WorldWideScience

Sample records for baculovirus nuclear localization

  1. Identification of novel residues involved in nuclear localization of a baculovirus polyhedrin protein.

    Science.gov (United States)

    Katsuma, S; Deng, D X; Zhou, C L; Iwanaga, M; Noguchi, Y; Kobayashi, M; Maeda, S

    2000-10-01

    A baculovirus polyhedrin protein has proven to possess a nuclear localization signal (NLS) sequence and a domain required for supramolecular assembly. Here we investigated five Bombyx mori nucleopolyhedrovirus (BmNPV) mutants that did not produce polyhedra. Two of five mutants were generated during routine baculoviral expression vector screening, and three were isolated by treatment with the mutagen 5-bromo-2'-deoxyuridine (BrdU). Marker rescue mapping and nucleotide sequence analysis showed that mutations in the polyhedrin gene caused the altered phenotype of these mutants. Biochemical fractionation indicated that cells infected with these mutants exhibited polyhedrin protein in both the nucleus and the cytoplasm. Electron microscopic observation revealed that polyhedrin produced by these mutants ocurred in both the nucleus and the cytoplasm, but did not form a crystalline lattice. Despite the incompleteness of polyhedrin nuclear localization, the NLSs of the five mutants were unchanged, although some of the mutations occurred within residues just outside of the domain reported to be required for polyhedron assembly (4). This result suggests that (a) the polyhedrin NLS directs polyhedrin to the nucleus, but the efficiency of this localization is regulated by regions other than the NLS (probably, polyhedrin conformation and its association with the nucleus are also involved), and (b) formation of a crystalline lattice may also be determined by several domains within polyhedrin. PMID:11129641

  2. Baculoviruses and nucleosome management

    Energy Technology Data Exchange (ETDEWEB)

    Volkman, Loy E., E-mail: lvolkman@berkeley.edu

    2015-02-15

    Negatively-supercoiled-ds DNA molecules, including the genomes of baculoviruses, spontaneously wrap around cores of histones to form nucleosomes when present within eukaryotic nuclei. Hence, nucleosome management should be essential for baculovirus genome replication and temporal regulation of transcription, but this has not been documented. Nucleosome mobilization is the dominion of ATP-dependent chromatin-remodeling complexes. SWI/SNF and INO80, two of the best-studied complexes, as well as chromatin modifier TIP60, all contain actin as a subunit. Retrospective analysis of results of AcMNPV time course experiments wherein actin polymerization was blocked by cytochalasin D drug treatment implicate actin-containing chromatin modifying complexes in decatenating baculovirus genomes, shutting down host transcription, and regulating late and very late phases of viral transcription. Moreover, virus-mediated nuclear localization of actin early during infection may contribute to nucleosome management. - Highlights: • Baculoviruses have negatively-supercoiled, circular ds DNA. • Negatively-supercoiled DNA spontaneously forms nucleosomes in the nucleus. • Nucleosomes must be mobilized for replication and transcription to proceed. • Actin-containing chromatin modifiers participate in baculovirus replication.

  3. A new mechanism for nuclear import by actin-based propulsion used by a baculovirus nucleocapsid.

    Science.gov (United States)

    Au, Shelly; Wu, Wei; Zhou, Lixin; Theilmann, David A; Panté, Nelly

    2016-08-01

    The transport of macromolecules into the nucleus is mediated by soluble cellular receptors of the importin β superfamily and requires the Ran-GTPase cycle. Several studies have provided evidence that there are exceptions to this canonical nuclear import pathway. Here, we report a new unconventional nuclear import mechanism exploited by the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV). We found that AcMNPV nucleocapsids entered the nucleus of digitonin-permeabilized cells in the absence of exogenous cytosol or under conditions that blocked the Ran-GTPase cycle. AcMNPV contains a protein that activates the Arp2/3 complex and induces actin polymerization at one end of the rod-shaped nucleocapsid. We show that inhibitors of Arp2/3 blocked nuclear import of nucleocapsids in semi-permeabilized cells. Nuclear import of nucleocapsids was also reconstituted in purified nuclei supplemented with G-actin and Arp2/3 under actin polymerization conditions. Thus, we propose that actin polymerization drives not only migration of baculovirus through the cytoplasm but also pushes the nucleocapsid through the nuclear pore complex to enter the cell nucleus. Our findings point to a very distinct role of actin-based motility during the baculovirus infection cycle. PMID:27284005

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

    International Nuclear Information System (INIS)

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

  5. Local Immune Stimulation by Intravesical Instillation of Baculovirus to Enable Bladder Cancer Therapy

    OpenAIRE

    Wei Xia Ang; Ying Zhao; Timothy Kwang; Chunxiao Wu; Can Chen; Han Chong Toh; Ratha Mahendran; Kesavan Esuvaranathan; Shu Wang

    2016-01-01

    Intravesical instillation of Bacillus Calmette-Guérin is currently used as adjuvant therapy for superficial, non-muscle invasive bladder cancer (NMIBC). However, nearly 40% of patients with NMIBC will fail Bacillus Calmette-Guérin therapy. In an attempt to investigate the feasibility of using insect baculovirus-based vectors for bladder cancer therapy, we observed that intravesical instillation of baculoviruses without transgene up-regulated a set of Th1-type of cytokines and increased the su...

  6. Baculovirus RNA Polymerase: Activities, Composition, and Evolution

    Institute of Scientific and Technical Information of China (English)

    A.Lorena Passarelli

    2007-01-01

    Baculoviruses are the only nuclear replicating DNA-containing viruses that encode their own DNA-directed RNA polymerase (RNAP). The baculovirus RNAP is specific for the transcription of genes expressed after virus DNA replication. It is composed of four subunits, making it the simplest multisubunit RNAP known. Two subunits contain motifs found at the catalytic center of other RNAPs and a third has capping enzyme functions. The function of the fourth subunit is not known. Structural studies on this unique RNAP will provide new insights into the functions of this enzyme and the regulation of viral genes and may be instrumental to optimize the baculovirus gene expression system.

  7. Baculovirus DNA replication.

    OpenAIRE

    Kool, M.

    1994-01-01

    Baculoviruses are attractive biological agents for the control of insect pests. They are highly specific for insects and cause a fatal disease (Granados and Federici, 1986). in addition, baculoviruses are successfully exploited as expression vectors for the production of heterologous proteins for various applications (Luckow and Summers, 1988; Luckow, 1991). In both cases large-scale systems for the production of baculoviruses are important. Production in insect larvae is difficult to scale u...

  8. Baculovirus DNA replication.

    NARCIS (Netherlands)

    Kool, M.

    1994-01-01

    Baculoviruses are attractive biological agents for the control of insect pests. They are highly specific for insects and cause a fatal disease (Granados and Federici, 1986). in addition, baculoviruses are successfully exploited as expression vectors for the production of heterologous proteins for va

  9. Baculovirus: Hospederos y especificidad

    Directory of Open Access Journals (Sweden)

    Juliana Gómez Valderrama

    2013-12-01

    Full Text Available Título corto: BaculovirusTítulo en ingles: Baculovirus: Hosts and specificityResumen: Los baculovirus son virus patógenos de insectos ampliamente empleados a nivel mundial como bioinsecticidas para el control de diferentes plagas de importancia agrícola y más recientemente como vectores de expresión de proteínas y vectores para terapia génica. Una de sus características principales es su alta especificidad de hospedero que incluye un rango muy estrecho de especies de insectos, que a menudo pertenecen a la misma familia. Sin embargo, es necesario entender los mecanismos involucrados en la definición del rango de hospederos de los baculovirus con el fin de evaluar la seguridad e inocuidad de su uso y determinar la posibilidad de mejorar sus propiedades para aplicaciones biotecnológicas mediante la construcción de baculovirus recombinantes con diferentes rangos de hospederos. En el presente artículo se revisarán los principales mecanismos comprendidos en la definición del rango de hospederos de los baculovirus, dentro de los que se destacan la especificidad para entrar en las células, la posibilidad de replicación del genoma viral, el control de los procesos bioquímicos y moleculares del insecto y las interacciones virus-hospedero que regulan la multiplicación del agente infeccioso, así como las perspectivas de la aplicación de este conocimiento.Palabras clave: infección viral, rango de hospederos, evolución, resistencia.Abstract: Baculoviruses are insect pathogenic viruses widely used as bioinsecticides for controlling of several agricultural important pests and more recently as protein expression vectors and gene therapy vectors. One of its main characteristics is its high host-specificity including a very narrow range of insect species, which often belong to the same family. However, to understand the mechanisms involved in the definition of baculoviruses host range is necessary in order to assess the security and safety

  10. Improving baculovirus recombination

    OpenAIRE

    Zhao, Yuguang; Chapman, David A. G.; Jones, Ian M.

    2003-01-01

    Recombinant baculoviruses have established themselves as a favoured technology for the high-level expression of recombinant proteins. The construction of recombinant viruses, however, is a time consuming step that restricts consideration of the technology for high throughput developments. Here we use a targeted gene knockout technology to inactivate an essential viral gene that lies adjacent to the locus used for recombination. Viral DNA prepared from the knockout fails to initiate an infecti...

  11. Identification of two functional nuclear localization signals in the capsid protein of duck circovirus

    International Nuclear Information System (INIS)

    The capsid protein (CP) of duck circovirus (DuCV) is the major immunogenic protein and has a high proportion of arginine residues concentrated at the N terminus of the protein, which inhibits efficient mRNA translation in prokaryotic expression systems. In this study, we investigated the subcellular distribution of DuCV CP expressed via recombinant baculoviruses in Sf9 cells and the DNA binding activities of the truncated recombinant DuCV CPs. The results showed that two independent bipartite nuclear localization signals (NLSs) situated at N-terminal 1–17 and 18–36 amino acid residue of the CP. Moreover, two expression level regulatory signals (ELRSs) and two DNA binding signals (DBSs) were also mapped to the N terminus of the protein and overlapped with the two NLSs. The ability of CP to bind DNA, coupled with the karyophilic nature of this protein, strongly suggests that it may be responsible for nuclear targeting of the viral genome.

  12. CO-OCCLUSION AND PERSISTENCE OF A BACULOVIRUS MUTANT LACKING THE POLYHEDRIN GENE

    Science.gov (United States)

    A co-occlusion process was evaluated as a commercially and ecologically acceptable strategy for the development of genetically improved baculovirus insecticides. oinfection of Spodoptera frugiperda (IPLB-SF-21) tissue culture cells with Aucographa californica nuclear polyhedrosis...

  13. HSP70 induction during baculovirus infection

    Science.gov (United States)

    Baculoviruses are arthropod-specific double-stranded DNA viruses that have been employed as bio-insecticides against crop pests and to produce heterologous proteins in baculovirus expression systems. Although a consensus has emerged on the dominant molecular events driving baculovirus replication i...

  14. Local computer network for nuclear research

    International Nuclear Information System (INIS)

    The logical and physical level description of the local area network for nuclear research is presented. The network architecture of the measurement center for U-240 cyclotron was considered. 3 refs., 1 tab., 3 figs

  15. Expression of adenovirus type 2 DNA polymerase in insect cells infected with a recombinant baculovirus.

    OpenAIRE

    Watson, C J; Hay, R T

    1990-01-01

    Sequences encoding adenovirus type 2 DNA polymerase were placed under control of the polyhedrin promoter and inserted into the baculovirus Autographa californica nuclear polyhedrosis virus by homologous recombination. Insect cells infected with the recombinant virus produced substantial amounts of the adenovirus type 2 DNA polymerase protein which was functional in both DNA polymerase and replication initiation reactions. Thus, the baculovirus expression system can provide active adenovirus t...

  16. The local community and the nuclear waste

    International Nuclear Information System (INIS)

    In this book social and political scientists discuss different aspects of the selection of a site for disposal of the Swedish nuclear waste. Special attention is given to the preliminary studies that have been performed at a few localities. The authors study the chain of events after a community is proposed for a site study. What powers are set in motion? How do different groups act in order to support or stop the study? Which is the role played by political parties, local environmentalist movements, media and experts? Why is there a forceful opposition in one community and not in another? Why does one local government invite the nuclear waste company to perform the study, while another refuses? The role of the local government has become crucial, since the nuclear waste company have chosen to perform studies only in municipalities that show a positive interest

  17. Localization of nuclear power plant technology

    International Nuclear Information System (INIS)

    Asia, and particularly China, has an enormous need for power and must deal with the practicalities of building large base load units. In China, as in other countries, there are limitations on the use of large quantities of fossil fuel. This raises the possibility of turning to nuclear power to satisfy their energy needs. Other issues tend to point to the nuclear option for these growing economies, including economic considerations, environmental concerns, energy independence and raising the technological capabilities of the country. When a country embarks on a nuclear power program with the intention of localizing the technology, a long-term commitment is necessary to achieve this objective. Localization of nuclear technology is not a new phenomenon. The nature of the industry from the early beginnings has always involved transfer of technology when a new country initiated a nuclear power construction program. In fact, most previous experiences with this localization process involved heavy governmental, political and financial support to drive the success of the program. Because of this strong governmental support, only the receiving nation's companies were generally allowed to participate in the local business operations of the technology recipient. What is new and different today is the retreat from heavy financial support by the receiving country's government. This change has created a strong emphasis on cost-effectiveness in the technology transfer process and opportunities for foreign companies to participate in local business activities. ABB is a world-wide company with two parent companies that have been very active over many years in establishing cost-justified local operations throughout the world. Today, ABB has become the largest electrical engineering company in the world with respected local operations in nearly every country. Lessons learned by ABB in their world-wide localization initiatives are being applied to the challenge of cost

  18. TRANSFECTION WITH BACULOVIRUS DNA

    Science.gov (United States)

    Purified DNA from the nuclear polyhedrosis viruses of Autographa californica (AcM NPV) and Rachiplusia ou (RoM NPV) were found to be infectious in TN-368 cells employing the calcium phosphate precipitation technique (F.L. Graham and A.J. van der Eb, Virology, 52, 456-467, 1973). ...

  19. Identification of two functional nuclear localization signals in the capsid protein of duck circovirus.

    Science.gov (United States)

    Xiang, Qi-Wang; Zou, Jin-Feng; Wang, Xin; Sun, Ya-Ni; Gao, Ji-Ming; Xie, Zhi-Jing; Wang, Yu; Zhu, Yan-Li; Jiang, Shi-Jin

    2013-02-01

    The capsid protein (CP) of duck circovirus (DuCV) is the major immunogenic protein and has a high proportion of arginine residues concentrated at the N terminus of the protein, which inhibits efficient mRNA translation in prokaryotic expression systems. In this study, we investigated the subcellular distribution of DuCV CP expressed via recombinant baculoviruses in Sf9 cells and the DNA binding activities of the truncated recombinant DuCV CPs. The results showed that two independent bipartite nuclear localization signals (NLSs) situated at N-terminal 1-17 and 18-36 amino acid residue of the CP. Moreover, two expression level regulatory signals (ELRSs) and two DNA binding signals (DBSs) were also mapped to the N terminus of the protein and overlapped with the two NLSs. The ability of CP to bind DNA, coupled with the karyophilic nature of this protein, strongly suggests that it may be responsible for nuclear targeting of the viral genome. PMID:23174505

  20. Book review: Baculovirus Molecular Biology, Second Edition

    Science.gov (United States)

    The application of cell culture and molecular biology methodologies to the study of baculoviruses has resulted in an explosion of information on this group of insect pathogens. The quantity of the corresponding literature on baculoviruses has reached a level difficult for any one researcher to mast...

  1. Localization of Nuclear Equipment and Reliability

    International Nuclear Information System (INIS)

    In Korea, the recent increase of the nuclear share of total power generation has inevitably raised the necessity of localization of nuclear equipment and as far as hardware is concerned, a remarkable improvement in the localization ratio has been achieved since the construction of KNU 5 and 6. Governmental support, such as the policy of unification of power plant suppliers, has also been effective in achieving the ultimate goal of localization of nuclear plant equipment. In this foundation stage, KHIC faced numerous problems arising from the lack of technology and skilled man-power. However, as a result of its 5 year endeavor, KHIC has not only successfully produced the equipment for KNU 7 and 8 in 1981, but has also attained an international reputation in such various fields as material production, precision fabrication and quality control. Now, KHIC is competitive enough to push the KNU 11 and 12 project forward as a main contractor of equipment supply, just as the foreign supplier did in previous projects. Consequently, the central idea of this paper will explored mainly on the status of localization we have performed and on the future planning will perform. In KNU 11 and 12 under preparation, KHIC is pushing business as a prime contractor of equipment supplier and also is giving their pioneering effort to the increasing of their localization ratio and to the safety under reliability of major component to be produced

  2. Nuclear technologies for local energy systems

    International Nuclear Information System (INIS)

    If nuclear energy is to realize its full potential as a safe and cost-effective alternative to fossil fuels, applications beyond those that are currently being serviced by large, central nuclear power stations must be identified and appropriate reactors developed. The Canadian program on reactor systems for local energy supply is at the forefront of these developments. This program emphasizes design simplicity, low power density and fuel rating, reliance on natural processes, passive systems, and reduced reliance on operator action. The first product, the SLOWPOKE Energy System, is a 10 MW heat source specifically designed to provide hot water to satisfy the needs of local heating systems for building complexes, institutions and municipal district heating systems. A demonstration heating reactor has been constructed at the Whiteshell Nuclear Research Establishment in Manitoba and has been undergoing an extensive test program since first operation in 1987 July. Based on the knowledge learned from the design, construction, licensing and operational testing of this facility, the design of the 10 MW commercial-size unit is well advanced, and Atomic Energy of Canada Limited is prepared to commit the construction of the first commercial unit. Although the technical demonstration of the concept is important, it is recognized that another crucial element is the public and regulatory acceptance of small nuclear systems in urban areas. The decision by a community to commit the construction of a SLOWPOKE Energy System brings to a sharp focus the current public apprehension about nuclear technologies

  3. Local AREA networks in advanced nuclear reactors

    International Nuclear Information System (INIS)

    The report assesses Local Area Network Communications with a view to their application in advanced nuclear reactor control and protection systems. Attention is focussed on commercially available techniques and systems for achieving the high reliability and availability required. A basis for evaluating network characteristics in terms of broadband or baseband type, medium, topology, node structure and access method is established. The reliability and availability of networks is then discussed. Several commercial networks are briefly assessed and a distinction made between general purpose networks and those suitable for process control. The communications requirements of nuclear reactor control and protection systems are compared with the facilities provided by current technology

  4. Local economic impact of nuclear power

    International Nuclear Information System (INIS)

    The local economic impact of nuclear installations is examined and the conclusion reached that much of the subsequent area growth may be coincidental to the facility. Nuclear siting criteria favor proximity to a regional power grid, abundant water for cooling, and extensive vacant land with a major access road. These criteria coincide with the characteristics of commuter suburbs, centers for retirement, and recreation areas. Clustering of nuclear units introduces an extraordinary level of new construction, office requirements, and capital. Economic changes will occur at the start and completion of the construction stage and at the time of decommissioning the facility. Past experiences are detailed in terms of employment, payroll, housing, public services, and procurement. When construction is completed, employment falls to a relatively low level. Proximity to the plant offers no advantage in terms of local power rates. While nuclear facilities do not preclude other development in the area, there are restrictions on access, regulatory agencies may reject absorbing the cost of public use as a business expense in the rate structure, and security measures may constrain public use. There is pressure for tax equalization laws to compensate communities for the loss of potential property tax revenues. Some agencies (e.g., the Tennessee Valley Authority) make in-lieu-of-tax payments, while some plants have produced tax benefits large enough to effect significant public improvements. 8 references

  5. RADIOIMMUNOASSAY ANALYSIS OF BACULOVIRUS GRANULINS AND POLYHEDRINS

    Science.gov (United States)

    Granulin and polyhedrin proteins were purified by preparative sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis from the baculoviruses Autographa californica, Rachiplusia ou, Heliothis zea, Heliothis armigera, Trichoplusia ni, and Spodotera frugiperda. Antisera were...

