dynamics. After polymerization of actin, various actin-crosslinking proteins including non-muscle myosin II facilitate the formation of spatially organized actin filament networks. The actin-myosin network is highly expanded beneath plasma membrane. The genome of influenza virus (vRNA) replicates in the cell nucleus. Then, newly synthesized vRNAs are nuclear-exported to the cytoplasm as ribonucleoprotein complexes (vRNPs), followed by transport to the beneath plasma membrane where virus particles assemble. Here, we found that, by inhibiting actin-myosin network formation, the virus titer tends to be reduced and HA viral spike protein is aggregated on the plasma membrane. These results indicate that the actin-myosin network plays an important role in the virus formation. - Highlights: • Actin-myosin network is important for the influenza virus production. • HA forms aggregations at the plasma membrane in the presence of blebbistatin. • M1 is recruited to the budding site through the actin-myosin network.

Actin-myosin network is required for proper assembly of influenza virus particles

International Nuclear Information System (INIS)

Kumakura, Michiko; Kawaguchi, Atsushi; Nagata, Kyosuke

2015-01-01

Actin filaments are known to play a central role in cellular dynamics. After polymerization of actin, various actin-crosslinking proteins including non-muscle myosin II facilitate the formation of spatially organized actin filament networks. The actin-myosin network is highly expanded beneath plasma membrane. The genome of influenza virus (vRNA) replicates in the cell nucleus. Then, newly synthesized vRNAs are nuclear-exported to the cytoplasm as ribonucleoprotein complexes (vRNPs), followed by transport to the beneath plasma membrane where virus particles assemble. Here, we found that, by inhibiting actin-myosin network formation, the virus titer tends to be reduced and HA viral spike protein is aggregated on the plasma membrane. These results indicate that the actin-myosin network plays an important role in the virus formation. - Highlights: • Actin-myosin network is important for the influenza virus production. • HA forms aggregations at the plasma membrane in the presence of blebbistatin. • M1 is recruited to the budding site through the actin-myosin network

Nuclear translocation and regulation of intranuclear distribution of cytoplasmic poly(A-binding protein are distinct processes mediated by two Epstein Barr virus proteins.

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Richard Park

Full Text Available Many viruses target cytoplasmic polyA binding protein (PABPC to effect widespread inhibition of host gene expression, a process termed viral host-shutoff (vhs. During lytic replication of Epstein Barr Virus (EBV we observed that PABPC was efficiently translocated from the cytoplasm to the nucleus. Translocated PABPC was diffusely distributed but was excluded from viral replication compartments. Vhs during EBV infection is regulated by the viral alkaline nuclease, BGLF5. Transfection of BGLF5 alone into BGLF5-KO cells or uninfected 293 cells promoted translocation of PAPBC that was distributed in clumps in the nucleus. ZEBRA, a viral bZIP protein, performs essential functions in the lytic program of EBV, including activation or repression of downstream viral genes. ZEBRA is also an essential replication protein that binds to viral oriLyt and interacts with other viral replication proteins. We report that ZEBRA also functions as a regulator of vhs. ZEBRA translocated PABPC to the nucleus, controlled the intranuclear distribution of PABPC, and caused global shutoff of host gene expression. Transfection of ZEBRA alone into 293 cells caused nuclear translocation of PABPC in the majority of cells in which ZEBRA was expressed. Co-transfection of ZEBRA with BGLF5 into BGLF5-KO cells or uninfected 293 cells rescued the diffuse intranuclear pattern of PABPC seen during lytic replication. ZEBRA mutants defective for DNA-binding were capable of regulating the intranuclear distribution of PABPC, and caused PABPC to co-localize with ZEBRA. One ZEBRA mutant, Z(S186E, was deficient in translocation yet was capable of altering the intranuclear distribution of PABPC. Therefore ZEBRA-mediated nuclear translocation of PABPC and regulation of intranuclear PABPC distribution are distinct events. Using a click chemistry-based assay for new protein synthesis, we show that ZEBRA and BGLF5 each function as viral host shutoff factors.

Comparison of two automated instruments for Epstein-Barr virus serology in a large adult hospital and implementation of an Epstein-Barr virus nuclear antigen-based testing algorithm.

Science.gov (United States)

Al Sidairi, Hilal; Binkhamis, Khalifa; Jackson, Colleen; Roberts, Catherine; Heinstein, Charles; MacDonald, Jimmy; Needle, Robert; Hatchette, Todd F; LeBlanc, Jason J

2017-11-01

Serology remains the mainstay for diagnosis of Epstein-Barr virus (EBV) infection. This study compared two automated platforms (BioPlex 2200 and Architect i2000SR) to test three EBV serological markers: viral capsid antigen (VCA) immunoglobulins of class M (IgM), VCA immunoglobulins of class G (IgG) and EBV nuclear antigen-1 (EBNA-1) IgG. Using sera from 65 patients at various stages of EBV disease, BioPlex demonstrated near-perfect agreement for all EBV markers compared to a consensus reference. The agreement for Architect was near-perfect for VCA IgG and EBNA-1 IgG, and substantial for VCA IgM despite five equivocal results. Since the majority of testing in our hospital was from adults with EBNA-1 IgG positive results, post-implementation analysis of an EBNA-based algorithm showed advantages over parallel testing of the three serologic markers. This small verification demonstrated that both automated systems for EBV serology had good performance for all EBV markers, and an EBNA-based testing algorithm is ideal for an adult hospital.

SUN1 Regulates HIV-1 Nuclear Import in a Manner Dependent on the Interaction between the Viral Capsid and Cellular Cyclophilin A.

Science.gov (United States)

Luo, Xinlong; Yang, Wei; Gao, Guangxia

2018-07-01

Human immunodeficiency virus type 1 (HIV-1) can infect nondividing cells via passing through the nuclear pore complex. The nuclear membrane-imbedded protein SUN2 was recently reported to be involved in the nuclear import of HIV-1. Whether SUN1, which shares many functional similarities with SUN2, is involved in this process remained to be explored. Here we report that overexpression of SUN1 specifically inhibited infection by HIV-1 but not that by simian immunodeficiency virus (SIV) or murine leukemia virus (MLV). Overexpression of SUN1 did not affect reverse transcription but led to reduced accumulation of the 2-long-terminal-repeat (2-LTR) circular DNA and integrated viral DNA, suggesting a block in the process of nuclear import. HIV-1 CA was mapped as a determinant for viral sensitivity to SUN1. Treatment of SUN1-expressing cells with cyclosporine (CsA) significantly reduced the sensitivity of the virus to SUN1, and an HIV-1 mutant containing CA-G89A, which does not interact with cyclophilin A (CypA), was resistant to SUN1 overexpression. Downregulation of endogenous SUN1 inhibited the nuclear entry of the wild-type virus but not that of the G89A mutant. These results indicate that SUN1 participates in the HIV-1 nuclear entry process in a manner dependent on the interaction of CA with CypA. IMPORTANCE HIV-1 infects both dividing and nondividing cells. The viral preintegration complex (PIC) can enter the nucleus through the nuclear pore complex. It has been well known that the viral protein CA plays an important role in determining the pathways by which the PIC enters the nucleus. In addition, the interaction between CA and the cellular protein CypA has been reported to be important in the selection of nuclear entry pathways, though the underlying mechanisms are not very clear. Here we show that both SUN1 overexpression and downregulation inhibited HIV-1 nuclear entry. CA played an important role in determining the sensitivity of the virus to SUN1: the regulatory

Inhibition of host protein synthesis by Sindbis virus: correlation with viral RNA replication and release of nuclear proteins to the cytoplasm.

Science.gov (United States)

Sanz, Miguel A; García-Moreno, Manuel; Carrasco, Luis

2015-04-01

Infection of mammalian cells by Sindbis virus (SINV) profoundly blocks cellular mRNA translation. Experimental evidence points to viral non-structural proteins (nsPs), in particular nsP2, as the mediator of this inhibition. However, individual expression of nsP1, nsP2, nsP3 or nsP1-4 does not block cellular protein synthesis in BHK cells. Trans-complementation of a defective SINV replicon lacking most of the coding region for nsPs by the co-expression of nsP1-4 propitiates viral RNA replication at low levels, and inhibition of cellular translation is not observed. Exit of nuclear proteins including T-cell intracellular antigen and polypyrimidine tract-binding protein is clearly detected in SINV-infected cells, but not upon the expression of nsPs, even when the defective replicon was complemented. Analysis of a SINV variant with a point mutation in nsP2, exhibiting defects in the shut-off of host protein synthesis, indicates that both viral RNA replication and the release of nuclear proteins to the cytoplasm are greatly inhibited. Furthermore, nucleoside analogues that inhibit cellular and viral RNA synthesis impede the blockade of host mRNA translation, in addition to the release of nuclear proteins. Prevention of the shut-off of host mRNA translation by nucleoside analogues is not due to the inhibition of eIF2α phosphorylation, as this prevention is also observed in PKR(-/-) mouse embryonic fibroblasts that do not phosphorylate eIF2α after SINV infection. Collectively, our observations are consistent with the concept that for the inhibition of cellular protein synthesis to occur, viral RNA replication must take place at control levels, leading to the release of nuclear proteins to the cytoplasm. © 2014 John Wiley & Sons Ltd.

High-Throughput Quantitative Proteomic Analysis of Dengue Virus Type 2 Infected A549 Cells

Science.gov (United States)

Chiu, Han-Chen; Hannemann, Holger; Heesom, Kate J.; Matthews, David A.; Davidson, Andrew D.

2014-01-01

Disease caused by dengue virus is a global health concern with up to 390 million individuals infected annually worldwide. There are no vaccines or antiviral compounds available to either prevent or treat dengue disease which may be fatal. To increase our understanding of the interaction of dengue virus with the host cell, we analyzed changes in the proteome of human A549 cells in response to dengue virus type 2 infection using stable isotope labelling in cell culture (SILAC) in combination with high-throughput mass spectrometry (MS). Mock and infected A549 cells were fractionated into nuclear and cytoplasmic extracts before analysis to identify proteins that redistribute between cellular compartments during infection and reduce the complexity of the analysis. We identified and quantified 3098 and 2115 proteins in the cytoplasmic and nuclear fractions respectively. Proteins that showed a significant alteration in amount during infection were examined using gene enrichment, pathway and network analysis tools. The analyses revealed that dengue virus infection modulated the amounts of proteins involved in the interferon and unfolded protein responses, lipid metabolism and the cell cycle. The SILAC-MS results were validated for a select number of proteins over a time course of infection by Western blotting and immunofluorescence microscopy. Our study demonstrates for the first time the power of SILAC-MS for identifying and quantifying novel changes in cellular protein amounts in response to dengue virus infection. PMID:24671231

High-throughput quantitative proteomic analysis of dengue virus type 2 infected A549 cells.

Directory of Open Access Journals (Sweden)

Han-Chen Chiu

Full Text Available Disease caused by dengue virus is a global health concern with up to 390 million individuals infected annually worldwide. There are no vaccines or antiviral compounds available to either prevent or treat dengue disease which may be fatal. To increase our understanding of the interaction of dengue virus with the host cell, we analyzed changes in the proteome of human A549 cells in response to dengue virus type 2 infection using stable isotope labelling in cell culture (SILAC in combination with high-throughput mass spectrometry (MS. Mock and infected A549 cells were fractionated into nuclear and cytoplasmic extracts before analysis to identify proteins that redistribute between cellular compartments during infection and reduce the complexity of the analysis. We identified and quantified 3098 and 2115 proteins in the cytoplasmic and nuclear fractions respectively. Proteins that showed a significant alteration in amount during infection were examined using gene enrichment, pathway and network analysis tools. The analyses revealed that dengue virus infection modulated the amounts of proteins involved in the interferon and unfolded protein responses, lipid metabolism and the cell cycle. The SILAC-MS results were validated for a select number of proteins over a time course of infection by Western blotting and immunofluorescence microscopy. Our study demonstrates for the first time the power of SILAC-MS for identifying and quantifying novel changes in cellular protein amounts in response to dengue virus infection.

First discovery of acetone extract from cottonseed oil sludge as a novel antiviral agent against plant viruses.

Science.gov (United States)

Zhao, Lei; Feng, Chaohong; Hou, Caiting; Hu, Lingyun; Wang, Qiaochun; Wu, Yunfeng

2015-01-01

A novel acetone extract from cottonseed oil sludge was firstly discovered against plant viruses including Tobacco mosaic virus (TMV), Rice stripe virus (RSV) and Southern rice black streaked dwarf virus (SRBSDV). Gossypol and β-sitosterol separated from the acetone extract were tested for their effects on anti-TMV and analysed by nuclear magnetic resonance (NMR) assay. In vivo and field trials in different geographic distributions and different host varieties declared that this extract mixture was more efficient than the commercial agent Ningnanmycin with a broad spectrum of anti-plant-viruses activity. No phytotoxic activity was observed in the treated plants and environmental toxicology showed that this new acetone extract was environmentally friendly, indicating that this acetone extract has potential application in the control of plant virus in the future.

Herpes simplex virus 1 induces de novo phospholipid synthesis

Energy Technology Data Exchange (ETDEWEB)

Sutter, Esther [Electron Microscopy, Institute of Veterinary Anatomy, University of Zuerich (Switzerland); Oliveira, Anna Paula de; Tobler, Kurt [Electron microscopy, Institute of Virology, University of Zuerich (Switzerland); Schraner, Elisabeth M. [Electron Microscopy, Institute of Veterinary Anatomy, University of Zuerich (Switzerland); Sonda, Sabrina [Institute of Parasitology, University of Zuerich (Switzerland); Kaech, Andres [Center for Microscopy and Image Analysis, University of Zuerich (Switzerland); Lucas, Miriam S. [Electron Microscopy ETH Zuerich (EMEZ), Swiss Federal Institute of Technology, Zuerich (Switzerland); Ackermann, Mathias [Electron microscopy, Institute of Virology, University of Zuerich (Switzerland); Wild, Peter, E-mail: pewild@access.uzh.ch [Electron Microscopy, Institute of Veterinary Anatomy, University of Zuerich (Switzerland)

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.

Hepatitis B virus HBx protein localized to the nucleus restores HBx-deficient virus replication in HepG2 cells and in vivo in hydrodynamically-injected mice

International Nuclear Information System (INIS)

Keasler, Victor V.; Hodgson, Amanda J.; Madden, Charles R.; Slagle, Betty L.

2009-01-01

Identifying the requirements for the regulatory HBx protein in hepatitis B virus (HBV) replication is an important goal. A plasmid-based HBV replication assay was used to evaluate whether HBx subcellular localization influences its ability to promote virus replication, as measured by real time PCR quantitation of viral capsid-associated DNA. HBx targeted to the nucleus by a nuclear localization signal (NLS-HBx) was able to restore HBx-deficient HBV replication, while HBx containing a nuclear export signal (NES-HBx) was not. Both NLS-HBx and NES-HBx were expressed at similar levels (by immunoprecipitation and Western blotting), and proper localization of the signal sequence-tagged proteins was confirmed by deconvolution microscopy using HBx, NLS-HBx, and NES-HBx proteins fused to GFP. Importantly, these findings were confirmed in vivo by hydrodynamic injection into mice. Our results demonstrate that in these HBV replication assays, at least one function of HBx requires its localization to the nucleus.

Identification of human hnRNP C1/C2 as a dengue virus NS1-interacting protein

International Nuclear Information System (INIS)

Noisakran, Sansanee; Sengsai, Suchada; Thongboonkerd, Visith; Kanlaya, Rattiyaporn; Sinchaikul, Supachok; Chen, Shui-Tein; Puttikhunt, Chunya

2008-01-01

Dengue virus nonstructural protein 1 (NS1) is a key glycoprotein involved in the production of infectious virus and the pathogenesis of dengue diseases. Very little is known how NS1 interacts with host cellular proteins and functions in dengue virus-infected cells. This study aimed at identifying NS1-interacting host cellular proteins in dengue virus-infected cells by employing co-immunoprecipitation, two-dimensional gel electrophoresis, and mass spectrometry. Using lysates of dengue virus-infected human embryonic kidney cells (HEK 293T), immunoprecipitation with an anti-NS1 monoclonal antibody revealed eight isoforms of dengue virus NS1 and a 40-kDa protein, which was subsequently identified by quadrupole time-of-flight tandem mass spectrometry (Q-TOF MS/MS) as human heterogeneous nuclear ribonucleoprotein (hnRNP) C1/C2. Further investigation by co-immunoprecipitation and co-localization confirmed the association of hnRNP C1/C2 and dengue virus NS1 proteins in dengue virus-infected cells. Their interaction may have implications in virus replication and/or cellular responses favorable to survival of the virus in host cells

The directionality of the nuclear transport of the influenza A genome is driven by selective exposure of nuclear localization sequences on nucleoprotein

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Panté Nelly

2009-06-01

Full Text Available Abstract Background Early in infection, the genome of the influenza A virus, consisting of eight complexes of RNA and proteins (termed viral ribonucleoproteins; vRNPs, enters the nucleus of infected cells for replication. Incoming vRNPs are imported into the nucleus of infected cells using at least two nuclear localization sequences on nucleoprotein (NP; NLS1 at the N terminus, and NLS2 in the middle of the protein. Progeny vRNP assembly occurs in the nucleus, and later in infection, these are exported from the nucleus to the cytoplasm. Nuclear-exported vRNPs are different from incoming vRNPs in that they are prevented from re-entering the nucleus. Why nuclear-exported vRNPs do not re-enter the nucleus is unknown. Results To test our hypothesis that the exposure of NLSs on the vRNP regulates the directionality of the nuclear transport of the influenza vRNPs, we immunolabeled the two NLSs of NP (NLS1 and NLS2 and analyzed their surface accessibility in cells infected with the influenza A virus. We found that the NLS1 epitope on NP was exposed throughout the infected cells, but the NLS2 epitope on NP was only exposed in the nucleus of the infected cells. Addition of the nuclear export inhibitor leptomycin B further revealed that NLS1 is no longer exposed in cytoplasmic NP and vRNPs that have already undergone nuclear export. Similar immunolabeling studies in the presence of leptomycin B and with cells transfected with the cDNA of NP revealed that the NLS1 on NP is hidden in nuclear exported-NP. Conclusion NLS1 mediates the nuclear import of newly-synthesized NP and incoming vRNPs. This NLS becomes hidden on nuclear-exported NP and nuclear-exported vRNPs. Thus the selective exposure of the NLS1 constitutes a critical mechanism to regulate the directionality of the nuclear transport of vRNPs during the influenza A viral life cycle.

Scaffold expulsion and genome packaging trigger stabilization of herpes simplex virus capsids

NARCIS (Netherlands)

Roos, W.H.; Radtke, K.; Kniesmeijer, E.G.R.; Geertsema, H.J.; Sodeik, B.; Wuite, G.J.L.

2009-01-01

Herpes simplex virus type 1 (HSV1) capsids undergo extensive structural changes during maturation and DNA packaging. As a result, they become more stable and competent for nuclear egress. To further elucidate this stabilization process, we used biochemical and nanoindentation approaches to analyze

Scaffold expulsion and genome packaging trigger stabilization of Herpes Simplex Virus capsids

NARCIS (Netherlands)

Roos, W.H.; Radtke, K.; Kniesmeijer, E.; Geertsema, H.J.; Sodeik, B.; Wuite, G.J.L.

2009-01-01

Herpes simplex virus type 1 (HSV1) capsids undergo extensive structural changes during maturation and DNA packaging. As a result, they become more stable and competent for nuclear egress. To further elucidate this stabilization process, we used biochemical and nanoindentation approaches to analyze

Role of a nuclear localization signal on the minor capsid Proteins VP2 and VP3 in BKPyV nuclear entry

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Bennett, Shauna M. [Cellular and Molecular Biology Program University of Michigan 1150W Medical Center Dr 5724 Medical Science Bldg II Ann Arbor, MI 48109 (United States); Zhao, Linbo [Doctoral Program in Cancer Biology Program University of Michigan 1150W Medical Center Dr 5724 Medical Science Bldg II Ann Arbor, MI 48109 (United States); Bosard, Catherine [Department of Microbiology and Immunology University of Michigan 1150W Medical Center Dr 5724 Medical Science Bldg II Ann Arbor, MI 48109 (United States); Imperiale, Michael J., E-mail: imperial@umich.edu [Cellular and Molecular Biology Program University of Michigan 1150W Medical Center Dr 5724 Medical Science Bldg II Ann Arbor, MI 48109 (United States); Doctoral Program in Cancer Biology Program University of Michigan 1150W Medical Center Dr 5724 Medical Science Bldg II Ann Arbor, MI 48109 (United States); Department of Microbiology and Immunology University of Michigan 1150W Medical Center Dr 5724 Medical Science Bldg II Ann Arbor, MI 48109 (United States)

2015-01-01

BK Polyomavirus (BKPyV) is a ubiquitous nonenveloped human virus that can cause severe disease in immunocompromised populations. After internalization into renal proximal tubule epithelial cells, BKPyV traffics through the ER and enters the cytosol. However, it is unclear how the virus enters the nucleus. In this study, we elucidate a role for the nuclear localization signal located on the minor capsid proteins VP2 and VP3 during infection. Site-directed mutagenesis of a single lysine in the basic region of the C-terminus of the minor capsid proteins abrogated their nuclear localization, and the analogous genomic mutation reduced infectivity. Additionally, through use of the inhibitor ivermectin and knockdown of importin β1, we found that the importin α/β pathway is involved during infection. Overall these data are the first to show the significance of the NLS of the BKPyV minor capsid proteins during infection in a natural host cell. - Highlights: • Polyomaviruses must deliver their genome to the nucleus to replicate. • The minor capsid proteins have a well-conserved nuclear localization signal. • Mutation of this NLS diminishes, but does not completely inhibit, infection.

Role of a nuclear localization signal on the minor capsid Proteins VP2 and VP3 in BKPyV nuclear entry

International Nuclear Information System (INIS)

Bennett, Shauna M.; Zhao, Linbo; Bosard, Catherine; Imperiale, Michael J.

2015-01-01

BK Polyomavirus (BKPyV) is a ubiquitous nonenveloped human virus that can cause severe disease in immunocompromised populations. After internalization into renal proximal tubule epithelial cells, BKPyV traffics through the ER and enters the cytosol. However, it is unclear how the virus enters the nucleus. In this study, we elucidate a role for the nuclear localization signal located on the minor capsid proteins VP2 and VP3 during infection. Site-directed mutagenesis of a single lysine in the basic region of the C-terminus of the minor capsid proteins abrogated their nuclear localization, and the analogous genomic mutation reduced infectivity. Additionally, through use of the inhibitor ivermectin and knockdown of importin β1, we found that the importin α/β pathway is involved during infection. Overall these data are the first to show the significance of the NLS of the BKPyV minor capsid proteins during infection in a natural host cell. - Highlights: • Polyomaviruses must deliver their genome to the nucleus to replicate. • The minor capsid proteins have a well-conserved nuclear localization signal. • Mutation of this NLS diminishes, but does not completely inhibit, infection

Crystal Structure of the Herpesvirus Nuclear Egress Complex Provides Insights into Inner Nuclear Membrane Remodeling

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Tzviya Zeev-Ben-Mordehai

2015-12-01

Full Text Available Although nucleo-cytoplasmic transport is typically mediated through nuclear pore complexes, herpesvirus capsids exit the nucleus via a unique vesicular pathway. Together, the conserved herpesvirus proteins pUL31 and pUL34 form the heterodimeric nuclear egress complex (NEC, which, in turn, mediates the formation of tight-fitting membrane vesicles around capsids at the inner nuclear membrane. Here, we present the crystal structure of the pseudorabies virus NEC. The structure revealed that a zinc finger motif in pUL31 and an extensive interaction network between the two proteins stabilize the complex. Comprehensive mutational analyses, characterized both in situ and in vitro, indicated that the interaction network is not redundant but rather complementary. Fitting of the NEC crystal structure into the recently determined cryoEM-derived hexagonal lattice, formed in situ by pUL31 and pUL34, provided details on the molecular basis of NEC coat formation and inner nuclear membrane remodeling.

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

International Nuclear Information System (INIS)

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

2009-01-01

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.

The interactomes of influenza virus NS1 and NS2 proteins identify new host factors and provide insights for ADAR1 playing a supportive role in virus replication.

Science.gov (United States)

de Chassey, Benoît; Aublin-Gex, Anne; Ruggieri, Alessia; Meyniel-Schicklin, Laurène; Pradezynski, Fabrine; Davoust, Nathalie; Chantier, Thibault; Tafforeau, Lionel; Mangeot, Philippe-Emmanuel; Ciancia, Claire; Perrin-Cocon, Laure; Bartenschlager, Ralf; André, Patrice; Lotteau, Vincent

2013-01-01

Influenza A NS1 and NS2 proteins are encoded by the RNA segment 8 of the viral genome. NS1 is a multifunctional protein and a virulence factor while NS2 is involved in nuclear export of viral ribonucleoprotein complexes. A yeast two-hybrid screening strategy was used to identify host factors supporting NS1 and NS2 functions. More than 560 interactions between 79 cellular proteins and NS1 and NS2 proteins from 9 different influenza virus strains have been identified. These interacting proteins are potentially involved in each step of the infectious process and their contribution to viral replication was tested by RNA interference. Validation of the relevance of these host cell proteins for the viral replication cycle revealed that 7 of the 79 NS1 and/or NS2-interacting proteins positively or negatively controlled virus replication. One of the main factors targeted by NS1 of all virus strains was double-stranded RNA binding domain protein family. In particular, adenosine deaminase acting on RNA 1 (ADAR1) appeared as a pro-viral host factor whose expression is necessary for optimal viral protein synthesis and replication. Surprisingly, ADAR1 also appeared as a pro-viral host factor for dengue virus replication and directly interacted with the viral NS3 protein. ADAR1 editing activity was enhanced by both viruses through dengue virus NS3 and influenza virus NS1 proteins, suggesting a similar virus-host co-evolution.

The interactomes of influenza virus NS1 and NS2 proteins identify new host factors and provide insights for ADAR1 playing a supportive role in virus replication.

Directory of Open Access Journals (Sweden)

Benoît de Chassey

Full Text Available Influenza A NS1 and NS2 proteins are encoded by the RNA segment 8 of the viral genome. NS1 is a multifunctional protein and a virulence factor while NS2 is involved in nuclear export of viral ribonucleoprotein complexes. A yeast two-hybrid screening strategy was used to identify host factors supporting NS1 and NS2 functions. More than 560 interactions between 79 cellular proteins and NS1 and NS2 proteins from 9 different influenza virus strains have been identified. These interacting proteins are potentially involved in each step of the infectious process and their contribution to viral replication was tested by RNA interference. Validation of the relevance of these host cell proteins for the viral replication cycle revealed that 7 of the 79 NS1 and/or NS2-interacting proteins positively or negatively controlled virus replication. One of the main factors targeted by NS1 of all virus strains was double-stranded RNA binding domain protein family. In particular, adenosine deaminase acting on RNA 1 (ADAR1 appeared as a pro-viral host factor whose expression is necessary for optimal viral protein synthesis and replication. Surprisingly, ADAR1 also appeared as a pro-viral host factor for dengue virus replication and directly interacted with the viral NS3 protein. ADAR1 editing activity was enhanced by both viruses through dengue virus NS3 and influenza virus NS1 proteins, suggesting a similar virus-host co-evolution.

Nuclear retention of multiply spliced HIV-1 RNA in resting CD4+ T cells.

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Kara G Lassen

2006-07-01

Full Text Available HIV-1 latency in resting CD4+ T cells represents a major barrier to virus eradication in patients on highly active antiretroviral therapy (HAART. We describe here a novel post-transcriptional block in HIV-1 gene expression in resting CD4+ T cells from patients on HAART. This block involves the aberrant localization of multiply spliced (MS HIV-1 RNAs encoding the critical positive regulators Tat and Rev. Although these RNAs had no previously described export defect, we show that they exhibit strict nuclear localization in resting CD4+ T cells from patients on HAART. Overexpression of the transcriptional activator Tat from non-HIV vectors allowed virus production in these cells. Thus, the nuclear retention of MS HIV-1 RNA interrupts a positive feedback loop and contributes to the non-productive nature of infection of resting CD4+ T cells. To define the mechanism of nuclear retention, proteomic analysis was used to identify proteins that bind MS HIV-1 RNA. Polypyrimidine tract binding protein (PTB was identified as an HIV-1 RNA-binding protein differentially expressed in resting and activated CD4+ T cells. Overexpression of PTB in resting CD4+ T cells from patients on HAART allowed cytoplasmic accumulation of HIV-1 RNAs. PTB overexpression also induced virus production by resting CD4+ T cells. Virus culture experiments showed that overexpression of PTB in resting CD4+ T cells from patients on HAART allowed release of replication-competent virus, while preserving a resting cellular phenotype. Whether through effects on RNA export or another mechanism, the ability of PTB to reverse latency without inducing cellular activation is a result with therapeutic implications.

Viruses and the nucleolus: the fatal attraction.

Science.gov (United States)

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. Copyright © 2014 Elsevier B.V. All rights reserved.

Analyses of pea necrotic yellow dwarf virus-encoded proteins.

Science.gov (United States)

Krenz, Björn; Schießl, Ingrid; Greiner, Eva; Krapp, Susanna

2017-06-01

Pea necrotic yellow dwarf virus (PNYDV) is a multipartite, circular, single-stranded DNA plant virus. PNYDV encodes eight proteins and the function of three of which remains unknown-U1, U2, and U4. PNYDV proteins cellular localization was analyzed by GFP tagging and bimolecular fluorescence complementation (BiFC) studies. The interactions of all eight PNYDV proteins were tested pairwise in planta (36 combinations in total). Seven interactions were identified and two (M-Rep with CP and MP with U4) were characterized further. MP and U4 complexes appeared as vesicle-like spots and were localized at the nuclear envelope and cell periphery. These vesicle-like spots were associated with the endoplasmatic reticulum. In addition, a nuclear localization signal (NLS) was mapped for U1, and a mutated U1 with NLS disrupted localized at plasmodesmata and therefore might also have a role in movement. Taken together, this study provides evidence for previously undescribed nanovirus protein-protein interactions and their cellular localization with novel findings not only for those proteins with unknown function, but also for characterized proteins such as the CP.

Nairobi sheep disease virus/Ganjam virus.

Science.gov (United States)

M D, Baron; B, Holzer

2015-08-01

Nairobi sheep disease virus (NSDV) is a tick-borne virus which causes a severe disease in sheep and goats, and has been responsible for several outbreaks of disease in East Africa. The virus is also found in the Indian subcontinent, where it is known as Ganjam virus. The virus only spreads through the feeding of competent infected ticks, and is therefore limited in its geographic distribution by the distribution of those ticks, Rhipicephalus appendiculata in Africa and Haemaphysalis intermedia in India. Animals bred in endemic areas do not normally develop disease, and the impact is therefore primarily on animals being moved for trade or breeding purposes. The disease caused by NSDV has similarities to several other ruminant diseases, and laboratory diagnosis is necessary for confirmation. There are published methods for diagnosis based on polymerase chain reaction, for virus growth in cell culture and for other simple diagnostic tests, though none has been commercialised. There is no established vaccine against NSDV, although cell-culture attenuated strains have been developed which show promise and could be put into field trials if it were deemed necessary. The virus is closely related to Crimean-Congo haemorrhagic fever virus, and studies on NSDV may therefore be useful in understanding this important human pathogen.

Interleukin-33 is expressed in the lesional epidermis in herpes virus infection but not in verruca vulgaris.

Science.gov (United States)

Jin, Meijuan; Komine, Mayumi; Tsuda, Hidetoshi; Oshio, Tomoyuki; Ohtsuki, Mamitaro

2018-04-25

Interleukin (IL)-33 is released on cell injury and activates the immune reaction. IL-33 is involved in antiviral reaction in herpes virus infection, but the source that secretes IL-33 has not been identified. We speculate that keratinocytes injured in herpes virus infection secrete IL-33. In order to detect IL-33 in the lesional epidermis of patients with herpes virus infection, we immunostained several cutaneous herpes virus infection samples with an anti-IL-33 antibody, and compared them with cutaneous human papilloma virus (HPV) infection samples. We observed strong nuclear and mild cytoplasmic staining in epidermal keratinocytes of the lesional skin samples with herpes simplex virus and varicella zoster virus infections. However, staining was not observed in the epidermis of verruca vulgaris (VV) samples. We assumed that the strong immune reaction to herpes virus infection may depend on strong IL-33 expression in the epidermis, while very weak immune reaction in samples from patients with VV may be due to low or no expression of IL-33 in the lesional epidermis. © 2018 Japanese Dermatological Association.

Residues R199H200 of prototype foamy virus transactivator Bel1 contribute to its binding with LTR and IP promoters but not its nuclear localization

International Nuclear Information System (INIS)

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

2014-01-01

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 199 H 200 and R 221 R 222 R 223 that likely function together as a bipartite nuclear localization signal. In this study, we report that mutating R 221 R 222 R 223 , but not R 199 H 200 , relocates Bel1 from the nucleus to the cytoplasm, suggesting an essential role for R 221 R 222 R 223 in the nuclear localization of Bel1. Although not affecting the nuclear localization of Bel1, mutating R 199 H 200 disables Bel1 from transactivating PFV promoters. Results of EMSA reveal that the R 199 H 200 residues are vital for the binding of Bel1 to viral promoter DNA. Moreover, mutating R 199 H 200 in Bel1 impairs PFV replication to a much greater extent than mutating R 221 R 222 R 223 . Collectively, our findings suggest that R 199 H 200 directly participate in Bel1 binding to viral promoter DNA and are indispensible for Bel1 transactivation activity. - Highlights: • The R 221 R 222 R 223 residues are essential for the nuclear localization of Bel1. • Although not affecting the nuclear localization of Bel1, mutating R 199 H 200 disables Bel1 from transactivating PFV promoters. • The R 199 H 200 residues directly participate in Bel1 binding to viral promoter DNA. • Mutating R 199 H 200 in Bel1 impairs PFV replication to a much greater extent than mutating R 221 R 222 R 223

Engineering of avian influenza virus detection system in a patient's body

International Nuclear Information System (INIS)

Budi Santoso; Romadhon; Sukandar, Istofa

2010-01-01

The avian influenza virus detection equipment in a patient's body has been made. Currently, detection of avian influenza virus carried out by expensive laboratory equipment's, so only certain hospitals can perform this detection. This developing equipment is expected to be cheaper than existing equipment and the diagnosis can be known immediately. The sensing device is made using the principle of nuclear radiation detection. Radiation comes from a drunk labelled tamiflu (oseltamivir) which is drunk to the patient. Tamiflu is a drug to catch H5N1 viruses in a patient's body. A labelled tamiflu is tamiflu which is labelled by 1-131 radioisotopes. The presence of virus in the body is proportional to the amount of radiation captured by the detector. The equipment is composed of a Geiger-Mueller (GM) pancake detector type, a signal processor, a counter, and a data processor (computer). The GM detector converts the radiation that comes into electrical signals. Electrical signal is then converted into TTL level pulses by the signal processor. Pulse counting results are processed by data processor. The total count is proportional to the amount of virus captured by labelled tamiflu. The measurement threshold can be set by medical officer through software. At a certain threshold can be inferred identified patients infected with avian influenza virus. If the measurement below the threshold means that the patient is still within safe limits. This equipment is expected to create avian influenza virus detection system that cheaply and quickly so that more and more hospitals are using to detect the avian influenza virus. (author)

Nuclear localization signal regulates porcine circovirus type 2 capsid protein nuclear export through phosphorylation.

Science.gov (United States)

Hou, Qiang; Hou, Shaohua; Chen, Qing; Jia, Hong; Xin, Ting; Jiang, Yitong; Guo, Xiaoyu; Zhu, Hongfei

2018-02-15

The open reading frame 2 (ORF2) of Porcine circovirus type 2 (PCV2) encodes the major Capsid (Cap) protein, which self-assembles into virus-like particle (VLP) of similar morphology to the PCV2 virion and accumulates in the nucleus through the N-terminal arginine-rich nuclear localization signal (NLS). In this study, PCV2 Cap protein and its derivates were expressed via the baculovirus expression system, and the cellular localization of the recombinant proteins were investigated using anti-Cap mAb by imaging flow cytometry. Analysis of subcellular localization of Cap protein and its variants demonstrated that NLS mediated Cap protein nuclear export as well as nuclear import, and a phosphorylation site (S17) was identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in the NLS domain to regulate Cap protein nuclear export. Phosphorylation of NLS regulating the PCV2 Cap protein nuclear export was also demonstrated in PK15 cells by fluorescence microscopy. Moreover, the influence of Rep and Rep' protein on Cap protein subcellular localization was investigated in PK15 cells. Phosphorylation of NLS regulating Cap protein nuclear export provides more detailed knowledge of the PCV2 viral life cycle. Copyright © 2018 Elsevier B.V. All rights reserved.

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

International Nuclear Information System (INIS)

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

2013-01-01

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

The nuclear inclusion a (NIa protease of turnip mosaic virus (TuMV cleaves amyloid-β.

Directory of Open Access Journals (Sweden)

Hye-Eun Han

Full Text Available BACKGROUND: The nuclear inclusion a (NIa protease of turnip mosaic virus (TuMV is responsible for the processing of the viral polyprotein into functional proteins. NIa was previously shown to possess a relatively strict substrate specificity with a preference for Val-Xaa-His-Gln↓, with the scissile bond located after Gln. The presence of the same consensus sequence, Val(12-His-His-Gln(15, near the presumptive α-secretase cleavage site of the amyloid-β (Aβ peptide led us to hypothesize that NIa could possess activity against Aβ. METHODOLOGY/PRINCIPAL FINDINGS: Western blotting results showed that oligomeric as well as monomeric forms of Aβ can be degraded by NIa in vitro. The specific cleavage of Aβ was further confirmed by mass spectrometry analysis. NIa was shown to exist predominantly in the cytoplasm as observed by immunofluorescence microscopy. The overexpression of NIa in B103 neuroblastoma cells resulted in a significant reduction in cell death caused by both intracellularly generated and exogenously added Aβ. Moreover, lentiviral-mediated expression of NIa in APP(sw/PS1 transgenic mice significantly reduced the levels of Aβ and plaques in the brain. CONCLUSIONS/SIGNIFICANCE: These results indicate that the degradation of Aβ in the cytoplasm could be a novel strategy to control the levels of Aβ, plaque formation, and the associated cell death.

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

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Liu, Cheng-Der; Cheng, Chi-Ping; Fang, Jia-Shih; Chen, Ling-Chih [Department of Life Sciences, Tzu-Chi University, 701 Chung-Yang Rd. Sec 3, Hualien 97004, Taiwan (China); Zhao, Bo; Kieff, Elliott [Department of Medicine and Microbiology and Molecular Genetics, Channing Laboratory, Brigham and Women’s Hospital and Harvard Medical School, 181 Longwood Ave., Boston 02115, MA (United States); Peng, Chih-Wen, E-mail: pengcw@mail.tcu.edu.tw [Department of Life Sciences, Tzu-Chi University, 701 Chung-Yang Rd. Sec 3, Hualien 97004, Taiwan (China)

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.

Molecular diagnosis for the silk worm Bombyx Mori L. Viral and bacterial diseases in the irradiated and non-irradiated individuals

International Nuclear Information System (INIS)

Abulyazid, I.; Elshafei, A.; El-Said, E.; Mousa, S.; Taha, R.

2007-01-01

Genetic maps for the Bombyx Mori infectious flacherrie virus (BmlFV) causing flacherrie (Fl) disease and nuclear polyhedrosis virus (BmNPV) causing grasserie (Gr) disease were built up in an attempt to diagnose diseases early in young larval stages. For the non-irradiated and irradiated viral RNA of IFV, no amplification was obtained by using RT-PCR and RAPD-PCR techniques. HcoRI, EcoRV, BamHI, Hind III and BamHI restriction enzymes were used to digest the non-irradiated and irradiated viral DNA of BmNPV. It was found that, the two viral DNA samples were genetically different; the similarity indexes were 0.14, 0, 0, 0.18 and 0.15, respectively. At the biochemical level, native protein electrophoresis showed 4 and 3 new proteins in non-irradiated and irradiated Fl diseased larvae, respectively, while Gr diseased larvae showed 1 and 3 new protein types. The similarity index (S.I) between all the tested samples was not exceeded 44%. For lipoprotein pattern, 2 and 3 new lipoprotein types were appeared due to Fl disease in the non-irradiated and irradiated haemolymph samples, respectively, while Gr disease showed 3 new lipoproteins in the non-irradiated samples only. The highest S.I recorded was 56%. Glycoprotein pattern revealed 3 and 5 new glycoprotein types appeared due to Fl disease while Gr disease showed 4 and 6 new types in the non-irradiated and irradiated samples, respectively. The highest S.I was 77%. Fractionated protein with SDS revealed 2 common bands shared between the tested samples with R f values 0.28 and 0.71. Fl disease increased the number of protein bands with the appearance of 5 and 4 new proteins types. Gr disease reduced the total number of proteins with the appearance of 2 and 3 new types. The highest S.I was 59%. Both diseases and irradiation may be mutagenic through the epigenetic level in silkworm larvae leading to death. Thus, the results of the biochemical and genetic characterization of IFV and BmNPV enable us to conclude that the

The Lysine Residues within the Human Ribosomal Protein S17 Sequence Naturally Inserted into the Viral Nonstructural Protein of a Unique Strain of Hepatitis E Virus Are Important for Enhanced Virus Replication

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Kenney, Scott P.

2015-01-01

ABSTRACT Hepatitis E virus (HEV) is an important but extremely understudied human pathogen. Due largely to the lack of an efficient cell culture system for HEV, the molecular mechanisms of HEV replication and pathogenesis are poorly understood. Recently, a unique genotype 3 strain of HEV recovered from a chronically infected patient was adapted for growth in HepG2C3A human hepatoma cells. The adaptation of the Kernow C-1 P6 HEV to propagate in HepG2C3A cells selected for a rare virus recombinant that contains an insertion of a 171-nucleotide sequence encoding amino acids 21 to 76 of the human ribosomal protein S17 (RPS17) within the hypervariable region (HVR) of the HEV ORF1 protein. When the RPS17 insertion was placed into a strain of genotype 1 HEV which infects only humans, it expanded the host range of the virus, allowing it to infect cell lines from multiple animal species, including cow, dog, cat, chicken, and hamster. In this study, we utilized forward and reverse genetics to attempt to define which aspects of the RPS17 insertion allow for the ability of the Kernow C-1 P6 HEV to adapt in cell culture and allow for expanded host tropism. We demonstrate that the RPS17 sequence insertion in HEV bestows novel nuclear/nucleolar trafficking capabilities to the ORF1 protein of Kernow P6 HEV and that lysine residues within the RPS17 insertion, but not nuclear localization of the ORF1 protein, correlate with the enhanced replication of the HEV Kernow C-1 P6 strain. The results from this study have important implications for understanding the mechanism of cross-species infection and replication of HEV. IMPORTANCE HEV is an important pathogen worldwide. The virus causes high mortality (up to 30%) in pregnant women and has been recognized to cause chronic hepatitis in immunocompromised populations. The life cycle of HEV has been understudied due to a lack of sufficient cell culture systems in which to propagate the virus. Recently, insertions and rearrangements of the

Ebola virus infection inversely correlates with the overall expression levels of promyelocytic leukaemia (PML protein in cultured cells

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Szekely Laszlo

2003-04-01

Full Text Available Abstract Background Ebola virus causes severe, often fatal hemorrhagic fever in humans. The mechanism of escape from cellular anti-viral mechanisms is not yet fully understood. The promyelocytic leukaemia (PML associated nuclear body is part of the interferon inducible cellular defense system. Several RNA viruses have been found to interfere with the anti-viral function of the PML body. The possible interaction between Ebola virus and the PML bodies has not yet been explored. Results We found that two cell lines, Vero E6 and MCF7, support virus production at high and low levels respectively. The expression of viral proteins was visualized and quantified using high resolution immunofluorescence microscopy. Ebola encoded NP and VP35 accumulated in cytoplasmic inclusion bodies whereas VP40 was mainly membrane associated but it was also present diffusely in the cytoplasm as well as in the euchromatic areas of the nucleus. The anti-VP40 antibody also allowed the detection of extracellular virions. Interferon-alpha treatment decreased the production of all three viral proteins and delayed the development of cytopathic effects in both cell lines. Virus infection and interferon-alpha treatment induced high levels of PML protein expression in MCF7 but much less in Vero E6 cells. No disruption of PML bodies, a common phenomenon induced by a variety of different viruses, was observed. Conclusion We have established a simple fixation and immunofluorescence staining procedure that allows specific co-detection and precise sub-cellular localization of the PML nuclear bodies and the Ebola virus encoded proteins NP, VP35 and VP40 in formaldehyde treated cells. Interferon-alpha treatment delays virus production in vitro. Intact PML bodies may play an anti-viral role in Ebola infected cells.

Parvovirus induced alterations in nuclear architecture and dynamics.

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Teemu O Ihalainen

2009-06-01

Full Text Available The nucleus of interphase eukaryotic cell is a highly compartmentalized structure containing the three-dimensional network of chromatin and numerous proteinaceous subcompartments. DNA viruses induce profound changes in the intranuclear structures of their host cells. We are applying a combination of confocal imaging including photobleaching microscopy and computational methods to analyze the modifications of nuclear architecture and dynamics in parvovirus infected cells. Upon canine parvovirus infection, expansion of the viral replication compartment is accompanied by chromatin marginalization to the vicinity of the nuclear membrane. Dextran microinjection and fluorescence recovery after photobleaching (FRAP studies revealed the homogeneity of this compartment. Markedly, in spite of increase in viral DNA content of the nucleus, a significant increase in the protein mobility was observed in infected compared to non-infected cells. Moreover, analysis of the dynamics of photoactivable capsid protein demonstrated rapid intranuclear dynamics of viral capsids. Finally, quantitative FRAP and cellular modelling were used to determine the duration of viral genome replication. Altogether, our findings indicate that parvoviruses modify the nuclear structure and dynamics extensively. Intranuclear crowding of viral components leads to enlargement of the interchromosomal domain and to chromatin marginalization via depletion attraction. In conclusion, parvoviruses provide a useful model system for understanding the mechanisms of virus-induced intranuclear modifications.

DIAGNOSTICS OF VIRUS PHYTOPATHOGENS FRUIT TREE PLUM POX VIRUS, PRUNUS NECROTIC RINGSPOT VIRUS AND PRUNUS DWARF VIRUS BY BIOLOGICAL AND MOLECULAR DIAGNOSTICS

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Július Rozák

2013-02-01

Full Text Available The aim of this study was to determine the incidence of viral phytopathogen Plum pox virus, Prunus necrotic ringspot virus and Prunus dwarf virus in selected localities of Slovakia and diagnose them using a molecular and biological methods. Forty samples of fruit trees of the genus Prunus, twenty samples from intensive plantings and twenty samples from wild subject were analysed. Biological diagnostic by using biological indicators Prunus persica cv. GF 305, Prunus serrulata cv. Schirofugen and molecular diagnostic by mRT-PCR were applied. Five samples with Plum pox virus were infected. The two samples positive for Prunus necrotic ringspot virus and one sample for Prunus dwarf virus were confirmed. The two samples were found to be infected with two viruses Prunus necrotic ringspot virus and Prunus dwarf virus. This work focuses on two techniques, their application to the diagnosis of stone fruit viruses and their routinely used for sanitary and certification programmes.

Antiviral Efficacy of Verdinexor In Vivo in Two Animal Models of Influenza A Virus Infection

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Perwitasari, Olivia; Johnson, Scott; Yan, Xiuzhen; Register, Emery; Crabtree, Jackelyn; Gabbard, Jon; Howerth, Elizabeth; Shacham, Sharon; Carlson, Robert; Tamir, Sharon; Tripp, Ralph A.

2016-01-01

Influenza A virus (IAV) causes seasonal epidemics of respiratory illness that can cause mild to severe illness and potentially death. Antiviral drugs are an important countermeasure against IAV; however, drug resistance has developed, thus new therapeutic approaches are being sought. Previously, we demonstrated the antiviral activity of a novel nuclear export inhibitor drug, verdinexor, to reduce influenza replication in vitro and pulmonary virus burden in mice. In this study, in vivo efficacy of verdinexor was further evaluated in two animal models or influenza virus infection, mice and ferrets. In mice, verdinexor was efficacious to limit virus shedding, reduce pulmonary pro-inflammatory cytokine expression, and moderate leukocyte infiltration into the bronchoalveolar space. Similarly, verdinexor-treated ferrets had reduced lung pathology, virus burden, and inflammatory cytokine expression in the nasal wash exudate. These findings support the anti-viral efficacy of verdinexor, and warrant its development as a novel antiviral therapeutic for influenza infection. PMID:27893810

Antiviral Efficacy of Verdinexor In Vivo in Two Animal Models of Influenza A Virus Infection.

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Olivia Perwitasari

Full Text Available Influenza A virus (IAV causes seasonal epidemics of respiratory illness that can cause mild to severe illness and potentially death. Antiviral drugs are an important countermeasure against IAV; however, drug resistance has developed, thus new therapeutic approaches are being sought. Previously, we demonstrated the antiviral activity of a novel nuclear export inhibitor drug, verdinexor, to reduce influenza replication in vitro and pulmonary virus burden in mice. In this study, in vivo efficacy of verdinexor was further evaluated in two animal models or influenza virus infection, mice and ferrets. In mice, verdinexor was efficacious to limit virus shedding, reduce pulmonary pro-inflammatory cytokine expression, and moderate leukocyte infiltration into the bronchoalveolar space. Similarly, verdinexor-treated ferrets had reduced lung pathology, virus burden, and inflammatory cytokine expression in the nasal wash exudate. These findings support the anti-viral efficacy of verdinexor, and warrant its development as a novel antiviral therapeutic for influenza infection.

Anti-sense expression of a metallopeptidase gene enhances nuclear entry of HBV-DNA

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Yeh, C.-T.; Lai, H.-Y.; Chu, S.-P.; Tseng, I-Chu

2004-01-01

Although several putative hepatitis B virus (HBV) receptors have been identified, none of them is capable of initiating HBV replication in a non-permissive human cell line. Using an Epstein-Barr virus-based extrachromosomal replication system, we have screened through a human liver cDNA library and successfully identified a clone capable of facilitating nuclear transport of HBV-DNA during the early phase of HBV infection. This clone contained a cDNA encoding a metallopeptidase-like protein in anti-sense orientation. Pretreatment of naive HepG2 cells with 1,10-phenanthroline, an inhibitor for liver metallopeptidases, led to nuclear entry of HBV-DNA after HBV infection. However, cccDNA was still undetectable in the nuclei, indicating other cellular factors required to complete the replication cycle were still missing. Our present data suggest that in the initial stage of HBV infection, liver metallopeptidase constitutes a barrier for effective nuclear entry of HBV genomic DNA. Attenuation of metallopeptidase activity may facilitate HBV infection

Interaction of influenza virus proteins with nucleosomes

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Garcia-Robles, Inmaculada; Akarsu, Hatice; Mueller, Christoph W.; Ruigrok, Rob W.H.; Baudin, Florence

2005-01-01

During influenza virus infection, transcription and replication of the viral RNA take place in the cell nucleus. Directly after entry in the nucleus the viral ribonucleoproteins (RNPs, the viral subunits containing vRNA, nucleoprotein and the viral polymerase) are tightly associated with the nuclear matrix. Here, we have analysed the binding of RNPs, M1 and NS2/NEP proteins to purified nucleosomes, reconstituted histone octamers and purified single histones. RNPs and M1 both bind to the chromatin components but at two different sites, RNP to the histone tails and M1 to the globular domain of the histone octamer. NS2/NEP did not bind to nucleosomes at all. The possible consequences of these findings for nuclear release of newly made RNPs and for other processes during the infection cycle are discussed

Kaposi sarcoma herpes virus latency associated nuclear antigen protein release the G2/M cell cycle blocks by modulating ATM/ATR mediated checkpoint pathway.

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Amit Kumar

Full Text Available The Kaposi's sarcoma-associated herpesvirus infects the human population and maintains latency stage of viral life cycle in a variety of cell types including cells of epithelial, mesenchymal and endothelial origin. The establishment of latent infection by KSHV requires the expression of an unique repertoire of genes among which latency associated nuclear antigen (LANA plays a critical role in the replication of the viral genome. LANA regulates the transcription of a number of viral and cellular genes essential for the survival of the virus in the host cell. The present study demonstrates the disruption of the host G2/M cell cycle checkpoint regulation as an associated function of LANA. DNA profile of LANA expressing human B-cells demonstrated the ability of this nuclear antigen in relieving the drug (Nocodazole induced G2/M checkpoint arrest. Caffeine suppressed nocodazole induced G2/M arrest indicating involvement of the ATM/ATR. Notably, we have also shown the direct interaction of LANA with Chk2, the ATM/ATR signalling effector and is responsible for the release of the G2/M cell cycle block.

Oncogenic Viruses and Breast Cancer: Mouse Mammary Tumor Virus (MMTV, Bovine Leukemia Virus (BLV, Human Papilloma Virus (HPV, and Epstein–Barr Virus (EBV

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James S. Lawson

2018-01-01

Full Text Available BackgroundAlthough the risk factors for breast cancer are well established, namely female gender, early menarche and late menopause plus the protective influence of early pregnancy, the underlying causes of breast cancer remain unknown. The development of substantial recent evidence indicates that a handful of viruses may have a role in breast cancer. These viruses are mouse mammary tumor virus (MMTV, bovine leukemia virus (BLV, human papilloma viruses (HPVs, and Epstein–Barr virus (EBV-also known as human herpes virus type 4. Each of these viruses has documented oncogenic potential. The aim of this review is to inform the scientific and general community about this recent evidence.The evidenceMMTV and human breast cancer—the evidence is detailed and comprehensive but cannot be regarded as conclusive. BLV and human breast cancer—the evidence is limited. However, in view of the emerging information about BLV in human breast cancer, it is prudent to encourage the elimination of BLV in cattle, particularly in the dairy industry. HPVs and breast cancer—the evidence is substantial but not conclusive. The availability of effective preventive vaccines is a major advantage and their use should be encouraged. EBV and breast cancer—the evidence is also substantial but not conclusive. Currently, there are no practical means of either prevention or treatment. Although there is evidence of genetic predisposition, and cancer in general is a culmination of events, there is no evidence that inherited genetic traits are causal.ConclusionThe influence of oncogenic viruses is currently the major plausible hypothesis for a direct cause of human breast cancer.

Oncogenic Viruses and Breast Cancer: Mouse Mammary Tumor Virus (MMTV), Bovine Leukemia Virus (BLV), Human Papilloma Virus (HPV), and Epstein-Barr Virus (EBV).

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Lawson, James S; Salmons, Brian; Glenn, Wendy K

2018-01-01

Although the risk factors for breast cancer are well established, namely female gender, early menarche and late menopause plus the protective influence of early pregnancy, the underlying causes of breast cancer remain unknown. The development of substantial recent evidence indicates that a handful of viruses may have a role in breast cancer. These viruses are mouse mammary tumor virus (MMTV), bovine leukemia virus (BLV), human papilloma viruses (HPVs), and Epstein-Barr virus (EBV-also known as human herpes virus type 4). Each of these viruses has documented oncogenic potential. The aim of this review is to inform the scientific and general community about this recent evidence. MMTV and human breast cancer-the evidence is detailed and comprehensive but cannot be regarded as conclusive. BLV and human breast cancer-the evidence is limited. However, in view of the emerging information about BLV in human breast cancer, it is prudent to encourage the elimination of BLV in cattle, particularly in the dairy industry. HPVs and breast cancer-the evidence is substantial but not conclusive. The availability of effective preventive vaccines is a major advantage and their use should be encouraged. EBV and breast cancer-the evidence is also substantial but not conclusive. Currently, there are no practical means of either prevention or treatment. Although there is evidence of genetic predisposition, and cancer in general is a culmination of events, there is no evidence that inherited genetic traits are causal. The influence of oncogenic viruses is currently the major plausible hypothesis for a direct cause of human breast cancer.

A small molecule for a big transformation: Topical application of a 20-nucleotide-long antisense fragment of the DIAP-2 gene inhibits the development of Drosophila melanogaster female imagos

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Nyadar Palmah M.