  6. Genetic Modification of Baculovirus Expression Vectors

    Institute of Scientific and Technical Information of China (English)

    Shu-fen Li; Hua-lin Wang; Zhi-hong Hu; Fei Deng

    2012-01-01

    As a protein expression vector,the baculovirus demonstrates many advantages over other vectors.With the development of biotechnology,baculoviral vectors have been genetically modified to facilitate high level expression of heterologous proteins in both insect and mammalian cells.These modifications include utilization of different promoters and signal peptides,deletion or replacement of viral genes for increasing protein secretion,integration of polycistronic expression cassette for producing protein complexes,and baculovirus pseudotyping,promoter accommodation or surface display for enhancing mammalian cell targeting gene delivery.This review summarizes the development and the current state of art of the baculovirus expression system.Further development of baculovirus expression systems will make them even more feasible and accessible for advanced applications.

  7. Classic nuclear localization signals and a novel nuclear localization motif are required for nuclear transport of porcine parvovirus capsid proteins.

    OpenAIRE

    Boisvert, Maude; Bouchard-Lévesque, Véronique; Fernandes, Sandra; Tijssen, Peter

    2014-01-01

    Nuclear targeting of capsid proteins (VPs) is important for genome delivery and precedes assembly in the replication cycle of porcine parvovirus (PPV). Clusters of basic amino acids, corresponding to potential nuclear localization signals (NLS), were found only in the unique region of VP1 (VP1up, for VP1 unique part). Of the five identified basic regions (BR), three were important for nuclear localization of VP1up: BR1 was a classic Pat7 NLS, and the combination of BR4 and BR5 was a classic b...

  8. Unattended nuclear systems for local energy supply

    International Nuclear Information System (INIS)

    This paper describes recent developments in a small nuclear heat and electricity production system - the SLOWPOKE Energy System - that make it possible to locate the system close to the load, and that could have a major impact on local energy supply. The most important unique features arising from these developments are walk-away safety and the ability to operate in an unattended mode. Walk-away safety means that radiological protection is provided by intrinsic characteristics and does not depend on either engineered safety systems or operator intervention. This, in our view, is essential to public acceptance. The capability for unattended operation results from self-regulation; however, the performance can be remotely monitored. The SLOWPOKE Energy System consists of a water-filled pool, operating at atmospheric pressure, which cools and moderates a beryllium-reflected thermal reactor that is fuelled with 100 to 400 kg of low-enriched uranium. The pool water also provides shielding from radioactive materials trapped in the fuel. Heat is drawn from the pool and transferred either to a building hot-water distribution system or to an organic liquid which is converted to vapour to drive a turbine-generator unit. Heating loads between 2 qnd 10 MWt, and electrical loads up to 1 MWe can be satisfied. SLOWPOKE is a dramatic departure from conventional nuclear power reactors. Its nuclear heat source is intrinsically simple, having only one moving part: a solid neutron absorber which is slowly withdrawn from the reactor to balance the fuel burnup. Its power is self-regulated and excessive heat production cannot occur, even for the most severe combinations of system failure. Cooling of the fuel is assured by natural physical processes that do not depend on mechanical components such as pumps. These intrinsic characteristics assure public safety and ultra high reliability

  9. Classic nuclear localization signals and a novel nuclear localization motif are required for nuclear transport of porcine parvovirus capsid proteins.

    Science.gov (United States)

    Boisvert, Maude; Bouchard-Lévesque, Véronique; Fernandes, Sandra; Tijssen, Peter

    2014-10-01

    Nuclear targeting of capsid proteins (VPs) is important for genome delivery and precedes assembly in the replication cycle of porcine parvovirus (PPV). Clusters of basic amino acids, corresponding to potential nuclear localization signals (NLS), were found only in the unique region of VP1 (VP1up, for VP1 unique part). Of the five identified basic regions (BR), three were important for nuclear localization of VP1up: BR1 was a classic Pat7 NLS, and the combination of BR4 and BR5 was a classic bipartite NLS. These NLS were essential for viral replication. VP2, the major capsid protein, lacked these NLS and contained no region with more than two basic amino acids in proximity. However, three regions of basic clusters were identified in the folded protein, assembled into a trimeric structure. Mutagenesis experiments showed that only one of these three regions was involved in VP2 transport to the nucleus. This structural NLS, termed the nuclear localization motif (NLM), is located inside the assembled capsid and thus can be used to transport trimers to the nucleus in late steps of infection but not for virions in initial infection steps. The two NLS of VP1up are located in the N-terminal part of the protein, externalized from the capsid during endosomal transit, exposing them for nuclear targeting during early steps of infection. Globally, the determinants of nuclear transport of structural proteins of PPV were different from those of closely related parvoviruses. Importance: Most DNA viruses use the nucleus for their replication cycle. Thus, structural proteins need to be targeted to this cellular compartment at two distinct steps of the infection: in early steps to deliver viral genomes to the nucleus and in late steps to assemble new viruses. Nuclear targeting of proteins depends on the recognition of a stretch of basic amino acids by cellular transport proteins. This study reports the identification of two classic nuclear localization signals in the minor capsid

  10. Local competence building and public information in European nuclear territories

    International Nuclear Information System (INIS)

    Opinion polls and case study analysis show that there is a strong public demand for more participation in decision-making processes relating to the environment and nuclear issues specifically (IGNA, 2007; EC DG TREN 2008). In this regard, the implementation of transparency and participation at local level in municipalities hosting nuclear facilities is being claimed by local representatives over Europe. The pilot project 'Local Competence Building and Public Information in European Nuclear Territories' (2007-2008) was promoted by the Group of European Municipalities with Nuclear Facilities (GMF) and partly financed by the Directorate General for Energy and Transport (DG TREN). The project aims at supporting the European Commission (EC) in the process of harmonising national practices in the field of governance in nuclear territories. The objectives of this pilot project were two-fold: firstly, to identify local good practices at European level with regards to governance in the nuclear field and, secondly, to provide a methodological framework to recommend good practices and establish a set of indicators. The results of the project are briefly explained under the five dimensions of local governance on nuclear matters: access to environmental information, participation, access to justice, competence building and local development. Firstly, access to information on nuclear issues is broadly acknowledged by local representatives to be of relevant importance in increasing public knowledge on nuclear issues, especially on nuclear safety and radiation protection. Secondly, most local representatives feel that the current participatory level on nuclear issues in municipalities is too low and they would appreciate a greater involvement of local communities in decision-making processes on nuclear issues through local commissions or other mechanisms. Thirdly, according to local authorities, access to justice should be provided not only for the public but also for themselves

  11. Pathogenesis induced by (recombinant) baculoviruses in insects.

    NARCIS (Netherlands)

    Flipsen, J.Th.M.

    1995-01-01

    Infection of insect larvae by a baculovirus leads to cessation of feeding and finally to the death of the larva. Under optimal conditions this process may take as little as five days during which the virus multiplies approximately a billion times and transforms 30% of the larval weight into viral pr

  12. Standard problem of nuclear power localization in China

    International Nuclear Information System (INIS)

    French nuclear power standard system includes the French nuclear laws, principal safety guides, RCC serial standard and technical documents. The RCC serial standard reaches on the design and construction of nuclear power plants, and is principally a closed standard system. The RCC serial standard have been used for the design and construction of Qinshan phase-II, the RCC-M have been used for the mechanical components of nuclear island. It is suggested that the Chinese nuclear power governing body research the problems relating nuclear power standard during the localization in China, the present condition of the Chinese nuclear power standards, and begin to develop the Chinese nuclear power standard based on the ASMEB and PVC, RCC serial standard and Chinese practical experienced in nuclear power engineering. Chinese nuclear power standard should be the strict, well applicable and advanced in the world, and should be continuously updated according to the accumulated operating experiences, technological progress and changes in regulatory requirement

  13. Japan's nuclear PA activity in local governments

    International Nuclear Information System (INIS)

    This presentation emphasises some points of PA activities, based on the experience of 'cooperation projects for local governments'. Local governments distribute the public information directly to the residents. This is very important because officers of the local government can be the opinion leaders of the region. Local government exist very close to the residents, while the central government is a distant and faceless existence for the local people. It is believed that the local governments play an imperative role in PA activities. In other words, we must further utilize the organizations and functions of the local governments to implement PA activities. In conclusion, three recommendations are offered. Firstly, enough budget and authority should be given to the local governments as far as PA activities in their areas are concerned, and most of such activities should be entrusted to the local governments. Local governments should place more public relations officers, and continue the manpower development. Second, with regard to highly technical or specialized issues which a local governments cannot treat alone, related organizations like JAERO should support their PA activities. Third, such related organizations should also cooperate with local government including assistance in providing know-how, when their public information activities focus on educators, journalists, or the women. These three points should be given due consideration in our cooperation projects for the local governments, and JAERO is doing its best every day

  14. Baculovirus Insecticides in Latin America: Historical Overview, Current Status and Future Perspectives

    Directory of Open Access Journals (Sweden)

    Santiago Haase

    2015-04-01

    Full Text Available Baculoviruses are known to regulate many insect populations in nature. Their host-specificity is very high, usually restricted to a single or a few closely related insect species. They are amongst the safest pesticides, with no or negligible effects on non-target organisms, including beneficial insects, vertebrates and plants. Baculovirus-based pesticides are compatible with integrated pest management strategies and the expansion of their application will significantly reduce the risks associated with the use of synthetic chemical insecticides. Several successful baculovirus-based pest control programs have taken place in Latin American countries. Sustainable agriculture (a trend promoted by state authorities in most Latin American countries will benefit from the wider use of registered viral pesticides and new viral products that are in the process of registration and others in the applied research pipeline. The success of baculovirus-based control programs depends upon collaborative efforts among government and research institutions, growers associations, and private companies, which realize the importance of using strategies that protect human health and the environment at large. Initiatives to develop new regulations that promote the use of this type of ecological alternatives tailored to different local conditions and farming systems are underway.

  15. Impacts of nuclear generating plants on local areas

    International Nuclear Information System (INIS)

    Impacts resulting from building and operating twelve nuclear generating stations were not generally characteristic of impacts associated with resource-based energy developments. Economic benefits to local areas were not large due to significant commutation levels, income leakage from the rural areas, and relatively few local purchases of construction material. Revenue impacts from nuclear plants have shown important gains to local jurisdictions. Nevertheless, revenue impacts were found to be wide ranging, depending on state utility tax structures. Socioeconomic changes due to nuclear generating plants have not resulted in public expressions of concern but have sensitized communities to growth management issues. 15 references, 1 figure, 4 tables

  16. Production of human c-myc protein in insect cells infected with a baculovirus expression vector.

    OpenAIRE

    Miyamoto, C.; Smith, G. E.; Farrell-Towt, J; Chizzonite, R.; Summers, M D; Ju, G.

    1985-01-01

    A cDNA fragment coding for human c-myc was inserted into the genome of the baculovirus Autographa californica nuclear polyhedrosis virus adjacent to the strong polyhedrin promoter. Insect cells infected with the recombinant virus produced significant amounts of c-myc protein, which constituted the major phosphoprotein component in these cells. By immunoprecipitation and immunoblot analysis, two proteins of 61 and 64 kilodaltons were detected with c-myc-specific antisera. The insect-derived pr...

  17. Insecticidal properties of genetically engineered baculoviruses expressing an insect juvenile hormone esterase gene.

    OpenAIRE

    Eldridge, R; O'Reilly, D R; Hammock, B D; Miller, L K

    1992-01-01

    Exploring the possibility of enhancing the properties of baculoviruses as biological control agents of insect pests, we tested the effect of expressing an insect gene (jhe) encoding juvenile hormone esterase. Juvenile hormone esterase inactivates juvenile hormone, which regulates the outcome of an insect molt. A cDNA encoding the juvenile hormone esterase of Heliothis virescens was inserted into the genome of Autographa californica nuclear polyhedrosis virus such that the gene was expressed u...

  18. Efficient expression of histidine-tagged large hepatitis delta antigen in baculovirus-transduced baby hamster kidney cells

    Institute of Scientific and Technical Information of China (English)

    Ying-Wei Chiang; Jaw-Chin Wu; Kuei-Chun Wang; Chia-Wei Lai; Yao-Chi Chung; Yu-Chen Hu

    2006-01-01

    AIM: To study the baculovirus/mammalian cell system for efficient expression of functional large hepatitis delta antigen (L-HDAg).METHODS: A recombinant baculovirus expressing histidine-tagged L-HDAg (L-HDAgH) was constructed to transduce baby hamster kidney (BHK) cells by a simplified transduction protocol.RESULTS: The recombinant baculovirus transduced BHK cells with efficiencies higher than 90% as determined by flow cytometry. The expression level was significantly higher than that obtained by plasmid transfection and was further enhanced 3-fold to around 19 pg/cell by the addition of 10 mmol/L sodium butyrate. Importantly,the expressed L-HDAgH was localized to the cell nucleus and correctly isoprenylated as determined by immunofluorescence labeling and confocal microscopy.Moreover, L-HDAgH interacted with hepatitis B surface antigen to form virus-like particles.CONCLUSION: The fusion with histidine tags as well as overexpression of L-HDAgH in the baculovirus-transduced BHK cells does not impair the biological functions. Taken together, the baculovirus/mammalian cell system offers an attractive alternative for high level expression of L-HDAgH or other proteins that require extensive posttranslational modifications.

  19. Fundamentals of Baculovirus Expression and Applications.

    Science.gov (United States)

    Kost, Thomas A; Kemp, Christopher W

    2016-01-01

    In 1982 E. coli produced human insulin, the world's first recombinant DNA drug, was approved by the FDA. Since this historical event, remarkable progress has been made in developing bacterial, yeast, mammalian and insect cell protein expression systems that are used to produce recombinant proteins for both research and clinical applications. Of the available approaches, the insect cell based baculovirus expression vector system (BEVS) has proven to be a particularly adaptable system for producing a diverse collection of proteins. Along with E. coli, the system has been valuable for the production of proteins for structural studies, including adequate quantities of difficult to produce G protein-coupled receptors. BEVS has also been used for production of the human papilloma virus vaccine, Cervarix, the first FDA approved insect cell produced product and FluBlok, a vaccine based on the influenza virus hemagglutinin protein. Baculoviruses, modified to contain mammalian promoters (BacMam viruses), have proven to be efficient gene delivery vectors for mammalian cells and provide an alternative transient mammalian cell based protein expression approach to that of plasmid DNA based transfection methodologies. Here we provide an update on recent advances in baculovirus vector development with a focus on the numerous applications of these viruses in basic research and biotechnology. PMID:27165326

  20. Understanding local residents of Korea using nuclear effective safety

    International Nuclear Information System (INIS)

    The risk perception gap between experts and lay people is based on the use of different concept on risk. It is getting increasingly important for nuclear practitioners to understand the lay people's subjective perception on nuclear safety. We proposed the nuclear effective safety index (NESI) which is based on data of the public survey of local inhabitants. We extracted the four factors for effective safety indicators; communication, trust, plant emergency response capability, and personal emergency coping skills. The latest NESI was 41.54, which was increased from 38.22 but still low. The three-year data of NESI showed the differences between genders and between sites as well as trend. The survey of antecedents of effective safety showed some meaningful events and profound differences between plant employees and local inhabitants. The NESI can be utilized as useful communication tool between the local inhabitants and nuclear practitioners. (authors)

  1. Nuclear power generation and co-existence with local area

    International Nuclear Information System (INIS)

    In addition to originally strong anxiety and unreliability on nuclear power generation, frequent accidents and troubles in the old Nuclear Reactor and Fuels Development Incorporation (the new Nuclear Fuel Cycle Development Incorporation), a fact that numbers of opposition group was over a half at citizen's vote conducted first in Japan and at Maki-machi in Niigata prefecture in August, 1996 on location of a nuclear power generation of the Tohoku Electric Co., Ltd., and so forth, affects some ripple effects to make difficult to form consensus on new location of a nuclear power generation. On making consensus of such location, every electric companies practise good communication with location areas, communication and other organizations, and so forth, promote positive information supplies and conversation type understanding works, and effort to acquire reliability for people of the location areas. Together with such efforts, they accumulate steadily safe operation results of the present nuclear power plants and develop nuclear PA (public acceptance) actions, local area promotion supporting countermeasures, and so forth. Here was introduced on a part of actions aiming at assistance to and co-existence with local areas promoted by every electric companies on various forms. (G.K.)