2018-01-01

Full Text Available Several genes have been identified to play important roles associated with sex selection in Drosophila melanogaster. An essential part is attributed to the sex-lethal gene that depends on the expression of the X:A (number of chromosomes to autosomes ratio signal controlling both sex selection and dosage compensation processes in D. melanogaster. Interestingly, for sex selection in D. melanogaster there are no documented data addressing the role of the inhibitor of apoptosis (IAP genes and their signaling influence on this biological process. In this study, we found that topical application of a 20-nucleotide-long antisense DNA fragment (oligoDIAP-2 from the death-associated inhibitor of apoptosis (DIAP-2 gene interferes with D. melanogaster development and significantly decreases the number of female imagos and their biomass. We show that the applied antisense oligoDIAP-2 fragment downregulates the target DIAP-2 gene whose normal concentration is necessary for the development of female D. melanogaster. These data correspond to the results on downregulation of the target host IAP-Z gene of Lymantria dispar L. female imagos after topical treatment with an 18-nucleotide-long antisense DNA fragment from the L. dispar multicapsid nuclear polyhedrosis virus IAP-3 gene at the larval stage. The observed novel phenomenon linking the downregulation of insect IAP genes and the low rate of female imago development could have practical application, especially in insect pest control and molecular pathology.

Serum-dependent expression of promyelocytic leukemia protein suppresses propagation of influenza virus

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Iki, Shigeo; Yokota, Shin-ichi; Okabayashi, Tamaki; Yokosawa, Noriko; Nagata, Kyosuke; Fujii, Nobuhiro

2005-01-01

The rate of propagation of influenza virus in human adenocarcinoma Caco-2 cells was found to negatively correlate with the concentration of fetal bovine serum (FBS) in the culture medium. Virus replicated more rapidly at lower FBS concentrations (0 or 2%) than at higher concentrations (10 or 20%) during an early stage of infection. Basal and interferon (IFN)-induced levels of typical IFN-inducible anti-viral proteins, such as 2',5'-oligoadenylate synthetase, dsRNA-activated protein kinase and MxA, were unaffected by variation in FBS concentrations. But promyelocytic leukemia protein (PML) was expressed in a serum-dependent manner. In particular, the 65 to 70 kDa isoform of PML was markedly upregulated following the addition of serum. In contrast, other isoforms were induced by IFN treatment, and weakly induced by FBS concentrations. Immunofluorescence microscopy indicated that PML was mainly formed nuclear bodies in Caco-2 cells at various FBS concentrations, and the levels of the PML-nuclear bodies were upregulated by FBS. Overexpression of PML isoform consisting of 560 or 633 amino acid residues by transfection of expression plasmid results in significantly delayed viral replication rate in Caco-2 cells. On the other hand, downregulation of PML expression by RNAi enhanced viral replication. These results indicate that PML isoforms which are expressed in a serum-dependent manner suppress the propagation of influenza virus at an early stage of infection

Conserved cell cycle regulatory properties within the amino terminal domain of the Epstein-Barr virus nuclear antigen 3C

International Nuclear Information System (INIS)

Sharma, Nikhil; Knight, Jason S.; Robertson, Erle S.

2006-01-01

The gammaherpesviruses Rhesus lymphocryptovirus (LCV) and Epstein-Barr virus (EBV) are closely related phylogenetically. Rhesus LCV efficiently immortalizes Rhesus B cells in vitro. However, despite a high degree of conservation between the Rhesus LCV and EBV genomes, Rhesus LCV fails to immortalize human B cells in vitro. This species restriction may, at least in part, be linked to the EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs), known to be essential for B cell transformation. We compared specific properties of EBNA3C, a well-characterized and essential EBV protein, with its Rhesus counterpart to determine whether EBNA3C phenotypes which contribute to cell cycle regulation are conserved in the Rhesus LCV. We show that both EBNA3C and Rhesus EBNA3C bind to a conserved region of mammalian cyclins, regulate pRb stability, and modulate SCF Skp2 -dependent ubiquitination. These results suggest that Rhesus LCV restriction from human B cell immortalization is independent of the conserved cell cycle regulatory functions of the EBNA3C protein

Evolutionary relationship of alfalfa mosaic virus with cucumber mosaic virus and brome mosaic virus

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Savithri, HS; Murthy, MRN

1983-01-01

The amino acid sequences of the non-structural protein (molecular weight 35,000; 3a protein) from three plant viruses - cucumber mosaic, brome mosaic and alfalfa mosaic have been systematically compared using the partial genomic sequences for these three viruses already available. The 3a protein of cucumber mosaic virus has an amino acid sequence homology of 33.7% with the corresponding protein of brome mosaic virus. A similar protein from alfalfa mosaic virus has a homology of 18.2% and 14.2...

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

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Dylan eFlather

2015-06-01

Full Text Available 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.

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

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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.

Infection cycles of large DNA viruses: emerging themes and underlying questions.

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Mutsafi, Yael; Fridmann-Sirkis, Yael; Milrot, Elad; Hevroni, Liron; Minsky, Abraham

2014-10-01

The discovery of giant DNA viruses and the recent realization that such viruses are diverse and abundant blurred the distinction between viruses and cells. These findings elicited lively debates on the nature and origin of viruses as well as on their potential roles in the evolution of cells. The following essay is, however, concerned with new insights into fundamental structural and physical aspects of viral replication that were derived from studies conducted on large DNA viruses. Specifically, the entirely cytoplasmic replication cycles of Mimivirus and Vaccinia are discussed in light of the highly limited trafficking of large macromolecules in the crowded cytoplasm of cells. The extensive spatiotemporal order revealed by cytoplasmic viral factories is described and contended to play an important role in promoting the efficiency of these 'nuclear-like' organelles. Generation of single-layered internal membrane sheets in Mimivirus and Vaccinia, which proceeds through a novel membrane biogenesis mechanism that enables continuous supply of lipids, is highlighted as an intriguing case study of self-assembly. Mimivirus genome encapsidation was shown to occur through a portal different from the 'stargate' portal that is used for genome release. Such a 'division of labor' is proposed to enhance the efficacy of translocation processes of very large viral genomes. Finally, open questions concerning the infection cycles of giant viruses to which future studies are likely to provide novel and exciting answers are discussed. Copyright © 2014 Elsevier Inc. All rights reserved.

Relationship between RNA polymerase II and efficiency of vaccinia virus replication

International Nuclear Information System (INIS)

Wilton, S.; Dales, S.

1989-01-01

It is clear from previous studies that host transcriptase or RNA polymerase II (pol II) has a role in poxvirus replication. To elucidate the participation of this enzyme further, in this study the authors examined several parameters related to pol II during the cycle of vaccinia virus infection in L-strain fibroblasts, HeLa cells, and L 6 H 9 rat myoblasts. Nucleocytoplasmic transposition of pol II into virus factories and virions was assessed by immunofluorescence and immunoblotting by using anti-pol II immunoglobulin G. RNA polymerase activities were compared in nuclear extracts containing cured enzyme preparations. Rates of translation into cellular or viral polypeptides were ascertained by labeling with [ 35 S]methionine. In L and HeLa cells, which produced vaccinia virus more abundantly, the rate of RNA polymerase and translation in controls and following infection were higher than in myoblasts. The data on synthesis and virus formation could be correlated with observations on transmigration of pol II, which was more efficient and complete in L and HeLa cells. The stimulus for pol II to leave the nucleus required the expression of both early and late viral functions. On the basis of current and past information, the authors suggest that mobilization of pol II depends on the efficiency of vaccinia virus replication and furthermore that control over vaccinia virus production by the host is related to the content or availability (or both) of pol II in different cell types

Immunofluorescent staining of nuclear antigen in lymphoid cells transformed by Herpesvirus papio (HVP).

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Schmitz, H

1981-01-01

An improved fixation method for antigen detection in lymphoblastoid cells is described. Herpesvirus papio nuclear antigen (HUPNA) could be stained in several transformed lymphoid cell lines by anti-complement immunofluorescence (ACIF). Antibody to HUPNA was detected in many human sera containing antibodies to Epstein-Barr virus capsid and nuclear antigen (EBNA). Rheumatoid arthritis sera showed a high incidence of both anti-EBNA and anti-HUPNA antibodies.

Comparative analysis of chrysanthemum transcriptome in response to three RNA viruses: Cucumber mosaic virus, Tomato spotted wilt virus and Potato virus X.

Science.gov (United States)

Choi, Hoseong; Jo, Yeonhwa; Lian, Sen; Jo, Kyoung-Min; Chu, Hyosub; Yoon, Ju-Yeon; Choi, Seung-Kook; Kim, Kook-Hyung; Cho, Won Kyong

2015-06-01

The chrysanthemum is one of popular flowers in the world and a host for several viruses. So far, molecular interaction studies between the chrysanthemum and viruses are limited. In this study, we carried out a transcriptome analysis of chrysanthemum in response to three different viruses including Cucumber mosaic virus (CMV), Tomato spotted wilt virus (TSWV) and Potato virus X (PVX). A chrysanthemum 135K microarray derived from expressed sequence tags was successfully applied for the expression profiles of the chrysanthemum at early stage of virus infection. Finally, we identified a total of 125, 70 and 124 differentially expressed genes (DEGs) for CMV, TSWV and PVX, respectively. Many DEGs were virus specific; however, 33 DEGs were commonly regulated by three viruses. Gene ontology (GO) enrichment analysis identified a total of 132 GO terms, and of them, six GO terms related stress response and MCM complex were commonly identified for three viruses. Several genes functioning in stress response such as chitin response and ethylene mediated signaling pathway were up-regulated indicating their involvement in establishment of host immune system. In particular, TSWV infection significantly down-regulated genes related to DNA metabolic process including DNA replication, chromatin organization, histone modification and cytokinesis, and they are mostly targeted to nucleosome and MCM complex. Taken together, our comparative transcriptome analysis revealed several genes related to hormone mediated viral stress response and DNA modification. The identified chrysanthemums genes could be good candidates for further functional study associated with resistant to various plant viruses.

A Heterogeneous Nuclear Ribonucleoprotein A/B-Related Protein Binds to Single-Stranded DNA near the 5′ End or within the Genome of Feline Parvovirus and Can Modify Virus Replication

Science.gov (United States)

Wang, Dai; Parrish, Colin R.

1999-01-01

Phage display of cDNA clones prepared from feline cells was used to identify host cell proteins that bound to DNA-containing feline panleukopenia virus (FPV) capsids but not to empty capsids. One gene found in several clones encoded a heterogeneous nuclear ribonucleoprotein (hnRNP)-related protein (DBP40) that was very similar in sequence to the A/B-type hnRNP proteins. DBP40 bound specifically to oligonucleotides representing a sequence near the 5′ end of the genome which is exposed on the outside of the full capsid but did not bind most other terminal sequences. Adding purified DBP40 to an in vitro fill-in reaction using viral DNA as a template inhibited the production of the second strand after nucleotide (nt) 289 but prior to nt 469. DBP40 bound to various regions of the viral genome, including a region between nt 295 and 330 of the viral genome which has been associated with transcriptional attenuation of the parvovirus minute virus of mice, which is mediated by a stem-loop structure of the DNA and cellular proteins. Overexpression of the protein in feline cells from a plasmid vector made them largely resistant to FPV infection. Mutagenesis of the protein binding site within the 5′ end viral genome did not affect replication of the virus. PMID:10438866

ICP27-dependent resistance of herpes simplex virus type 1 to leptomycin B is associated with enhanced nuclear localization of ICP4 and ICP0

International Nuclear Information System (INIS)

Lengyel, Joy; Strain, Anna K.; Perkins, Keith D.; Rice, Stephen A.

2006-01-01

It was previously shown that herpes simplex virus type 1 (HSV-1) is sensitive to leptomycin B (LMB), an inhibitor of nuclear export factor CRM1, and that a single methionine to threonine change at residue 50 (M50T) of viral immediate-early (IE) protein ICP27 can confer LMB resistance. In this work, we show that deletion of residues 21-63 from ICP27 can also confer LMB resistance. We further show that neither the M50T mutation nor the presence of LMB affects the nuclear shuttling activity of ICP27, suggesting that another function of ICP27 determines LMB resistance. A possible clue to this function emerged when it was discovered that LMB treatment of HSV-1-infected cells dramatically enhances the cytoplasmic accumulation of two other IE proteins, ICP0 and ICP4. This effect is completely dependent on ICP27 and is reversed in cells infected with LMB-resistant mutants. Moreover, LMB-resistant mutations in ICP27 enhance the nuclear localization of ICP0 and ICP4 even in the absence of LMB, and this effect can be discerned in transfected cells. Thus, the same amino (N)-terminal region of ICP27 that determines sensitivity to LMB also enhances ICP27's previously documented ability to promote the cytoplasmic accumulation of ICP4 and ICP0. We speculate that ICP27's effects on ICP4 and ICP0 may contribute to HSV-1 LMB sensitivity

Drug-induced hypersensitivity syndrome associated with Epstein-Barr virus infection.

Science.gov (United States)

Descamps, V; Mahe, E; Houhou, N; Abramowitz, L; Rozenberg, F; Ranger-Rogez, S; Crickx, B

2003-05-01

Association of drug-induced hypersensitivity syndrome with viral infection is debated. Human herpesvirus 6 (HHV-6) reactivation has been the most frequently reported infection associated with this syndrome. However, a case of cytomegalovirus (CMV) infection was recently described associated with anticonvulsant-induced hypersensitivity syndrome. We report a case of severe allopurinol-induced hypersensitivity syndrome with pancreatitis associated with Epstein-Barr virus (EBV) infection. Active EBV infection was demonstrated in two consecutive serum samples by the presence of anti-EBV early antigen (EA) IgM antibodies and an increase in anti-EBV EA IgG antibodies, whereas no anti-EBV nuclear antigen IgG antibodies were detected. EBV DNA was detected by polymerase chain reaction (PCR) in peripheral blood mononuclear cells. Reactivation of HHV-6 was suggested only by the presence of anti-HHV-6 IgM antibodies, but HHV-6 DNA was not detected by PCR in the serum. Other viral investigations showed previous infection (CMV, rubella, measles, parvovirus B19), immunization after vaccination (hepatitis B virus), or absence of previous infection (hepatitis C virus, human immunodeficiency virus). We suggest that EBV infection may participate in some cases, as do the other herpesviruses HHV-6 or CMV, in the development of drug-induced hypersensitivity syndrome.

Infection cycles of large DNA viruses: Emerging themes and underlying questions

International Nuclear Information System (INIS)

Mutsafi, Yael; Fridmann-Sirkis, Yael; Milrot, Elad; Hevroni, Liron; Minsky, Abraham

2014-01-01

The discovery of giant DNA viruses and the recent realization that such viruses are diverse and abundant blurred the distinction between viruses and cells. These findings elicited lively debates on the nature and origin of viruses as well as on their potential roles in the evolution of cells. The following essay is, however, concerned with new insights into fundamental structural and physical aspects of viral replication that were derived from studies conducted on large DNA viruses. Specifically, the entirely cytoplasmic replication cycles of Mimivirus and Vaccinia are discussed in light of the highly limited trafficking of large macromolecules in the crowded cytoplasm of cells. The extensive spatiotemporal order revealed by cytoplasmic viral factories is described and contended to play an important role in promoting the efficiency of these ‘nuclear-like’ organelles. Generation of single-layered internal membrane sheets in Mimivirus and Vaccinia, which proceeds through a novel membrane biogenesis mechanism that enables continuous supply of lipids, is highlighted as an intriguing case study of self-assembly. Mimivirus genome encapsidation was shown to occur through a portal different from the ‘stargate’ portal that is used for genome release. Such a ‘division of labor’ is proposed to enhance the efficacy of translocation processes of very large viral genomes. Finally, open questions concerning the infection cycles of giant viruses to which future studies are likely to provide novel and exciting answers are discussed. - Highlights: • The discovery of giant DNA viruses blurs the distinction between viruses and cells. • Mimivirus and Vaccinia replicate exclusively in their host cytoplasm. • Mimivirus genome is delivered through a unique portal coined the Stargate. • Generation of Mimivirus internal membrane proceeds through a novel pathway

Infection cycles of large DNA viruses: Emerging themes and underlying questions

Energy Technology Data Exchange (ETDEWEB)

Mutsafi, Yael, E-mail: yael.mutsafi@weizmann.ac.il; Fridmann-Sirkis, Yael; Milrot, Elad; Hevroni, Liron; Minsky, Abraham, E-mail: avi.minsky@weizmann.ac.il

2014-10-15

The discovery of giant DNA viruses and the recent realization that such viruses are diverse and abundant blurred the distinction between viruses and cells. These findings elicited lively debates on the nature and origin of viruses as well as on their potential roles in the evolution of cells. The following essay is, however, concerned with new insights into fundamental structural and physical aspects of viral replication that were derived from studies conducted on large DNA viruses. Specifically, the entirely cytoplasmic replication cycles of Mimivirus and Vaccinia are discussed in light of the highly limited trafficking of large macromolecules in the crowded cytoplasm of cells. The extensive spatiotemporal order revealed by cytoplasmic viral factories is described and contended to play an important role in promoting the efficiency of these ‘nuclear-like’ organelles. Generation of single-layered internal membrane sheets in Mimivirus and Vaccinia, which proceeds through a novel membrane biogenesis mechanism that enables continuous supply of lipids, is highlighted as an intriguing case study of self-assembly. Mimivirus genome encapsidation was shown to occur through a portal different from the ‘stargate’ portal that is used for genome release. Such a ‘division of labor’ is proposed to enhance the efficacy of translocation processes of very large viral genomes. Finally, open questions concerning the infection cycles of giant viruses to which future studies are likely to provide novel and exciting answers are discussed. - Highlights: • The discovery of giant DNA viruses blurs the distinction between viruses and cells. • Mimivirus and Vaccinia replicate exclusively in their host cytoplasm. • Mimivirus genome is delivered through a unique portal coined the Stargate. • Generation of Mimivirus internal membrane proceeds through a novel pathway.

Identification of rep-associated factors in herpes simplex virus type 1-induced adeno-associated virus type 2 replication compartments.

Science.gov (United States)

Nicolas, Armel; Alazard-Dany, Nathalie; Biollay, Coline; Arata, Loredana; Jolinon, Nelly; Kuhn, Lauriane; Ferro, Myriam; Weller, Sandra K; Epstein, Alberto L; Salvetti, Anna; Greco, Anna

2010-09-01

Adeno-associated virus (AAV) is a human parvovirus that replicates only in cells coinfected with a helper virus, such as adenovirus or herpes simplex virus type 1 (HSV-1). We previously showed that nine HSV-1 factors are able to support AAV rep gene expression and genome replication. To elucidate the strategy of AAV replication in the presence of HSV-1, we undertook a proteomic analysis of cellular and HSV-1 factors associated with Rep proteins and thus potentially recruited within AAV replication compartments (AAV RCs). This study resulted in the identification of approximately 60 cellular proteins, among which factors involved in DNA and RNA metabolism represented the largest functional categories. Validation analyses indicated that the cellular DNA replication enzymes RPA, RFC, and PCNA were recruited within HSV-1-induced AAV RCs. Polymerase delta was not identified but subsequently was shown to colocalize with Rep within AAV RCs even in the presence of the HSV-1 polymerase complex. In addition, we found that AAV replication is associated with the recruitment of components of the Mre11/Rad50/Nbs1 complex, Ku70 and -86, and the mismatch repair proteins MSH2, -3, and -6. Finally, several HSV-1 factors were also found to be associated with Rep, including UL12. We demonstrated for the first time that this protein plays a role during AAV replication by enhancing the resolution of AAV replicative forms and AAV particle production. Altogether, these analyses provide the basis to understand how AAV adapts its replication strategy to the nuclear environment induced by the helper virus.

Changes in Cellular mRNA Stability, Splicing, and Polyadenylation through HuR Protein Sequestration by a Cytoplasmic RNA Virus

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Michael D. Barnhart

2013-11-01

Full Text Available The impact of RNA viruses on the posttranscriptional regulation of cellular gene expression is unclear. Sindbis virus causes a dramatic relocalization of the cellular HuR protein from the nucleus to the cytoplasm in infected cells. This is to the result of the expression of large amounts of viral RNAs that contain high-affinity HuR binding sites in their 3′ UTRs effectively serving as a sponge for the HuR protein. Sequestration of HuR by Sindbis virus is associated with destabilization of cellular mRNAs that normally bind HuR and rely on it to regulate their expression. Furthermore, significant changes can be observed in nuclear alternative polyadenylation and splicing events on cellular pre-mRNAs as a result of sequestration of HuR protein by the 3′ UTR of transcripts of this cytoplasmic RNA virus. These studies suggest a molecular mechanism of virus-host interaction that probably has a significant impact on virus replication, cytopathology, and pathogenesis.

Interaction study of rice stripe virus proteins reveals a region of the nucleocapsid protein (NP) required for NP self-interaction and nuclear localization.

Science.gov (United States)

Lian, Sen; Cho, Won Kyong; Jo, Yeonhwa; Kim, Sang-Min; Kim, Kook-Hyung

2014-04-01

Rice stripe virus (RSV), which belongs to the genus Tenuivirus, is an emergent virus problem. The RSV genome is composed of four single-strand RNAs (RNA1-RNA4) and encodes seven proteins. We investigated interactions between six of the RSV proteins by yeast-two hybrid (Y2H) assay in vitro and by bimolecular fluorescence complementation (BiFC) in planta. Y2H identified self-interaction of the nucleocapsid protein (NP) and NS3, while BiFC revealed self-interaction of NP, NS3, and NCP. To identify regions(s) and/or crucial amino acid (aa) residues required for NP self-interaction, we generated various truncated and aa substitution mutants. Y2H assay showed that the N-terminal region of NP (aa 1-56) is necessary for NP self-interaction. Further analysis with substitution mutants demonstrated that additional aa residues located at 42-47 affected their interaction with full-length NP. These results indicate that the N-terminal region (aa 1-36 and 42-47) is required for NP self-interaction. BiFC and co-localization studies showed that the region required for NP self-interaction is also required for NP localization at the nucleus. Overall, our results indicate that the N-terminal region (aa 1-47) of the NP is important for NP self-interaction and that six aa residues (42-47) are essential for both NP self-interaction and nuclear localization. Copyright © 2014 Elsevier B.V. All rights reserved.

Competitive virus assay method for titration of noncytopathogenic bovine viral diarrhea viruses (END⁺ and END⁻ viruses).

Science.gov (United States)

Muhsen, Mahmod; Ohi, Kota; Aoki, Hiroshi; Ikeda, Hidetoshi; Fukusho, Akio

2013-03-01

A new, reliable and secure virus assay method, named the competitive virus assay (CVA) method, has been established for the titration of bovine viral diarrhea viruses (BVDVs) that either show the exaltation of Newcastle disease virus (END) phenomenon or heterologous interference phenomenon (but not the END phenomenon). This method is based on the principle of (1) homologous interference between BVDVs, by using BVDV RK13/E(-) or BVDV RK13/E(+) strains as competitor virus, and (2) END phenomenon and heterologous interference, by using attenuated Newcastle disease virus (NDV) TCND strain as challenge virus. In titration of BVDV END(+) and BVDV END(-) viruses, no significant difference in estimated virus titer was observed between CVA and conventional methods. CVA method demonstrated comparable levels of sensitivity and accuracy as conventional END and interference methods, which require the use of a velogenic Miyadera strain of NDV and vesicular stomatitis virus (VSV), both of which are agents of high-risk diseases. As such, the CVA method is a safer alternative, with increased bio-safety and bio-containment, through avoidance of virulent strains that are commonly employed with conventional methods. Copyright © 2012 Elsevier B.V. All rights reserved.

Establishing cyber security programs for I and C systems at nuclear facilities

International Nuclear Information System (INIS)

Waedt, Karl

2012-01-01

In recent years, across the international nuclear community, cyber security issues have quickly gained significant attention from safety authorities and plant designers alike. This increased attention was accelerated by news of the Stuxnet virus, which impaired control systems at Iranian nuclear facilities in 2010, but is also fueled by regular news about cyber security breaches of data systems at large business corporations. This paper discusses key aspects of establishing a cyber security program for Instrumentation and Control (I and C) systems at a nuclear facility, and identifies inherent aspects of nuclear power plant (NPP) design, that differentiate the needs of such a cyber security program from those of typical corporate data systems. (orig.)

Establishing cyber security programs for I and C systems at nuclear facilities

Energy Technology Data Exchange (ETDEWEB)

Waedt, Karl [AREVA NP GmbH (Germany)

2012-11-01

In recent years, across the international nuclear community, cyber security issues have quickly gained significant attention from safety authorities and plant designers alike. This increased attention was accelerated by news of the Stuxnet virus, which impaired control systems at Iranian nuclear facilities in 2010, but is also fueled by regular news about cyber security breaches of data systems at large business corporations. This paper discusses key aspects of establishing a cyber security program for Instrumentation and Control (I and C) systems at a nuclear facility, and identifies inherent aspects of nuclear power plant (NPP) design, that differentiate the needs of such a cyber security program from those of typical corporate data systems. (orig.)

Virus wars: using one virus to block the spread of another

Directory of Open Access Journals (Sweden)

Matthew L. Paff

2016-06-01

Full Text Available The failure of traditional interventions to block and cure HIV infections has led to novel proposals that involve treating infections with therapeutic viruses–infectious viruses that specifically inhibit HIV propagation in the host. Early efforts in evaluating these proposals have been limited chiefly to mathematical models of dynamics, for lack of suitable empirical systems. Here we propose, develop and analyze an empirical system of a therapeutic virus that protects a host cell population against a lethal virus. The empirical system uses E. coli bacteria as the host cell population, an RNA phage as the lethal virus and a filamentous phage as the therapeutic virus. Basic dynamic properties are established for each virus alone and then together. Observed dynamics broadly agree with those predicted by a computer simulation model, although some differences are noted. Two cases of dynamics are contrasted, differing in whether the therapeutic virus is introduced before the lethal virus or after the lethal virus. The therapeutic virus increases in both cases but by different mechanisms. With the therapeutic virus introduced first, it spreads infectiously without any appreciable change in host dynamics. With the therapeutic virus introduced second, host abundance is depressed at the time therapy is applied; following an initial period of therapeutic virus spread by infection, the subsequent rise of protection is through reproduction by hosts already protected. This latter outcome is due to inheritance of the therapeutic virus state when the protected cell divides. Overall, the work establishes the feasibility and robustness to details of a viral interference using a therapeutic virus.

Zika virus disease

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Adel I Al-Afaleq

2017-01-01

Full Text Available The Zika virus is an arbovirus belonging to the virus family Flaviviridae. The virus was isolated in 1947 from a rhesus monkey in the Zika Forest of Uganda. The virus causes sporadic mild human infections in Africa and later in Asia. However, by 2007 a major shift in its infection pattern was noticed and thousands of human infections were reported in the State of Yap and Federated States of Micronesia. In the last 3 years, major outbreaks have continued to occur and the virus has spread to several Pacific and American countries. These outbreaks were mostly asymptomatic; however, there were more severe clinical signs associated with the infections. Those signs included microcephaly and Guillain–Barre syndrome. It is believed that various species of mosquitoes can biologically transmit the virus. However, Aedes aegypti is most widely associated with the Zika virus. Recently, new modes of virus transmission have been reported, including mother-to-fetus, sexual, blood transfusion, animal bites, laboratory exposure and breast milk. Differential diagnosis is very important as some other arboviruses such as yellow fever virus, West Nile virus, dengue virus, and chikungunya virus have similar clinical manifestations to the Zika virus infection as well as relating serologically to some of these viruses. Established laboratory diagnostic tests to detect the Zika virus are limited, with reverse transcription polymerase chain reaction being the most widely used test. Taking into consideration the quickness of the spread of infection, size of the infected population and change of the infection severity pattern, the Zika virus infection merits collective efforts on all levels to prevent and control the disease. Limited research work and data, concurrent infection with other arboviruses, involvement of biological vectors, mass crowd events, human and trade movements and lack of vaccines are some of the challenges that we face in our efforts to prevent and

Dengue virus receptor

OpenAIRE

Hidari, Kazuya I.P.J.; Suzuki, Takashi

2011-01-01

Dengue virus is an arthropod-borne virus transmitted by Aedes mosquitoes. Dengue virus causes fever and hemorrhagic disorders in humans and non-human primates. Direct interaction of the virus introduced by a mosquito bite with host receptor molecule(s) is crucial for virus propagation and the pathological progression of dengue diseases. Therefore, elucidation of the molecular mechanisms underlying the interaction between dengue virus and its receptor(s) in both humans and mosquitoes is essent...

Japanese encephalitis virus non-coding RNA inhibits activation of interferon by blocking nuclear translocation of interferon regulatory factor 3.

Science.gov (United States)

Chang, Ruey-Yi; Hsu, Ta-Wen; Chen, Yen-Lin; Liu, Shu-Fan; Tsai, Yi-Jer; Lin, Yun-Tong; Chen, Yi-Shiuan; Fan, Yi-Hsin

2013-09-27

Noncoding RNA (ncRNA) plays a critical role in modulating a broad range of diseases. All arthropod-borne flaviviruses produce short fragment ncRNA (sfRNA) collinear with highly conserved regions of the 3'-untranslated region (UTR) in the viral genome. We show that the molar ratio of sfRNA to genomic RNA in Japanese encephalitis virus (JEV) persistently infected cells is greater than that in acutely infected cells, indicating an sfRNA role in establishing persistent infection. Transfecting excess quantities of sfRNA into JEV-infected cells reduced interferon-β (IFN-β) promoter activity by 57% and IFN-β mRNA levels by 52%, compared to mock-transfected cells. Transfection of sfRNA into JEV-infected cells also reduced phosphorylation of interferon regulatory factor-3 (IRF-3), the IFN-β upstream regulator, and blocked roughly 30% of IRF-3 nuclear localization. Furthermore, JEV-infected sfRNA transfected cells produced 23% less IFN-β-stimulated apoptosis than mock-transfected groups did. Taken together, these results suggest that sfRNA plays a role against host-cell antiviral responses, prevents cells from undergoing apoptosis, and thus contributes to viral persistence. Copyright © 2013 Elsevier B.V. All rights reserved.

Vulnerabilities of the security of nuclear material due to cyber attacks

International Nuclear Information System (INIS)

Daschil, F.

2002-01-01

Full text: The spread of new communication and computer technologies is now the potential for a dedicated, sophisticated adversary to conduct coordinated strikes against the computers, communications systems, and databases of nuclear material security systems. All weapons of cyber war, like viruses, Trojans, access to computer systems are potential risks for the security of nuclear material. New forms of destruction of electronic devices, e.g. Transient Electromagnetic Devices (TED), that could, in the hands of enemies, criminals, or terrorist pose a significant threat to nuclear safety infrastructure components that are based on micro-circuits and computer or micro-processor control. These examples shows the necessity to clearly identify possible risks as: information and communication based disorders and other disruptions and attacks; partial or complete failure of systems causing the risk of unexpected reaction or unmanageable situations; cracking of access codes and elimination of alarm equipment could give easy access to nuclear material; computer manipulation or complete computer and system takeover due to brute force attacks, viruses, Trojans and worms could lead to malfunctions, disruptions and nuclear disasters; computer spying and brute force attacks could give detailed information about technical, organizational and organizational data; combined actions of physical, electronic and computer attacks. The paper gives an overview of potential computer and communication safety gaps and points out strategic implications as the identification of the critical infrastructure, surveillance of computer systems, data access and communication paths. The basic necessity of the development of computer and communication breakdown backup systems as well as measures of precaution against cyber attacks is shown in the paper. (author)

ECHO virus

Science.gov (United States)

... page: //medlineplus.gov/ency/article/001340.htm ECHO virus To use the sharing features on this page, please enable JavaScript. Enteric cytopathic human orphan (ECHO) viruses are a group of viruses that can lead ...

Hepatitis E virus coinfection with hepatotropic viruses in Egyptian children.

Science.gov (United States)

Zaki, Maysaa El Sayed; Salama, Osama Saad; Mansour, Fathy Awaad; Hossein, Shaimaa

2008-06-01

Major hepatotropic viruses continue to be important causes of acute viral hepatitis in developing countries. This work was carried out to detect the seroprevalence of hepatitis E virus (HEV) markers in children with acute viral hepatitis due to hepatotropic viruses (A, B and C) and non-A, non-B, non-C acute hepatitis, and to ascertain the influence of HEV superinfection in individuals infected with hepatitis viruses (A, B and C). We studied prospectively 162 children with sporadic acute hepatitis who reported to our hospital. Thirteen healthy controls were also included in the study. Laboratory investigations were performed, including complete liver function tests. Complete serological profiles for hepatitis viruses A, B, C and E were evaluated. HEV immunoglobulin G was detected with highest percentage among patients with hepatitis B (56.7%), followed by patients with hepatitis C virus (52.0%), hepatitis A virus (34.1%) and combined hepatitis B and C viruses (30.0%). The detection rate among patients with non-A, non-B, non-C hepatitis was 7.1%. HEV immunoglobulin M was found in 4.5% of hepatitis A virus patients and in 3.3% of hepatitis B patients. The prevalence of HEV immunoglobulin G and immunoglobulin M correlated with the levels of hepatic aspartate aminotransferase and alanine aminotransferase in patients with dual markers of infection with hepatitis E and other viruses compared to patients with acute hepatitis due to A and C viruses. HEV serological markers are common among children with acute viral hepatitis, especially from hepatitis C and B viruses. There may be increased sensitivity to HEV coinfection in association with hepatitis B and C infections. Dual infection with HEV and other hepatotropic viruses was associated with greater elevation of aspartate and alanine aminotransferases.

Cytoplasmic ATR Activation Promotes Vaccinia Virus Genome Replication

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Antonio Postigo

2017-05-01

Full Text Available In contrast to most DNA viruses, poxviruses replicate their genomes in the cytoplasm without host involvement. We find that vaccinia virus induces cytoplasmic activation of ATR early during infection, before genome uncoating, which is unexpected because ATR plays a fundamental nuclear role in maintaining host genome integrity. ATR, RPA, INTS7, and Chk1 are recruited to cytoplasmic DNA viral factories, suggesting canonical ATR pathway activation. Consistent with this, pharmacological and RNAi-mediated inhibition of canonical ATR signaling suppresses genome replication. RPA and the sliding clamp PCNA interact with the viral polymerase E9 and are required for DNA replication. Moreover, the ATR activator TOPBP1 promotes genome replication and associates with the viral replisome component H5. Our study suggests that, in contrast to long-held beliefs, vaccinia recruits conserved components of the eukaryote DNA replication and repair machinery to amplify its genome in the host cytoplasm.

Phytophthora viruses.

Science.gov (United States)

Cai, Guohong; Hillman, Bradley I

2013-01-01

Phytophthora sp. is a genus in the oomycetes, which are similar to filamentous fungi in morphology and habitat, but phylogenetically more closely related to brown algae and diatoms and fall in the kingdom Stramenopila. In the past few years, several viruses have been characterized in Phytophthora species, including four viruses from Phytophthora infestans, the late blight pathogen, and an endornavirus from an unnamed Phytophthora species from Douglas fir. Studies on Phytophthora viruses have revealed several interesting systems. Phytophthora infestans RNA virus 1 (PiRV-1) and PiRV-2 are likely the first members of two new virus families; studies on PiRV-3 support the establishment of a new virus genus that is not affiliated with established virus families; PiRV-4 is a member of Narnaviridae, most likely in the genus Narnavirus; and Phytophthora endornavirus 1 (PEV1) was the first nonplant endornavirus at the time of reporting. Viral capsids have not been found in any of the above-mentioned viruses. PiRV-1 demonstrated a unique genome organization that requires further examination, and PiRV-2 may have played a role in late blight resurgence in 1980s-1990s. Copyright © 2013 Elsevier Inc. All rights reserved.

Viruses infecting reptiles.

Science.gov (United States)

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.

Viruses Infecting Reptiles

Directory of Open Access Journals (Sweden)

Rachel E. Marschang

2011-11-01

Full Text Available 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.

A novel chromosome region maintenance 1-independent nuclear export signal of the large form of hepatitis delta antigen that is required for the viral assembly.

Science.gov (United States)

Lee, C H; Chang, S C; Wu, C H; Chang, M F

2001-03-16

Hepatitis delta virus (HDV) is a satellite virus of hepatitis B virus, as it requires hepatitis B virus for virion production and transmission. We have previously demonstrated that sequences within the C-terminal 19-amino acid domain flanking the isoprenylation motif of the large hepatitis delta antigen (HDAg-L) are important for virion assembly. In this study, site-directed mutagenesis and immunofluorescence staining demonstrated that in the absence of hepatitis B virus surface antigen (HBsAg), the wild-type HDAg-L was localized in the nuclei of transfected COS7 cells. Nevertheless, in the presence of HBsAg, the HDAg-L became both nuclei- and cytoplasm-distributed in about half of the cells. An HDAg-L mutant with a substitution of Pro-205 to alanine could neither form HDV-like particles nor shift the subcellular localization in the presence of HBsAg. In addition, nuclear trafficking of HDAg-L in heterokaryons indicated that HDAg-L is a nucleocytoplasmic shuttling protein. A proline-rich HDAg peptide spanning amino acid residues 198 to 210, designated NES(HDAg-L), can function as a nuclear export signal (NES) in Xenopus oocytes. Pro-205 is critical for the NES function. Furthermore, assembly of HDV is insensitive to leptomycin B, indicating that the NES(HDAg-L) directs nuclear export of HDAg-L to the cytoplasm via a chromosome region maintenance 1-independent pathway.

Characterization of uncultivable bat influenza virus using a replicative synthetic virus.

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Bin Zhou

2014-10-01

Full Text Available Bats harbor many viruses, which are periodically transmitted to humans resulting in outbreaks of disease (e.g., Ebola, SARS-CoV. Recently, influenza virus-like sequences were identified in bats; however, the viruses could not be cultured. This discovery aroused great interest in understanding the evolutionary history and pandemic potential of bat-influenza. Using synthetic genomics, we were unable to rescue the wild type bat virus, but could rescue a modified bat-influenza virus that had the HA and NA coding regions replaced with those of A/PR/8/1934 (H1N1. This modified bat-influenza virus replicated efficiently in vitro and in mice, resulting in severe disease. Additional studies using a bat-influenza virus that had the HA and NA of A/swine/Texas/4199-2/1998 (H3N2 showed that the PR8 HA and NA contributed to the pathogenicity in mice. Unlike other influenza viruses, engineering truncations hypothesized to reduce interferon antagonism into the NS1 protein didn't attenuate bat-influenza. In contrast, substitution of a putative virulence mutation from the bat-influenza PB2 significantly attenuated the virus in mice and introduction of a putative virulence mutation increased its pathogenicity. Mini-genome replication studies and virus reassortment experiments demonstrated that bat-influenza has very limited genetic and protein compatibility with Type A or Type B influenza viruses, yet it readily reassorts with another divergent bat-influenza virus, suggesting that the bat-influenza lineage may represent a new Genus/Species within the Orthomyxoviridae family. Collectively, our data indicate that the bat-influenza viruses recently identified are authentic viruses that pose little, if any, pandemic threat to humans; however, they provide new insights into the evolution and basic biology of influenza viruses.

Characterization of uncultivable bat influenza virus using a replicative synthetic virus.

Science.gov (United States)

Zhou, Bin; Ma, Jingjiao; Liu, Qinfang; Bawa, Bhupinder; Wang, Wei; Shabman, Reed S; Duff, Michael; Lee, Jinhwa; Lang, Yuekun; Cao, Nan; Nagy, Abdou; Lin, Xudong; Stockwell, Timothy B; Richt, Juergen A; Wentworth, David E; Ma, Wenjun

2014-10-01

Bats harbor many viruses, which are periodically transmitted to humans resulting in outbreaks of disease (e.g., Ebola, SARS-CoV). Recently, influenza virus-like sequences were identified in bats; however, the viruses could not be cultured. This discovery aroused great interest in understanding the evolutionary history and pandemic potential of bat-influenza. Using synthetic genomics, we were unable to rescue the wild type bat virus, but could rescue a modified bat-influenza virus that had the HA and NA coding regions replaced with those of A/PR/8/1934 (H1N1). This modified bat-influenza virus replicated efficiently in vitro and in mice, resulting in severe disease. Additional studies using a bat-influenza virus that had the HA and NA of A/swine/Texas/4199-2/1998 (H3N2) showed that the PR8 HA and NA contributed to the pathogenicity in mice. Unlike other influenza viruses, engineering truncations hypothesized to reduce interferon antagonism into the NS1 protein didn't attenuate bat-influenza. In contrast, substitution of a putative virulence mutation from the bat-influenza PB2 significantly attenuated the virus in mice and introduction of a putative virulence mutation increased its pathogenicity. Mini-genome replication studies and virus reassortment experiments demonstrated that bat-influenza has very limited genetic and protein compatibility with Type A or Type B influenza viruses, yet it readily reassorts with another divergent bat-influenza virus, suggesting that the bat-influenza lineage may represent a new Genus/Species within the Orthomyxoviridae family. Collectively, our data indicate that the bat-influenza viruses recently identified are authentic viruses that pose little, if any, pandemic threat to humans; however, they provide new insights into the evolution and basic biology of influenza viruses.

Epstein-Barr virus nuclear antigen 2 specifically induces expression of the B-cell activation antigen CD23

International Nuclear Information System (INIS)

Wang, F.; Gregory, C.D.; Rowe, M.; Rickinson, A.B.; Wang, D.; Birkenbach, M.; Kikutani, H.; Kishimoto, T.; Kieff, E.

1987-01-01

Epstein-Barr virus (EBV) infection of EBV-negative Burkitt lymphoma (BL) cells includes some changes similar to those seen in normal B lymphocytes that have been growth transformed by EBV. The role of individual EBV genes in this process was evaluated by introducing each of the viral genes that are normally expressed in EBV growth-transformed and latently infected lymphoblasts into an EBV-negative BL cell line, using recombinant retrovirus-mediated transfer. Clones of cells were derived that stably express the EBV nuclear antigen 1 (EBNA-1), EBNA-2, EBNA-3, EBNA-leader protein, or EBV latent membrane protein (LMP). These were compared with control clones infected with the retrovirus vector. All 10 clones converted to EBNA-2 expression differed from control clones or clones expressing other EBV proteins by growth in tight clumps and by markedly increased expression of one particular surface marker of B-cell activation, CD23. Other activation antigens were unaffected by EBNA-2 expression, as were markers already expressed on the parent BL cell line. The results indicate that EBNA-2 is a specific direct or indirect trans-activator of CD23. This establishes a link between an EBV gene and cell gene expression. Since CD23 has been implicated in the transduction of B-cell growth signals, its specific induction by EBNA-2 could be important in EBV induction of B-lymphocyte transformation

Degradation of the encephalomyocarditis virus and hepatitis A virus 3C proteases by the ubiquitin/26S proteasome system in vivo

International Nuclear Information System (INIS)

Schlax, Peter E.; Zhang Jin; Lewis, Elizabeth; Planchart, Antonio; Lawson, T. Glen

2007-01-01

We have isolated stably transfected mouse embryonic fibroblast cell lines that inducibly express either the mature encephalomyocarditis virus (EMCV) or hepatitis A virus (HAV) 3C protease and have used these cells to demonstrate that both proteins are subject to degradation in vivo by the ubiquitin/26S proteasome system. The detection of 3C protease expression in these cells requires inducing conditions and the presence of one of several proteasome inhibitors. Both 3C proteases are incorporated into conjugates with ubiquitin in vivo. HAV 3C protease expression has deleterious effects on cell viability, as determined by observation and counting of cells cultured in the absence or presence of inducing conditions. The EMCV 3C protease was found to be preferentially localized to the nucleus of induced cells, while the HAV 3C protease remains in the cytoplasm. The absence of polyubiquitinated EMCV 3C protease conjugates in nuclear fraction preparations suggests that localization to the nucleus can protect this protein from ubiquitination

Herpes Simplex Virus 1 Us3 Deletion Mutant is Infective Despite Impaired Capsid Translocation to the Cytoplasm

Directory of Open Access Journals (Sweden)

Peter Wild

2015-01-01

Full Text Available Herpes simplex virus 1 (HSV-1 capsids are assembled in the nucleus bud at the inner nuclear membrane into the perinuclear space, acquiring envelope and tegument. In theory, these virions are de-enveloped by fusion of the envelope with the outer nuclear membrane and re-enveloped by Golgi membranes to become infective. Us3 enables the nucleus to cytoplasm capsid translocation. Nevertheless, Us3 is not essential for the production of infective progeny viruses. Determination of phenotype distribution by quantitative electron microscopy, and calculation per mean nuclear or cell volume revealed the following: (i The number of R7041(∆US3 capsids budding at the inner nuclear membrane was significantly higher than that of wild type HSV-1; (ii The mean number of R7041(∆US3 virions per mean cell volume was 2726, that of HSV-1 virions 1460 by 24 h post inoculation; (iii 98% of R7041(∆US3 virions were in the perinuclear space; (iv The number of R7041(∆US3 capsids in the cytoplasm, including those budding at Golgi membranes, was significantly reduced. Cell associated R7041(∆US3 yields were 2.37 × 108 and HSV-1 yields 1.57 × 108 PFU/mL by 24 h post inoculation. We thus conclude that R7041(∆US3 virions, which acquire envelope and tegument by budding at the inner nuclear membrane into the perinuclear space, are infective.

Leukaemia near british nuclear installations

International Nuclear Information System (INIS)

Hubert, D.

1991-01-01

An excess of childhood leukaemia has been seen near some British nuclear installations, especially near the Sellafield reprocessing plant. The same result was found in a more general study including a large number of nuclear sites. Similar studies made in USA, Canada and France have been negative. Moreover, epidemiological studies made in England have discovered other childhood leukaemia clusters in areas far from nuclear facilities, and especially near potential sites of nuclear installations. Several explanations are suggested but no definite conclusion is yet possible. Doses from radioactive releases seem to be too low to account for the additional deaths from leukaemia by environmental contamination. A virus activation, which might be associated with population influx into rural isolated areas, has been considered. The hypothesis of genetic mutation induced by ionising radiation in the fathers of children with leukaemia has been made because a higher risk of leukaemia was observed for children of fathers employed at Sellafield. No firm conclusion is possible considering the small number of observed cases and the lack of excess leukaemias in the offspring of Hiroshima and Nagasaki survivors. The possibility of internal contamination, chemicals or even radon is discussed as other causes. Studies in progress might allow to find an answer to the problem of leukaemia in the vicinity of British nuclear installations [fr

Prevalence of human immunodeficiency virus, hepatitis C virus ...

African Journals Online (AJOL)

Background. Human immunodeficiency virus (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV) and syphilis remain major infections around the world. In Angola, about 166 000 individuals are living with HIV, representing a prevalence of 1.98% in adults between 15 and 49 years of age. In a 2003 study in Luanda, 4.5% ...

Nuclear export of human hepatitis B virus core protein and pregenomic RNA depends on the cellular NXF1-p15 machinery.

Science.gov (United States)

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.

The herpes simplex virus 1 U{sub S}3 regulates phospholipid synthesis

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Wild, Peter, E-mail: pewild@access.uzh.ch [Institute of Veterinary Anatomy, University of Zuerich (Switzerland); Institute of Virology, University of Zuerich (Switzerland); Oliveira, Anna Paula de [Institute of Virology, University of Zuerich (Switzerland); Sonda, Sabrina [Institute for Parasitology, University of Zuerich (Switzerland); Schraner, Elisabeth M. [Institute of Veterinary Anatomy, University of Zuerich (Switzerland); Institute of Virology, University of Zuerich (Switzerland); Ackermann, Mathias; Tobler, Kurt [Institute of Virology, University of Zuerich (Switzerland)

2012-10-25

Herpes simplex virus type 1 capsids bud at nuclear and Golgi membranes for envelopment by phospholipid bilayers. In the absence of U{sub S}3, nuclear membranes form multiple folds harboring virions that suggests disturbance in membrane turnover. Therefore, we investigated phospholipid metabolism in cells infected with the U{sub S}3 deletion mutant R7041({Delta}U{sub S}3), and quantified membranes involved in viral envelopment. We report that (i) [{sup 3}H]-choline incorporation into nuclear membranes and cytoplasmic membranes was enhanced peaking at 12 or 20 h post inoculation with wild type HSV-1 and R7041({Delta}U{sub S}3), respectively, (ii) the surface area of nuclear membranes increased until 24 h of R7041({Delta}U{sub S}3) infection forming folds that equaled {approx}45% of the nuclear surface, (iii) the surface area of viral envelopes between nuclear membranes equaled {approx}2400 R7041({Delta}U{sub S}3) virions per cell, and (iv) during R7041({Delta}U{sub S}3) infection, the Golgi complex expanded dramatically. The data indicate that U{sub S}3 plays a significant role in regulation of membrane biosynthesis.

Epstein - Barr Virus

OpenAIRE

Štorkánová, Lenka

2011-01-01

Epstein-Barr virus Bachelor thesis summarizes the findings of Epstein-Barr virus (EBV), its general characteristics, transmission and spread of the virus, symptoms of disease and subsequent therapy and recovery. More specifically, it focuses on infectious mononucleosis, as well as more generally to other diseases, which the Epstein-Barr virus causes. It includes details of the vaccine against EB virus. There are the statistics on the incidence of infectious mononucleosis.

B23/nucleophosmin interacts with bovine immunodeficiency virus Rev protein and facilitates viral replication.

Science.gov (United States)

Passos-Castilho, Ana Maria; Marchand, Claude; Archambault, Denis

2018-02-01

The bovine immunodeficiency virus (BIV) Rev shuttling protein contains nuclear/nucleolar localization signals and nuclear import/export mechanisms that are novel among lentivirus Rev proteins. Several viral proteins localize to the nucleolus, which may play a role in processes that are essential to the outcome of viral replication. Although BIV Rev localizes to the nucleoli of transfected/infected cells and colocalizes with one of its major proteins, nucleophosmin (NPM1, also known as B23), the role of the nucleolus and B23 in BIV replication remains to be determined. Here, we demonstrate for the first time that BIV Rev interacts with nucleolar phosphoprotein B23 in cells. Using small interfering RNA (siRNA) technology, we show that depletion of B23 expression inhibits virus production by BIV-infected cells, indicating that B23 plays an important role in BIV replication. The interaction between Rev and B23 may represent a potential new target for the development of antiviral drugs against lentiviruses. Copyright © 2017 Elsevier Inc. All rights reserved.

Studies on immune responses to Epstein-Barr virus among A-bomb survivors

International Nuclear Information System (INIS)

Kusunoki, Y.; Kyoizumi, S.; Ozaki, K.; Cologne, J.B.; Akiyama, M.

1992-01-01

Previous studies revealed that reactivity of T-lymphocytes to phytohemag-glutinin and allo-antigens as well as the number of mature CD5 + T cells are decreased among atomic bomb survivors. Possible radiation effects were suggested for impairment of antibody production to certain type A influenza viruses and for an increased prevalence rate of hepatitis B virus surface antigen in sera among survivors. These findings lead to research of effects of A-bomb radiation on immune responses to certain ubiquitous viruses such as Epstein-Barr Virus. Reactivation of EBV induced by depression of immune competence might be reflected by changes in serum titers of these antibodies. Significant increases in titers of antiviral capsed antigen IgC or anti-early antigen (EA) IgC and frequent absence o.r low levels of anti- EBV-associated nuclear antigen antibodies were observed in immunologically compromised individuals. Without regard to diseases, occurrence of significant titers of anti-EA IgC in healthy sero-positive individuals has been ascribed to reactivation of the viral carrier stage. This study examines serum titers of these anti-EBV antibodies to investigate whether any alteration of immune competence to the virus was detectable in relation to the previous A-bomb radiation exposure. Also, an attempt was made to evaluated T-cell responses to EBV in A-bomb survivors for the purpose of understanding involvement of T-cell function in reactivation of the virus, using the precursor frequency analysis of cytotoxic lymphocytes against autologous B cell transformed in vitro with EBV. (author). 13 refs., 2 figs., 1 tab

Emerging influenza viruses and the prospect of a universal influenza virus vaccine.

Science.gov (United States)

Krammer, Florian

2015-05-01

Influenza viruses cause annual seasonal epidemics and pandemics at irregular intervals. Several cases of human infections with avian and swine influenza viruses have been detected recently, warranting enhanced surveillance and the development of more effective countermeasures to address the pandemic potential of these viruses. The most effective countermeasure against influenza virus infection is the use of prophylactic vaccines. However, vaccines that are currently in use for seasonal influenza viruses have to be re-formulated and re-administered in a cumbersome process every year due to the antigenic drift of the virus. Furthermore, current seasonal vaccines are ineffective against novel pandemic strains. This paper reviews zoonotic influenza viruses with pandemic potential and technological advances towards better vaccines that induce broad and long lasting protection from influenza virus infection. Recent efforts have focused on the development of broadly protective/universal influenza virus vaccines that can provide immunity against drifted seasonal influenza virus strains but also against potential pandemic viruses. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Virus-host interaction in feline immunodeficiency virus (FIV) infection.

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Taniwaki, Sueli Akemi; Figueiredo, Andreza Soriano; Araujo, João Pessoa

2013-12-01

Feline immunodeficiency virus (FIV) infection has been the focus of several studies because this virus exhibits genetic and pathogenic characteristics that are similar to those of the human immunodeficiency virus (HIV). FIV causes acquired immunodeficiency syndrome (AIDS) in cats, nevertheless, a large fraction of infected cats remain asymptomatic throughout life despite of persistent chronic infection. This slow disease progression may be due to the presence of factors that are involved in the natural resistance to infection and the immune response that is mounted by the animals, as well as due to the adaptation of the virus to the host. Therefore, the study of virus-host interaction is essential to the understanding of the different patterns of disease course and the virus persistence in the host, and to help with the development of effective vaccines and perhaps the cure of FIV and HIV infections. Copyright © 2013 Elsevier Ltd. All rights reserved.

Involvement of UL24 in herpes-simplex-virus-1-induced dispersal of nucleolin

International Nuclear Information System (INIS)

Lymberopoulos, Maria H.; Pearson, Angela

2007-01-01

UL24 of herpes simplex virus 1 is important for efficient viral replication, but its function is unknown. We generated a recombinant virus, vHA-UL24, encoding UL24 with an N-terminal hemagglutinin tag. By indirect immunofluorescence at 9 h post-infection (hpi), we detected HA-UL24 in nuclear foci and in cytoplasmic speckles. HA-UL24 partially co-localized with nucleolin, but not with ICP8 or coilin, markers for nucleoli, viral replication compartments, and Cajal bodies respectively. HA-UL24 staining was often juxtaposed to that of another nucleolar protein, fibrillarin. Analysis of HSV-1-induced nucleolar modifications revealed that by 18 hpi, nucleolin staining had dispersed, and fibrillarin staining went from clusters of small spots to a few separate but prominent spots. Fibrillarin redistribution appeared to be independent of UL24. In contrast, cells infected with a UL24-deficient virus retained foci of nucleolin staining. Our results demonstrate involvement of UL24 in dispersal of nucleolin during infection

Systematic analysis of protein identity between Zika virus and other arthropod-borne viruses.

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Chang, Hsiao-Han; Huber, Roland G; Bond, Peter J; Grad, Yonatan H; Camerini, David; Maurer-Stroh, Sebastian; Lipsitch, Marc

2017-07-01

To analyse the proportions of protein identity between Zika virus and dengue, Japanese encephalitis, yellow fever, West Nile and chikungunya viruses as well as polymorphism between different Zika virus strains. We used published protein sequences for the Zika virus and obtained protein sequences for the other viruses from the National Center for Biotechnology Information (NCBI) protein database or the NCBI virus variation resource. We used BLASTP to find regions of identity between viruses. We quantified the identity between the Zika virus and each of the other viruses, as well as within-Zika virus polymorphism for all amino acid k -mers across the proteome, with k ranging from 6 to 100. We assessed accessibility of protein fragments by calculating the solvent accessible surface area for the envelope and nonstructural-1 (NS1) proteins. In total, we identified 294 Zika virus protein fragments with both low proportion of identity with other viruses and low levels of polymorphisms among Zika virus strains. The list includes protein fragments from all Zika virus proteins, except NS3. NS4A has the highest number (190 k -mers) of protein fragments on the list. We provide a candidate list of protein fragments that could be used when developing a sensitive and specific serological test to detect previous Zika virus infections.

Blueberry (Vaccinium corymbosum)-Virus Diseases

Science.gov (United States)

At least six viruses have been found in highbush blueberry plantings in the Pacific Northwest: Blueberry mosaic virus, Blueberry red ringspot virus, Blueberry scorch virus, Blueberry shock virus, Tobacco ringspot virus, and Tomato ringspot virus. Six other virus and virus-like diseases of highbush b...