  2. Initial operations in local nuclear emergency response headquarter

    International Nuclear Information System (INIS)

    As a result of the Fukushima nuclear accident due to the Great East Japan Earthquake and the tsunami that occurred thereafter, local nuclear emergency response headquarters (local headquarters) was set up at off-site center (OFC). However, several obstacles such as the collapse of means of communication resulting from severed communication lines, food and fuel shortage resulting from stagnant physical distribution, and increasing radiation dose around the center significantly restricted originally intended operation of local headquarters. In such severe situation, the personnel gathered at the OFC from the government, local public bodies and electric companies from March 11 to 15 acted without sufficient food, sleep or rest and did all they could against successively occurring unexpected challenges by using limited means of communication. However, issues requiring further consideration were activities of each functional group, location of OFC and the functions of equipment, machines and materials and reflecting the consideration results into future protective measures and revision of the manual for nuclear emergency response were greatly important. This report described investigated results on initial operations in local headquarters such as situation of activities conducted by local headquarters and operations at functional groups. (T. Tanaka)

  3. Hormone activation of baculovirus expressed progesterone receptors.

    Science.gov (United States)

    Elliston, J F; Beekman, J M; Tsai, S Y; O'Malley, B W; Tsai, M J

    1992-03-15

    Human and chicken progesterone receptors (A form) were overproduced in a baculovirus expression system. These recombinant progesterone receptors were full-length bound progesterone specifically and were recognized by monoclonal antibodies, AB52 and PR22, specific for human and chicken progesterone receptor, respectively. In gel retardation studies, binding of recombinant human and chicken progesterone receptors to their progesterone response element (PRE) was specific and was enhanced in the presence of progesterone. Binding of human progesterone receptor to the PRE was also enhanced in the presence of the antiprogestin, RU486, but very little effect was observed in the presence of estradiol, dexamethasone, testosterone, and vitamin D. In our cell-free transcription system, human progesterone receptor induced transcription in a receptor-dependent and hormone-activable manner. Receptor-stimulated transcription required the presence of the PRE in the test template and could be specifically inhibited by excess PRE oligonucleotides. Furthermore, chicken progesterone receptor also induced in vitro transcription in a hormone-activable manner. These results demonstrate that steroid receptors overexpressed in a baculovirus expression system are functional and exhibit steroid-responsive binding and transcription. These observations support our present understanding of the mechanism of steroid receptor-regulated gene expression and provide a technological format for studies of the role of hormone and antihormone in altering gene expression. PMID:1544902

  4. State and local government regulation of nuclear waste transportation

    International Nuclear Information System (INIS)

    The Hazardous Materials Transportation Act (HMTA), authorizes the U.S. Department of Transportation (DOT) to promulgate rules governing the safe transportation in commerce of hazardous materials, including nuclear waste. The HMTA further provides that any state or local government requirement is preempted and thus invalid if it is inconsistent with a DOT requirement issued pursuant to the HMTA

  5. Fauna in nuclear power plant localities on Czechoslovak territory. I

    International Nuclear Information System (INIS)

    The territory of the CSSR comprises seven types of fauna: 1. arboreal, 2. steppe-eremial, 3. oreo-tundral, 3. fauna spread over large areas, 5. special fauna of the limnic biocycle, 6. epiareal elements of terrestrial fauna and 7. anthropogenically conditioned fauna. Each group has its characteristics and examples are given of animals belonging to the said groups. On the basis of a survey of local fauna and data on a nuclear power plant sited in the respective locality measures may be taken to protect local fauna and to predict its further development. (E.S.). 8 refs

  6. The effects of the nuclear disaster at Fukushima Daiichi Nuclear Power Station on local governments

    International Nuclear Information System (INIS)

    All Japan council of local governments with atomic power stations consisted of 24 reactor site and 6 neighboring local governments to solve reactor site related problems. Nuclear disaster at Fukushima Daiichi Nuclear Power Station affected member local governments to be forced in severe conditions such as 'removal of administrative function' or 'refuge over a wide district beyond local government area', not imagined before. The council set up working group for thirteen local governments themselves to investigate this disaster and find safety and prevention of disaster measures to be deployed in nuclear administration, which published report in March 2012. This article described outline of investigation and derived problems and direction of their solution. Main items were related with communication, resident evacuation, prevention of disaster system, and management of refuge site. (T. Tanaka)

  7. The ANCLI White Paper on Local Governance of Nuclear Sites

    International Nuclear Information System (INIS)

    The 1981 circular, 'Information Commissions attached to Power Stations', was an important regulatory step, which enabled local communities to come together to discuss nuclear issues with the site operators and official inspectorates. However the Local Information Commissions (CLI), have encountered numerous obstacles, both practical and legislative, in fulfilling their role as information providers and, particularly, as monitors. It is now acknowledged in France both by members of commissions at local level and by public authorities that CLI would benefit from a more statutory role in the decision-making process as well as from a specific legal statute associated with independent resources. The forthcoming Nuclear Safety and Transparency Bill should establish a new legal and institutional framework for local commissions. The National Association of CLI (ANCLI) gathers and represents the CLI at national level. In Spring 2005, in the perspective of the Nuclear Safety and Transparency bill, the ANCLI initiated a participatory process involving about 30 delegates from a dozen CLI (elected representatives, NGOs, local experts) in order to raise a common expertise and make recommendations on this new legal statute, on the basis of their own return of experience. The White Paper is structured around four main subject areas: Structure and status of CLIs, local powers; - Role of CLIs and ANCLI in the national and international decision-making process; Local communities' access to specialist expertise; and Waste management. The CLIs reaffirm their local powers and ability to best assure the continued monitoring of nuclear sites and any other nuclear issues in their area. In particular, this local authority allows them to balance different risks as well as political and economic decisions which have a bearing on their region's future. It implies a certain autonomy on the part of CLIs in the exercise of their scrutiny functions, evaluating all the factors impacting both on the

  8. Regulation of Nuclear Localization of Signaling Proteins by Cytokinin

    Energy Technology Data Exchange (ETDEWEB)

    Kieber, J.J.

    2010-05-01

    Cytokinins are a class of mitogenic plant hormones that play an important role in most aspects of plant development, including shoot and root growth, vascular and photomorphogenic development and leaf senescence. A model for cytokinin perception and signaling has emerged that is similar to bacterial two-component phosphorelays. In this model, binding of cytokinin to the extracellular domain of the Arabidopsis histidine kinase (AHKs) receptors induces autophosphorylation within the intracellular histidine-kinase domain. The phosphoryl group is subsequently transferred to cytosolic Arabidopsis histidine phosphotransfer proteins (AHPs), which have been suggested to translocate to the nucleus in response to cytokinin treatment, where they then transfer the phosphoryl group to nuclear-localized response regulators (Type-A and Type-B ARRs). We examined the effects of cytokinin on AHP subcellular localization in Arabidopsis and, contrary to expectations, the AHPs maintained a constant nuclear/cytosolic distribution following cytokinin treatment. Furthermore, mutation of the conserved phosphoacceptor histidine residue of the AHP, as well as disruption of multiple cytokinin signaling elements, did not affect the subcellular localization of the AHP proteins. Finally, we present data indicating that AHPs maintain a nuclear/cytosolic distribution by balancing active transport into and out of the nucleus. Our findings suggest that the current models indicating relocalization of AHP protein into the nucleus in response to cytokinin are incorrect. Rather, AHPs actively maintain a consistent nuclear/cytosolic distribution regardless of the status of the cytokinin response pathway.

  9. Nuclear materials transportation workshops: USDOE outreach to local governments

    International Nuclear Information System (INIS)

    To provide direct outreach to local governments, the Transportation Management Division of the United States Department of Energy asked the Urban Consortium and its Energy Task Force to assemble representatives for two workshops focusing on the transport of nuclear materials. The first session, for jurisdictions east of the Mississippi River, was held in New Orleans on May 5--6, 1988; the second was conducted on June 6--7, 1988 in Denver for jurisdictions to the west. Twenty local government professionals with management or operational responsibility for hazardous materials transportation within their jurisdictions were selected to attend each workshop. The discussions identified five major areas of concern to local government professionals; coordination; training; information resources; marking and placarding; and responder resources. Integrated federal, state, and local levels of government emerged as a priority coordination issue along with the need for expanded availability of training and training resources for first-reponders

  10. Ancient Coevolution of Baculoviruses and Their Insect Hosts

    OpenAIRE

    Elisabeth A Herniou; Olszewski, Julie A.; O'Reilly, David R.; Jenny S Cory

    2004-01-01

    If the relationships between baculoviruses and their insect hosts are subject to coevolution, this should lead to long-term evolutionary effects such as the specialization of these pathogens for their hosts. To test this hypothesis, a phylogeny of the Baculoviridae, including 39 viruses from hosts of the orders Lepidoptera, Diptera, and Hymenoptera, was reconstructed based on sequences from the genes lef-8 and ac22. The tree showed a clear division of the baculoviruses according to the order ...

  11. Baculoviruses as Vectors for Gene Therapy against Human Prostate Cancer

    OpenAIRE

    Stanbridge Lindsay J.; Dussupt Vincent; Maitland Norman J.

    2003-01-01

    Current curative strategies for prostate cancer are restricted to the primary tumour, and the effect of treatments to control metastatic disease is not sustained. Therefore, the application of gene therapy to prostate cancer is an attractive alternative. Baculoviruses are highly restricted insect viruses, which can enter, but not replicate in mammalian cells. Baculoviruses can incorporate large amounts of extra genetic material, and will express transgenes in mammalian cells when under the co...

  12. Functional analysis of a novel baculovirus envelope fusion protein

    OpenAIRE

    Westenberg, M.

    2004-01-01

    Baculoviridae are a family of large double stranded DNA viruses that are exclusively pathogenic to arthropods. Baculoviruses have been studied i) with the aim to develop alternatives to chemical pest control, ii) for their application asaneukaryotic expression system to express heterologous proteins, and recently iii) as gene delivery vehicle in gene therapy. Baculoviruses cluster into two distinct genera on the basis of the occlusion body (OB) morphology: Nucleopolyhedrovirus (NPV) and Granu...

  13. Seismic test of local control panel for nuclear power plant

    International Nuclear Information System (INIS)

    The safety related equipment installed in Nuclear Power Plant are required to perform a safety function during and after a seismic event. To accomplish this safety function, they must be seismically qualified in accordance with the intent and requirements of the USNRC Regulatory Guide 1.100 and IEEE Std. 344. The local control panel is a safety related equipment which provide operators with control, display and monitoring for plant operating conditions. This paper describes test procedure for seismic qualification test of local control panel which is installed in Ulchin units 3 and 4. (author)

  14. Application of a nuclear localization signal gene in transgene mice

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Efficient gene transfer by cytoplasm co-injec- tion will offer a powerful means for transgenic animals. Using co-injection in cytoplasm, two independent gene constructs, including bovine (?-s1-casein-hG-CSF and a mammal expression vector expressing a nuclear localization signal (mNLS), were introduced into fertilized mouse eggs. The target gene construct was docked into host nucleus probably by the nuclear localization signal. Transgene mice have been obtained at 58% (29/50) of integration ratio. Expression level of the positive transgene mice was detected by Western blotting. Maximal expression of human G-CSF was estimated about 540 mg/L of milk. The expression ratio was up to 75% (9/12). The results here have important practical implications for the generation of mammary gland bioreactors and other transgene studies. Co-injection of a target gene with an expression vector of a mammal nuclear localization signal by cytoplasm appears to be a useful, efficient and easy strategy for generating transgenic animals, which may be able to substitute the routine method of pronucleus-injection of fertilized eggs.

  15. Nuclear surface localization of preequilibrium reactions at low energies

    International Nuclear Information System (INIS)

    Average quantities related to characteristics of nucleon-nucleon (NN) interaction along the trajectory of a projectile in preequilibrium reactions are calculated using the semiclassical method to follow the incoming particle path in the nuclear target. The radial dependences of the nucleon's mean free path and the probability for the first NN collision have pointed out the surface character of the first NN interaction in multistep reactions even at low energies. In the local density approximation an average Fermi energy and an average strength of the effective NN interaction Vo along the trajectory of the incident nucleon are obtained with respect to both the nuclear nuclear density and the first NN collision probability. A good agreement is found between the average strengths obtained with the Hartree-Fock potential plus the dispersive component and by using the parametrization based on Brueckner - Hartree - Fock nuclear matter calculations. It is also shown that the nuclear-density dependence of the effective NN interaction may account for the low-energy phenomenological Vo - values which are much more increased in comparison with any predictions

  16. Cellular stress stimulates nuclear localization signal (NLS) independent nuclear transport of MRJ

    International Nuclear Information System (INIS)

    HSP40 family member MRJ (DNAJB6) has been in the spot light for its relevance to Huntington's, Parkinson's diseases, limb-girdle muscular dystrophy, placental development, neural stem cells, cell cycle and malignancies such as breast cancer and melanoma. This gene has two spliced variants coding for 2 distinct proteins with significant homology. However, MRJ(L) (large variant) is predominantly localized to the nucleus whereas MRJ(S) (small variant) is predominantly cytoplasmic. Interestingly MRJ(S) translocates to the nucleus in response to heat shock. The classical heat shock proteins respond to crises (stress) by increasing the number of molecules, usually by transcriptional up-regulation. Our studies imply that a quick increase in the molar concentration of MRJ in the nuclear compartment is a novel method by which MRJ responds to stress. We found that MRJ(S) shows NLS (nuclear localization signal) independent nuclear localization in response to heat shock and hypoxia. The specificity of this response is realized due to lack of such response by MRJ(S) when challenged by other stressors, such as some cytokines or UV light. Deletion analysis has allowed us to narrow down on a 20 amino acid stretch at the C-terminal region of MRJ(S) as a potential stress sensing region. Functional studies indicated that constitutive nuclear localization of MRJ(S) promoted attributes of malignancy such as proliferation and invasiveness overall indicating distinct phenotypic characteristics of nuclear MRJ(S).

  17. Cellular stress stimulates nuclear localization signal (NLS) independent nuclear transport of MRJ

    Energy Technology Data Exchange (ETDEWEB)

    Andrews, Joel F.; Sykora, Landon J.; Barik Letostak, Tiasha; Menezes, Mitchell E.; Mitra, Aparna [Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL (United States); Barik, Sailen [Center for Gene Regulation in Health and Disease, Department of Biological, Geological, and Environmental Sciences, College of Science, Cleveland State University, Cleveland, OH (United States); Shevde, Lalita A. [Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL (United States); Samant, Rajeev S., E-mail: rsamant@usouthal.edu [Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL (United States)

    2012-06-10

    HSP40 family member MRJ (DNAJB6) has been in the spot light for its relevance to Huntington's, Parkinson's diseases, limb-girdle muscular dystrophy, placental development, neural stem cells, cell cycle and malignancies such as breast cancer and melanoma. This gene has two spliced variants coding for 2 distinct proteins with significant homology. However, MRJ(L) (large variant) is predominantly localized to the nucleus whereas MRJ(S) (small variant) is predominantly cytoplasmic. Interestingly MRJ(S) translocates to the nucleus in response to heat shock. The classical heat shock proteins respond to crises (stress) by increasing the number of molecules, usually by transcriptional up-regulation. Our studies imply that a quick increase in the molar concentration of MRJ in the nuclear compartment is a novel method by which MRJ responds to stress. We found that MRJ(S) shows NLS (nuclear localization signal) independent nuclear localization in response to heat shock and hypoxia. The specificity of this response is realized due to lack of such response by MRJ(S) when challenged by other stressors, such as some cytokines or UV light. Deletion analysis has allowed us to narrow down on a 20 amino acid stretch at the C-terminal region of MRJ(S) as a potential stress sensing region. Functional studies indicated that constitutive nuclear localization of MRJ(S) promoted attributes of malignancy such as proliferation and invasiveness overall indicating distinct phenotypic characteristics of nuclear MRJ(S).

  18. Additive effect of calreticulin and translation initiation factor eIF4E on secreted protein production in the baculovirus expression system

    NARCIS (Netherlands)

    Teng, C.Y.; Oers, van M.M.; Wu, T.Y.

    2013-01-01

    The baculovirus expression vector system is widely used for the production of recombinant proteins. However, the yield of membrane-bound or secreted proteins is relatively low when compared with intracellular or nuclear proteins. In a previous study, we had demonstrated that the co-expression of the

  19. ANCLI's white book about the local governance of nuclear activities

    International Nuclear Information System (INIS)

    This white book is the result of the work carried out by the local commissions of information about nuclear activities (CLI) with the objective of making a common status and recommendations about 4 main topics: the clear definition of the structure and juridical status of CLI's and the acknowledgement of their competence at the local scale, the role and participation of CLI's, and of the national association of CLI's (ANCLI), in the decision processes at the national and international scale, the access of local CLI actors to expertise, and the consideration of ANCLI's opinion in the solutions proposed for the management of radioactive wastes. This last point is taken as an illustration of the previous 3 topics. (J.S.)

  20. An N-terminal nuclear localization sequence but not the calmodulin-binding domain mediates nuclear localization of nucleomorphin, a protein that regulates nuclear number in Dictyostelium.