MNF, an ankyrin repeat protein of myxoma virus, is part of a native cellular SCF complex during viral infection

Directory of Open Access Journals (Sweden)

Gelfi Jacqueline

2010-03-01

Full Text Available Abstract Myxoma virus (MYXV, a member of the Poxviridae family, is the agent responsible for myxomatosis, a fatal disease in the European rabbit (Oryctolagus cuniculus. Like all poxviruses, MYXV is known for encoding multiple proteins that regulate cellular signaling pathways. Among them, four proteins share the same ANK/PRANC structure: M148R, M149R, MNF (Myxoma Nuclear factor and M-T5, all of them described as virulence factors. This family of poxvirus proteins, recently identified, has drawn considerable attention for its potential role in modulating the host ubiquitin-proteasome system during viral infection. To date, many members of this novel protein family have been shown to interact with SCF components, in vitro. Here, we focus on MNF gene, which has been shown to express a nuclear protein presenting nine ANK repeats, one of which has been identified as a nuclear localization signal. In transfection, MNF has been shown to colocalise with the transcription factor NF-κB in the nucleus of TNFα-stimulated cells. Functionally, MNF is a critical virulence factor since its deletion generates an almost apathogenic virus. In this study, to pursue the investigation of proteins interacting with MNF and of its mechanism of action, we engineered a recombinant MYXV expressing a GFP-linked MNF under the control of MNF native promoter. Infection of rabbits with MYXV-GFPMNF recombinant virus provided the evidence that the GFP fusion does not disturb the main function of MNF. Hence, cells were infected with MYXV-GFPMNF and immunoprecipitation of the GFPMNF fusion protein was performed to identify MNF's partners. For the first time, endogenous components of SCF (Cullin-1 and Skp1 were co-precipitated with an ANK myxoma virus protein, expressed in an infectious context, and without over-expression of any protein.

MNF, an ankyrin repeat protein of myxoma virus, is part of a native cellular SCF complex during viral infection

Science.gov (United States)

2010-01-01

Myxoma virus (MYXV), a member of the Poxviridae family, is the agent responsible for myxomatosis, a fatal disease in the European rabbit (Oryctolagus cuniculus). Like all poxviruses, MYXV is known for encoding multiple proteins that regulate cellular signaling pathways. Among them, four proteins share the same ANK/PRANC structure: M148R, M149R, MNF (Myxoma Nuclear factor) and M-T5, all of them described as virulence factors. This family of poxvirus proteins, recently identified, has drawn considerable attention for its potential role in modulating the host ubiquitin-proteasome system during viral infection. To date, many members of this novel protein family have been shown to interact with SCF components, in vitro. Here, we focus on MNF gene, which has been shown to express a nuclear protein presenting nine ANK repeats, one of which has been identified as a nuclear localization signal. In transfection, MNF has been shown to colocalise with the transcription factor NF-κB in the nucleus of TNFα-stimulated cells. Functionally, MNF is a critical virulence factor since its deletion generates an almost apathogenic virus. In this study, to pursue the investigation of proteins interacting with MNF and of its mechanism of action, we engineered a recombinant MYXV expressing a GFP-linked MNF under the control of MNF native promoter. Infection of rabbits with MYXV-GFPMNF recombinant virus provided the evidence that the GFP fusion does not disturb the main function of MNF. Hence, cells were infected with MYXV-GFPMNF and immunoprecipitation of the GFPMNF fusion protein was performed to identify MNF's partners. For the first time, endogenous components of SCF (Cullin-1 and Skp1) were co-precipitated with an ANK myxoma virus protein, expressed in an infectious context, and without over-expression of any protein. PMID:20211013

Role of CD137 signaling in dengue virus-mediated apoptosis

International Nuclear Information System (INIS)

Nagila, Amar; Netsawang, Janjuree; Srisawat, Chatchawan; Noisakran, Sansanee; Morchang, Atthapan; Yasamut, Umpa; Puttikhunt, Chunya; Kasinrerk, Watchara

2011-01-01

Highlights: → For the first time the role of CD137 in dengue virus (DENV) infection. → Induction of DENV-mediated apoptosis by CD137 signaling. → Sensitization to CD137-mediated apoptosis by dengue virus capsid protein (DENV C). → Nuclear localization of DENV C is required for CD137-mediated apoptosis. -- Abstract: Hepatic dysfunction is a well recognized feature of dengue virus (DENV) infection. However, molecular mechanisms of hepatic injury are still poorly understood. A complex interaction between DENV and the host immune response contributes to DENV-mediated tissue injury. DENV capsid protein (DENV C) physically interacts with the human death domain-associated protein Daxx. A double substitution mutation in DENV C (R85A/K86A) abrogates Daxx interaction, nuclear localization and apoptosis. Therefore we compared the expression of cell death genes between HepG2 cells expressing DENV C and DENV C (R85A/K86A) using a real-time PCR array. Expression of CD137, which is a member of the tumor necrosis factor receptor family, increased significantly in HepG2 cells expressing DENV C compared to HepG2 cells expressing DENV C (R85A/K86A). In addition, CD137-mediated apoptotic activity in HepG2 cells expressing DENV C was significantly increased by anti-CD137 antibody compared to that of HepG2 cells expressing DENV C (R85A/K86A). In DENV-infected HepG2 cells, CD137 mRNA and CD137 positive cells significantly increased and CD137-mediated apoptotic activity was increased by anti-CD137 antibody. This work is the first to demonstrate the contribution of CD137 signaling to DENV-mediated apoptosis.

Role of CD137 signaling in dengue virus-mediated apoptosis

Energy Technology Data Exchange (ETDEWEB)

Nagila, Amar [Medical Molecular Biology Unit, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Department of Biochemistry, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Netsawang, Janjuree [Faculty of Medical Technology, Rangsit University, Bangkok (Thailand); Srisawat, Chatchawan [Department of Biochemistry, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Noisakran, Sansanee [Dengue Hemorrhagic Fever Research Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok (Thailand); Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok (Thailand); Morchang, Atthapan; Yasamut, Umpa [Medical Molecular Biology Unit, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Department of Immunology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Puttikhunt, Chunya [Dengue Hemorrhagic Fever Research Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok (Thailand); Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok (Thailand); Kasinrerk, Watchara [Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai (Thailand); Biomedical Technology Research Center, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency at Chiang Mai University, Chiang Mai (Thailand); and others

2011-07-08

Highlights: {yields} For the first time the role of CD137 in dengue virus (DENV) infection. {yields} Induction of DENV-mediated apoptosis by CD137 signaling. {yields} Sensitization to CD137-mediated apoptosis by dengue virus capsid protein (DENV C). {yields} Nuclear localization of DENV C is required for CD137-mediated apoptosis. -- Abstract: Hepatic dysfunction is a well recognized feature of dengue virus (DENV) infection. However, molecular mechanisms of hepatic injury are still poorly understood. A complex interaction between DENV and the host immune response contributes to DENV-mediated tissue injury. DENV capsid protein (DENV C) physically interacts with the human death domain-associated protein Daxx. A double substitution mutation in DENV C (R85A/K86A) abrogates Daxx interaction, nuclear localization and apoptosis. Therefore we compared the expression of cell death genes between HepG2 cells expressing DENV C and DENV C (R85A/K86A) using a real-time PCR array. Expression of CD137, which is a member of the tumor necrosis factor receptor family, increased significantly in HepG2 cells expressing DENV C compared to HepG2 cells expressing DENV C (R85A/K86A). In addition, CD137-mediated apoptotic activity in HepG2 cells expressing DENV C was significantly increased by anti-CD137 antibody compared to that of HepG2 cells expressing DENV C (R85A/K86A). In DENV-infected HepG2 cells, CD137 mRNA and CD137 positive cells significantly increased and CD137-mediated apoptotic activity was increased by anti-CD137 antibody. This work is the first to demonstrate the contribution of CD137 signaling to DENV-mediated apoptosis.

Single virus genomics: a new tool for virus discovery.

Directory of Open Access Journals (Sweden)

Lisa Zeigler Allen

Full Text Available Whole genome amplification and sequencing of single microbial cells has significantly influenced genomics and microbial ecology by facilitating direct recovery of reference genome data. However, viral genomics continues to suffer due to difficulties related to the isolation and characterization of uncultivated viruses. We report here on a new approach called 'Single Virus Genomics', which enabled the isolation and complete genome sequencing of the first single virus particle. A mixed assemblage comprised of two known viruses; E. coli bacteriophages lambda and T4, were sorted using flow cytometric methods and subsequently immobilized in an agarose matrix. Genome amplification was then achieved in situ via multiple displacement amplification (MDA. The complete lambda phage genome was recovered with an average depth of coverage of approximately 437X. The isolation and genome sequencing of uncultivated viruses using Single Virus Genomics approaches will enable researchers to address questions about viral diversity, evolution, adaptation and ecology that were previously unattainable.

Resistance to Two Heterologous Neurotropic Oncolytic Viruses, Semliki Forest Virus and Vaccinia Virus, in Experimental Glioma

Science.gov (United States)

Le Boeuf, Fabrice; Lemay, Chantal; De Silva, Naomi; Diallo, Jean-Simon; Cox, Julie; Becker, Michelle; Choi, Youngmin; Ananth, Abhirami; Sellers, Clara; Breton, Sophie; Roy, Dominic; Falls, Theresa; Brun, Jan; Hemminki, Akseli; Hinkkanen, Ari; Bell, John C.

2013-01-01

Attenuated Semliki Forest virus (SFV) may be suitable for targeting malignant glioma due to its natural neurotropism, but its replication in brain tumor cells may be restricted by innate antiviral defenses. We attempted to facilitate SFV replication in glioma cells by combining it with vaccinia virus, which is capable of antagonizing such defenses. Surprisingly, we found parenchymal mouse brain tumors to be refractory to both viruses. Also, vaccinia virus appears to be sensitive to SFV-induced antiviral interference. PMID:23221568

Engineering development of avian influenza virus detection system in a patient's body

International Nuclear Information System (INIS)

Budi Santoso; Romadhon; Sukandar; Istofa

2010-01-01

The avian influenza virus detection equipment in a patient's body has been made. Currently, detection of avian influenza virus carried out by expensive laboratory equipment's, so only certain hospitals can perform this detection. This developing equipment is expected to be cheaper than existing equipment and the diagnosis can be known immediately. The sensing device is made using the principle of nuclear radiation detection. Radiation comes from a drunk labelled tamiflu (oseltamivir) which is drunk to the patient. Tamiflu is a drug to catch, H5N1 viruses in a patient's body. A labelled tamiflu is tamiflu which is labelled by I-131 radioisotopes. The presence of virus in the body is proportional to the amount of radiation captured by the detector. The equipment is composed of a Geiger-Mueller (GM) pancake detector type, a signal processor, a counter, and a data processor (computer). The GM detector converts the radiation that comes into electrical signals. Electrical signal is then converted into TTL level pulses by the signal processor. Pulse counting results are processed by data processor. The total count is proportional to the amount of virus captured by labelled tamiflu. The measurement threshold can be set by medical officer through software. At a certain threshold can be inferred identified patients infected with avian influenza virus. If the measurement below the threshold means that the patient is still within safe limits. This equipment is expected to create avian influenza virus detection system that cheaply and quickly so that more and more hospitals are using to detect the avian influenza virus. (author)

Detection of sweet potato virus C, sweet potato virus 2 and sweet potato feathery mottle virus in Portugal.

Science.gov (United States)

Varanda, Carla M R; Santos, Susana J; Oliveira, Mônica D M; Clara, Maria Ivone E; Félix, Maria Rosário F

2015-06-01

Field sweet potato plants showing virus-like symptoms, as stunting, leaf distortion, mosaic and chlorosis, were collected in southwest Portugal and tested for the presence of four potyviruses, sweet potato virus C (SPVC), sweet potato virus 2 (SPV2), sweet potato feathery mottle virus (SPFMV), sweet potato virus G (SPVG), and the crinivirus sweet potato chlorotic stunt virus (SPCSV). DsRNA fractions were extracted from symptomatic leaves and used as templates in single and multiplex RT-PCR assays using previously described specific primers for each analyzed virus. The amplified reaction products for SPVC, SPV2 and SPFMV were of expected size, and direct sequencing of PCR products revealed that they correspond to the coat protein gene (CP) and showed 98%, 99% and 99% identity, respectively, to those viruses. Comparison of the CP genomic and amino acid sequences of the Portuguese viral isolates recovered here with those of ten other sequences of isolates obtained in different countries retrieved from the GenBank showed very few differences. The application of the RT-PCR assays revealed for the first time the presence of SPVC and SPFMV in the sweet potato crop in Portugal, the absence of SPVG and SPCSV in tested plants, as well as the occurrence of triple virus infections under field conditions.

Hepatitis E virus and fulminant hepatitis--a virus or host-specific pathology?

Science.gov (United States)

Smith, Donald B; Simmonds, Peter

2015-04-01

Fulminant hepatitis is a rare outcome of infection with hepatitis E virus. Several recent reports suggest that virus variation is an important determinant of disease progression. To critically examine the evidence that virus-specific factors underlie the development of fulminant hepatitis following hepatitis E virus infection. Published sequence information of hepatitis E virus isolates from patients with and without fulminant hepatitis was collected and analysed using statistical tests to identify associations between virus polymorphisms and disease outcome. Fulminant hepatitis has been reported following infection with all four hepatitis E virus genotypes that infect humans comprising multiple phylogenetic lineages within genotypes 1, 3 and 4. Analysis of virus sequences from individuals infected by a common source did not detect any common substitutions associated with progression to fulminant hepatitis. Re-analysis of previously reported associations between virus substitutions and fulminant hepatitis suggests that these were probably the result of sampling biases. Host-specific factors rather than virus genotype, variants or specific substitutions appear to be responsible for the development of fulminant hepatitis. © 2014 The Authors. Liver International Published by John Wiley & Sons Ltd.

Interaction of HTLV-1 Tax protein with the calreticulin: Implications for Tax nuclear export and secretion

OpenAIRE

Alefantis, Timothy; Flaig, Katherine E.; Wigdahl, Brian; Jain, Pooja

2007-01-01

Human T cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The HTLV-1 transcriptional transactivator protein Tax plays an integral role in virus replication and disease progression. Traditionally, Tax is described as a nuclear protein where it performs its primary role as a transcriptional transactivator. However, recent studies have clearly shown that Tax can also be localized to t...

Circulating avian influenza viruses closely related to the 1918 virus have pandemic potential

Science.gov (United States)

Watanabe, Tokiko; Zhong, Gongxun; Russell, Colin A.; Nakajima, Noriko; Hatta, Masato; Hanson, Anthony; McBride, Ryan; Burke, David F.; Takahashi, Kenta; Fukuyama, Satoshi; Tomita, Yuriko; Maher, Eileen A.; Watanabe, Shinji; Imai, Masaki; Neumann, Gabriele; Hasegawa, Hideki; Paulson, James C.; Smith, Derek J.; Kawaoka, Yoshihiro

2014-01-01

Summary Wild birds harbor a large gene pool of influenza A viruses that have the potential to cause influenza pandemics. Foreseeing and understanding this potential is important for effective surveillance. Our phylogenetic and geographic analyses revealed the global prevalence of avian influenza virus genes whose proteins differ only a few amino acids from the 1918 pandemic influenza virus, suggesting that 1918-like pandemic viruses may emerge in the future. To assess this risk, we generated and characterized a virus composed of avian influenza viral segments with high homology to the 1918 virus. This virus exhibited higher pathogenicity in mice and ferrets than an authentic avian influenza virus. Further, acquisition of seven amino acid substitutions in the viral polymerases and the hemagglutinin surface glycoprotein conferred respiratory droplet transmission to the 1918-like avian virus in ferrets, demonstrating that contemporary avian influenza viruses with 1918 virus-like proteins may have pandemic potential. PMID:24922572

Lack of Durable Cross-Neutralizing Antibodies Against Zika Virus from Dengue Virus Infection.

Science.gov (United States)

Collins, Matthew H; McGowan, Eileen; Jadi, Ramesh; Young, Ellen; Lopez, Cesar A; Baric, Ralph S; Lazear, Helen M; de Silva, Aravinda M

2017-05-01

Cross-reactive antibodies elicited by dengue virus (DENV) infection might affect Zika virus infection and confound serologic tests. Recent data demonstrate neutralization of Zika virus by monoclonal antibodies or human serum collected early after DENV infection. Whether this finding is true in late DENV convalescence (>6 months after infection) is unknown. We studied late convalescent serum samples from persons with prior DENV or Zika virus exposure. Despite extensive cross-reactivity in IgG binding, Zika virus neutralization was not observed among primary DENV infections. We observed low-frequency (23%) Zika virus cross-neutralization in repeat DENV infections. DENV-immune persons who had Zika virus as a secondary infection had distinct populations of antibodies that neutralized DENVs and Zika virus, as shown by DENV-reactive antibody depletion experiments. These data suggest that most DENV infections do not induce durable, high-level Zika virus cross-neutralizing antibodies. Zika virus-specific antibody populations develop after Zika virus infection irrespective of prior DENV immunity.

Bombyx mori nucleopolyhedrovirus BM5 protein regulates progeny virus production and viral gene expression

International Nuclear Information System (INIS)

Kokusho, Ryuhei; Koh, Yoshikazu; Fujimoto, Masaru; Shimada, Toru; Katsuma, Susumu

2016-01-01

Bombyx mori nucleopolyhedrovirus (BmNPV) orf5 (Bm5) is a core gene of lepidopteran baculoviruses and encodes the protein with the conserved amino acid residues (DUF3627) in its C-terminus. Here, we found that Bm5 disruption resulted in lower titers of budded viruses and fewer numbers of occlusion bodies (OBs) in B. mori cultured cells and larvae, although viral genome replication was not affected. Bm5 disruption also caused aberrant expression of various viral genes at the very late stage of infection. Immunocytochemical analysis revealed that BM5 localized to the nuclear membrane. We also found that DUF3627 is important for OB production, transcriptional regulation of viral genes, and subcellular localization of BM5. Compared with wild-type BmNPV infection, larval death was delayed when B. mori larvae were infected with Bm5 mutants. These results suggest that BM5 is involved in progeny virus production and regulation of viral gene expression at the very late stage of infection. -- Highlights: •The role of BmNPV BM5 protein was examined in B. mori cultured cells and larvae. •BM5 contributes to efficient production of budded viruses and occlusion bodies. •BM5 regulates viral gene expression at the very late stage of infection. •BM5 dominantly localizes to the nuclear membrane. •Bm5 mutant showed v-cath down-regulation and resulting delay of larval death.

Bombyx mori nucleopolyhedrovirus BM5 protein regulates progeny virus production and viral gene expression

Energy Technology Data Exchange (ETDEWEB)

Kokusho, Ryuhei, E-mail: kokusho@ss.ab.a.u-tokyo.ac.jp; Koh, Yoshikazu; Fujimoto, Masaru; Shimada, Toru; Katsuma, Susumu, E-mail: katsuma@ss.ab.a.u-tokyo.ac.jp

2016-11-15

Bombyx mori nucleopolyhedrovirus (BmNPV) orf5 (Bm5) is a core gene of lepidopteran baculoviruses and encodes the protein with the conserved amino acid residues (DUF3627) in its C-terminus. Here, we found that Bm5 disruption resulted in lower titers of budded viruses and fewer numbers of occlusion bodies (OBs) in B. mori cultured cells and larvae, although viral genome replication was not affected. Bm5 disruption also caused aberrant expression of various viral genes at the very late stage of infection. Immunocytochemical analysis revealed that BM5 localized to the nuclear membrane. We also found that DUF3627 is important for OB production, transcriptional regulation of viral genes, and subcellular localization of BM5. Compared with wild-type BmNPV infection, larval death was delayed when B. mori larvae were infected with Bm5 mutants. These results suggest that BM5 is involved in progeny virus production and regulation of viral gene expression at the very late stage of infection. -- Highlights: •The role of BmNPV BM5 protein was examined in B. mori cultured cells and larvae. •BM5 contributes to efficient production of budded viruses and occlusion bodies. •BM5 regulates viral gene expression at the very late stage of infection. •BM5 dominantly localizes to the nuclear membrane. •Bm5 mutant showed v-cath down-regulation and resulting delay of larval death.

Ganjam virus.

Science.gov (United States)

Sudeep, A B; Jadi, R S; Mishra, A C

2009-11-01

Ganjam virus (GANV), a member of genus Nairovirus of family Bunyavirdae is of considerable veterinary importance in India. Though, predominantly tick borne, GANV was also isolated from mosquitoes, man and sheep. Neutralizing and complement fixing antibodies to GANV have been detected in animal and human sera collected from different parts of the country. Thirty three strains of GANV have been isolated from India, mainly from Haemaphysalis ticks. The virus replicated in certain vertebrate and mosquito cell lines and found pathogenic to laboratory animals. One natural infection and five laboratory-acquired infections in men were also reported. GANV is antigenically related to Nairobi sheep disease virus (NSDV) of Africa, which is highly pathogenic for sheep and goats causing 70-90 per cent mortality among the susceptible population. Recent molecular studies have demonstrated that GANV is an Asian variant of NSDV and both these viruses are related to the dreaded Crimean Congo haemorrhagic fever (CCHF) group viruses. The versatility of the virus to replicate in different arthropod species, its ability to infect sheep, goat and man makes it an important zoonotic agent.

Generation and comprehensive analysis of an influenza virus polymerase cellular interaction network.

Science.gov (United States)

Tafforeau, Lionel; Chantier, Thibault; Pradezynski, Fabrine; Pellet, Johann; Mangeot, Philippe E; Vidalain, Pierre-Olivier; Andre, Patrice; Rabourdin-Combe, Chantal; Lotteau, Vincent

2011-12-01

The influenza virus transcribes and replicates its genome inside the nucleus of infected cells. Both activities are performed by the viral RNA-dependent RNA polymerase that is composed of the three subunits PA, PB1, and PB2, and recent studies have shown that it requires host cell factors to transcribe and replicate the viral genome. To identify these cellular partners, we generated a comprehensive physical interaction map between each polymerase subunit and the host cellular proteome. A total of 109 human interactors were identified by yeast two-hybrid screens, whereas 90 were retrieved by literature mining. We built the FluPol interactome network composed of the influenza virus polymerase (PA, PB1, and PB2) and the nucleoprotein NP and 234 human proteins that are connected through 279 viral-cellular protein interactions. Analysis of this interactome map revealed enriched cellular functions associated with the influenza virus polymerase, including host factors involved in RNA polymerase II-dependent transcription and mRNA processing. We confirmed that eight influenza virus polymerase-interacting proteins are required for virus replication and transcriptional activity of the viral polymerase. These are involved in cellular transcription (C14orf166, COPS5, MNAT1, NMI, and POLR2A), translation (EIF3S6IP), nuclear transport (NUP54), and DNA repair (FANCG). Conversely, we identified PRKRA, which acts as an inhibitor of the viral polymerase transcriptional activity and thus is required for the cellular antiviral response.

Borna disease virus nucleoprotein inhibits type I interferon induction through the interferon regulatory factor 7 pathway

International Nuclear Information System (INIS)

Song, Wuqi; Kao, Wenping; Zhai, Aixia; Qian, Jun; Li, Yujun; Zhang, Qingmeng; Zhao, Hong; Hu, Yunlong; Li, Hui; Zhang, Fengmin

2013-01-01

Highlights: •IRF7 nuclear localisation was inhibited by BDV persistently infected. •BDV N protein resistant to IFN induction both in BDV infected OL cell and N protein plasmid transfected OL cell. •BDV N protein is related to the inhibition of IRF7 nuclear localisation. -- Abstract: The expression of type I interferon (IFN) is one of the most potent innate defences against viral infection in higher vertebrates. Borna disease virus (BDV) establishes persistent, noncytolytic infections in animals and in cultured cells. Early studies have shown that the BDV phosphoprotein can inhibit the activation of type I IFN through the TBK1–IRF3 pathway. The function of the BDV nucleoprotein in the inhibition of IFN activity is not yet clear. In this study, we demonstrated IRF7 activation and increased IFN-α/β expression in a BDV-persistently infected human oligodendroglia cell line following RNA interference-mediated BDV nucleoprotein silencing. Furthermore, we showed that BDV nucleoprotein prevented the nuclear localisation of IRF7 and inhibited endogenous IFN induction by poly(I:C), coxsackie virus B3 and IFN-β. Our findings provide evidence for a previously undescribed mechanism by which the BDV nucleoprotein inhibits type I IFN expression by interfering with the IRF7 pathway

Presence and characterization of Zucchini yellow mosaic virus in watermelon in Serbia

Directory of Open Access Journals (Sweden)

Vučurović Ana

2012-01-01

Full Text Available The presence of Zucchini yellow mosaic virus (ZYMV in two out of seven watermelon production localities in Serbia during 2011 was investigated by analyzing leaves sampled from symptomatic and asymptomatic watermelon plants and utilizing DAS-ELISA test. In the locality of Gornji Tavankut, ZYMV was detected in 23.08% of tested plants in single infections, and in the locality of Silbas it was detected in 35.29% of tested plants in mixed infections with Cucumber mosaic virus and Alfalfa mosaic virus. ZYMV was successfully mechanically transmitted from naturally infected watermelon plants to Cucurbita pepo 'Ezra F1'. Molecular detection was performed by RT-PCR and amplification of part of the gene for nuclear inclusions, gene of coat protein and part of 3' non-coding region, which confirmed the identification of the ZYMV isolates. Phylogenetic analysis revealed grouping of the isolate originating from watermelon with other isolates from Serbia and Central Europe within A-I subgroup. Analysis of amino acid sequences of the N terminal end of the CP gene revealed that isolate 550-11 belongs to the Central European branch.

Selective inhibition of influenza virus protein synthesis by inhibitors of DNA function

International Nuclear Information System (INIS)

Minor, P.D.; Dimmock, N.J.

1977-01-01

Various known inhibitors of cellular DNA function were shown to inhibit cellular RNA synthesis and influenza (fowl plague) virus multiplication. The drugs were investigated for their effect upon the synthesis of influenza virus proteins. According to this effect they could be classified with previously studied compounds as follows: Group I (ethidium bromide, proflavine, and N-nitroquinoline-N-oxide) inhibited both viral and cellular protein synthesis; Group II (nogalomycin, daunomycin and α-amanitin) inhibited viral but not cellular protein synthesis, and all viral proteins were inhibited coordinately; Group III (mithramycin, echinomycin, and actinomycin D) inhibited all viral but not cellular protein synthesis at high concentrations, but at a lower critical concentration inhibited the synthesis of viral haemagglutinin, neuraminidase, and M protein preferentially; Group IV(uv irradiation and camptothecin) inhibited the synthesis of viral haemagglutinin, neuraminidase, and M protein, but not other viral proteins, even at high doses. The mode of action of these inhibitors is discussed in relation to the mechanism of the nuclear events upon which influenza virus multiplication is dependent

The Drosophila Nora virus is an enteric virus, transmitted via feces.

Science.gov (United States)

Habayeb, Mazen S; Cantera, Rafael; Casanova, Gabriela; Ekström, Jens-Ola; Albright, Shannon; Hultmark, Dan

2009-04-01

The biology of the Drosophila viruses has not been intensely investigated. Here we have investigated the biology of the Nora virus, a persistent Drosophila virus. We find that injected Nora virus is able to replicate in the files, reaching a high titer that is maintained in the next generation. There is a remarkable variation in the viral loads of individual flies in persistently infected stocks; the titers can differ by three orders of magnitude. The Nora virus is mainly found in the intestine of infected flies, and the histology of these infected intestines show increased vacuolization. The virus is excreted in the feces and is horizontally transmitted. The Nora virus infection has a very mild effect on the longevity of the flies, and no significant effect on the number of eggs laid and the percent of eggs that develop to adults.

Herpes viruses and human papilloma virus in nasal polyposis and controls

Directory of Open Access Journals (Sweden)

Dimitrios Ioannidis

2015-12-01

Full Text Available ABSTRACT INTRODUCTION: Chronic rhinosinusitis with nasal polyps is a multifactorial disease entity with an unclear pathogenesis. Contradictory data exist in the literature on the potential implication of viral elements in adult patients with chronic rhinosinusitis. OBJECTIVE: To compare the prevalence of human herpes viruses (1-6 and Human Papilloma Virus in adult patients with chronic rhinosinusitis with nasal polyps and healthy controls. METHODS: Viral DNA presence was evaluated by real-time polymerase chain reaction application to nasal polyps specimens from 91 chronic rhinosinusitis with nasal polyps patients and nasal turbinate mucosa from 38 healthy controls. RESULTS: Epstein-Barr virus positivity was higher in nasal polyps (24/91; 26.4% versus controls (4/38; 10.5%, but the difference did not reach significance (p = 0.06. Human herpes virus-6 positivity was lower in nasal polyps (13/91; 14.29% versus controls (10/38; 26.32%,p = 0.13. In chronic rhinosinusitis with nasal polyps group, 1 sample was herpes simplex virus-1-positive (1/91; 1.1%, and another was cytomegalovirus-positive (1/91; 1.1%, versus none in controls. No sample was positive for herpes simplex virus-2, varicella-zoster virus, high-risk-human papilloma viruses (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and low-risk-human papilloma viruses (6, 11. CONCLUSION: Differences in Epstein-Barr virus and human herpes virus-6 positivity among patients with chronic rhinosinusitis with nasal polyps and healthy controls are not statistically significant, weakening the likelihood of their implication in chronic rhinosinusitis with nasal polyps pathogenesis.

Evaluation of recombinant influenza virus-simian immunodeficiency virus vaccines in macaques.

Science.gov (United States)

Sexton, Amy; De Rose, Robert; Reece, Jeanette C; Alcantara, Sheilajen; Loh, Liyen; Moffat, Jessica M; Laurie, Karen; Hurt, Aeron; Doherty, Peter C; Turner, Stephen J; Kent, Stephen J; Stambas, John

2009-08-01

There is an urgent need for human immunodeficiency virus (HIV) vaccines that induce robust mucosal immunity. Influenza A viruses (both H1N1 and H3N2) were engineered to express simian immunodeficiency virus (SIV) CD8 T-cell epitopes and evaluated following administration to the respiratory tracts of 11 pigtail macaques. Influenza virus was readily detected from respiratory tract secretions, although the infections were asymptomatic. Animals seroconverted to influenza virus and generated CD8 and CD4 T-cell responses to influenza virus proteins. SIV-specific CD8 T-cell responses bearing the mucosal homing marker beta7 integrin were induced by vaccination of naïve animals. Further, SIV-specific CD8 T-cell responses could be boosted by recombinant influenza virus-SIV vaccination of animals with already-established SIV infection. Sequential vaccination with influenza virus-SIV recombinants of different subtypes (H1N1 followed by H3N2 or vice versa) produced only a limited boost in immunity, probably reflecting T-cell immunity to conserved internal proteins of influenza A virus. SIV challenge of macaques vaccinated with an influenza virus expressing a single SIV CD8 T cell resulted in a large anamnestic recall CD8 T-cell response, but immune escape rapidly ensued and there was no impact on chronic SIV viremia. Although our results suggest that influenza virus-HIV vaccines hold promise for the induction of mucosal immunity to HIV, broader antigen cover will be needed to limit cytotoxic T-lymphocyte escape.

Schmallenberg virus – et nyt virus hos drøvtyggere

DEFF Research Database (Denmark)

Lohse, Louise

undersøgelser på laboratoriet, blev alle prøver i en længere periode testet negative for kendte patogener, der evt. kunne tænkes at knytte sig til disse sygdomsudbrud. Efterfølgende fandt man ved sekvensanalyse på DNA/RNA oprenset fra prøvematerialet lighed med virus fra orthobunyavirusgruppen. Disse virus er...... normalt vektorbårne og er kendt som årsag til sygdom i drøvtyggere i Australien, Asien, Afrika og Mellemøsten. Det nye virus blev kaldt Schmallenberg virus (SBV) efter stedet hvor det første tilfælde blev påvist. Siden har Schmallenberg virus spredt sig og er nu påvist i får, kvæg og geder i Tyskland......, Holland, Belgien, Luxembourg, Frankrig, Spanien, Italien og Storbritannien (status april 2012). Det antages, at virus overføres med mitter og myg og inkubationstiden har fra eksperimentelle forsøg vist sig at være kort (1-4 dage). Virus cirkulerer få dage i blodet og manifesterer sig ved milde uspecifikke...

Sensitive radioimmunosorbent assay for the detection of plant viruses. [Cauliflower mosaic virus, lettuce mosaic virus

Energy Technology Data Exchange (ETDEWEB)

Ghabrial, S A; Shepherd, R J [Kentucky Univ., Lexington (USA); California Univ., Davis (USA))

1980-06-01

A simple and highly sensitive radioimmunosorbent assay (RISA) for the detection of plant viruses is described. The RISA procedure is a microplate method based on the principle of 'double-antibody sandwich' and follows essentially the protocol of the enzyme-linked immunosorbent assay (ELISA) (Clark and Adams, 1977), with the exception that /sup 125/I-labelled ..gamma..-globulin is substituted for the ..gamma..-globulin enzyme conjugate; the bound /sup 125/I-..gamma..-globulin is dissociated by acidification from the double-antibody sandwich. The radioactivity is proportional to virus concentration, and cauliflower mosaic virus (CaMV) and lettuce mosaic virus (LMV) could be detected at concentrations as low as 5 and 2 ng/ml, respectively. Direct evidence of the adverse effects of conjugation with enzyme on the binding abilities of antibodies is presented. The RISA procedure should prove valuable with viruses for which the ELISA values are too low to be dependable.

Inhibition of CRM1-mediated nuclear export of influenza A nucleoprotein and nuclear export protein as a novel target for antiviral drug development.

Science.gov (United States)

Chutiwitoonchai, Nopporn; Mano, Takafumi; Kakisaka, Michinori; Sato, Hirotaka; Kondoh, Yasumitsu; Osada, Hiroyuki; Kotani, Osamu; Yokoyama, Masaru; Sato, Hironori; Aida, Yoko

2017-07-01

An anti-influenza compound, DP2392-E10 based on inhibition of the nuclear export function of the viral nucleoprotein-nuclear export signal 3 (NP-NES3) domain was successfully identified by our previous high-throughput screening system. Here, we demonstrated that DP2392-E10 exerts its antiviral effect by inhibiting replication of a broad range of influenza A subtypes. In regard to the molecular mechanism, we revealed that DP2392-E10 inhibits nuclear export of both viral NP and nuclear export protein (NEP). More specifically, in vitro pull-down assays revealed that DP2392-E10 directly binds cellular CRM1, which mediates nuclear export of NP and NEP. In silico docking suggested that DP2392-E10 binds at a region close to the HEAT9 and HEAT10 domains of CRM1. Together, these results indicate that the CRM1-mediated nuclear export function of influenza virus represents a new potential target for antiviral drug development, and also provide a core structure for a novel class of inhibitors that target this function. Copyright © 2017 Elsevier Inc. All rights reserved.

Epstein-Barr virus-derived EBNA2 regulates STAT3 activation

International Nuclear Information System (INIS)

Muromoto, Ryuta; Ikeda, Osamu; Okabe, Kanako; Togi, Sumihito; Kamitani, Shinya; Fujimuro, Masahiro; Harada, Shizuko; Oritani, Kenji; Matsuda, Tadashi

2009-01-01

The Epstein-Barr virus (EBV)-encoded latency protein EBNA2 is a nuclear transcriptional activator that is essential for EBV-induced cellular transformation. Here, we show that EBNA2 interacts with STAT3, a signal transducer for an interleukin-6 family cytokine, and enhances the transcriptional activity of STAT3 by influencing its DNA-binding activity. Furthermore, EBNA2 cooperatively acts on STAT3 activation with LMP1. These data demonstrate that EBNA2 acts as a transcriptional coactivator of STAT3.

Nuclear Cyber Security Issues and Policy Recommendations

Energy Technology Data Exchange (ETDEWEB)

Lee, Cheol-Kwon; Lee, Dong-Young [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of); Lee, Na-Young; Hwang, Young-Soo [Korea Institute of Nuclear Nonproliferation and Control, Daejeon (Korea, Republic of)

2015-10-15

The cyber-attack against computer systems causes the loss of function which brings about the big economic loss, and it becomes a national-wide issue. In recent days the cyber threat has occurred in the national critical infrastructure around the world. In the nuclear industry, while discussing responses to various threats against nuclear facilities since 2006, cyber-terrorism was also discussed. But at that time, cyber-attacks against control networks in nuclear facilities were not seriously considered because those networks were isolated from the Internet thoroughly and it was evaluated that cyber penetration would not be possible. However Stuxnet worm virus which attacked Iran's nuclear facilities confirmed that the cyber security problem could occur even in other nuclear facilities. The facilities were isolated from the Internet. After the cyber incident, we began to discuss the topic of NPP cyber security. It is very difficult to predict whether or when or how the cyber-attack will be occurred, which is a characteristic of cyber-attack. They could be always detected only after when an incident had occurred. This paper summarizes the report, 'Nuclear Cyber Security Issues and Policy Recommendations' by issue committee in the Korea Nuclear Society, which reviewed the cyber security framework for nuclear facilities in the Republic of Korea being established to prevent nuclear facilities from cyber-attacks and to respond systematically. As a result this paper proposes several comments to improve the security and furthermore safety of nuclear facilities Digital technology will be used more widely at the national critical infrastructure including nuclear facilities in the future, and moreover wireless technologies and mobile devices will be soon introduced to nuclear industry. It is therefore anticipated that the rapid advance in digital technology will accelerate the opportunity of hacking these facilities.

Nuclear Cyber Security Issues and Policy Recommendations

International Nuclear Information System (INIS)

Lee, Cheol-Kwon; Lee, Dong-Young; Lee, Na-Young; Hwang, Young-Soo

2015-01-01

The cyber-attack against computer systems causes the loss of function which brings about the big economic loss, and it becomes a national-wide issue. In recent days the cyber threat has occurred in the national critical infrastructure around the world. In the nuclear industry, while discussing responses to various threats against nuclear facilities since 2006, cyber-terrorism was also discussed. But at that time, cyber-attacks against control networks in nuclear facilities were not seriously considered because those networks were isolated from the Internet thoroughly and it was evaluated that cyber penetration would not be possible. However Stuxnet worm virus which attacked Iran's nuclear facilities confirmed that the cyber security problem could occur even in other nuclear facilities. The facilities were isolated from the Internet. After the cyber incident, we began to discuss the topic of NPP cyber security. It is very difficult to predict whether or when or how the cyber-attack will be occurred, which is a characteristic of cyber-attack. They could be always detected only after when an incident had occurred. This paper summarizes the report, 'Nuclear Cyber Security Issues and Policy Recommendations' by issue committee in the Korea Nuclear Society, which reviewed the cyber security framework for nuclear facilities in the Republic of Korea being established to prevent nuclear facilities from cyber-attacks and to respond systematically. As a result this paper proposes several comments to improve the security and furthermore safety of nuclear facilities Digital technology will be used more widely at the national critical infrastructure including nuclear facilities in the future, and moreover wireless technologies and mobile devices will be soon introduced to nuclear industry. It is therefore anticipated that the rapid advance in digital technology will accelerate the opportunity of hacking these facilities

Release of Virus from Lymphoid Tissue Affects Human Immunodeficiency Virus Type 1 and Hepatitis C Virus Kinetics in the Blood

NARCIS (Netherlands)

Müller, Viktor; Marée, Athanasius F.M.; Boer, R.J. de

2000-01-01

Kinetic parameters of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) infections have been estimated from plasma virus levels following perturbation of the chronically infected (quasi-) steady state. We extend previous models by also considering the large pool of virus

The metabolic activator FOXO1 binds hepatitis B virus DNA and activates its transcription

International Nuclear Information System (INIS)

Shlomai, Amir; Shaul, Yosef

2009-01-01

Hepatitis B virus (HBV) is a small DNA virus that targets the liver and infects humans worldwide. Recently we have shown that the metabolic regulator PGC-1α coactivates HBV transcription thereby rendering the virus susceptible to fluctuations in the nutritional status of the liver. PGC-1α coactivation of HBV is mediated through the liver-enriched nuclear receptor HNF4α and through another yet unknown transcription factor(s). Here we show that the forkhead transcription factor FOXO1, a known target for PGC-1α coactivation and a central mediator of glucose metabolism in the liver, binds HBV core promoter and activates its transcription. This activation is further enhanced in the presence of PGC-1α, implying that FOXO1 is a target for PGC-1α coactivation of HBV transcription. Thus, our results identify another key metabolic regulator as an activator of HBV transcription, thereby supporting the principle that HBV gene expression is regulated in a similar way to key hepatic metabolic genes.

Hepatitis E Virus and Related Viruses in Animals.

Science.gov (United States)

Thiry, D; Mauroy, A; Pavio, N; Purdy, M A; Rose, N; Thiry, E; de Oliveira-Filho, E F

2017-02-01

Hepatitis E is an acute human liver disease in healthy individuals which may eventually become chronic. It is caused by the hepatitis E virus (HEV) and can have a zoonotic origin. Nearly 57,000 people die yearly from hepatitis E-related conditions. The disease is endemic in both developing and developed countries with distinct epidemiologic profiles. In developing countries, the disease is associated with inadequate water treatment, while in developed countries, transmission is associated with animal contact and the ingestion of raw or uncooked meat, especially liver. All human HEV are grouped into at least four genotypes, while HEV or HEV-related viruses have been identified in an increasing number of domestic and wild animal species. Despite a high genetic diversity, only one single HEV serotype has been described to date for HEV genotypes 1-4. The discovery of new HEV or HEV-related viruses leads to a continuing increase in the number of genotypes. In addition, the genome organization of all these viruses is variable with overlapping open reading frames (ORF) and differences in the location of ORF3. In spite of the role of some domestic and wild animals as reservoir, the origin of HEV and HEV-related viruses in humans and animals is still unclear. This review discusses aspects of the detection, molecular virology, zoonotic transmission and origin of HEV and HEV-related viruses in the context of 'One Health' and establishes a link between the previous and the new taxonomy of this growing virus family. © 2015 Blackwell Verlag GmbH.

"The evil virus cell": Students' knowledge and beliefs about viruses.

Science.gov (United States)

Simon, Uwe K; Enzinger, Sonja M; Fink, Andreas

2017-01-01

Education about virus biology at school is of pivotal interest to raise public awareness concerning means of disease transmission and, thus, methods to prevent infection, and to reduce unnecessary antibiotic treatment due to patient pressure on physicians in case of viral diseases such as influenza. This study aimed at making visible the knowledge of Austrian high school and university students with respect to virus biology, virus structure and health-education issues. The data presented here stem from comprehensive questionnaire analyses, including the task to draw a virus, from a cross-sectional study with 133 grade 7 and 199 grade 10 high school students, and 133 first-year biology and 181 first-year non-biology university students. Analyses were performed both quantitatively and qualitatively. ANOVA revealed a highly significant group effect for total knowledge relating to virus biology and health issues (F(3, 642) = 44.17, p students and grade 10 high school students. Students enrolled in university-level biology outperformed all other groups, even though they had not yet encountered this topic at their courses; part of this phenomenon might be due to their affinity for learning about biological topics. However, even many first-year biology students had a high number of severe misconceptions, e.g., defining a virus as a pro- or eukaryotic cell, or falsely naming malaria as a viral disease. Since there was no significant difference in virus-related knowledge between high schools, virus biology seems to have been taught similarly among the tested schools. However, the majority of participants stated that the virus-related knowledge they had acquired at school was not sufficient. Based on the results presented here we urgently suggest improving and intensifying teaching this topic at school, since virus-related knowledge was by far too fragmentary among many participants. Such lack of health-relevant knowledge may contribute to pressure on physicians by patients

Region of Nipah virus C protein responsible for shuttling between the cytoplasm and nucleus

Energy Technology Data Exchange (ETDEWEB)

Horie, Ryo; Yoneda, Misako, E-mail: yone@ims.u-tokyo.ac.jp; Uchida, Shotaro; Sato, Hiroki; Kai, Chieko

2016-10-15

Nipah virus (NiV) causes severe encephalitis in humans, with high mortality. NiV nonstructural C protein (NiV-C) is essential for its pathogenicity, but its functions are unclear. In this study, we focused on NiV-C trafficking in cells and found that it localizes predominantly in the cytoplasm but partly in the nucleus. An analysis of NiV-C mutants showed that amino acids 2, 21–24 and 110–139 of NiV-C are important for its localization in the cytoplasm. Inhibitor treatment indicates that the nuclear export determinant is not a classical CRM1-dependent nuclear export signal. We also determined that amino acids 60–75 and 72–75 were important for nuclear localization of NiV-C. Furthermore, NiV-C mutants that had lost their capacity for nuclear localization inhibited the interferon (IFN) response more strongly than complete NiV-C. These results indicate that the IFN-antagonist activity of NiV-C occurs in the cytoplasm. -- Highlights: •Nipah virus (NiV) infection resulted in high mortality, but effective treatment has not been established. •Several reports revealed that NiV nonstructural C protein (NiV-C) was essential for NiV pathogenicity, however, whole of NiV-C function is still unknown. •Although nonstructural C proteins of other Paramyxoviruses are expressed in similar mechanism and exert similar activity, subcellular localization and cellular targets are different. In this study, we evaluated the subcellular localization of NiV-C. •To our knowledge, this is the first report showing that NiV-C shuttles between the nucleus and cytoplasm. We also clarified that NiV-C has nuclear export signal and nuclear localization signal using NiV-C deleted, alanine substitution mutants and enhanced green fluorescent protein (EGFP) fused proteins. •And we also showed that interferon (IFN) antagonist activity of NiV-C related to its subcellular localization. Our results indicate that NiV-C exert IFN antagonist activity in the cytoplasm.

Herpes viruses and human papilloma virus in nasal polyposis and controls.

Science.gov (United States)

Ioannidis, Dimitrios; Lachanas, Vasileios A; Florou, Zoe; Bizakis, John G; Petinaki, Efthymia; Skoulakis, Charalampos E

2015-01-01

Chronic rhinosinusitis with nasal polyps is a multifactorial disease entity with an unclear pathogenesis. Contradictory data exist in the literature on the potential implication of viral elements in adult patients with chronic rhinosinusitis. To compare the prevalence of human herpes viruses (1-6) and Human Papilloma Virus in adult patients with chronic rhinosinusitis with nasal polyps and healthy controls. Viral DNA presence was evaluated by real-time polymerase chain reaction application to nasal polyps specimens from 91 chronic rhinosinusitis with nasal polyps patients and nasal turbinate mucosa from 38 healthy controls. Epstein-Barr virus positivity was higher in nasal polyps (24/91; 26.4%) versus controls (4/38; 10.5%), but the difference did not reach significance (p=0.06). Human herpes virus-6 positivity was lower in nasal polyps (13/91; 14.29%) versus controls (10/38; 26.32%, p=0.13). In chronic rhinosinusitis with nasal polyps group, 1 sample was herpes simplex virus-1-positive (1/91; 1.1%), and another was cytomegalovirus-positive (1/91; 1.1%), versus none in controls. No sample was positive for herpes simplex virus-2, varicella-zoster virus, high-risk-human papilloma viruses (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59) and low-risk-human papilloma viruses (6, 11). Differences in Epstein-Barr virus and human herpes virus-6 positivity among patients with chronic rhinosinusitis with nasal polyps and healthy controls are not statistically significant, weakening the likelihood of their implication in chronic rhinosinusitis with nasal polyps pathogenesis. Copyright © 2015 Associação Brasileira de Otorrinolaringologia e Cirurgia Cérvico-Facial. Published by Elsevier Editora Ltda. All rights reserved.

Computer Virus and Trends

OpenAIRE

Tutut Handayani; Soenarto Usna,Drs.MMSI

2004-01-01

Since its appearance the first time in the mid-1980s, computer virus has invited various controversies that still lasts to this day. Along with the development of computer systems technology, viruses komputerpun find new ways to spread itself through a variety of existing communications media. This paper discusses about some things related to computer viruses, namely: the definition and history of computer viruses; the basics of computer viruses; state of computer viruses at this time; and ...

Viruses in reptiles

Directory of Open Access Journals (Sweden)

Ariel Ellen

2011-09-01

Full Text Available Abstract 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

Genomic characterisation of Almpiwar virus, Harrison Dam virus and Walkabout Creek virus; three novel rhabdoviruses from northern Australia

Directory of Open Access Journals (Sweden)

Jane McAllister

2014-09-01

Full Text Available Rhabdoviridae represent a diverse group of viruses with the potential to cause disease in humans, animals and plants. Currently there are nine genera in the family; however a large number of rhabdoviruses remain unassigned. Here we characterise three novel rhabdoviruses genomes. Almpiwar virus (ALMV, isolated from skinks in northern Queensland, is the first completely sequenced rhabdovirus from squamates, with serological studies indicating multiple animal host species. Harrison Dam virus (HARDV and Walkabout Creek virus (WACV were isolated from mosquitoes in the Northern Territory and biting midges in southern Queensland respectively and their vertebrate hosts remain unknown. Serological cross-neutralisation tests with other Australian rhabdoviruses indicate that ALMV, WACV and HARDV are distinct viruses with little antigenic cross-reactivity. Next-generation sequencing revealed that all viruses encode the core proteins common to rhabdoviruses (N, P, M, G and L, plus additional ORFs between the M and G genes. HARDV also contains a small ORF between the G and L genes. Phylogenetic analysis of N and L proteins suggests that HARDV and WACV share a common lineage with the tupaviruses and Sandjimba group, whereas ALMV is a distinct and divergent virus showing no clear relationship to any rhabdovirus except the recently characterised Niahka virus (NIAV.

A conserved RNA structural element within the hepatitis B virus post-transcriptional regulatory element enhance nuclear export of intronless transcripts and repress the splicing mechanism.

Science.gov (United States)

Visootsat, Akasit; Payungporn, Sunchai; T-Thienprasert, Nattanan P

2015-12-01

Hepatitis B virus (HBV) infection is a primary cause of hepatocellular carcinoma and liver cirrhosis worldwide. To develop novel antiviral drugs, a better understanding of HBV gene expression regulation is vital. One important aspect is to understand how HBV hijacks the cellular machinery to export unspliced RNA from the nucleus. The HBV post-transcriptional regulatory element (HBV PRE) has been proposed to be the HBV RNA nuclear export element. However, the function remains controversial, and the core element is unclear. This study, therefore, aimed to identify functional regulatory elements within the HBV PRE and investigate their functions. Using bioinformatics programs based on sequence conservation and conserved RNA secondary structures, three regulatory elements were predicted, namely PRE 1151-1410, PRE 1520-1620 and PRE 1650-1684. PRE 1151-1410 significantly increased intronless and unspliced luciferase activity in both HepG2 and COS-7 cells. Likewise, PRE 1151-1410 significantly elevated intronless and unspliced HBV surface transcripts in liver cancer cells. Moreover, motif analysis predicted that PRE 1151-1410 contains several regulatory motifs. This study reported the roles of PRE 1151-1410 in intronless transcript nuclear export and the splicing mechanism. Additionally, these results provide knowledge in the field of HBV RNA regulation. Moreover, PRE 1151-1410 may be used to enhance the expression of other mRNAs in intronless reporter plasmids.

Block effect on HCV infection by HMGB1 released from virus-infected cells: An insight from mathematical modeling

Science.gov (United States)

Wang, Wei; Ma, Wanbiao

2018-06-01

The nuclear protein high-mobility group box 1 (HMGB1) can have an active role in deoxyribonucleic acid (DNA) organization and the regulation of transcription. Based on the new findings from a recent experimental study, the blocking effect on HCV infection by HMGB1 released from virus-infected cells is investigated using a diffusive model for viral infection dynamics. In the model, the diffusion of the virus depends not only on its concentration gradient, but also on the concentration of HMGB1. The basic reproduction number, threshold dynamics, stability properties of the steady states, travelling wave solutions, and spreading speed for the proposed model are studied. We show that the HMGB1-induced blocking of HCV infection slows the spread of virus compared with random diffusion only. Numerically, it is shown that a high concentration of HMGB1 can block the spread of virus and this confirms, not only qualitatively but also quantitatively, the experimental result.

Powassan (POW) Virus Basics

Science.gov (United States)

... Health Professionals Related Topics For International Travelers Powassan Virus Disease Basics Download this fact sheet formatted for ... Virus Disease Fact Sheet (PDF) What is Powassan virus? Powassan virus is a tickborne flavivirus that is ...

Initial characterization of Vaccinia Virus B4 suggests a role in virus spread

Energy Technology Data Exchange (ETDEWEB)

Burles, Kristin; Irwin, Chad R.; Burton, Robyn-Lee [Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada T6G 2S2 (Canada); Schriewer, Jill [Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, MO (United States); Evans, David H. [Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada T6G 2S2 (Canada); Buller, R. Mark [Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, MO (United States); Barry, Michele, E-mail: michele.barry@ualberta.ca [Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada T6G 2S2 (Canada)

2014-05-15

Currently, little is known about the ankyrin/F-box protein B4. Here, we report that B4R-null viruses exhibited reduced plaque size in tissue culture, and decreased ability to spread, as assessed by multiple-step growth analysis. Electron microscopy indicated that B4R-null viruses still formed mature and extracellular virions; however, there was a slight decrease of virions released into the media following deletion of B4R. Deletion of B4R did not affect the ability of the virus to rearrange actin; however, VACV811, a large vaccinia virus deletion mutant missing 55 open reading frames, had decreased ability to produce actin tails. Using ectromelia virus, a natural mouse pathogen, we demonstrated that virus devoid of EVM154, the B4R homolog, showed decreased spread to organs and was attenuated during infection. This initial characterization suggests that B4 may play a role in virus spread, and that other unidentified mediators of actin tail formation may exist in vaccinia virus. - Highlights: • B4R-null viruses show reduced plaque size, and decreased ability to spread. • B4R-null viruses formed mature and extracellular virions; and rearranged actin. • Virus devoid of EVM154, the B4R homolog, was attenuated during infection. • Initial characterization suggests that B4 may play a role in virus spread. • Unidentified mediators of actin tail formation may exist in vaccinia virus.

Initial characterization of Vaccinia Virus B4 suggests a role in virus spread

International Nuclear Information System (INIS)

Burles, Kristin; Irwin, Chad R.; Burton, Robyn-Lee; Schriewer, Jill; Evans, David H.; Buller, R. Mark; Barry, Michele

2014-01-01

Currently, little is known about the ankyrin/F-box protein B4. Here, we report that B4R-null viruses exhibited reduced plaque size in tissue culture, and decreased ability to spread, as assessed by multiple-step growth analysis. Electron microscopy indicated that B4R-null viruses still formed mature and extracellular virions; however, there was a slight decrease of virions released into the media following deletion of B4R. Deletion of B4R did not affect the ability of the virus to rearrange actin; however, VACV811, a large vaccinia virus deletion mutant missing 55 open reading frames, had decreased ability to produce actin tails. Using ectromelia virus, a natural mouse pathogen, we demonstrated that virus devoid of EVM154, the B4R homolog, showed decreased spread to organs and was attenuated during infection. This initial characterization suggests that B4 may play a role in virus spread, and that other unidentified mediators of actin tail formation may exist in vaccinia virus. - Highlights: • B4R-null viruses show reduced plaque size, and decreased ability to spread. • B4R-null viruses formed mature and extracellular virions; and rearranged actin. • Virus devoid of EVM154, the B4R homolog, was attenuated during infection. • Initial characterization suggests that B4 may play a role in virus spread. • Unidentified mediators of actin tail formation may exist in vaccinia virus

How Hepatitis D Virus Can Hinder the Control of Hepatitis B Virus

NARCIS (Netherlands)

Xiridiou, M.; Borkent-Raven, B.; Hulshof, J.; Wallinga, J.

2009-01-01

Background: Hepatitis D (or hepatitis delta) virus is a defective virus that relies on hepatitis B virus (HBV) for transmission; infection with hepatitis D can occur only as coinfection with HBV or superinfection of an existing HBV infection. Because of the bond between the two viruses, control

9 CFR 113.215 - Bovine Virus Diarrhea Vaccine, Killed Virus.

Science.gov (United States)

2010-01-01

... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Bovine Virus Diarrhea Vaccine, Killed Virus. 113.215 Section 113.215 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD...

Influenza (Flu) Viruses

Science.gov (United States)

... Types Seasonal Avian Swine Variant Pandemic Other Influenza (Flu) Viruses Language: English (US) Español Recommend on Facebook ... influenza circulate and cause illness. More Information about Flu Viruses Types of Influenza Viruses Influenza A and ...

Quantitative Epstein-Barr virus (EBV) serology in lung transplant recipients with primary EBV infection and/or post-transplant lymphoproliferative disease.

NARCIS (Netherlands)

Verschuuren, E; van der Bij, W; Boer, W.; Timens, W.; Middeldorp, J.M.; The, T.H.