    Science.gov (United States)

    Myre, Michael A; O'Day, Danton H

    2005-06-24

    Nucleomorphin is a novel nuclear calmodulin (CaM)-binding protein (CaMBP) containing an extensive DEED (glu/asp repeat) domain that regulates nuclear number. GFP-constructs of the 38 kDa NumA1 isoform localize as intranuclear patches adjacent to the inner nuclear membrane. The translocation of CaMBPs into nuclei has previously been shown by others to be mediated by both classic nuclear localization sequences (NLSs) and CaM-binding domains (CaMBDs). Here we show that NumA1 possesses a CaMBD (171EDVSRFIKGKLLQKQQKIYKDLERF195) containing both calcium-dependent-binding motifs and an IQ-like motif for calcium-independent binding. GFP-constructs containing only NumA1 residues 1-129, lacking the DEED and CaMBDs, still localized as patches at the internal periphery of nuclei thus ruling out a direct role for the CaMBD in nuclear import. These constructs contained the amino acid residues 48KKSYQDPEIIAHSRPRK64 that include both a putative bipartite and classical NLS. GFP-bipartite NLS constructs localized uniformly within nuclei but not as patches. As with previous work, removal of the DEED domain resulted in highly multinucleate cells. However as shown here, multinuclearity only occurred when the NLS was present allowing the protein to enter nuclei. Site-directed mutation analysis in which the NLS was changed to 48EF49 abolished the stability of the GFP fusion at the protein but not RNA level preventing subcellular analyses. Cells transfected with the 48EF49 construct exhibited slowed growth when compared to parental AX3 cells and other GFP-NumA1 deletion mutants. In addition to identifying an NLS that is sufficient for nuclear translocation of nucleomorphin and ruling out CaM-binding in this event, this work shows that the nuclear localization of NumA1 is crucial to its ability to regulate nuclear number in Dictyostelium. PMID:15896312

  1. Localization of Vibrating Noise Sources in Nuclear Reactor Cores

    International Nuclear Information System (INIS)

    In this thesis the possibility of locating vibrating noise sources in a nuclear reactor core from the neutron noise has been investigated using different localization methods. The influence of the vibrating noise source has been considered to be a small perturbation of the neutron flux inside the reactor. Linear perturbation theory has been used to construct the theoretical framework upon which the localization methods are based. Two different cases have been considered: one where a one-dimensional one-group model has been used and another where a two-dimensional two-energy group noise simulator has been used. In the first case only one localization method is able to determine the position with good accuracy. This localization method is based on finding roots of an equation and is sensitive to other perturbations of the neutron flux. It will therefore work better with the assistance of approximative methods that reconstruct the noise source to determine if the results are reliable or not. In the two-dimensional case the results are more promising. There are several different localization techniques that reproduce both the vibrating noise source position and the direction of vibration with enough precision. The approximate methods that reconstruct the noise source are substantially better and are able to support the root finding method in a more constructive way. By combining the methods, the results will be more reliable

  2. Installation for accident localization at a nuclear power plant

    International Nuclear Information System (INIS)

    The installation for accident localization at a nuclear power plant with multiloop structure of the first circuit is described. The installation represents a pressurized shell inside which the reactor with the first circuit equipment are located. The whole volume under the shell is divided into autonomous boxes for one or several circulation loops. In case of accident as capacity for condensate discharge not the whole volume under pressurized shell is used, but only one of the autonomous boxes. This circumstance decreases the accident effect zone and expenditures related to the liquidation of accident consequences

  3. Lead localized corrosion evaluation in high activity nuclear waste containers

    International Nuclear Information System (INIS)

    In Argentina, the conceptual basis for the final disposal of high activity nuclear waste was set according to the multiple barrier concept. A lead layer was preselected due to its good radiological protection and corrosion resistance. The focus of present experiments is to assess on the high purity lead corrosion kinetics due to localized attack by aggressive anions, such as chlorides, nitrates and acetates. Hence, high purity lead corrosion potentials with time, potential kinetic and current time measurements at constant potential were performed in synthetic groundwater contaminated with such anions. (author)

  4. AcMNPV As A Model for Baculovirus DNA Replication

    Institute of Scientific and Technical Information of China (English)

    Eric B. Carstens

    2009-01-01

    Baculoviruses were first identified as insect-specific pathogens, and it was this specificity that lead to their use as safe, target specific biological pesticides. For the past 30 years, AcMNPV has served as the subject of intense basic molecular research into the baculovirus infectious cycle including the interaction of the virus with a continuous insect cell line derived from Spodoptera frugiperda. The studies on baculoviruese have led to an in-depth understanding of the physical organization of the viral genomes including many complete genomic sequences, the time course of gene expression, and the application of this basic research to the use of baculoviruses not only as insecticides, but also as a universal eukaryotic protein expression system, and a potential vector in gene therapy. A great deal has also been discovered about the viral genes required for the replication of the baculovirus genome, while much remains to be learned about the mechanism of viral DNA replication. This report outlines the current knowledge of the factors involved in baculovirus DNA replication, using data on AcMNPV as a model for most members of the Baculoviridae.

  5. Gene Acquisition Convergence between Entomopoxviruses and Baculoviruses

    Directory of Open Access Journals (Sweden)

    Julien Thézé

    2015-04-01

    Full Text Available Organisms from diverse phylogenetic origins can thrive within the same ecological niches. They might be induced to evolve convergent adaptations in response to a similar landscape of selective pressures. Their genomes should bear the signature of this process. The study of unrelated virus lineages infecting the same host panels guarantees a clear identification of phyletically independent convergent adaptation. Here, we investigate the evolutionary history of genes in the accessory genome shared by unrelated insect large dsDNA viruses: the entomopoxviruses (EPVs, Poxviridae and the baculoviruses (BVs. EPVs and BVs have overlapping ecological niches and have independently evolved similar infection processes. They are, in theory, subjected to the same selective pressures from their host’s immune responses. Their accessory genomes might, therefore, bear analogous genomic signatures of convergent adaption and could point out key genomic mechanisms of adaptation hitherto undetected in viruses. We uncovered 32 homologous, yet independent acquisitions of genes originating from insect hosts, different eukaryotes, bacteria and viruses. We showed different evolutionary levels of gene acquisition convergence in these viruses, underlining a continuous evolutionary process. We found both recent and ancient gene acquisitions possibly involved to the adaptation to both specific and distantly related hosts. Multidirectional and multipartite gene exchange networks appear to constantly drive exogenous gene assimilations, bringing key adaptive innovations and shaping the life histories of large DNA viruses. This evolutionary process might lead to genome level adaptive convergence.

  6. Ligands specify estrogen receptor alpha nuclear localization and degradation

    Directory of Open Access Journals (Sweden)

    Caze-Subra Stéphanie

    2010-12-01

    Full Text Available Abstract Background The estrogen receptor alpha (ERα is found predominately in the nucleus, both in hormone stimulated and untreated cells. Intracellular distribution of the ERα changes in the presence of agonists but the impact of different antiestrogens on the fate of ERα is a matter of debate. Results A MCF-7 cell line stably expressing GFP-tagged human ERα (SK19 cell line was created to examine the localization of ligand-bound GFP-ERα. We combined digitonin-based cell fractionation analyses with fluorescence and immuno-electron microscopy to determine the intracellular distribution of ligand-bound ERα and/or GFP-ERα. Using fluorescence- and electron microscopy we demonstrate that both endogenous ERα and GFP-ERα form numerous nuclear focal accumulations upon addition of agonist, 17β-estradiol (E2, and pure antagonists (selective estrogen regulator disruptor; SERD, ICI 182,780 or RU58,668, while in the presence of partial antagonists (selective estrogen regulator modulator; SERM, 4-hydroxytamoxifen (OHT or RU39,411, diffuse nuclear staining persisted. Digitonin based cell fractionation analyses confirmed that endogenous ERα and GFP-ERα predominantly reside in the nuclear fraction. Overall ERα protein levels were reduced after estradiol treatment. In the presence of SERMs ERα was stabilized in the nuclear soluble fraction, while in the presence of SERDs protein levels decreased drastically and the remaining ERα was largely found in a nuclear insoluble fraction. mRNA levels of ESR1 were reduced compared to untreated cells in the presence of all ligands tested, including E2. E2 and SERDs induced ERα degradation occurred in distinct nuclear foci composed of ERα and the proteasome providing a simple explanation for ERα sequestration in the nucleus. Conclusions Our results indicate that chemical structure of ligands directly affect the nuclear fate and protein turnover of the estrogen receptor alpha independently of their impact on

  7. Recombinant baculovirus vectors expressing glutathione-S-transferase fusion proteins.

    Science.gov (United States)

    Davies, A H; Jowett, J B; Jones, I M

    1993-08-01

    Recombinant baculoviruses are a popular means of producing heterologous protein in eukaryotic cells. Purification of recombinant proteins away from the insect cell background can, however, remain an obstacle for many developments. Recently, prokaryotic fusion protein expression systems have been developed allowing single-step purification of the heterologous protein and specific proteolytic cleavage of the affinity tag moiety from the desired antigen. Here we report the introduction of these attributes to the baculovirus system. "Baculo-GEX" vectors enable baculovirus production of fusion proteins with the above advantages, but in a eukaryotic post-translational processing environment. Glutathione-S-transferase (GST) fusions are stable cytoplasmic proteins in insect cells and may therefore be released by sonication alone, avoiding the solubility problems and detergent requirements of bacterial systems. Thus large amounts of authentic antigen may be purified in a single, non-denaturing step. PMID:7763917

  8. Baculoviruses as Vectors for Gene Therapy against Human Prostate Cancer

    Directory of Open Access Journals (Sweden)

    Lindsay J. Stanbridge

    2003-01-01

    Full Text Available Current curative strategies for prostate cancer are restricted to the primary tumour, and the effect of treatments to control metastatic disease is not sustained. Therefore, the application of gene therapy to prostate cancer is an attractive alternative. Baculoviruses are highly restricted insect viruses, which can enter, but not replicate in mammalian cells. Baculoviruses can incorporate large amounts of extra genetic material, and will express transgenes in mammalian cells when under the control of a mammalian or strong viral promoter. Successful gene delivery has been achieved both in vitro and in vivo and into both dividing and nondividing cells, which is important since prostate cancers divide relatively slowly. In addition, the envelope protein gp64 is sufficiently mutable to allow targeted transduction of particular cell types. In this review, the advantages of using baculoviruses for prostate cancer gene therapy are explored, and the mechanisms of viral entry and transgene expression are described.

  9. Baculovirus p35 gene is oppositely regulated by P53 and AP-1 like factors in Spodoptera frugiperda

    Energy Technology Data Exchange (ETDEWEB)

    Mohareer, Krishnaveni [Laboratory of Molecular and Cell Biology, Center for DNA Fingerprinting and Diagnostics, Hyderabad 500001 (India); Institute of Life Sciences, University of Hyderabad Campus, Prof. C.R. Rao Road, Gachibowli, Hyderabad 500046 (India); Sahdev, Sudhir [Laboratory of Molecular and Cell Biology, Center for DNA Fingerprinting and Diagnostics, Hyderabad 500001 (India); Ranbaxy Pharmaceuticals, Gurgaon, New Delhi (India); Hasnain, Seyed E., E-mail: seh@bioschool.iitd.ac.in [Institute of Life Sciences, University of Hyderabad Campus, Prof. C.R. Rao Road, Gachibowli, Hyderabad 500046 (India); Kusuma School of Biological Sciences, IIT Delhi, New Delhi 110016 (India); ILBS, Vasant Kunj, New Delhi (India); King Saud University, Riyadh, KSA (Saudi Arabia)

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer Baculovirus p35 is regulated by both viral and host factors. Black-Right-Pointing-Pointer Baculovirus p35 is negatively regulated by SfP53-like factor. Black-Right-Pointing-Pointer Baculovirus p35 is positively regulated by SfAP-1-like factor. -- Abstract: Baculovirus p35 belongs to the early class of genes of AcMNPV and requires viral factors like Immediate Early protein-1 for its transcription. To investigate the role of host factors in regulating p35 gene expression, the putative transcription factor binding sites were examined in silico and the role of these factors in influencing the transcription of p35 gene was assessed. We focused our studies on AP-1 and P53-like factors, which are activated under oxidative stress conditions. The AP-1 motif is located at -1401 while P53 motif is at -1912 relative to p35 translation start site. The predicted AP-1 and P53 elements formed specific complexes with Spodoptera frugiperda nuclear extracts. Both AP-1 and P53 motif binding proteins were down regulated as a function of AcMNPV infection in Spodoptera cells. To address the question whether during an oxidative outburst, the p35 transcription is enhanced; we investigated the role of these oxidative stress induced host transcription factors in influencing p35 gene transcription. Reporter assays revealed that AP-1 element enhances the transcription of p35 by a factor of two. Interestingly, P53 element appears to repress the transcription of p35 gene.

  10. Baculovirus p35 gene is oppositely regulated by P53 and AP-1 like factors in Spodoptera frugiperda

    International Nuclear Information System (INIS)

    Highlights: ► Baculovirus p35 is regulated by both viral and host factors. ► Baculovirus p35 is negatively regulated by SfP53-like factor. ► Baculovirus p35 is positively regulated by SfAP-1-like factor. -- Abstract: Baculovirus p35 belongs to the early class of genes of AcMNPV and requires viral factors like Immediate Early protein-1 for its transcription. To investigate the role of host factors in regulating p35 gene expression, the putative transcription factor binding sites were examined in silico and the role of these factors in influencing the transcription of p35 gene was assessed. We focused our studies on AP-1 and P53-like factors, which are activated under oxidative stress conditions. The AP-1 motif is located at −1401 while P53 motif is at −1912 relative to p35 translation start site. The predicted AP-1 and P53 elements formed specific complexes with Spodoptera frugiperda nuclear extracts. Both AP-1 and P53 motif binding proteins were down regulated as a function of AcMNPV infection in Spodoptera cells. To address the question whether during an oxidative outburst, the p35 transcription is enhanced; we investigated the role of these oxidative stress induced host transcription factors in influencing p35 gene transcription. Reporter assays revealed that AP-1 element enhances the transcription of p35 by a factor of two. Interestingly, P53 element appears to repress the transcription of p35 gene.

  11. Dynamic mitochondrial localization of nuclear transcription factor HMGA1

    International Nuclear Information System (INIS)

    It has been well established that high mobility group A1 (HMGA1) proteins act within the nucleus of mammalian cells as architectural transcription factors that regulate the expression of numerous genes. Here, however, we report on the unexpected cytoplasmic/mitochondrial localization of the HMGA1 proteins within multiple cell types. Indirect immunofluorescence, electron microscopic immunolocalization, and Western blot studies revealed that, in addition to the nucleus, HMGA1 proteins could also be found in both the cytoplasm and mitochondria of randomly dividing populations of wild-type murine NIH3T3 cells and transgenic human MCF-7 breast cancer epithelial cells expressing a hemagglutinin tagged-HMGA1a fusion protein. While the molecular mechanisms underlying these novel subcellular localization patterns have not yet been determined, initial synchronization studies revealed a dynamic, cell cycle-dependent translocation of HMGA1 proteins from the nucleus into the cytoplasm and mitochondria of NIH3T3 cells. Furthermore, preliminary functionality studies utilizing a modified 'chromatin' immunoprecipitation protocol revealed that HMGA1 retains its DNA binding capabilities within the mitochondria and associates with the regulatory D-loop region in vivo. We discuss potential new biological roles for the classically nuclear HMGA1 proteins with regard to the observed nucleocytoplasmic translocation, mitochondrial internalization, and regulatory D-loop DNA binding

  12. Nuclear warhead verification system based on local area network

    International Nuclear Information System (INIS)

    The authors built up a nuclear warhead verification system based on Local Area Network (LAN). The hardware of the system consists of an Intranet server, two personnel computers and a portable high pure germanium gamma-ray spectrometer. The software, Blanking-out, is made of two modules designed for the inspecting and inspected parties respectively. Both of the two modules are interactive and communicate through the Intranet. When the system operates, any sensitive information carried by the high energy resolved gamma-ray spectrum collected by the detector will be meddled by the inspected module before the package of spectral data is sent to the inspecting module to prevent a disclosure of sensitive information. After receiving of the spectral data, the inspecting module will display on the screen of the inspecting PC terminal the verification results in a from of blanked spectrum (spectral blanking-out form) or a form of a piece of phrase (phrase form). The guideline to the spectral blanking-out is that: for those ranges of energy obligated to be inspected, the inspected module should ensure that the inspecting module can display their truly and subjective spectrum; and for the other ranges of energy, the inspected module can blank-out certain parts of the spectra based on the knowledge of sensitive information. Phrase form is rather straight-forward, answering the question whether the inspected warhead is a 'uranium-type', or 'Plutonium-type' or 'non-nuclear type'. The authors conducted a demonstration on some surrogates for nuclear warheads to see whether the nuclear warhead verification system possesses that capability of identifying the type of a warhead and blanking sensitive spectral information. The demonstration was carried out successfully. Phrase form is especially recommendable due to its stronger capability to prevent sensitive information from disclosure and its higher verification credibility. The demonstration also disclosed some deficiencies of the

  13. Gene gymnastics: Synthetic biology for baculovirus expression vector system engineering.

    Science.gov (United States)

    Vijayachandran, Lakshmi S; Thimiri Govinda Raj, Deepak B; Edelweiss, Evelina; Gupta, Kapil; Maier, Josef; Gordeliy, Valentin; Fitzgerald, Daniel J; Berger, Imre

    2013-01-01

    Most essential activities in eukaryotic cells are catalyzed by large multiprotein assemblies containing up to ten or more interlocking subunits. The vast majority of these protein complexes are not easily accessible for high resolution studies aimed at unlocking their mechanisms, due to their low cellular abundance and high heterogeneity. Recombinant overproduction can resolve this bottleneck and baculovirus expression vector systems (BEVS) have emerged as particularly powerful tools for the provision of eukaryotic multiprotein complexes in high quality and quantity. Recently, synthetic biology approaches have begun to make their mark in improving existing BEVS reagents by de novo design of streamlined transfer plasmids and by engineering the baculovirus genome. Here we present OmniBac, comprising new custom designed reagents that further facilitate the integration of heterologous genes into the baculovirus genome for multiprotein expression. Based on comparative genome analysis and data mining, we herein present a blueprint to custom design and engineer the entire baculovirus genome for optimized production properties using a bottom-up synthetic biology approach. PMID:23328086

  14. Studies of the silencing of Baculovirus DNA binding protein

    NARCIS (Netherlands)

    Quadt, I.; Lent, van J.W.M.; Knebel-Morsdorf, D.