2003-01-01

The Epstein-Barr virus (EBV)-specific antibody response was studied in lung transplant patients to assess their value in the diagnosis and prognosis of post-transplant lymphoproliferative disease. Recently developed synthetic peptides representing Epstein-Barr nuclear antigen-1 (EBNA-1), diffuse

Quantitative Epstein-Barr virus (EBV) serology in lung transplant recipients with primary EBV infection and/or post-transplant lymphoproliferative disease

NARCIS (Netherlands)

Verschuuren, E; van der Bij, W; de Boer, W; Timens, W; Middeldorp, J; The, TH

The Epstein-Barr virus (EBV)-specific antibody response was studied in lung transplant patients to assess their value in the diagnosis and prognosis of post-transplant lymphoproliferative disease. Recently developed synthetic peptides representing Epstein-Barr nuclear antigen-1 (EBNA-1), diffuse

[The growth of attenuated strains of canine parvovirus, mink enteritis virus, feline panleukopenia virus, and rabies virus on various types of cell cultures].

Science.gov (United States)

Zuffa, T

1987-10-01

The growth characteristics were studied in the attenuated strains of canine parvovirus CPVA-BN 80/82, mink enteritis virus MEVA-BN 63/82 and feline panleucopenia virus FPVA-BN 110/83 on the stable feline kidney cell line FE, and in the attenuated canine distemper virus CDV-F-BN 10/83 on chicken embryo cell cultures (KEB) and cultures of the stable cell line VERO. When the FE cultures were infected with different parvoviruses in cell suspension at MOI 2-4 TKID50 per cell, the first multiplication of the intracellular virus was recorded 20 hours p. i. In the canine parvovirus, the content of intracellular and extracellular virus continued increasing parallelly until the fourth day; then, from the fourth to the sixth day, the content of extracellular virus still increased whereas that of intracellular virus fell rapidly. In the case of the mink enteritis virus the release of the virus into the culture medium continued parallelly with the production of the cellular virus until the sixth day. In the case of the feline panleucopenia virus the values concerning free virus and virus bound to cells were lower, starting from the second day p. i. When KEB or VERO cultures were infected in cell suspension with the canine distemper virus at MOI about 0.004 per 1 cell, the replicated intracellular virus was first recorded in the KEB cultures five hours after infection but in the VERO cultures only 20 hours after infection, with a timely release of the virus into the culture medium in both kinds of tissue. In the KEB and VERO cultures the highest values of infection titres were recorded on the fourth day p. i., the course of virus multiplication on the cells being parallel with its release into the culture medium.

Viruses of asparagus.

Science.gov (United States)

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. Copyright © 2012 Elsevier Inc. All rights reserved.

Viruses and Breast Cancer

Science.gov (United States)

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

Viruses and Breast Cancer

International Nuclear Information System (INIS)

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

Viruses and Breast Cancer

Energy Technology Data Exchange (ETDEWEB)

Lawson, James S., E-mail: james.lawson@unsw.edu.au; Heng, Benjamin [School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney (Australia)

2010-04-30

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.

"The evil virus cell": Students‘ knowledge and beliefs about viruses

Science.gov (United States)

Enzinger, Sonja M.; Fink, Andreas

2017-01-01

Education about virus biology at school is of pivotal interest to raise public awareness concerning means of disease transmission and, thus, methods to prevent infection, and to reduce unnecessary antibiotic treatment due to patient pressure on physicians in case of viral diseases such as influenza. This study aimed at making visible the knowledge of Austrian high school and university students with respect to virus biology, virus structure and health-education issues. The data presented here stem from comprehensive questionnaire analyses, including the task to draw a virus, from a cross-sectional study with 133 grade 7 and 199 grade 10 high school students, and 133 first-year biology and 181 first-year non-biology university students. Analyses were performed both quantitatively and qualitatively. ANOVA revealed a highly significant group effect for total knowledge relating to virus biology and health issues (F(3, 642) = 44.17, p students and grade 10 high school students. Students enrolled in university-level biology outperformed all other groups, even though they had not yet encountered this topic at their courses; part of this phenomenon might be due to their affinity for learning about biological topics. However, even many first-year biology students had a high number of severe misconceptions, e.g., defining a virus as a pro- or eukaryotic cell, or falsely naming malaria as a viral disease. Since there was no significant difference in virus-related knowledge between high schools, virus biology seems to have been taught similarly among the tested schools. However, the majority of participants stated that the virus-related knowledge they had acquired at school was not sufficient. Based on the results presented here we urgently suggest improving and intensifying teaching this topic at school, since virus-related knowledge was by far too fragmentary among many participants. Such lack of health-relevant knowledge may contribute to pressure on physicians by patients

Influenza virus inactivated by artificial ribonucleases as a prospective killed virus vaccine.

Science.gov (United States)

Fedorova, Antonina A; Goncharova, Elena P; Kovpak, Mikhail P; Vlassov, Valentin V; Zenkova, Marina A

2012-04-19

The inactivation of viral particles with agents causing minimal damage to the structure of surface epitopes is a well-established approach for the production of killed virus vaccines. Here, we describe new agents for the inactivation of influenza virus, artificial ribonucleases (aRNases), which are chemical compounds capable of cleaving RNA molecules. Several aRNases were identified, exhibiting significant virucidal activity against the influenza A virus and causing a minimal effect on the affinity of monoclonal antibodies for the inactivated virus. Using a murine model of the influenza virus infection, a high protective activity of the aRNase-inactivated virus as a vaccine was demonstrated. The results of the experiments demonstrate the efficacy of novel chemical agents in the preparation of vaccines against influenza and, perhaps, against other infections caused by RNA viruses. Copyright © 2012 Elsevier Ltd. All rights reserved.

[The great virus comeback].

Science.gov (United States)

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

Protoplasts and plant viruses

International Nuclear Information System (INIS)

Murakishi, H.; Lesney, M.S.; Carlson, P.

1984-01-01

The use of protoplasts in the study of plant viruses has attracted considerable attention since its inception in the late 1960s. This article is an attempt to assess the current status of protoplasts (primarily) and all cell cultures (in some instances) in studies of virus infection, virus replication, cytopathology, cross-protection, virus resistance, and the use of in vitro methods and genetic engineering to recover virus-resistant plants. These areas of study proved difficult to do entirely with whole plants or plant parts. However, because protoplasts could be synchronously infected with virus, they provided a valuable alternative means of following biochemical and cytological events in relation to the virus growth cycle in a more precise manner than previously possible

Hepatitis A Virus and Hepatitis E Virus: Emerging and Re-Emerging Enterically Transmitted Hepatitis Viruses.

Science.gov (United States)

Lemon, Stanley M; Walker, Christopher M

2018-05-07

Over the past two decades, progress in understanding human infections with hepatitis A virus (HAV) and hepatitis E virus (HEV) has been eclipsed by the priority of combating persistent hepatitis B virus (HBV) and hepatitis C virus (HCV) infections. During that time, the global burden of liver disease caused by enteric hepatitis viruses has not abated. Because of vaccines, hepatitis A has become increasingly a disease of adults instead of early childhood in many regions of the world, resulting in an age-related shift toward more severe disease. HEV has remained endemic in many developing countries, and in well-developed, economically advanced countries it is now recognized as a cause of chronic, progressive liver disease in individuals with compromised immunity. The goal of this collection of articles is to review recent progress and to shine a bright light on gaps in our understanding of how these viruses replicate, cause disease, interact with the liver and host immune system, and are transmitted, along with prospects for improved control in human populations. Renewed efforts to study and compare HAV and HEV biology in humans and animal models have high potential to enhance our understanding of host-pathogen balance in the liver, and may contribute ultimately to the control of other infectious diseases of the liver. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

Evolutionary ecology of virus emergence.

Science.gov (United States)

Dennehy, John J

2017-02-01

The cross-species transmission of viruses into new host populations, termed virus emergence, is a significant issue in public health, agriculture, wildlife management, and related fields. Virus emergence requires overlap between host populations, alterations in virus genetics to permit infection of new hosts, and adaptation to novel hosts such that between-host transmission is sustainable, all of which are the purview of the fields of ecology and evolution. A firm understanding of the ecology of viruses and how they evolve is required for understanding how and why viruses emerge. In this paper, I address the evolutionary mechanisms of virus emergence and how they relate to virus ecology. I argue that, while virus acquisition of the ability to infect new hosts is not difficult, limited evolutionary trajectories to sustained virus between-host transmission and the combined effects of mutational meltdown, bottlenecking, demographic stochasticity, density dependence, and genetic erosion in ecological sinks limit most emergence events to dead-end spillover infections. Despite the relative rarity of pandemic emerging viruses, the potential of viruses to search evolutionary space and find means to spread epidemically and the consequences of pandemic viruses that do emerge necessitate sustained attention to virus research, surveillance, prophylaxis, and treatment. © 2016 New York Academy of Sciences.

A virus of hyperthermophilic archaea with a unique architecture among DNA viruses.

Science.gov (United States)

Rensen, Elena Ilka; Mochizuki, Tomohiro; Quemin, Emmanuelle; Schouten, Stefan; Krupovic, Mart; Prangishvili, David

2016-03-01

Viruses package their genetic material in diverse ways. Most known strategies include encapsulation of nucleic acids into spherical or filamentous virions with icosahedral or helical symmetry, respectively. Filamentous viruses with dsDNA genomes are currently associated exclusively with Archaea. Here, we describe a filamentous hyperthermophilic archaeal virus, Pyrobaculum filamentous virus 1 (PFV1), with a type of virion organization not previously observed in DNA viruses. The PFV1 virion, 400 ± 20 × 32 ± 3 nm, contains an envelope and an inner core consisting of two structural units: a rod-shaped helical nucleocapsid formed of two 14-kDa major virion proteins and a nucleocapsid-encompassing protein sheath composed of a single major virion protein of 18 kDa. The virion organization of PFV1 is superficially similar to that of negative-sense RNA viruses of the family Filoviridae, including Ebola virus and Marburg virus. The linear dsDNA of PFV1 carries 17,714 bp, including 60-bp-long terminal inverted repeats, and contains 39 predicted ORFs, most of which do not show similarities to sequences in public databases. PFV1 is a lytic virus that completely disrupts the host cell membrane at the end of the infection cycle.

DIAGNOSTICS OF VIRUS PHYTOPATHOGENS FRUIT TREE PLUM POX VIRUS, PRUNUS NECROTIC RINGSPOT VIRUS AND PRUNUS DWARF VIRUS BY BIOLOGICAL AND MOLECULAR DIAGNOSTICS

OpenAIRE

Július Rozák; Zdenka Gálová

2013-01-01

The aim of this study was to determine the incidence of viral phytopathogen Plum pox virus, Prunus necrotic ringspot virus and Prunus dwarf virus in selected localities of Slovakia and diagnose them using a molecular and biological methods. Forty samples of fruit trees of the genus Prunus, twenty samples from intensive plantings and twenty samples from wild subject were analysed. Biological diagnostic by using biological indicators Prunus persica cv. GF 305, Prunus serrulata cv. Schirofugen a...

Contribution to case study session 1, PIME 89[Nuclear energy in Sweden

Energy Technology Data Exchange (ETDEWEB)

Sokolowski, Evelyn [KSU (Sweden)

1989-07-01

Analysing the arguments of the antinuclear movement in general, one is suprized to find that they are mostly of a pseudo-scientific nature. There is no hesitation to talk about 'half-life', 'kilowatts' or 'critical mass', let alone'Down's syndrome' or 'childhood leukemia'. Obviously, 'facts' are believed to impress more than ideological arguments. Judging from the success of the anti-nuclear movement, the hypothesis is correct. Information from nuclear industry lacks the doomsday note and drama that catches the attention of the media and the public. While the anti-nuclear message has the character of a virus, 'nucler' might instead be considered a vaccine - it does not cause much commotion when administered but has an extended protective effect. Most anti-nuclear arguments are variations of a few predictable themes: radiation, accidents, waste, proliferation, and economy. This could be considered to be in favour of 'nuclear'. It should be considred that there is frequent need for quick response.

The Antiviral Mechanism of an Influenza A Virus Nucleoprotein-Specific Single-Domain Antibody Fragment

Energy Technology Data Exchange (ETDEWEB)

Hanke, Leo; Knockenhauer, Kevin E.; Brewer, R. Camille; van Diest, Eline; Schmidt, Florian I.; Schwartz, Thomas U.; Ploegh, Hidde L. (Whitehead); (MIT)

2016-12-13

Alpaca-derived single-domain antibody fragments (VHHs) that target the influenza A virus nucleoprotein (NP) can protect cells from infection when expressed in the cytosol. We found that one such VHH, αNP-VHH1, exhibits antiviral activity similar to that of Mx proteins by blocking nuclear import of incoming viral ribonucleoproteins (vRNPs) and viral transcription and replication in the nucleus. We determined a 3.2-Å crystal structure of αNP-VHH1 in complex with influenza A virus NP. The VHH binds to a nonconserved region on the body domain of NP, which has been associated with binding to host factors and serves as a determinant of host range. Several of the NP/VHH interface residues determine sensitivity of NP to antiviral Mx GTPases. The structure of the NP/αNP-VHH1 complex affords a plausible explanation for the inhibitory properties of the VHH and suggests a rationale for the antiviral properties of Mx proteins. Such knowledge can be leveraged for much-needed novel antiviral strategies.

IMPORTANCEInfluenza virus strains can rapidly escape from protection afforded by seasonal vaccines or acquire resistance to available drugs. Additional ways to interfere with the virus life cycle are therefore urgently needed. The influenza virus nucleoprotein is one promising target for antiviral interventions. We have previously isolated alpaca-derived single-domain antibody fragments (VHHs) that protect cells from influenza virus infection if expressed intracellularly. We show here that one such VHH exhibits antiviral activities similar to those of proteins of the cellular antiviral defense (Mx proteins). We determined the three-dimensional structure of this VHH in complex with the influenza virus nucleoprotein and identified the interaction site, which overlaps regions that determine sensitivity of the virus to Mx proteins. Our data define a new vulnerability of influenza virus, help us to better understand the cellular antiviral mechanisms, and

Functional properties of Virus-Encoded and Virus-Regulated 7TM Receptors

DEFF Research Database (Denmark)

Spiess, Katja; Rosenkilde, Mette Marie

2014-01-01

During co-evolution with their hosts, viruses have developed several survival strategies that involve exploitation of 7TM receptors. These include virus-encoded 7TM receptors and ligands and viral regulation of endogenous receptors. Many functional properties have been ascribed to virus-exploited...

Operation technology of air treatment system in nuclear facilities

CERN Document Server

Chun, Y B; Hwong, Y H; Lee, H K; Min, D K; Park, K J; Uom, S H; Yang, S Y

2001-01-01

Effective operation techniques were reviewed on the air treatment system to protect the personnel in nuclear facilities from the contamination of radio-active particles and to keep the environment clear. Nuclear air treatment system consisted of the ventilation and filtering system was characterized by some test. Measurement of air velocity of blowing/exhaust fan in the ventilation system, leak tests of HEPA filters in the filtering, and measurement of pressure difference between the areas defined by radiation level were conducted. The results acquired form the measurements were reflected directly for the operation of air treatment. In the abnormal state of virus parts of devices composted of the system, the repairing method, maintenance and performance test were also employed in operating effectively the air treatment system. These measuring results and techniques can be available to the operation of air treatment system of PIEF as well as the other nuclear facilities in KAERI.

Electron microscopic identification of Zinga virus as a strain of Rift Valley fever virus.

Science.gov (United States)

Olaleye, O D; Baigent, C L; Mueller, G; Tomori, O; Schmitz, H

1992-01-01

Electron microscopic examination of a negatively stained suspension of Zinga virus showed particles 90-100 nm in diameter, enveloped with spikes 12-20 nm in length and 5 nm in diameter. Further identification of the virus by immune electron microscopy showed the reactivity of human Rift Valley fever virus-positive serum with Zinga virus. Results of this study are in agreement with earlier reports that Zinga virus is a strain of Rift Valley fever virus.

Nuclear medicine and AIDS

International Nuclear Information System (INIS)

O'Doherty, M.J.; Kent and Canterbury Hospital, Canterbury; Nunan, T.O.

1993-01-01

The human immunodeficiency virus (HIV) infection and its associated illnesses in a relatively young population of patients provides an expanding role for nuclear medicine. The disease enforces a review of each department's infection control procedures. It has also resulted in an increase in the number of patients presenting with diseases such as Pneumocystis carinii pneumonia, Kaposi's sarcoma etc. which prior to the HIV epidemic were extremely rare. Thus in high risk patients the interpretation of abnormalities in nuclear medicine scans needs to include the spectrum of opportunistic infections and unusual tumours. The presence of opportunistic infections in the severely immunocompromised patient has led to the development of techniques not normally used, i.e. lung 99 Tc m -diethylenetriamine pentaacetate (DTPA) transfer/clearance, donor leukocyte scanning to allow rapid diagnosis of an abnormality. Radionuclide techniques are also used to monitor the effect of therapy directed at the HIV itself or against opportunistic infections. This review covers aspects of infection control as well as the use of radionuclides to investigate specific problems related to HIV infection and therapy of the associated disease processes. (author)

Nonhuman Primate Models of Hepatitis A Virus and Hepatitis E Virus Infections.

Science.gov (United States)

Lanford, Robert E; Walker, Christopher M; Lemon, Stanley M

2018-04-23

Although phylogenetically unrelated, human hepatitis viruses share an exclusive or near exclusive tropism for replication in differentiated hepatocytes. This narrow tissue tropism may contribute to the restriction of the host ranges of these viruses to relatively few host species, mostly nonhuman primates. Nonhuman primate models thus figure prominently in our current understanding of the replication and pathogenesis of these viruses, including the enterically transmitted hepatitis A virus (HAV) and hepatitis E virus (HEV), and have also played major roles in vaccine development. This review draws comparisons of HAV and HEV infection from studies conducted in nonhuman primates, and describes how such studies have contributed to our current understanding of the biology of these viruses. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

Zika Virus

Science.gov (United States)

... Funding CDC Activities For Healthcare Providers Clinical Evaluation & Disease Sexual Transmission HIV Infection & Zika Virus Testing for Zika Test Specimens – At Time of Birth Diagnostic Tests Understanding Zika Virus Test Results ...

Apple latent spherical virus vectors for reliable and effective virus-induced gene silencing among a broad range of plants including tobacco, tomato, Arabidopsis thaliana, cucurbits, and legumes

International Nuclear Information System (INIS)

Igarashi, Aki; Yamagata, Kousuke; Sugai, Tomokazu; Takahashi, Yukari; Sugawara, Emiko; Tamura, Akihiro; Yaegashi, Hajime; Yamagishi, Noriko; Takahashi, Tsubasa; Isogai, Masamichi; Takahashi, Hideki; Yoshikawa, Nobuyuki

2009-01-01

Apple latent spherical virus (ALSV) vectors were evaluated for virus-induced gene silencing (VIGS) of endogenous genes among a broad range of plant species. ALSV vectors carrying partial sequences of a subunit of magnesium chelatase (SU) and phytoene desaturase (PDS) genes induced highly uniform knockout phenotypes typical of SU and PDS inhibition on model plants such as tobacco and Arabidopsis thaliana, and economically important crops such as tomato, legume, and cucurbit species. The silencing phenotypes persisted throughout plant growth in these plants. In addition, ALSV vectors could be successfully used to silence a meristem gene, proliferating cell nuclear antigen and disease resistant N gene in tobacco and RCY1 gene in A. thaliana. As ALSV infects most host plants symptomlessly and effectively induces stable VIGS for long periods, the ALSV vector is a valuable tool to determine the functions of interested genes among a broad range of plant species.

Hepatitis C virus infection in the human immunodeficiency virus infected patient

DEFF Research Database (Denmark)

Clausen, Louise Nygaard; Lundbo, Lene Fogt; Benfield, Thomas

2014-01-01

Human immunodeficiency virus (HIV) and hepatitis C virus (HCV) share the same transmission routes; therefore, coinfection is frequent. An estimated 5-10 million individuals alone in the western world are infected with both viruses. The majority of people acquire HCV by injection drug use and...

Immunological responses against human papilloma virus and human papilloma virus induced laryngeal cancer.

Science.gov (United States)

Chitose, Shun-ichi; Sakazaki, T; Ono, T; Kurita, T; Mihashi, H; Nakashima, T

2010-06-01

This study aimed to clarify the local immune status in the larynx in the presence of infection or carcinogenesis associated with human papilloma virus. Cytological samples (for human papilloma virus detection) and laryngeal secretions (for immunoglobulin assessment) were obtained from 31 patients with laryngeal disease, during microscopic laryngeal surgery. On histological examination, 12 patients had squamous cell carcinoma, four had laryngeal papilloma and 15 had other benign laryngeal disease. Cytological samples were tested for human papilloma virus DNA using the Hybrid Capture 2 assay. High risk human papilloma virus DNA was detected in 25 per cent of patients (three of 12) with laryngeal cancer. Low risk human papilloma virus DNA was detected only in three laryngeal papilloma patients. The mean laryngeal secretion concentrations of immunoglobulins M, G and A and secretory immunoglobulin A in human papilloma virus DNA positive patients were more than twice those in human papilloma virus DNA negative patients. A statistically significant difference was observed between the secretory immunoglobulin A concentrations in the two groups. Patients with laryngeal cancer had higher laryngeal secretion concentrations of each immunoglobulin type, compared with patients with benign laryngeal disease. The study assessed the mean laryngeal secretion concentrations of each immunoglobulin type in the 12 laryngeal cancer patients, comparing human papilloma virus DNA positive patients (n = 3) and human papilloma virus DNA negative patients (n = 9); the mean concentrations of immunoglobulins M, G and A and secretory immunoglobulin A tended to be greater in human papilloma virus DNA positive cancer patients, compared with human papilloma virus DNA negative cancer patients. These results suggest that the local laryngeal immune response is activated by infection or carcinogenesis due to human papilloma virus. The findings strongly suggest that secretory IgA has inhibitory activity

Autoantibodies in infectious mononucleosis have specificity for the glycine-alanine repeating region of the Epstein-Barr virus nuclear antigen

Science.gov (United States)

1987-01-01

Viruses have been postulated to be involved in the induction of autoantibodies by: autoimmunization with tissue proteins released by virally induced tissue damage; immunization with virally encoded antigens bearing molecular similarities to normal tissue proteins; or nonspecific (polyclonal) B cell stimulation by the infection. Infectious mononucleosis (IM) is an experiment of nature that provides the opportunity for examining these possibilities. We show here that IgM antibodies produced in this disease react with at least nine normal tissue proteins, in addition to the virally encoded Epstein-Barr nuclear antigen (EBNA-1). The antibodies are generated to configurations in the glycine-alanine repeat region of EBNA-1 and are crossreactive with the normal tissue proteins through similar configurations, as demonstrated by the effectiveness of a synthetic glycine-alanine peptide in inhibiting the reactions. The antibodies are absent in preillness sera and gradually disappear over a period of months after illness, being replaced by IgG anti-EBNA-1 antibodies that do not crossreact with the normal tissue proteins but that are still inhibited by the glycine-alanine peptide. These findings are most easily explained by either a molecular mimicry model of IgM autoantibody production or by the polyclonal activation of a germline gene for a crossreactive antibody. It also indicates a selection of highly specific, non-crossreactive anti-EBNA-1 antibodies during IgM to IgG isotype switching. PMID:2435830

Deep-Sea Hydrothermal Vent Viruses Compensate for Microbial Metabolism in Virus-Host Interactions.

Science.gov (United States)

He, Tianliang; Li, Hongyun; Zhang, Xiaobo

2017-07-11

Viruses are believed to be responsible for the mortality of host organisms. However, some recent investigations reveal that viruses may be essential for host survival. To date, it remains unclear whether viruses are beneficial or harmful to their hosts. To reveal the roles of viruses in the virus-host interactions, viromes and microbiomes of sediment samples from three deep-sea hydrothermal vents were explored in this study. To exclude the influence of exogenous DNAs on viromes, the virus particles were purified with nuclease (DNase I and RNase A) treatments and cesium chloride density gradient centrifugation. The metagenomic analysis of viromes without exogenous DNA contamination and microbiomes of vent samples indicated that viruses had compensation effects on the metabolisms of their host microorganisms. Viral genes not only participated in most of the microbial metabolic pathways but also formed branched pathways in microbial metabolisms, including pyrimidine metabolism; alanine, aspartate, and glutamate metabolism; nitrogen metabolism and assimilation pathways of the two-component system; selenocompound metabolism; aminoacyl-tRNA biosynthesis; and amino sugar and nucleotide sugar metabolism. As is well known, deep-sea hydrothermal vent ecosystems exist in relatively isolated environments which are barely influenced by other ecosystems. The metabolic compensation of hosts mediated by viruses might represent a very important aspect of virus-host interactions. IMPORTANCE Viruses are the most abundant biological entities in the oceans and have very important roles in regulating microbial community structure and biogeochemical cycles. The relationship between virus and host microbes is broadly thought to be that of predator and prey. Viruses can lyse host cells to control microbial population sizes and affect community structures of hosts by killing specific microbes. However, viruses also influence their hosts through manipulation of bacterial metabolism. We found

Viruses in reptiles.

Science.gov (United States)

Ariel, Ellen

2011-09-21

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.

A REVIEW ON ZIKA VIRUS (ZIKV) -A DREADFUL MEMBER OF THE VIRUS FAMILY FLAVIVIRIDAE

OpenAIRE

1Rafiya Begum, 2 Raafia Aseena, 3Nuha Rasheed and 4 Abdul Saleem Mohammad

2017-01-01

Research on zika virus examine the virus that is spread to humans through a mosquito bite, with symptoms that include fever, rash, joint pain, and conjuctivities. For most people zika virus is not necessarily anything to worry, as it is not fatal and symptoms are generally mild for period up to a week. Hospitalization because of zika virus is almost always not necessary. However, the zika virus can be extremely dangerous to pregnant womes. Key Words: Zika, virus, transmission, fatal, flavivir...

The influenza fingerprints: NS1 and M1 proteins contribute to specific host cell ultrastructure signatures upon infection by different influenza A viruses

Energy Technology Data Exchange (ETDEWEB)

Terrier, Olivier; Moules, Vincent; Carron, Coralie; Cartet, Gaeelle [Equipe VirCell, Laboratoire de Virologie et Pathologie Humaine, VirPath EMR 4610, Universite de Lyon, Universite Claude Bernard Lyon 1, Hospices Civils de Lyon, Faculte de medecine RTH Laennec, rue Guillaume Paradin, F-69008 Lyon (France); Frobert, Emilie [Laboratoire de Virologie, Centre de Biologie et de Pathologie Est, Hospices Civils de Lyon, 59 boulevard Pinel, F-69677 Bron Cedex, Lyon (France); Yver, Matthieu; Traversier, Aurelien [Equipe VirCell, Laboratoire de Virologie et Pathologie Humaine, VirPath EMR 4610, Universite de Lyon, Universite Claude Bernard Lyon 1, Hospices Civils de Lyon, Faculte de medecine RTH Laennec, rue Guillaume Paradin, F-69008 Lyon (France); Wolff, Thorsten [Division of Influenza/Respiratory Viruses, Robert Koch Institute, Nordufer 20, D-13353 Berlin (Germany); Riteau, Beatrice [Laboratoire de Virologie et Pathologie Humaine, VirPath EMR 4610, Universite de Lyon, Universite Claude Bernard Lyon 1, Hospices Civils de Lyon, Faculte de medecine RTH Laennec, rue Guillaume Paradin, F-69008 Lyon (France); Naffakh, Nadia [Institut Pasteur, Unite de Genetique Moleculaire des Virus Respiratoires, URA CNRS 3015, EA302 Universite Paris Diderot, Paris (France); and others

2012-10-10

Influenza A are nuclear replicating viruses which hijack host machineries in order to achieve optimal infection. Numerous functional virus-host interactions have now been characterized, but little information has been gathered concerning their link to the virally induced remodeling of the host cellular architecture. In this study, we infected cells with several human and avian influenza viruses and we have analyzed their ultrastructural modifications by using electron and confocal microscopy. We discovered that infections lead to a major and systematic disruption of nucleoli and the formation of a large number of diverse viral structures showing specificity that depended on the subtype origin and genomic composition of viruses. We identified NS1 and M1 proteins as the main actors in the remodeling of the host ultra-structure and our results suggest that each influenza A virus strain could be associated with a specific cellular fingerprint, possibly correlated to the functional properties of their viral components.

Type C virus activation in nontransformed mouse cells by uv-irradiated herpes simplex virus

Energy Technology Data Exchange (ETDEWEB)

Hampar, B. (National Institutes of Health, Bethesda, MD); Hatanaka, M.; Aulakh, G.; Derge, J.G.; Lee, L.; Showalter, S.