    2007-01-01

    Baculovirus DNA binding protein (DBP) binds preferentially single-stranded DNA in vitro and colocalizes with viral DNA replication sites. Here, its putative role as viral replication factor has been addressed by RNA interference. Silencing of DBP in Autographa californica multiple nucleopolyhedrovir

  15. Vaccines for viral and parasitic diseases produced with baculovirus vectors

    NARCIS (Netherlands)

    Oers, van M.M.

    2006-01-01

    The baculovirus¿insect cell expression system is an approved system for the production of viral antigens with vaccine potential for humans and animals and has been used for production of subunit vaccines against parasitic diseases as well. Many candidate subunit vaccines have been expressed in this

  16. Characterization of cell-surface determinants important for baculovirus infection.

    Science.gov (United States)

    Tani, H; Nishijima, M; Ushijima, H; Miyamura, T; Matsuura, Y

    2001-01-01

    Baculovirus gp64 envelope glycoprotein is a major component of the envelope of the budded virus and is involved in virus entry into the host cells by endocytosis. To investigate the cell-surface molecules important for infection of baculovirus into mammalian cells, we constructed a recombinant baculovirus, Ac64-CAluc, which has gp64 and luciferase genes under the polyhedrin and the CAG promoter, respectively. For controls, we constructed recombinant viruses possessing vesicular stomatitis virus (VSV) G protein, mouse hepatitis virus (MHV) S protein, or green fluorescent protein (GFP) gene under the polyhedrin promoter and the luciferase gene under the CAG promoter (AcVSVG-CAluc, AcMHVS-CAluc, and AcGFP-CAluc). Treatment of HepG2 cells with phospholipase C markedly reduced the reporter gene expression by Ac64-CAluc or AcVSVG-CAluc in a dose-dependent manner, whereas AcMHVS-CAluc was shown to be resistant to the treatment. Inhibition with purified lipids and susceptibility to the mutant CHO hamster cell lines deficient in phospholipids synthesis suggest that the interaction of gp64 and phospholipids on the cell surface might play an important role in baculovirus infection into mammalian cells. PMID:11145915

  17. Construction of a host range-expanded hybrid baculovirus of BmNPV and AcNPV,and knockout of cysteinase gene for more efficient expression

    Institute of Scientific and Technical Information of China (English)

    WU Xiaofeng; CAO Cuiping; XU Yaxiang; LU Xingmeng

    2004-01-01

    AcNPV(Autographa californica nuclear polyhedrosis virus)and BmNPV(Bombyx mori nuclear polyhedrosis virus)are two principal insect-baculovirus expression systems,each having different characteristics.AcNPV has a wider host range and can infect a series of cell lines thus making it suitable for cell suspension culture expression,but the small size of the host insect,A.californica,makes AcNPV less suitable for large scale protein synthesis.In contrast,BmNPV can only infect the silkworm,Bornbyx rnori,which is well-known for its easy rearing and large size.These characteristics make the BmNPV system especially suitable for large-scale industrial expression.To utilize the advantages of both AcNPV and BmNPV,we tried to expand their host range through homologous recombination and successfully constructed a hybrid baculovirus of AcNPV and BmNPV,designated as HyNPV.The hybrid baculovirus can infect the hosts of both AcNPV and BmNPV.Taking the human basic fibroblast growth factor(Bfgf)gene as an application example,we constructed a recombinant,HyNPV-Bfgf.This construct is able to express the Bfgf protein both in silkworm larvae and in common-use cell lines,sf21,sf9 and High-five.Moreover,to reduce the loss of recombinant protein due to degradation by proteases that are simultaneously expressed by the baculovirus,we knocked out the cysteinase gene coding for one of the most important baculovirus proteases.This knockout mutation improves the production efficiency of the Bfgf recombinant protein.

  18. Display of VP1 on the surface of baculovirus and its immunogenicity against heterologous human enterovirus 71 strains in mice.

    Directory of Open Access Journals (Sweden)

    Tao Meng

    Full Text Available BACKGROUND: Human Enterovirus 71 (EV71 is a common cause of hand, foot and mouth disease (HFMD in young children. It is often associated with severe neurological diseases and has caused high mortalities in recent outbreaks across the Asia Pacific region. Currently, there is no effective vaccine and antiviral agents available against EV71 infections. VP1 is one of the major immunogenic capsid protein of EV71 and plays a crucial role in viral infection. Antibodies against VP1 are important for virus neutralization. METHODOLOGY/PRINCIPAL FINDING: In the present study, infectious EV71 viruses were generated from their synthetic complementary DNA using the human RNA polymerase I reverse genetics system. Secondly, the major immunogenic capsid protein (VP1 of EV71-Fuyang (subgenogroup C4 was displayed on the surface of recombinant baculovirus Bac-Pie1-gp64-VP1 as gp64 fusion protein under a novel White Spot Syndrome Virus (WSSV immediate early ie1 promoter. Baculovirus expressed VP1 was able to maintain its structural and antigenic conformity as indicated by immunofluorescence assay and western blot analysis. Interestingly, our results with confocal microscopy revealed that VP1 was able to localize on the plasma membrane of insect cells infected with recombinant baculovirus. In addition, we demonstrated with transmission electron microscopy that baculovirus successfully acquired VP1 from the insect cell membrane via the budding process. After two immunizations in mice, Bac-Pie1-gp64-VP1 elicited neutralization antibody titer of 1∶64 against EV71 (subgenogroup C4 in an in vitro neutralization assay. Furthermore, the antisera showed high cross-neutralization activities against all 11 subgenogroup EV71 strains. CONCLUSION: Our results illustrated that Bac-Pie1-gp64-VP1 retained native epitopes of VP1 and acted as an effective EV71 vaccine candidate which would enable rapid production without any biosafety concerns.

  19. Preparing of local contingency planning for nuclear accident of nuclear submarine at port

    International Nuclear Information System (INIS)

    Full text: The presence of a nuclear submarine in port involves a radiological risk which requires to be taken into account appropriately. The implementation of radiological monitoring and prevention measures are basic requisites to assure this visit being reasonably safer. In normal condition, the radiological monitoring of some marine environmental samples are necessary to be performed before the arrival and during the staying and after the departure of the nuclear ship. In the other side, prevention measures involve the preparedness of local contingency plans to protect the people in the port in any nuclear accidental event. The plan is designed by assuming an accident where 10% of core inventory of a 30 MW(e) nuclear power reactor are released to environment. The plan basically takes into account immediate measures inside a zone of 2,5 km around the submarine - from the normal place for anchor - being some of them the clearing of the zone from ships and people and taking the damaged submarine out at sea in order to reduce the scope of possible emissions. Likewise, previsions are made for protecting people up to 10 km in the coast. All these measures are performed by an organization commanded by the Port Captain and supported by other state institutions in land. (orig.)

  20. Nuclear localization of Annexin A7 during murine brain development

    Directory of Open Access Journals (Sweden)

    Noegel Angelika A

    2005-04-01

    Full Text Available Abstract Background Annexin A7 is a member of the annexin protein family, which is characterized by its ability to interact with phospholipids in the presence of Ca2+-ions and which is thought to function in Ca2+-homeostasis. Results from mutant mice showed altered Ca2+-wave propagation in astrocytes. As the appearance and distribution of Annexin A7 during brain development has not been investigated so far, we focused on the distribution of Annexin A7 protein during mouse embryogenesis in the developing central nervous system and in the adult mouse brain. Results Annexin A7 is expressed in cells of the developing brain where a change in its subcellular localization from cytoplasm to nucleus was observed. In the adult CNS, the subcellular distribution of Annexin A7 depends on the cell type. By immunohistochemistry analysis Annexin A7 was detected in the cytosol of undifferentiated cells at embryonic days E5–E8. At E11–E15 the protein is still present in the cytosol of cells predominantly located in the ventricular germinative zone surrounding the lateral ventricle. Later on, at embryonic day E16, Annexin A7 in cells of the intermediate and marginal zone of the neopallium translocates to the nucleus. Neuronal cells of all areas in the adult brain present Annexin A7 in the nucleus, whereas glial fibrillary acidic protein (GFAP-positive astrocytes exhibit both, a cytoplasmic and nuclear staining. The presence of nuclear Annexin A7 was confirmed by extraction of the nucleoplasm from isolated nuclei obtained from neuronal and astroglial cell lines. Conclusion We have demonstrated a translocation of Annexin A7 to nuclei of cells in early murine brain development and the presence of Annexin A7 in nuclei of neuronal cells in the adult animal. The role of Annexin A7 in nuclei of differentiating and mature neuronal cells remains elusive.

  1. Genetic Variation in Field Populations of Baculoviruses: Mechanisms for Generating Variation and Its Potential Role in Baculovirus Epizootiology

    Institute of Scientific and Technical Information of China (English)

    Martin A. Erlandson

    2009-01-01

    Baculoviridae is a family of insect-specific DNA viruses that have been used as biological control agents for insect pest control. In most cases these baculovirus control agents are natural field isolates that have been selected based on their infectivity and virulence. The advent of molecular tools such as restriction endonucleases, targeted polymerase chain reaction and new DNA sequencing strategies have allowed for efficient detection and characterization of genotypic variants within and among geographic and temporal isolates of baculovirus species. It has become evident that multiple genotypic variants occur even within individual infected larvae. Clonal strains of baculovirus species derived either by in vitro or in vivo approaches have been shown to vary with respect to infectivity and virulence. Many of the cell culture derived plague-purified strains have deletions that interrupt egt expression leading to virus strains that kill infected hosts more quickly. As well, in vitro clones often involve larger genomic deletions with the loss of pif gene function, resulting in strains deficient for oral infectivity. There are an increasing number of baculovirus species for which complete genome sequences are available for more than one strain or field isolate. Results of comparative analysis of these strains indicated that hr regions and bro genes often mark "hot spots" of genetic variability between strains and of potential recombination events. In addition, the degree of nucleotide polymorphisms between and within strains and their role in amino acid substitutions within ORFs and changes in promoter motifs is also beginning to be appreciated. In this short review the potential mechanisms that generate and maintain this genetic diversity within baculovirus populations is discussed, as is the potential role of genetic variation in host-pathogen interactions.

  2. Role of 'commission locale d'information' in France from the viewpoint of nuclear regulatory governance

    International Nuclear Information System (INIS)

    In France, there exist organizations called 'Commission Locale d'Information' (CLI) in all the siting areas where nuclear facilities located. Previously, the CLI organizations were established voluntarily by some local governments or nuclear utilities. Since 2006, however, the Nuclear Transparency and Safety Act has obliged the establishment of CLI in all the siting areas in conjunction with reforming the nuclear regulatory agencies. This means that the concerned local governments are officially part of nuclear safety regulation. In this study, we investigated present conditions of the CLI organizations through some interviews in France and consider their roles from the standpoint of nuclear regulatory governance. As a result, we found that the CLI plays the following roles: (1) medium of communication among concerned parties (not only between nuclear utilities and local habitants but also between the national nuclear regulatory agency and various local governments) and (2) implementing various activities in accordance with each local condition through the participation of local assembly members. In addition, we clarified that CLI's activities are supported by related institutional infrastructures, such as cost burden between central and local governments, and some other systems of citizen participation in building or expanding nuclear facilities. (author)

  3. The Parasitoid Factor in the Virulence and Spread of Lepidopteran Baculoviruses

    Institute of Scientific and Technical Information of China (English)

    J. E. Cossentine

    2009-01-01

    Insect parasitoids and baculoviruses play important roles in the natural and strategic biological control of insects. The two parasites are frequent competitors within common hosts and much research has focused on the negative impact that baculoviral host infections have on parasitoids. This review summarizes the impacts that parasitoids may have on the virulence and spread of lepidopteran baculoviruses. By changing host behavior and development, parasitoids have been shown to decrease baculovirus virulence and productivity within parasitized baculovirus-susceptible hosts; however, studies of the tools used by hymenopteran parasitoids to overcome their hosts' immune systems, suggest that parasitoids may, in some cases, facilitate baculoviral infections in less susceptible hosts. Laboratory and field research have demonstrated that parasitoids can mechanically transmit bacuioviruses between insects, and in this way, increase the efficacy of the viruses. Instances of new, more virulent isolates of baculoviruses have been recorded from specifically parasitoid-targeted hosts suggesting other possible benefits from the transmission or activation of baculoviruses by parasitoids.

  4. Efficient and dynamic nuclear localization of green fluorescent protein via RNA binding

    International Nuclear Information System (INIS)

    Classical nuclear localization signal (NLS) sequences have been used for artificial localization of green fluorescent protein (GFP) in the nucleus as a positioning marker or for measurement of the nuclear-cytoplasmic shuttling rate in living cells. However, the detailed mechanism of nuclear retention of GFP-NLS remains unclear. Here, we show that a candidate mechanism for the strong nuclear retention of GFP-NLS is via the RNA-binding ability of the NLS sequence. GFP tagged with a classical NLS derived from Simian virus 40 (GFP-NLSSV40) localized not only in the nucleoplasm, but also to the nucleolus, the nuclear subdomain in which ribosome biogenesis takes place. GFP-NLSSV40 in the nucleolus was mobile, and intriguingly, the diffusion coefficient, which indicates the speed of diffusing molecules, was 1.5-fold slower than in the nucleoplasm. Fluorescence correlation spectroscopy (FCS) analysis showed that GFP-NLSSV40 formed oligomers via RNA binding, the estimated molecular weight of which was larger than the limit for passive nuclear export into the cytoplasm. These findings suggest that the nuclear localization of GFP-NLSSV40 likely results from oligomerization mediated via RNA binding. The analytical technique used here can be applied for elucidating the details of other nuclear localization mechanisms, including those of several types of nuclear proteins. In addition, GFP-NLSSV40 can be used as an excellent marker for studying both the nucleoplasm and nucleolus in living cells. - Highlights: • Nuclear localization signal-tagged GFP (GFP-NLS) showed clear nuclear localization. • The GFP-NLS dynamically localized not only in the nucleoplasm, but also to the nucleolus. • The nuclear localization of GFP-NLS results from transient oligomerization mediated via RNA binding. • Our NLS-tagging procedure is ideal for use in artificial sequestration of proteins in the nucleus

  5. Efficient and dynamic nuclear localization of green fluorescent protein via RNA binding

    Energy Technology Data Exchange (ETDEWEB)

    Kitamura, Akira; Nakayama, Yusaku; Kinjo, Masataka, E-mail: kinjo@sci.hokudai.ac.jp

    2015-07-31

    Classical nuclear localization signal (NLS) sequences have been used for artificial localization of green fluorescent protein (GFP) in the nucleus as a positioning marker or for measurement of the nuclear-cytoplasmic shuttling rate in living cells. However, the detailed mechanism of nuclear retention of GFP-NLS remains unclear. Here, we show that a candidate mechanism for the strong nuclear retention of GFP-NLS is via the RNA-binding ability of the NLS sequence. GFP tagged with a classical NLS derived from Simian virus 40 (GFP-NLS{sup SV40}) localized not only in the nucleoplasm, but also to the nucleolus, the nuclear subdomain in which ribosome biogenesis takes place. GFP-NLS{sup SV40} in the nucleolus was mobile, and intriguingly, the diffusion coefficient, which indicates the speed of diffusing molecules, was 1.5-fold slower than in the nucleoplasm. Fluorescence correlation spectroscopy (FCS) analysis showed that GFP-NLS{sup SV40} formed oligomers via RNA binding, the estimated molecular weight of which was larger than the limit for passive nuclear export into the cytoplasm. These findings suggest that the nuclear localization of GFP-NLS{sup SV40} likely results from oligomerization mediated via RNA binding. The analytical technique used here can be applied for elucidating the details of other nuclear localization mechanisms, including those of several types of nuclear proteins. In addition, GFP-NLS{sup SV40} can be used as an excellent marker for studying both the nucleoplasm and nucleolus in living cells. - Highlights: • Nuclear localization signal-tagged GFP (GFP-NLS) showed clear nuclear localization. • The GFP-NLS dynamically localized not only in the nucleoplasm, but also to the nucleolus. • The nuclear localization of GFP-NLS results from transient oligomerization mediated via RNA binding. • Our NLS-tagging procedure is ideal for use in artificial sequestration of proteins in the nucleus.

  6. Empirical Proton-Neutron Interactions and Nuclear Density Functional Theory: Global, Regional and Local Comparisons

    OpenAIRE

    Stoitsov, M.; Cakirli, R. B.; Casten, R. F.; W. Nazarewicz; Satula, W.

    2006-01-01

    Calculations of nuclear masses, using nuclear density functional theory, are presented for even-even nuclei spanning the nuclear chart. The resulting binding energy differences can be interpreted in terms of valence proton-neutron interactions. These are compared globally, regionally, and locally with empirical values. Overall, excellent agreement is obtained. Discrepancies highlight neglected degrees of freedom and can point to improved density functionals.

  7. Expression from baculovirus and serological reactivity of the nucleocapsid protein of dolphin morbillivirus.

    Science.gov (United States)

    Grant, Rebecca J; Kelley, Karen L; Maruniak, James E; Garcia-Maruniak, Alejandra; Barrett, Tom; Manire, Charles A; Romero, Carlos H

    2010-07-14

    The nucleocapsid (N) protein of dolphin morbillivirus (DMV) was expressed from a baculovirus (Autographa californica nuclear polyhedrosis virus) vector and shown by SDS-PAGE and Western blot analysis to be about 57 kDa. Transmission electron microscopy revealed fully assembled nucleocapsid-like particles (NLPs) exhibiting the typical helical herringbone morphology. These NLPs were approximately 20-22 nm in diameter and varied in length from 50 to 100 nm. Purified DMV-N protein was used as antigen in an indirect ELISA (iELISA) and shown to react with rabbit and human antisera to measles virus (MV) and dog sera with antibodies to canine distemper virus (CDV). The iELISA was used for the demonstration of morbillivirus antibodies in the serum of cetaceans and manatees, showing potential as a serological tool for the mass screening of morbillivirus antibodies in marine mammals. PMID:20005643

  8. An analysis on the market of instrumentation in nuclear power plant in China and its localization

    International Nuclear Information System (INIS)

    The Instrumentation plays a critical role in guaranteeing the safety and efficiency of the nuclear power plant. The instrumentations used in the working nuclear power plants in China, especially the Class 1E ones, are mainly imported products. This has been one of the key elements restricting the localized development of the Chinese nuclear power industry. Based on a fundamental survey of the Chinese nuclear instrumentation market, this paper introduces the definition, classification of nuclear instrumentations, states the market status, analyzes the cause of how this situation was formed, and makes some proposals on how to handle the localization job in the future. (authors)

  9. Highly efficient baculovirus-mediated multigene delivery in primary cells

    OpenAIRE

    Maysam Mansouri, Maysam; Bellon-Echeverria, Itxaso; Rizk, Aurélien; Ehsaei, Zahra; Cianciolo Cosentino, Chiara; Silva, Catarina S.; Berger, Imre

    2016-01-01

    Multigene delivery and subsequent cellular expression is emerging as a key technology required in diverse research fields including, synthetic and structural biology, cellular reprogramming and functional pharmaceutical screening. Current viral delivery systems such as retro- and adenoviruses suffer from limited DNA cargo capacity, thus impeding unrestricted multigene expression. We developed MultiPrime, a modular, non-cytotoxic, non-integrating, baculovirus-based vector system expediting hig...

  10. Sources of evaluation of nuclear and renewable energy contained in the local press

    OpenAIRE

    Pligt, van der, J.; Spears, R.; Eiser,, J.R.

    1987-01-01

    Examined the sources of evaluative coverage concerning nuclear power and renewable alternatives contained in local UK daily press coverage. 10 categories of source were defined for their relevance to the nuclear debate and energy issues. Out of these, only pronuclear industries and national government produced more positive than negative appraisals of nuclear power. However, detractors of nuclear power were more varied, the most prolific category being the general public. Alternative technolo...

  11. Importin α1 Mediates Yorkie Nuclear Import via an N-terminal Non-canonical Nuclear Localization Signal.

    Science.gov (United States)

    Wang, Shimin; Lu, Yi; Yin, Meng-Xin; Wang, Chao; Wu, Wei; Li, Jinhui; Wu, Wenqing; Ge, Ling; Hu, Lianxin; Zhao, Yun; Zhang, Lei

    2016-04-01

    The Hippo signaling pathway controls organ size by orchestrating cell proliferation and apoptosis. When the Hippo pathway was inactivated, the transcriptional co-activator Yorkie translocates into the nucleus and forms a complex with transcription factor Scalloped to promote the expression of Hippo pathway target genes. Therefore, the nuclear translocation of Yorkie is a critical step in Hippo signaling. Here, we provide evidence that the N-terminal 1-55 amino acids of Yorkie, especially Arg-15, were essential for its nuclear localization. By mass spectrometry and biochemical analyses, we found that Importin α1 can directly interact with the Yorkie N terminus and drive Yorkie into the nucleus. Further experiments show that the upstream component Hippo can inhibit Importin α1-mediated Yorkie nuclear import. Taken together, we identified a potential nuclear localization signal at the N-terminal end of Yorkie as well as a critical role for Importin α1 in Yorkie nuclear import. PMID:26887950

  12. A functional nuclear localization sequence in the VP1 capsid protein of coxsackievirus B3

    International Nuclear Information System (INIS)

    The capsid proteins of some RNA viruses can translocate to the nucleus and interfere with cellular phenotypes. In this study we found that the VP1 capsid protein of coxsackievirus B3 (CVB3) was dominantly localized in the nucleus of the cells transfected with VP1-expressing plasmid. The VP1 nuclear localization also occurred in the cells infected with CVB3. Truncation analysis indicated that the VP1 nuclear localization sequence located near the C-terminal. The substitution of His220 with threonine completely abolished its translocation. The VP1 proteins of other CVB types might have the nuclear localization potential because this region was highly conserved. Moreover, the VP1 nuclear localization induced cell cycle deregulation, including a prolonged S phase and shortened G2-M phase. Besides these findings, we also found a domain between Ala72 and Phe106 that caused the VP1 truncates dotted distributed in the cytoplasm. Our results suggest a new pathogenic mechanism of CVB. - Highlights: ► The VP1 protein of coxsackievirus B3 can specifically localize in the nucleus. ► The nuclear localization signal of coxsackievirus B3 VP1 protein locates near its C-terminal. ► The VP1 nuclear localization of coxsackievirus B3 can deregulate cell cycle. ► There is a domain in the VP1 that determines it dotted distributed in the cytoplasm.

  13. A functional nuclear localization sequence in the VP1 capsid protein of coxsackievirus B3

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Tianying; Yu, Bohai; Lin, Lexun; Zhai, Xia; Han, Yelu; Qin, Ying; Guo, Zhiwei; Wu, Shuo; Zhong, Xiaoyan; Wang, Yan; Tong, Lei; Zhang, Fengmin; Si, Xiaoning [Department of Microbiology, Harbin Medical University, Harbin 150081 (China); Zhao, Wenran, E-mail: wenran.zhao@gmail.com [Department of Cell Biology, Harbin Medical University, Harbin 150081 (China); Zhong, Zhaohua, E-mail: zhonghmu@gmail.com [Department of Microbiology, Harbin Medical University, Harbin 150081 (China)

    2012-11-25

    The capsid proteins of some RNA viruses can translocate to the nucleus and interfere with cellular phenotypes. In this study we found that the VP1 capsid protein of coxsackievirus B3 (CVB3) was dominantly localized in the nucleus of the cells transfected with VP1-expressing plasmid. The VP1 nuclear localization also occurred in the cells infected with CVB3. Truncation analysis indicated that the VP1 nuclear localization sequence located near the C-terminal. The substitution of His220 with threonine completely abolished its translocation. The VP1 proteins of other CVB types might have the nuclear localization potential because this region was highly conserved. Moreover, the VP1 nuclear localization induced cell cycle deregulation, including a prolonged S phase and shortened G2-M phase. Besides these findings, we also found a domain between Ala72 and Phe106 that caused the VP1 truncates dotted distributed in the cytoplasm. Our results suggest a new pathogenic mechanism of CVB. - Highlights: Black-Right-Pointing-Pointer The VP1 protein of coxsackievirus B3 can specifically localize in the nucleus. Black-Right-Pointing-Pointer The nuclear localization signal of coxsackievirus B3 VP1 protein locates near its C-terminal. Black-Right-Pointing-Pointer The VP1 nuclear localization of coxsackievirus B3 can deregulate cell cycle. Black-Right-Pointing-Pointer There is a domain in the VP1 that determines it dotted distributed in the cytoplasm.

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

    International Nuclear Information System (INIS)

    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

  15. 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.

  16. Effect of spray drying processing parameters on the insecticidal activity of two encapsulated formulations of baculovirus

    Science.gov (United States)

    The aim of this work was to evaluate the effect of spray dryer processing parameters on the process yield and insecticidal activity of baculovirus to support the development of this beneficial group of microbes as biopesticides. For each of two baculoviruses [granulovirus (GV) from Pieris rapae (L....

  17. Nuclear data uncertainties for local power densities in the Martin-Hoogenboom benchmark

    International Nuclear Information System (INIS)

    The recently developed method of fast Total Monte Carlo to propagate nuclear data uncertainties was applied to the Martin-Hoogenboom benchmark. This Martin- Hoogenboom benchmark prescribes that one calculates local pin powers (of light water cooled reactor) with a statistical uncertainty lower than 1% everywhere. Here we report, for the first time, an estimate of the nuclear data uncertainties for these local pin powers. For each of the more than 6 million local power tallies, the uncertainty due to nuclear data uncertainties was calculated, based on random variation of data for 235U, 238U, 239Pu and H in H2O thermal scattering. In the center of the core region, the nuclear data uncertainty is 0.9%. Towards the edges of the core, this uncertainty increases to roughly 3%. The nuclear data uncertainties have been shown to be larger than the statistical uncertainties that the benchmark prescribes

  18. Early localization of NPA58, a rat nuclear pore-associated protein, to the reforming nuclear envelope during mitosis

    Indian Academy of Sciences (India)

    Radhika Ganeshan; Nandini Rangaraj; Veena K Parnaik

    2001-03-01

    We have studied the mitotic reassembly of the nuclear envelope, using antibodies to nuclear marker proteins and NPA58 in F-111 rat fibroblast cells. In earlier studies we have proposed that NPA58, a 58 kDa rat nuclear protein, is involved in nuclear protein import. In this report, NPA58 is shown to be localized on the cytoplasmic face of the envelope in interphase cells, in close association with nuclear pores. In mitotic cells NPA58 is dispersed in the cytoplasm till anaphase. The targeting of NPA58 to the reforming nuclear envelope in early telophase coincides with the recruitment of a well-characterized class of nuclear pore proteins recognized by the antibody mAb 414, and occurs prior to the incorporation of lamin B1 into the envelope. Significant protein import activity is detectable only after localization of NPA58 in the newly-formed envelope. The early targeting of NPA58 is consistent with its proposed role in nuclear transport.

  19. Nuclear localization of Merkel cell polyomavirus large T antigen in Merkel cell carcinoma

    International Nuclear Information System (INIS)

    To clarify whether mutations in the large T gene encoded by Merkel cell polyomavirus affect the expression and function of large T antigen in Merkel cell carcinoma cases, we investigated the expression of large T antigen in vitro and in vivo. Immunohistochemistry using a rabbit polyclonal antibody revealed that large T antigen was expressed in the nuclei of Merkel cell carcinoma cells with Merkel cell polyomavirus infection. Deletion mutant analyses identified an Arg-Lys-Arg-Lys sequence (amino acids 277-280) as a nuclear localization signal in large T antigen. Sequence analyses revealed that there were no mutations in the nuclear localization signal in any of the eleven Merkel cell polyomavirus strains examined. Furthermore, stop codons were not observed in the upstream of the nuclear localization signal in any of the Merkel cell carcinoma cases examined. These data suggest that the nuclear localization signal is highly conserved and functional in Merkel cell carcinoma cases.

  20. Pharmacokinetics local model and Its application in nuclear medicine

    Institute of Scientific and Technical Information of China (English)

    曹国宪; 李卫一; 等

    1996-01-01

    The Pharmacokinetics local model for studying kinetic action of pharmaceuticals in the specific part of the body is established on the basis of the compartment model.A series of formulae is deduced,and with them the pharmacokinetic equation and several important parameters can be obtained.The local model successfully expanded the area of pharmacokinetics from the compartment model ,which is mainly dealing with kinetic action of pharmaceuticals in blood,to the local model,which can study kinetic action of pharmaceuticals in any organ,tissue or fluid.

  1. Atypical nuclear localization of VIP receptors in glioma cell lines and patients

    International Nuclear Information System (INIS)

    Highlights: • The VIP receptor VPAC1 contains a putative NLS signal. • VPAC1 is predominantly nuclear in GBM cell lines but not VPAC2. • Non-nuclear VPAC1/2 protein expression is correlated with glioma grade. • Nuclear VPAC1 is observed in 50% of stage IV glioma (GBM). - Abstract: An increasing number of G protein-coupled receptors, like receptors for vasoactive intestinal peptide (VIP), are found in cell nucleus. As VIP receptors are involved in the regulation of glioma cell proliferation and migration, we investigated the expression and the nuclear localization of the VIP receptors VPAC1 and VPAC2 in this cancer. First, by applying Western blot and immunofluorescence detection in three human glioblastoma (GBM) cell lines, we observed a strong nuclear staining for the VPAC1 receptor and a weak nuclear VPAC2 receptor staining. Second, immunohistochemical staining of VPAC1 and VPAC2 on tissue microarrays (TMA) showed that the two receptors were expressed in normal brain and glioma tissues. Expression in the non-nuclear compartment of the two receptors significantly increased with the grade of the tumors. Analysis of nuclear staining revealed a significant increase of VPAC1 staining with glioma grade, with up to 50% of GBM displaying strong VPAC1 nuclear staining, whereas nuclear VPAC2 staining remained marginal. The increase in VPAC receptor expression with glioma grades and the enhanced nuclear localization of the VPAC1 receptors in GBM might be of importance for glioma progression

  2. Atypical nuclear localization of VIP receptors in glioma cell lines and patients

    Energy Technology Data Exchange (ETDEWEB)

    Barbarin, Alice; Séité, Paule [Equipe Récepteurs, Régulations et Cellules Tumorales, Université de Poitiers, PBS bât 36, 1 rue Georges Bonnet, TSA 51106, 86073 Poitiers Cedex 9 (France); Godet, Julie [Laboratoire d’anatomie et de cytologie pathologiques, CHU de Poitiers, 2 rue de la Milétrie, 86000 Poitiers (France); Bensalma, Souheyla; Muller, Jean-Marc [Equipe Récepteurs, Régulations et Cellules Tumorales, Université de Poitiers, PBS bât 36, 1 rue Georges Bonnet, TSA 51106, 86073 Poitiers Cedex 9 (France); Chadéneau, Corinne, E-mail: corinne.chadeneau@univ-poitiers.fr [Equipe Récepteurs, Régulations et Cellules Tumorales, Université de Poitiers, PBS bât 36, 1 rue Georges Bonnet, TSA 51106, 86073 Poitiers Cedex 9 (France)

    2014-11-28

    Highlights: • The VIP receptor VPAC1 contains a putative NLS signal. • VPAC1 is predominantly nuclear in GBM cell lines but not VPAC2. • Non-nuclear VPAC1/2 protein expression is correlated with glioma grade. • Nuclear VPAC1 is observed in 50% of stage IV glioma (GBM). - Abstract: An increasing number of G protein-coupled receptors, like receptors for vasoactive intestinal peptide (VIP), are found in cell nucleus. As VIP receptors are involved in the regulation of glioma cell proliferation and migration, we investigated the expression and the nuclear localization of the VIP receptors VPAC1 and VPAC2 in this cancer. First, by applying Western blot and immunofluorescence detection in three human glioblastoma (GBM) cell lines, we observed a strong nuclear staining for the VPAC1 receptor and a weak nuclear VPAC2 receptor staining. Second, immunohistochemical staining of VPAC1 and VPAC2 on tissue microarrays (TMA) showed that the two receptors were expressed in normal brain and glioma tissues. Expression in the non-nuclear compartment of the two receptors significantly increased with the grade of the tumors. Analysis of nuclear staining revealed a significant increase of VPAC1 staining with glioma grade, with up to 50% of GBM displaying strong VPAC1 nuclear staining, whereas nuclear VPAC2 staining remained marginal. The increase in VPAC receptor expression with glioma grades and the enhanced nuclear localization of the VPAC1 receptors in GBM might be of importance for glioma progression.

  3. Localization of Manufacturing Capabilities in Setting Up Nuclear Power Plants

    International Nuclear Information System (INIS)

    Nuclear renaissance is now imminent and is inevitable in view of rapidly increasing global warming concerns. A steep shift towards environmentally benign sources of energy remains an unavoidable choice as continents are warming up pushing seas into human habitation and disturbing global ecology. Accordingly, Indian government in its integrated energy policy document has planned for raising nuclear power capacity to generate 63 GWe by 2030. This envisages estimated investments of US$22 billion in the next 15 to 20 years. Setting up of nuclear energy generation capacity, however, remains a painstakingly slow process primarily due to complex, multidisciplinary efforts required to crank up a reactor. A robust supply chain remains key to expediting this process. In the light of this, it is critically important to ensure supply-chain for materials and components and putting in place cost effective project management to complete the projects on time and within the budgets. In this context, the participation of industries and their preparedness to meet the challenges are necessary. This would also require investments towards up gradation of manufacturing technology, training of manpower and mobilization of resources at the construction site. The industry would also need to enhance detailing and design engineering capabilities for the plants. It is only when such capabilities have been brought up that the possibilities of time-bound setting up of nuclear plants can be realized. In this paper, various issues with regard to project cost, regulatory and licensing, technology and gestation period etc for new build plants relevant to manufacturing industry are discussed. The plans for enhancing manufacturing capabilities for the critical path items of the project schedule with viable business, ensuring returns to stakeholders and financing and investment cycle are brought out. The various steps and initiatives being taken by Bharat Forge Ltd, the flagship company of Kalyani

  4. Localization of manufacturing capabilities in setting up nuclear power plants

    International Nuclear Information System (INIS)

    Full text: Nuclear renaissance is now imminent and is inevitable in view of the global warming concerns .Indian government in its integrated energy policy document has planed for raising nuclear power capacity to generate 63 GWe by 2030. This envisages estimated investments of US $22 billion in next 15 to 20 years. In the light of this, for setting up Nuclear Power Plants, it becomes very important to ensure supply-chain for materials and components and putting in place cost effective project management to complete the projects on time and within the budgets. In this context, the participation of industries and their preparedness to meet the challenges are necessary. This would also require investments towards up gradation of manufacturing technology, training of manpower and mobilization of resources at the construction site. The industry would also need to enhance detailing and design engineering capabilities for the plants. In this paper, various issues with regard to project cost, regulatory and licensing, technology and gestation period etc for new build plants relevant to manufacturing industry are discussed. The plans for enhancing manufacturing capabilities for the critical path items of the project schedule with viable business, ensuring returns to stakeholders and financing and investment cycle are brought out. The various steps and initiatives being taken by Bharat Forge Ltd a flagship company of Kalyani group for supply of forgings for nuclear reactor, vessels, steam generators , turbine generators and other safety critical components including pumps, valves, pipes and tubes, and other integration work for the balance of plant are summarized. The company has planned significant investments for setting up an integrated manufacturing facility to clear the global bottleneck in the supply of ultra-heavy forgings for light water reactors. Civil structuring involves execution by prior qualification with proven skills for specialized civil works needed in the

  5. Immunofluorescent localization of the proteins of nuclear ribonucleoprotein complexes

    OpenAIRE

    1980-01-01

    Antibodies were raised in chickens against heterogeneous nuclear RNA (hnRNA)-binding proteins from 30S ribonucleoprotein (RNP) complexes of mouse Taper hepatoma ascites cell nuclei. The antibody preparations were characterized for immunological specificity and purity by double- diffusion gels, binding to specific bands in SDS polyacrylamide gels, and crossed immunoelectrophoresis. Antibodies raised against either whole 30S RNP complexes or purified RNP core proteins had a strong selective aff...