1977-02-01

Infection of nontransformed mouse cells with uv-irradiated herpes simplex virus (uv-HSV) resulted in the activation of an endogenous xenotropic (x-tropic) type C virus. Synthesis of type C virus persisted for only a few days, with most of the virus remaining cell associated. The levels of type C virus activated by uv-HSV varied depending on the multiplicity of infection (m.o.i.) and the uv dose. At low uv doses, where cell killing occurred, little or no type C virus synthesis was observed. Maximum levels of type C virus synthesis were observed with the minimum uv dose which eliminated cell killing by HSV. Synthesis of type C virus, albeit at lower levels, was still observed at uv doses beyond those required to prevent cell killing.

Type C virus activation in nontransformed mouse cells by uv-irradiated herpes simplex virus

International Nuclear Information System (INIS)

Hampar, B.; Hatanaka, M.; Aulakh, G.; Derge, J.G.; Lee, L.; Showalter, S.

1977-01-01

Infection of nontransformed mouse cells with uv-irradiated herpes simplex virus (uv-HSV) resulted in the activation of an endogenous xenotropic (x-tropic) type C virus. Synthesis of type C virus persisted for only a few days, with most of the virus remaining cell associated. The levels of type C virus activated by uv-HSV varied depending on the multiplicity of infection (m.o.i.) and the uv dose. At low uv doses, where cell killing occurred, little or no type C virus synthesis was observed. Maximum levels of type C virus synthesis were observed with the minimum uv dose which eliminated cell killing by HSV. Synthesis of type C virus, albeit at lower levels, was still observed at uv doses beyond those required to prevent cell killing

Chikungunya virus

Science.gov (United States)

Chikungunya virus infection; Chikungunya ... Where Chikungunya is Found Before 2013, the virus was found in Africa, Asia, Europe, and the Indian and Pacific oceans. In late 2013, outbreaks occurred for the first time in the ...

Perinatal hepatitis B virus detection by hepatitis B virus-DNA analysis.

OpenAIRE

De Virgiliis, S; Frau, F; Sanna, G; Turco, M P; Figus, A L; Cornacchia, G; Cao, A

1985-01-01

Maternal transmission of hepatitis B virus infection in relation to the hepatitis B e antigen/antibody system and serum hepatitis B virus-DNA were evaluated. Results indicate that hepatitis B virus-DNA analysis can identify hepatitis B serum antigen positive mothers who may transmit infection to their offspring.

Virus like particle-based vaccines against emerging infectious disease viruses.

Science.gov (United States)

Liu, Jinliang; Dai, Shiyu; Wang, Manli; Hu, Zhihong; Wang, Hualin; Deng, Fei

2016-08-01

Emerging infectious diseases are major threats to human health. Most severe viral disease outbreaks occur in developing regions where health conditions are poor. With increased international travel and business, the possibility of eventually transmitting infectious viruses between different countries is increasing. The most effective approach in preventing viral diseases is vaccination. However, vaccines are not currently available for numerous viral diseases. Virus-like particles (VLPs) are engineered vaccine candidates that have been studied for decades. VLPs are constructed by viral protein expression in various expression systems that promote the selfassembly of proteins into structures resembling virus particles. VLPs have antigenicity similar to that of the native virus, but are non-infectious as they lack key viral genetic material. VLP vaccines have attracted considerable research interest because they offer several advantages over traditional vaccines. Studies have shown that VLP vaccines can stimulate both humoral and cellular immune responses, which may offer effective antiviral protection. Here we review recent developments with VLP-based vaccines for several highly virulent emerging or re-emerging infectious diseases. The infectious agents discussed include RNA viruses from different virus families, such as the Arenaviridae, Bunyaviridae, Caliciviridae, Coronaviridae, Filoviridae, Flaviviridae, Orthomyxoviridae, Paramyxoviridae, and Togaviridae families.

Immunogenicity of a modified-live virus vaccine against bovine viral diarrhea virus types 1 and 2, infectious bovine rhinotracheitis virus, bovine parainfluenza-3 virus, and bovine respiratory syncytial virus when administered intranasally in young calves.

Science.gov (United States)

Xue, Wenzhi; Ellis, John; Mattick, Debra; Smith, Linda; Brady, Ryan; Trigo, Emilio

2010-05-14

The immunogenicity of an intranasally-administered modified-live virus (MLV) vaccine in 3-8 day old calves was evaluated against bovine viral diarrhea virus (BVDV) types 1 and 2, infectious bovine rhinotracheitis (IBR) virus, parainfluenza-3 (PI-3) virus and bovine respiratory syncytial virus (BRSV). Calves were intranasally vaccinated with a single dose of a multivalent MLV vaccine and were challenged with one of the respective viruses three to four weeks post-vaccination in five separate studies. There was significant sparing of diseases in calves intranasally vaccinated with the MLV vaccine, as indicated by significantly fewer clinical signs, lower rectal temperatures, reduced viral shedding, greater white blood cell and platelet counts, and less severe pulmonary lesions than control animals. This was the first MLV combination vaccine to demonstrate efficacy against BVDV types 1 and 2, IBR, PI-3 and BRSV in calves 3-8 days of age. Copyright 2010 Elsevier Ltd. All rights reserved.

Ocular Tropism of Respiratory Viruses

Science.gov (United States)

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

Understanding Ebola Virus Transmission

Directory of Open Access Journals (Sweden)

Seth Judson

2015-02-01

Full Text Available 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.

Ganjam virus/Nairobi sheep disease virus induces a pro-inflammatory response in infected sheep

OpenAIRE

bin Tarif, Abid; Lasecka, Lidia; Holzer, Barbara; Baron, Michael D

2012-01-01

Abstract Partly due to climate change, and partly due to changes of human habitat occupation, the impact of tick-borne viruses is increasing. Nairobi sheep disease virus (NSDV) and Ganjam virus (GV) are two names for the same virus, which causes disease in sheep and goats and is currently known to be circulating in India and East Africa. The virus is transmitted by ixodid ticks and causes a severe hemorrhagic disease. We have developed a real-time PCR assay for the virus genome and validated ...

Zika Virus

Science.gov (United States)

... through blood transfusions. There have been outbreaks of Zika virus in the United States, Africa, Southeast Asia, the ... not travel to areas where there is a Zika virus outbreak. If you do decide to travel, first ...

Generation and Comprehensive Analysis of an Influenza Virus Polymerase Cellular Interaction Network▿†§

Science.gov (United States)

Tafforeau, Lionel; Chantier, Thibault; Pradezynski, Fabrine; Pellet, Johann; Mangeot, Philippe E.; Vidalain, Pierre-Olivier; Andre, Patrice; Rabourdin-Combe, Chantal; Lotteau, Vincent

2011-01-01

The influenza virus transcribes and replicates its genome inside the nucleus of infected cells. Both activities are performed by the viral RNA-dependent RNA polymerase that is composed of the three subunits PA, PB1, and PB2, and recent studies have shown that it requires host cell factors to transcribe and replicate the viral genome. To identify these cellular partners, we generated a comprehensive physical interaction map between each polymerase subunit and the host cellular proteome. A total of 109 human interactors were identified by yeast two-hybrid screens, whereas 90 were retrieved by literature mining. We built the FluPol interactome network composed of the influenza virus polymerase (PA, PB1, and PB2) and the nucleoprotein NP and 234 human proteins that are connected through 279 viral-cellular protein interactions. Analysis of this interactome map revealed enriched cellular functions associated with the influenza virus polymerase, including host factors involved in RNA polymerase II-dependent transcription and mRNA processing. We confirmed that eight influenza virus polymerase-interacting proteins are required for virus replication and transcriptional activity of the viral polymerase. These are involved in cellular transcription (C14orf166, COPS5, MNAT1, NMI, and POLR2A), translation (EIF3S6IP), nuclear transport (NUP54), and DNA repair (FANCG). Conversely, we identified PRKRA, which acts as an inhibitor of the viral polymerase transcriptional activity and thus is required for the cellular antiviral response. PMID:21994455

Avian influenza A virus PB2 promotes interferon type I inducing properties of a swine strain in porcine dendritic cells

International Nuclear Information System (INIS)

Ocaña-Macchi, Manuela; Ricklin, Meret E.; Python, Sylvie; Monika, Gsell-Albert; Stech, Jürgen; Stech, Olga; Summerfield, Artur

2012-01-01

The 2009 influenza A virus (IAV) pandemic resulted from reassortment of avian, human and swine strains probably in pigs. To elucidate the role of viral genes in host adaptation regarding innate immune responses, we focussed on the effect of genes from an avian H5N1 and a porcine H1N1 IAV on infectivity and activation of porcine GM-CSF-induced dendritic cells (DC). The highest interferon type I responses were achieved by the porcine virus reassortant containing the avian polymerase gene PB2. This finding was not due to differential tropism since all viruses infected DC equally. All viruses equally induced MHC class II, but porcine H1N1 expressing the avian viral PB2 induced more prominent nuclear NF-κB translocation compared to its parent IAV. The enhanced activation of DC may be detrimental or beneficial. An over-stimulation of innate responses could result in either pronounced tissue damage or increased resistance against IAV reassortants carrying avian PB2.

Avian influenza A virus PB2 promotes interferon type I inducing properties of a swine strain in porcine dendritic cells

Energy Technology Data Exchange (ETDEWEB)

Ocana-Macchi, Manuela; Ricklin, Meret E.; Python, Sylvie; Monika, Gsell-Albert [Institute of Virology and Immunoprophylaxis, Mittelhaeusern (Switzerland); Stech, Juergen; Stech, Olga [Friedrich-Loeffler Institut, Greifswald-Insel Riems (Germany); Summerfield, Artur, E-mail: artur.summerfield@ivi.admin.ch [Institute of Virology and Immunoprophylaxis, Mittelhaeusern (Switzerland)

2012-05-25

The 2009 influenza A virus (IAV) pandemic resulted from reassortment of avian, human and swine strains probably in pigs. To elucidate the role of viral genes in host adaptation regarding innate immune responses, we focussed on the effect of genes from an avian H5N1 and a porcine H1N1 IAV on infectivity and activation of porcine GM-CSF-induced dendritic cells (DC). The highest interferon type I responses were achieved by the porcine virus reassortant containing the avian polymerase gene PB2. This finding was not due to differential tropism since all viruses infected DC equally. All viruses equally induced MHC class II, but porcine H1N1 expressing the avian viral PB2 induced more prominent nuclear NF-{kappa}B translocation compared to its parent IAV. The enhanced activation of DC may be detrimental or beneficial. An over-stimulation of innate responses could result in either pronounced tissue damage or increased resistance against IAV reassortants carrying avian PB2.

Ganjam virus/Nairobi sheep disease virus induces a pro-inflammatory response in infected sheep

Directory of Open Access Journals (Sweden)

bin Tarif Abid

2012-10-01

Full Text Available Abstract Partly due to climate change, and partly due to changes of human habitat occupation, the impact of tick-borne viruses is increasing. Nairobi sheep disease virus (NSDV and Ganjam virus (GV are two names for the same virus, which causes disease in sheep and goats and is currently known to be circulating in India and East Africa. The virus is transmitted by ixodid ticks and causes a severe hemorrhagic disease. We have developed a real-time PCR assay for the virus genome and validated it in a pilot study of the pathogenicity induced by two different isolates of NSDV/GV. One isolate was highly adapted to tissue culture, grew in most cell lines tested, and was essentially apathogenic in sheep. The second isolate appeared to be poorly adapted to cell culture and retained pathogenicity in sheep. The real-time PCR assay for virus easily detected 4 copies or less of the viral genome, and allowed a quantitative measure of the virus in whole blood. Measurement of the changes in cytokine mRNAs showed similar changes to those observed in humans infected by the closely related virus Crimean Congo hemorrhagic fever virus.

Ganjam virus/Nairobi sheep disease virus induces a pro-inflammatory response in infected sheep.

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Bin Tarif, Abid; Lasecka, Lidia; Holzer, Barbara; Baron, Michael D

2012-10-19

Partly due to climate change, and partly due to changes of human habitat occupation, the impact of tick-borne viruses is increasing. Nairobi sheep disease virus (NSDV) and Ganjam virus (GV) are two names for the same virus, which causes disease in sheep and goats and is currently known to be circulating in India and East Africa. The virus is transmitted by ixodid ticks and causes a severe hemorrhagic disease. We have developed a real-time PCR assay for the virus genome and validated it in a pilot study of the pathogenicity induced by two different isolates of NSDV/GV. One isolate was highly adapted to tissue culture, grew in most cell lines tested, and was essentially apathogenic in sheep. The second isolate appeared to be poorly adapted to cell culture and retained pathogenicity in sheep. The real-time PCR assay for virus easily detected 4 copies or less of the viral genome, and allowed a quantitative measure of the virus in whole blood. Measurement of the changes in cytokine mRNAs showed similar changes to those observed in humans infected by the closely related virus Crimean Congo hemorrhagic fever virus.

Transmission of Influenza A Viruses

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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

New frontiers in oncolytic viruses: optimizing and selecting for virus strains with improved efficacy

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Lundstrom K

2018-02-01

Full Text Available Kenneth Lundstrom PanTherapeutics, Lutry, Switzerland Abstract: Oncolytic viruses have demonstrated selective replication and killing of tumor cells. Different types of oncolytic viruses – adenoviruses, alphaviruses, herpes simplex viruses, Newcastle disease viruses, rhabdoviruses, Coxsackie viruses, and vaccinia viruses – have been applied as either naturally occurring or engineered vectors. Numerous studies in animal-tumor models have demonstrated substantial tumor regression and prolonged survival rates. Moreover, clinical trials have confirmed good safety profiles and therapeutic efficacy for oncolytic viruses. Most encouragingly, the first cancer gene-therapy drug – Gendicine, based on oncolytic adenovirus type 5 – was approved in China. Likewise, a second-generation oncolytic herpes simplex virus-based drug for the treatment of melanoma has been registered in the US and Europe as talimogene laherparepvec. Keywords: immunotherapy, viral vectors, clinical trials, drug approval

Postmortem stability of Ebola virus.

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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.

A Foxtail mosaic virus Vector for Virus-Induced Gene Silencing in Maize.

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Mei, Yu; Zhang, Chunquan; Kernodle, Bliss M; Hill, John H; Whitham, Steven A

2016-06-01

Plant viruses have been widely used as vectors for foreign gene expression and virus-induced gene silencing (VIGS). A limited number of viruses have been developed into viral vectors for the purposes of gene expression or VIGS in monocotyledonous plants, and among these, the tripartite viruses Brome mosaic virus and Cucumber mosaic virus have been shown to induce VIGS in maize (Zea mays). We describe here a new DNA-based VIGS system derived from Foxtail mosaic virus (FoMV), a monopartite virus that is able to establish systemic infection and silencing of endogenous maize genes homologous to gene fragments inserted into the FoMV genome. To demonstrate VIGS applications of this FoMV vector system, four genes, phytoene desaturase (functions in carotenoid biosynthesis), lesion mimic22 (encodes a key enzyme of the porphyrin pathway), iojap (functions in plastid development), and brown midrib3 (caffeic acid O-methyltransferase), were silenced and characterized in the sweet corn line Golden × Bantam. Furthermore, we demonstrate that the FoMV infectious clone establishes systemic infection in maize inbred lines, sorghum (Sorghum bicolor), and green foxtail (Setaria viridis), indicating the potential wide applications of this viral vector system for functional genomics studies in maize and other monocots. © 2016 American Society of Plant Biologists. All Rights Reserved.

Resveratrol treatment reveals a novel role for HMGB1 in regulation of the type 1 interferon response in dengue virus infection.

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Zainal, Nurhafiza; Chang, Chih-Peng; Cheng, Yi-Lin; Wu, Yan-Wei; Anderson, Robert; Wan, Shu-Wen; Chen, Chia-Ling; Ho, Tzong-Shiann; AbuBakar, Sazaly; Lin, Yee-Shin

2017-02-20

Dengue is one of the most significant mosquito-borne virus diseases worldwide, particularly in tropical and subtropical regions. This study sought to examine the antiviral activity of resveratrol (RESV), a phytoalexin secreted naturally by plants, against dengue virus (DENV) infection. Our data showed that RESV inhibits the translocation of high mobility group box 1 (HMGB1), a DNA binding protein that normally resides in the nucleus, into the cytoplasm and extracellular milieu. HMGB1 migrates out of the nucleus during DENV infection. This migration is inhibited by RESV treatment and is mediated by induction of Sirt1 which leads to the retention of HMGB1 in the nucleus and consequently helps in the increased production of interferon-stimulated genes (ISGs). Nuclear HMGB1 was found to bind to the promoter region of the ISG and positively regulated the expression of ISG. The enhanced transcription of ISGs by nuclear HMGB1 thus contributes to the antiviral activity of RESV against DENV. To the best of our knowledge, this is the first report to demonstrate that RESV antagonizes DENV replication and that nuclear HMGB1 plays a role in regulating ISG production.

Characterization of Sendai virus persistently infected L929 cells and Sendai virus pi strain: recombinant Sendai viruses having Mpi protein shows lower cytotoxicity and are incapable of establishing persistent infection

International Nuclear Information System (INIS)

Nishio, Machiko; Tsurudome, Masato; Ito, Morihiro; Kawano, Mitsuo; Komada, Hiroshi; Ito, Yasuhiko

2003-01-01

It is commonly accepted that the temperature-sensitive phenotype of Sendai virus (SeV) persistently infected cells is caused by the M and/or HN proteins. Expression level of the L, M, HN, and V proteins is extremely low in L929 cells persistently infected with SeVpi (L929/SeVpi cells) incubated at 38 deg. C. The HN protein quickly disappears in L929/SeVpi cells following a temperature shift up to 38 deg. C, and pulse-chase experiments show that the Lpi, HNpi, and Mpi proteins are unstable at 38 deg. C. Following a temperature shift either upward or downward, M protein is translocated into the nucleus and then localizes to the perinuclear region. None of virus-specific polypeptides are detected in the cells primarily infected with SeVpi and incubated at 38 deg. C and virus proteins are not pulse-labeled at 38 deg. C, indicating that temperature-sensitive step is at an early stage of infection. The Mpi protein is transiently located in the nucleus of the SeVpi primarily infected cells. Recombinant SeVs possessing the HNpi or/and Mpi proteins are not temperature-sensitive. The HN protein is expressed at very low levels and the F protein localizes to the perinuclear region in rSeV(Mpi)-infected cells incubated at 38 deg. C for 18 h. rSeVs having the Mpi protein exhibit lower cytotoxicity and are incapable of establishing persistent infection. Amino acid 116 of the Mpi protein is related to the nuclear translocation and lower cytopathogenesis, whereas aa183 is involved in the interaction between M protein and viral glycoproteins

Atoms for peace and development: Contributing to global progress through nuclear science and technology

International Nuclear Information System (INIS)

Amano, Yukiya

2015-01-01

Cultivating new crop varieties, reducing soil erosion and helping African countries respond to Ebola Virus Disease are just some of the areas in which the IAEA helps Member States to benefit from nuclear technology. Assisting countries in the safe and secure use of nuclear techniques for development is as important to the IAEA as its non-proliferation work. For many developing countries, it is the most important thing we do. Our mandate has been summarized as Atoms for Peace. Today, I feel that our mandate could be better understood as Atoms for Peace and Development.

Nuclear medicine in the management of the AIDS patient

International Nuclear Information System (INIS)

Kramer, E.L.; Sanger, J.J.

1990-01-01

For the medical diagnostic imaging specialist in general, and for the nuclear medicine physician specifically, the AIDS epidemic has generated an enormous demand to develop a means of making early diagnoses of the complications of AIDS. For the most part this has meant the early detection, and when possible, the characterization of the opportunistic infections and neoplasms that are a major source of morbidity and mortality for the AIDS patient. Detection of opportunistic infections has been helpful in reclassifying HIV-seropositive patients as having AIDS. This paper reports on nuclear medicine used to evaluate the efficacy and the complications of treatment in human immunodeficiency virus infection. Most recently, functional brain imaging has been used for the diagnosis and follow-up of the AIDS dementia complex

Release of the herpes simplex virus 1 protease by self cleavage is required for proper conformation of the portal vertex

International Nuclear Information System (INIS)

Yang, Kui; Wills, Elizabeth G.; Baines, Joel D.

2012-01-01

We identify an NLS within herpes simplex virus scaffold proteins that is required for optimal nuclear import of these proteins into infected or uninfected nuclei, and is sufficient to mediate nuclear import of GFP. A virus lacking this NLS replicated to titers reduced by 1000-fold, but was able to make capsids containing both scaffold and portal proteins suggesting that other functions can complement the NLS in infected cells. We also show that Vp22a, the major scaffold protein, is sufficient to mediate the incorporation of portal protein into capsids, whereas proper portal immunoreactivity in the capsid requires the larger scaffold protein pU L 26. Finally, capsid angularization in infected cells did not require the HSV-1 protease unless full length pU L 26 was expressed. These data suggest that the HSV-1 portal undergoes conformational changes during capsid maturation, and reveal that full length pU L 26 is required for this conformational change.

Release of the herpes simplex virus 1 protease by self cleavage is required for proper conformation of the portal vertex

Energy Technology Data Exchange (ETDEWEB)

Yang, Kui; Wills, Elizabeth G. [Department of Microbiology and Immunology, Cornell University, Ithaca, NY 14853 (United States); Baines, Joel D., E-mail: jdb11@cornell.edu [Department of Microbiology and Immunology, Cornell University, Ithaca, NY 14853 (United States)

2012-07-20

We identify an NLS within herpes simplex virus scaffold proteins that is required for optimal nuclear import of these proteins into infected or uninfected nuclei, and is sufficient to mediate nuclear import of GFP. A virus lacking this NLS replicated to titers reduced by 1000-fold, but was able to make capsids containing both scaffold and portal proteins suggesting that other functions can complement the NLS in infected cells. We also show that Vp22a, the major scaffold protein, is sufficient to mediate the incorporation of portal protein into capsids, whereas proper portal immunoreactivity in the capsid requires the larger scaffold protein pU{sub L}26. Finally, capsid angularization in infected cells did not require the HSV-1 protease unless full length pU{sub L}26 was expressed. These data suggest that the HSV-1 portal undergoes conformational changes during capsid maturation, and reveal that full length pU{sub L}26 is required for this conformational change.

Efficient production of infectious viruses requires enzymatic activity of Epstein-Barr virus protein kinase.

Science.gov (United States)

Murata, Takayuki; Isomura, Hiroki; Yamashita, Yoriko; Toyama, Shigenori; Sato, Yoshitaka; Nakayama, Sanae; Kudoh, Ayumi; Iwahori, Satoko; Kanda, Teru; Tsurumi, Tatsuya

2009-06-20

The Epstein-Barr virus (EBV) BGLF4 gene product is the only protein kinase encoded by the virus genome. In order to elucidate its physiological roles in viral productive replication, we here established a BGLF4-knockout mutant and a revertant virus. While the levels of viral DNA replication of the deficient mutant were equivalent to those of the wild-type and the revertant, virus production was significantly impaired. Expression of the BGLF4 protein in trans fully complemented the low yield of the mutant virus, while expression of a kinase-dead (K102I) form of the protein failed to restore the virus titer. These results demonstrate that BGLF4 plays a significant role in production of infectious viruses and that the kinase activity is crucial.

Propagation Effect of a Virus Outbreak on a Network with Limited Anti-Virus Ability.

Directory of Open Access Journals (Sweden)

Yonghong Xu

Full Text Available This paper describes a new computer virus spreading model which takes into account the possibility of a virus outbreak on a network with limited anti-virus ability. Then, the model is investigated for the existence of equilibria and their stabilities are proved and illustrated. Moreover, it is found that these two factors are not only relative to the threshold value determining whether the virus becomes extinct or not, but that they are also relative to the virus epidemic levels. Theoretical and experimental results indicate that, in some ways, it would be practically possible to eradicate the virus or suppress its prevalence below a suitable level. Consequently, some suggestions are proposed that may help eradicate or suppress virus propagation over a real computer network.

An introduction to computer viruses

Energy Technology Data Exchange (ETDEWEB)

Brown, D.R.

1992-03-01

This report on computer viruses is based upon a thesis written for the Master of Science degree in Computer Science from the University of Tennessee in December 1989 by David R. Brown. This thesis is entitled An Analysis of Computer Virus Construction, Proliferation, and Control and is available through the University of Tennessee Library. This paper contains an overview of the computer virus arena that can help the reader to evaluate the threat that computer viruses pose. The extent of this threat can only be determined by evaluating many different factors. These factors include the relative ease with which a computer virus can be written, the motivation involved in writing a computer virus, the damage and overhead incurred by infected systems, and the legal implications of computer viruses, among others. Based upon the research, the development of a computer virus seems to require more persistence than technical expertise. This is a frightening proclamation to the computing community. The education of computer professionals to the dangers that viruses pose to the welfare of the computing industry as a whole is stressed as a means of inhibiting the current proliferation of computer virus programs. Recommendations are made to assist computer users in preventing infection by computer viruses. These recommendations support solid general computer security practices as a means of combating computer viruses.

Viruses infecting marine molluscs.

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Arzul, Isabelle; Corbeil, Serge; Morga, Benjamin; Renault, Tristan

2017-07-01

Although a wide range of viruses have been reported in marine molluscs, most of these reports rely on ultrastructural examination and few of these viruses have been fully characterized. The lack of marine mollusc cell lines restricts virus isolation capacities and subsequent characterization works. Our current knowledge is mostly restricted to viruses affecting farmed species such as oysters Crassostrea gigas, abalone Haliotis diversicolor supertexta or the scallop Chlamys farreri. Molecular approaches which are needed to identify virus affiliation have been carried out for a small number of viruses, most of them belonging to the Herpesviridae and birnaviridae families. These last years, the use of New Generation Sequencing approach has allowed increasing the number of sequenced viral genomes and has improved our capacity to investigate the diversity of viruses infecting marine molluscs. This new information has in turn allowed designing more efficient diagnostic tools. Moreover, the development of experimental infection protocols has answered some questions regarding the pathogenesis of these viruses and their interactions with their hosts. Control and management of viral diseases in molluscs mostly involve active surveillance, implementation of effective bio security measures and development of breeding programs. However factors triggering pathogen development and the life cycle and status of the viruses outside their mollusc hosts still need further investigations. Copyright © 2017 Elsevier Inc. All rights reserved.

Antiviral Activities of Several Oral Traditional Chinese Medicines against Influenza Viruses.

Science.gov (United States)

Ma, Lin-Lin; Ge, Miao; Wang, Hui-Qiang; Yin, Jin-Qiu; Jiang, Jian-Dong; Li, Yu-Huan

2015-01-01

Influenza is still a serious threat to human health with significant morbidity and mortality. The emergence of drug-resistant influenza viruses poses a great challenge to existing antiviral drugs. Traditional Chinese medicines (TCMs) may be an alternative to overcome the challenge. Here, 10 oral proprietary Chinese medicines were selected to evaluate their anti-influenza activities. These drugs exhibit potent inhibitory effects against influenza A H1N1, influenza A H3N2, and influenza B virus. Importantly, they demonstrate potent antiviral activities against drug-resistant strains. In the study of mechanisms, we found that Xiaoqinglong mixture could increase antiviral interferon production by activating p38 MAPK, JNK/SAPK pathway, and relative nuclear transcription factors. Lastly, our studies also indicate that some of these medicines show inhibitory activities against EV71 and CVB strains. In conclusion, the 10 traditional Chinese medicines, as kind of compound combination medicines, show broad-spectrum antiviral activities, possibly also including inhibitory activities against strains resistant to available antiviral drugs.

Coinfection with Epstein–Barr Virus (EBV), Human Papilloma Virus (HPV) and Polyoma BK Virus (BKPyV) in Laryngeal, Oropharyngeal and Oral Cavity Cancer

OpenAIRE

Drop, Bartłomiej; Strycharz-Dudziak, Małgorzata; Kliszczewska, Ewa; Polz-Dacewicz, Małgorzata

2017-01-01

Most research providing evidence for the role of oncogenic viruses in head and neck squamous cell carcinoma (SCC) development is focused on one type of virus without analyzing possible interactions between two or more types of viruses. The aim of this study was to analyse the prevalence of co-infection with human papillomavirus (HPV), Epstein–Barr virus (EBV) and polyoma BK virus (BKPyV) in oral, oropharyngeal and laryngeal squamous cell carcinomas in Polish patients. The correlations between...

RNA viruses in the sea.

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