  6. Local civic forum. An experimental study promoting public acceptance on nuclear energy

    International Nuclear Information System (INIS)

    In order to promote public acceptance on nuclear energy, an experimental study named 'Local Civic Forum' has been performed as one of the research activities on relationship between nuclear energy and society promoted by the 21st Century Center of Excellence Program: 'Innovative Nuclear Energy Systems for Sustainable Development of the World' conducted in Tokyo Institute of Technology (Tokyo Tech). The Local Civic Forum was designed to implement a dialogue among doctoral students of Tokyo Tech and multi-stakeholders of society. The framework of the forum looks like a stakeholder meeting or a consensus conference, however, it is different from these because no nuclear specialists directly join the dialogue. The specialists actively supported the doctoral students. The multi-stakeholders were selected from a viewpoint of social responsibility. Subjects in the Local Civic Forum were selected by the students from problems of nuclear energy. The participants of the forum discussed the selected subject based on the same eyes, and tried to get findings for hidden problems or issues on public acceptance of nuclear energy. We have made seven experiments of the forums between FY2005 and 2006. Some valuable comments were obtained from the participants to improve the scheme and management of the forum. It was found that the selection of multi-stakeholders and the facilitator of the Local Civic Forum played an important role. In this paper, the experimental results of the Local Civic Forum and the findings obtained by the participants will be presented. (author)

  7. Identification of novel nuclear localization signal within the ErbB-2 protein

    Institute of Scientific and Technical Information of China (English)

    Qiao Qiao CHEN; Xiao Ying CHEN; Yun Yun JIANG; Jing LIU

    2005-01-01

    ErbB2,a member of the receptor tyrosine kinase family,is frequently over-expressed in breast cancer.Proteolysis of the extracellular domain of ErbB2 results in constitutive activation of ErbB2 kinase.Recent study reported that ErbB2 is found in the nucleus.Here,we showed that ErbB2 is imported into the nucleus through a nuclear localization signal (NLS)-mediated mechanism.The NLS sequence KRRQQKIRKYTMRR (aa655-668) contains three clusters of basic amino acids and it is sufficient to target GFP into the nucleus.However,mutation in any basic amino acid cluster of this NLS sequence significantly affects its nuclear localization.Furthermore,it was found that this NLS is essential for the nuclear localization of ErbB2 since the intracellular domain of Erb2 lacking NLS completely abrogates its nuclear translocation.Taken together,our study identified a novel nuclear localization signal and reveals a novel mechanism underlying ErbB2 nuclear trafficking and localization.

  8. To the non-local theory of cold nuclear fusion

    OpenAIRE

    Alexeev, Boris V.

    2014-01-01

    In this paper, we revisit the cold fusion (CF) phenomenon using the generalized Bolzmann kinetics theory which can represent the non-local physics of this CF phenomenon. This approach can identify the conditions when the CF can take place as the soliton creation under the influence of the intensive sound waves. The vast mathematical modelling leads to affirmation that all parts of soliton move with the same velocity and with the small internal change of the pressure. The zone of the high dens...

  9. Baculovirus vector-mediated transfer of NIS gene into colon tumor cells for radionuclide therapy

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    AIM:To investigate the feasibility of radionuclide therapy of colon tumor cells by baculovirus vector-mediated transfer of the sodium/iodide symporter(NIS) gene.METHODS:A recombinant baculovirus plasmid carrying the NIS gene was constructed,and the viruses(BacNIS) were prepared using the Bac-to-Bac system.The infection efficiency in the colon cancer cell line SW1116 of a green fluorescent protein(GFP) expressing baculovirus(Bac-GFP) at different multiplicities of infection(MOI) with various concentrations o...

  10. Reflections on the development of local suppliers for the Argentine nuclear industry

    International Nuclear Information System (INIS)

    Argentina has given recently a new start to its nuclear power activities. Looking for background and experiences that can be useful under the new reality, the paper is a survey of the past development of local suppliers for the national nuclear industry. Based on the intention to answer the questions: Why so early it was decided to build a nuclear power plant? Why it was decided to buy it under a turnkey basis rather than developing an indigenous design? and what was the meaning of the 'opening of the technology package' at that time?, the paper describes the actions that led to the purchase of the Atucha I, Embalse and Atucha II nuclear power plants and how these decisions were implemented in order to maximize local participation and the technology transfer. It also analyzes the influence of the Argentine Nuclear Plan of the late seventies on the development of endogenous technology and describes the facts that helped to preserve until now the technological nuclear capabilities of the country in spite of the stopping of the Atucha II construction, and to create positive expectations regarding the revival of the local industry as a supplier of nuclear goods and services. (author)

  11. The bovine papillomavirus type 1 E2 transactivator and repressor proteins use different nuclear localization signals.

    Science.gov (United States)

    Skiadopoulos, M H; McBride, A A

    1996-02-01

    The E2 gene of bovine papillomavirus type 1 encodes at least three nuclear phosphoproteins that regulate viral transcription and DNA replication. All three proteins have a common C-terminal domain that has DNA-binding and dimerization activities. A basic region in this domain forms an alpha helix which makes direct contact with the DNA target. In this study, it is shown that in addition to its role in DNA binding, this basic region functions as a nuclear localization signal both in the E2 DNA-binding domain and in a heterologous protein. Deletion of this signal sequence resulted in increased accumulation of the E2 transactivator and repressor proteins in the cytoplasm, but nuclear localization was not eliminated. In the full-length transactivator protein, another signal, present in the N-terminal transactivation domain, is used for transport to the nucleus, and the C-terminal nuclear localization signal(s) are masked. The use of different nuclear localization signals could potentially allow differential regulation of the subcellular localization of the E2 transactivator and repressor proteins at some stage in the viral life cycle. PMID:8551571

  12. The Political Styles of Local Anti-Nuclear Waste Movements in Finland

    International Nuclear Information System (INIS)

    This paper aims to analyse the political styles of local anti-nuclear waste movements in Finland. The main focus is on the tension between the environmental impact assessment process (EIA) and the activity of local opposing groups. According to the EIA Act the purpose of the EIA process is to provide information and opportunities for citizens to participate in planning. It therefore aims to enhance the transparency of the decision-making process. The Finnish nuclear waste company, Posiva Oy, made great efforts locally to create opportunities for participation, but was much criticised by local activists. My questions are the following: How did these local movements participate in Posiva's EIA process? What kind of local differences were there in participating and how can these differences be explained? And finally; what did the EIA process mean to the political styles of movements? EIA meant a temporary change in political style on local level. Firstly; because it brought science into local politics very clearly. Secondly EIA with the Nuclear Energy Act framed decision-making with a timetable, which Posiva emphasised when local groups wanted to make a decision immediately. Thirdly EIA enabled the problem to be defined according to local needs but on the other hand plan level EIA separated nuclear waste issue from the use of nuclear power. Fourthly dialogue in the EIA process favoured rational discourse. Although local views were heard, the process did try to teach the people how to speak with the decision-making system, in the system's language. Thus the purpose of EIA was to make the local discussion controllable. All the local groups, the Romuvaara, Kivetty and Loviisa Movements and the Friends of the Earth in Pori region, took part in EIA dialogue but the reactions were different. The Romuvaara Movement was very active in participating and succeeded in exploiting EIA locally whereas the Loviisa Movement tried to displace the whole process. The Loviisa Movement

  13. The Political Styles of Local Anti-Nuclear Waste Movements in Finland

    Energy Technology Data Exchange (ETDEWEB)

    Kojo, Matti [Univ. of Tampere (Finland). Dept. of Political Science and International Relations

    2001-07-01

    This paper aims to analyse the political styles of local anti-nuclear waste movements in Finland. The main focus is on the tension between the environmental impact assessment process (EIA) and the activity of local opposing groups. According to the EIA Act the purpose of the EIA process is to provide information and opportunities for citizens to participate in planning. It therefore aims to enhance the transparency of the decision-making process. The Finnish nuclear waste company, Posiva Oy, made great efforts locally to create opportunities for participation, but was much criticised by local activists. My questions are the following: How did these local movements participate in Posiva's EIA process? What kind of local differences were there in participating and how can these differences be explained? And finally; what did the EIA process mean to the political styles of movements? EIA meant a temporary change in political style on local level. Firstly; because it brought science into local politics very clearly. Secondly EIA with the Nuclear Energy Act framed decision-making with a timetable, which Posiva emphasised when local groups wanted to make a decision immediately. Thirdly EIA enabled the problem to be defined according to local needs but on the other hand plan level EIA separated nuclear waste issue from the use of nuclear power. Fourthly dialogue in the EIA process favoured rational discourse. Although local views were heard, the process did try to teach the people how to speak with the decision-making system, in the system's language. Thus the purpose of EIA was to make the local discussion controllable. All the local groups, the Romuvaara, Kivetty and Loviisa Movements and the Friends of the Earth in Pori region, took part in EIA dialogue but the reactions were different. The Romuvaara Movement was very active in participating and succeeded in exploiting EIA locally whereas the Loviisa Movement tried to displace the whole process. The Loviisa

  14. Local anesthetics and nuclear medical bone images of the equine fore limb

    International Nuclear Information System (INIS)

    The effects of two local anesthetic agents on the diagnostic quality of nuclear medical bone images (NMBIs) of distal parts of the equine fore limb were investigated. Local effects on bone uptake of technetium 99m methylene diphosphonate (99mTc-MDP) 4 and 24 hours after perineural and intraarticular injection of mepivacaine hydrochloride and bupivacaine hydrochloride were evaluated in the carpal and metacarpophalangeal regions of 12 horses and ponies. Neither mepivacaine hydrochloride nor bupivacaine hydrochloride significantly altered the diagnostic quality of the NMBIs. The injection and subsequent action of local anesthetics do not appear to influence local bone uptake of 99mTc-MDP significantly

  15. Inhibition of HIV-1 integrase nuclear import and replication by a peptide bearing integrase putative nuclear localization signal

    Directory of Open Access Journals (Sweden)

    Waigmann Elisabeth

    2009-12-01

    Full Text Available Abstract Background The integrase (IN of human immunodeficiency virus type 1 (HIV-1 has been implicated in different steps during viral replication, including nuclear import of the viral pre-integration complex. The exact mechanisms underlying the nuclear import of IN and especially the question of whether it bears a functional nuclear localization signal (NLS remain controversial. Results Here, we studied the nuclear import pathway of IN by using multiple in vivo and in vitro systems. Nuclear import was not observed in an importin α temperature-sensitive yeast mutant, indicating an importin α-mediated process. Direct interaction between the full-length IN and importin α was demonstrated in vivo using bimolecular fluorescence complementation assay (BiFC. Nuclear import studies in yeast cells, with permeabilized mammalian cells, or microinjected cultured mammalian cells strongly suggest that the IN bears a NLS domain located between residues 161 and 173. A peptide bearing this sequence -NLS-IN peptide- inhibited nuclear accumulation of IN in transfected cell-cycle arrested cells. Integration of viral cDNA as well as HIV-1 replication in viral cell-cycle arrested infected cells were blocked by the NLS-IN peptide. Conclusion Our present findings support the view that nuclear import of IN occurs via the importin α pathway and is promoted by a specific NLS domain. This import could be blocked by NLS-IN peptide, resulting in inhibition of viral infection, confirming the view that nuclear import of the viral pre-integration complex is mediated by viral IN.

  16. Identification of the nuclear localization and export signals of high risk HPV16 E7 oncoprotein

    International Nuclear Information System (INIS)

    The E7 oncoprotein of high risk human papillomavirus type 16 (HPV16) binds and inactivates the retinoblastoma (RB) family of proteins. Our previous studies suggested that HPV16 E7 enters the nucleus via a novel Ran-dependent pathway independent of the nuclear import receptors (Angeline, M., Merle, E., and Moroianu, J. (2003). The E7 oncoprotein of high-risk human papillomavirus type 16 enters the nucleus via a nonclassical Ran-dependent pathway. Virology 317(1), 13-23.). Here, analysis of the localization of specific E7 mutants revealed that the nuclear localization of E7 is independent of its interaction with pRB or of its phosphorylation by CKII. Fluorescence microscopy analysis of enhanced green fluorescent protein (EGFP) and 2xEGFP fusions with E7 and E7 domains in HeLa cells revealed that E7 contains a novel nuclear localization signal (NLS) in the N-terminal domain (aa 1-37). Interestingly, treatment of transfected HeLa cells with two specific nuclear export inhibitors, Leptomycin B and ratjadone, changed the localization of 2xEGFP-E738-98 from cytoplasmic to mostly nuclear. These data suggest the presence of a leucine-rich nuclear export signal (NES) and a second NLS in the C-terminal domain of E7 (aa 38-98). Mutagenesis of critical amino acids in the putative NES sequence (76IRTLEDLLM84) changed the localization of 2xEGFP-E738-98 from cytoplasmic to mostly nuclear suggesting that this is a functional NES. The presence of both NLSs and an NES suggests that HPV16 E7 shuttles between the cytoplasm and nucleus which is consistent with E7 having functions in both of these cell compartments

  17. Nuclear localization of DMP1 proteins suggests a role in intracellular signaling

    Energy Technology Data Exchange (ETDEWEB)

    Siyam, Arwa [Department of Biomedical Sciences, Baylor College of Dentistry, Texas A and M Health Science Center, 3302 Gaston Ave., Dallas, TX 75246-2013 (United States); Department of Endodontology, Kornberg School of Dentistry, Temple University, 3223 North Broad Street, Philadelphia, PA 19140-5007 (United States); Wang, Suzhen; Qin, Chunlin; Mues, Gabriele [Department of Biomedical Sciences, Baylor College of Dentistry, Texas A and M Health Science Center, 3302 Gaston Ave., Dallas, TX 75246-2013 (United States); Stevens, Roy [Department of Endodontology, Kornberg School of Dentistry, Temple University, 3223 North Broad Street, Philadelphia, PA 19140-5007 (United States); D' Souza, Rena N. [Department of Biomedical Sciences, Baylor College of Dentistry, Texas A and M Health Science Center, 3302 Gaston Ave., Dallas, TX 75246-2013 (United States); Lu, Yongbo, E-mail: ylu@bcd.tamhsc.edu [Department of Biomedical Sciences, Baylor College of Dentistry, Texas A and M Health Science Center, 3302 Gaston Ave., Dallas, TX 75246-2013 (United States)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer Nuclear localization of DMP1 in various cell lines. Black-Right-Pointing-Pointer Non-synchronized cells show either nuclear or cytoplasmic localization of DMP1. Black-Right-Pointing-Pointer Nuclear DMP1 is restricted to the nucleoplasm but absent in the nucleolus. -- Abstract: Dentin matrix protein 1 (DMP1) is highly expressed in odontoblasts and osteoblasts/osteocytes and plays an essential role in tooth and bone mineralization and phosphate homeostasis. It is debatable whether DMP1, in addition to its function in the extracellular matrix, can enter the nucleus and function as a transcription factor. To better understand its function, we examined the nuclear localization of endogenous and exogenous DMP1 in C3H10T1/2 mesenchymal cells, MC3T3-E1 preosteoblast cells and 17IIA11 odontoblast-like cells. RT-PCR analyses showed the expression of endogenous Dmp1 in all three cell lines, while Western-blot analysis detected a major DMP1 protein band corresponding to the 57 kDa C-terminal fragment generated by proteolytic processing of the secreted full-length DMP1. Immunofluorescent staining demonstrated that non-synchronized cells presented two subpopulations with either nuclear or cytoplasmic localization of endogenous DMP1. In addition, cells transfected with a construct expressing HA-tagged full-length DMP1 also showed either nuclear or cytoplasmic localization of the exogenous DMP1 when examined with an antibody against the HA tag. Furthermore, nuclear DMP1 was restricted to the nucleoplasm but was absent in the nucleolus. In conclusion, these findings suggest that, apart from its role as a constituent of dentin and bone matrix, DMP1 might play a regulatory role in the nucleus.

  18. Nuclear localization of DMP1 proteins suggests a role in intracellular signaling

    International Nuclear Information System (INIS)

    Highlights: ► Nuclear localization of DMP1 in various cell lines. ► Non-synchronized cells show either nuclear or cytoplasmic localization of DMP1. ► Nuclear DMP1 is restricted to the nucleoplasm but absent in the nucleolus. -- Abstract: Dentin matrix protein 1 (DMP1) is highly expressed in odontoblasts and osteoblasts/osteocytes and plays an essential role in tooth and bone mineralization and phosphate homeostasis. It is debatable whether DMP1, in addition to its function in the extracellular matrix, can enter the nucleus and function as a transcription factor. To better understand its function, we examined the nuclear localization of endogenous and exogenous DMP1 in C3H10T1/2 mesenchymal cells, MC3T3-E1 preosteoblast cells and 17IIA11 odontoblast-like cells. RT-PCR analyses showed the expression of endogenous Dmp1 in all three cell lines, while Western-blot analysis detected a major DMP1 protein band corresponding to the 57 kDa C-terminal fragment generated by proteolytic processing of the secreted full-length DMP1. Immunofluorescent staining demonstrated that non-synchronized cells presented two subpopulations with either nuclear or cytoplasmic localization of endogenous DMP1. In addition, cells transfected with a construct expressing HA-tagged full-length DMP1 also showed either nuclear or cytoplasmic localization of the exogenous DMP1 when examined with an antibody against the HA tag. Furthermore, nuclear DMP1 was restricted to the nucleoplasm but was absent in the nucleolus. In conclusion, these findings suggest that, apart from its role as a constituent of dentin and bone matrix, DMP1 might play a regulatory role in the nucleus.

  19. Study on Fusion Protein and Its gene in Baculovirus Specificity

    International Nuclear Information System (INIS)

    Baculoviruses are subdivided into two groups depending on the type of budded virus envelop fusion protein; group I utilized gp64 which include the most of nucleopolyhedroviruses (NPVs), group II utilized F protein which include the remnants of NPVs and all Granuloviruses (GVs). Recent studies reported the viral F protein coding gene as a host cellular sourced gene and may evolutionary acquired from the host genome referring to phylogeny analysis of fusion proteins. Thus, it was deduced that F protein coding gene is species- specific nucleotide sequence related to the type of the specific host and if virus could infect an unexpected host, the resulted virus may encode a vary F gene. In this regard, the present study utilized the mentioned properties of F gene in an attempt to produce a model of specific and more economic wider range granulovirus bio- pesticide able to infect both Spodoptera littoralis and Phthorimaea operculella larvae. Multiple sequence alignment and phylogeny analysis were performed on six members of group II baculovirus, novel universal PCR primers were manually designed from the conserved regions in the alignment graph, targeted to amplify species- specific sequence entire F gene open reading frame (ORF) which is useful in molecular identification of baculovirus in unknown samples. So, the PCR product of SpliGV used to prepare a specific probe for the F gene of this type of virus. Results reflected that it is possible to infect S. littoralis larvae by PhopGV if injected into larval haemocoel, the resulted virus of this infection showed by using DNA hybridization technique to be encode to F gene homologous with the F gene of Spli GV, which is revealed that the resulted virus acquired this F gene sequence from the host genome after infection. Consequently, these results may infer that if genetic aberrations occur in the host genome, this may affect in baculoviral infectivity. So, this study aimed to investigate the effect of gamma radiation at

  20. New computer simulation technology of WSPEEDI for local and regional environmental assessment during nuclear emergency

    Energy Technology Data Exchange (ETDEWEB)

    Chino, Masamichi; Furuno, Akiko; Terada, Hiroaki; Kitabata, Hideyuki [Japan Atomic Energy Research Institute, Ibaraki-Ken (Japan)

    2002-07-01

    The increase of nuclear power plants in the Asian region necessitates the capability to predict long-range atmospheric dispersions of radionuclides and radiological impacts due to a nuclear accident. For this purpose, we have developed a computer-based emergency response system WSPEEDI. This paper aims to expanding the capability of WSPEEDI so that it can be applied to simultaneous multi-scale predictions of local and regional scales in the Asian region.

  1. Evaluation of nuclear power and renewable alternatives as portrayed in UK local press coverage title

    OpenAIRE

    Spears, R.; Eiser, J R; van der Pligt, J.

    1986-01-01

    A content analysis was conducted of all UK local daily newspaper articles appearing in the first half of 1981, concerned with nuclear power or renewable alternatives. Evaluative coverage of these technologies was compared on dimensions found to characterise energy issues (economic, environmental, technological future/political, physical and psychological risks). In addition, comparisons were drawn between coverage in areas 'threatened' with the potential siting of a new nuclear power station ...

  2. Framing the national nuclear legacy at the local level: Implications for the future of federal facilities

    International Nuclear Information System (INIS)

    There are several major federal nuclear facilities located in small towns and rural areas of the United States. While many of these facilities were developed in the 1950s to support national defense, in the 1960s and 1970s, some of these shifted their mission to focus on national energy infrastructure. Now, many of these facilities are in a clean-up phase, and local communities are becoming increasingly engaged in influencing decisions about the future of the sites. Communicating with the public in rural communities is challenging when it involves a complicated environmental issue that could have widespread economic impacts. The local media reflect public understanding, so getting a sense of how these media frame issues can be a crucial first step to developing an effective community engagement strategy. A media content analysis of one local newspaper was completed in relation to a major federal nuclear facility. The content analysis is compared to the results of a telephone survey in the region served by the paper and the results suggest that there is a relationship between how the facility is portrayed in local media and public concern. This study has important implications for other nuclear facilities because of the role of local citizens in decision-making. - Highlights: ► Decisions about federal nuclear facilities include local citizen participation. ► Local media can play an important role in public perception of environmental risk. ► Local print media rely on a limited number in of sources for their stories. ► Effective risk communication should begin by understanding local public concerns.

  3. Characterization of a beta-catenin nuclear localization defect in MCF-7 breast cancer cells.

    Science.gov (United States)

    Jamieson, Cara; Mills, Kate M; Lui, Christina; Semaan, Crystal; Molloy, Mark P; Sharma, Manisha; Forwood, Jade K; Henderson, Beric R

    2016-02-15

    Beta-catenin plays a key role in transducing Wnt signals from the plasma membrane to the nucleus. Here we characterize an unusual subcellular distribution of beta-catenin in MCF-7 breast cancer cells, wherein beta-catenin localizes to the cytoplasm and membrane but atypically did not relocate to the nucleus after Wnt treatment. The inability of Wnt or the Wnt agonist LiCl to induce nuclear localization of beta-catenin was not due to defective nuclear transport, as the transport machinery was intact and ectopic GFP-beta-catenin displayed rapid nuclear entry in living cells. The mislocalization is explained by a shift in the retention of beta-catenin from nucleus to cytoplasm. The reduced nuclear retention is caused by unusually low expression of lymphoid enhancer factor/T-cell factor (LEF/TCF) transcription factors. The reconstitution of LEF-1 or TCF4 expression rescued nuclear localization of beta-catenin in Wnt treated cells. In the cytoplasm, beta-catenin accumulated in recycling endosomes, golgi and beta-COP-positive coatomer complexes. The peripheral association with endosomes diminished after Wnt treatment, potentially releasing β-catenin into the cytoplasm for nuclear entry. We propose that in MCF-7 and perhaps other breast cancer cells, beta-catenin may contribute to cytoplasmic functions such as ER-golgi transport, in addition to its transactivation role in the nucleus. PMID:26844628

  4. Characterization of the Nuclear Localization Signal of High Risk HPV16 E2 Protein

    OpenAIRE

    Klucevsek, Kristin; Wertz, Mary; Lucchi, John; Leszczynski, Anna; Moroianu, Junona

    2006-01-01

    The E2 protein of high risk human papillomavirus type 16 (HPV16) contains an amino-terminal (N) domain, a hinge (H) region and a carboxyl-terminal (C) DNA binding domain. Using enhanced green fluorescent protein (EGFP) fusions with full length E2 and E2 domains in transfection assays in HeLa cells we found that the C domain is responsible for the nuclear localization of E2 in vivo, whereas the N and H domains do not contain additional nuclear localization signals (NLSs). Deletion analysis of ...

  5. The RelA nuclear localization signal folds upon binding to IκBα

    OpenAIRE

    Cervantes, Carla F.; Bergqvist, Simon; Kjaergaard, Magnus; Kroon, Gerard; Sue, Shih-Che; Dyson, H Jane; Komives, Elizabeth A.

    2010-01-01

    The nuclear localization signal (NLS) polypeptide of RelA, the canonical NFκB family member, is responsible for regulating the nuclear localization of RelA-containing NFκB dimers. The RelA NLS polypeptide also plays a crucial role in mediating the high affinity and specificity of the interaction of RelA-containing dimers with the inhibitor IκBα, forming two helical motifs according to the published X-ray crystal structure. In order to define the nature of the interaction between the RelA NLS ...

  6. Local nuclear slope and curvature in high energy pp and pp-bar elastic scattering

    Energy Technology Data Exchange (ETDEWEB)

    Desgrolard, P. [Lyon-1 Univ., 69 - Villeurbanne (France). Inst. de Physique Nucleaire; Kontros, J.; Lengyel, A.I. [Inst. of Electron Physics, Uzhgorod (Ukraine); Martynov, E.S. [National Academy of Sciences of Ukraine, Kiev (Ukraine). Bogolyubov Inst. for Theoretical Physics

    1997-05-01

    The local nuclear slope is reconstructed from the experimental angular distributions with a procedure that uses overlapping t-bins, for an energy that ranges from the ISR to the Sp-bar pS and the Tevatron. Predictions of several models of (p-bar,p) elastic scattering at high energy are tested. Only a model with two-components Pomeron and Odderon gives a satisfactory agreement with the (non fitted) slope data. The extreme sensitivity of the local nuclear curvature with the choice for a Pomeron model is emphasized. (author). 30 refs.

  7. Local nuclear slope and curvature in high energy pp and pp-bar elastic scattering

    International Nuclear Information System (INIS)

    The local nuclear slope is reconstructed from the experimental angular distributions with a procedure that uses overlapping t-bins, for an energy that ranges from the ISR to the Sp-bar pS and the Tevatron. Predictions of several models of (p-bar,p) elastic scattering at high energy are tested. Only a model with two-components Pomeron and Odderon gives a satisfactory agreement with the (non fitted) slope data. The extreme sensitivity of the local nuclear curvature with the choice for a Pomeron model is emphasized. (author)

  8. The development and localization of nuclear fuel technology for KMRR

    International Nuclear Information System (INIS)

    This project was implemented aiming at localizing the fabrication of the KMRR fuel by october 1993. The contents of this project were divided into three parts: fuel design, fuel fabrication and process criticality analysis. In the fuel design, the radial power distribution in the fuel core was modeled and formulated taking account of the neutron flux depression in the radial direction. It was also performed to model and formulate the thermal characteristics such as the thermal conductivity and specific heat of the fuel core, U3Si-Al, the swelling and the film coefficient of heat transfer between the aluminum clad and light water coolant. The two dimensional heat transfer in the finned fuel element was equated based on the general equation governing the heat transfer in materials in order to develope a computer code, TEMP2D. TEMP2D solves finite differenced equations to calculate a two dimensional fuel temperature distribution under the steady and transient states. In the fuel fabrication, the technologies of fabricating uranium silicide fuel meat were tried by using depleted uranium as a raw material. These were extended to find the problems in technologies and to establish the ways of approach. The end product, so called fuel meat, was a metallic powder compound, U3Six(1≤x≤2), dispersed in Al matrix. The fuel meat was fabricated by the horizontal extrusion technique, and powder extrusion technique. Fabrication technologies comprise five different continuous processes: melting and casting of metallic uranium with silicon and aluminum, heat treatment, chipping and crushing, pulverizing, and extrusion. In the process criticality analysis, AMPX-KENO benchmark calculation was performed and calculational error of AMPX-KENO system was established. (Author)

  9. Development of Necessary Technology for localizing of Nuclear Safety Grade I and C System

    International Nuclear Information System (INIS)

    Through KNICS and Nu-Tech 2012, a MMIS(Man-Machine Interface System) package for nuclear plants was localized and is expected to be applied to SUN 1,2 plants. This study is aimed to support the application of the MMIS package including the following technologies that meet strengthened regulation requirements and enhanced utility's performance requirements to nuclear plants. - Dedication to safety-grade computers and real time operating systems - The broadband communication network for safety information - Application of the automatic test for safety systems - Application of programmable logic controllers to nuclear plants - Development strategy for an integrated SW development tool for control device

  10. P220-S Dual-Purpose Insect Cell Expression Vector for Transient Transfection and Baculovirus Generation

    OpenAIRE

    Loomis, K.; ROCKWELL, C; Sternard, H.; Novy, R.

    2007-01-01

    Baculovirus-mediated expression has proven to be a robust method of generating recombinant proteins from insect cells. However, generating baculovirus recombinants using traditional techniques is time consuming and tedious. To accelerate the process of insect cell expression, EMD developed a rapid transient transfection-based approach, the InsectDirect System. This approach is well suited for the rapid generation of small to moderate amounts of recombinant protein. For situations that demand ...

  11. Utilizing the virus-induced blocking of apoptosis in an easy baculovirus titration method

    OpenAIRE

    Athanasios Niarchos; George Lagoumintzis; Konstantinos Poulas

    2015-01-01

    Baculovirus-mediated protein expression is a robust experimental technique for producing recombinant higher-eukaryotic proteins because it combines high yields with considerable post-translational modification capabilities. In this expression system, the determination of the titer of recombinant baculovirus stocks is important to achieve the correct multiplicity of infection for effective amplification of the virus and high expression of the target protein. To overcome the drawbacks of existi...

  12. Protective Efficacy of Baculovirus Dual Expression System Vaccine Expressing Plasmodium falciparum Circumsporozoite Protein

    OpenAIRE

    Iyori, Mitsuhiro; Nakaya, Hiroki; Inagaki, Katsuya; Pichyangkul, Sathit; Yamamoto, Daisuke S.; Kawasaki, Masanori; Kwak, Kyungtak; Mizukoshi, Masami; Goto, Yoshihiro; Matsuoka, Hiroyuki; Matsumoto, Makoto; Yoshida, Shigeto

    2013-01-01

    We have previously developed a new malaria vaccine delivery system based on the baculovirus dual expression system (BDES). In this system, expression of malaria antigens is driven by a dual promoter consisting of the baculovirus-derived polyhedrin and mammal-derived cytomegalovirus promoters. To test this system for its potential as a vaccine against human malaria parasites, we investigated immune responses against the newly developed BDES-based Plasmodium falciparum circumsporozoite protein ...

  13. Optimization of canine interleukin-12 production using a baculovirus insect cell expression system

    OpenAIRE

    de Pinheiro, Cristiane Garboggini Melo; Pedrosa, Mayara de Oliveira; Teixeira, Naiara Carvalho; Ano Bom, Ana Paula Dinis; van Oers, Monique M.; Oliveira, Geraldo Gileno de Sá

    2016-01-01

    Background Interleukin-12 is an important cytokine in mediating cellular immune responses. Results Recombinant single-chain canine IL-12 was produced in a baculovirus-insect cell system with the aim of conducting further studies on modulation of immune responses in dogs. To optimize the production of recombinant canine IL-12, a classical baculovirus and a modified vector (chitinase A and v-cathepsin knockout) were used containing a native or an optimized insert of canine IL-12. The optimized ...

  14. A tubular segmented-flow bioreactor for the infection of insect cells with recombinant baculovirus

    OpenAIRE

    Hu, Yu-Chen; Wang, Ming-Ying; Bentley, William E.

    1997-01-01

    A continuous process of insect cell (S f9) growth and baculovirus infection is tested with the sequential combination of a CSTR and a tubular reactor. A tubular infection reactor enables continuous introduction of baculovirus and therefore avoids the ‘passage effect’ observed in two-stage CSTR systems. Moreover, a tubular reactor can be used to test cell infection kinetics and the subsequent metabolism of infected insect cells. Unlike batch and CSTR culture, cells in a horizontally positioned...

  15. Mapping Signals That Are Important for Nuclear and Nucleolar Localization in MCRS2

    OpenAIRE

    Kuo, Chung-Wen; Wang, Wen-Hung; Liu, Shih-Tung

    2011-01-01

    MCRS2 is an oncoprotein that is sequestered in the nucleolus. When in the nucleolus, it promotes the transcription of the rRNA gene. MCRS2 also brings proteins into the nucleolus to change their function. This study analyzes the sequence of MCRS2 and determines that the nuclear localization signal, which has the sequence KRKK, is situated between amino acids 66 and 69. Meanwhile, MCRS2 contains a bipartite nucleolar localization signal, which comprises a KKSK motif, located between amino acid...

  16. Anomalous organic magnetoresistance from competing carrier-spin-dependent interactions with localized electronic and nuclear spins

    OpenAIRE

    Wang, Y.; Harmon, N. J.; Sahin-Tiras, K.; Wohlgenannt, M.; Flatté, M. E.

    2014-01-01

    We describe a new regime for low-field magnetoresistance in organic semiconductors, in which the spin-relaxing effects of localized nuclear spins and electronic spins interfere. The regime is studied by the controlled addition of localized electronic spins to a material that exhibits substantial room-temperature magnetoresistance ($\\sim 20$\\%). Although initially the magnetoresistance is suppressed by the doping, at intermediate doping there is a regime where the magnetoresistance is insensit...

  17. Nuclear localization of the dehydrin OpsDHN1 is determined by histidine-rich motif.

    Science.gov (United States)

    Hernández-Sánchez, Itzell E; Maruri-López, Israel; Ferrando, Alejandro; Carbonell, Juan; Graether, Steffen P; Jiménez-Bremont, Juan F

    2015-01-01

    The cactus OpsDHN1 dehydrin belongs to a large family of disordered and highly hydrophilic proteins known as Late Embryogenesis Abundant (LEA) proteins, which accumulate during the late stages of embryogenesis and in response to abiotic stresses. Herein, we present the in vivo OpsDHN1 subcellular localization by N-terminal GFP translational fusion; our results revealed a cytoplasmic and nuclear localization of the GFP::OpsDHN1 protein in Nicotiana benthamiana epidermal cells. In addition, dimer assembly of OpsDHN1 in planta using a Bimolecular Fluorescence Complementation (BiFC) approach was demonstrated. In order to understand the in vivo role of the histidine-rich motif, the OpsDHN1-ΔHis version was produced and assayed for its subcellular localization and dimer capability by GFP fusion and BiFC assays, respectively. We found that deletion of the OpsDHN1 histidine-rich motif restricted its localization to cytoplasm, but did not affect dimer formation. In addition, the deletion of the S-segment in the OpsDHN1 protein affected its nuclear localization. Our data suggest that the deletion of histidine-rich motif and S-segment show similar effects, preventing OpsDHN1 from getting into the nucleus. Based on these results, the histidine-rich motif is proposed as a targeting element for OpsDHN1 nuclear localization. PMID:26442018

  18. Nuclear localization of the dehydrin OpsDHN1 is determined by histidine-rich motif

    Directory of Open Access Journals (Sweden)

    Itzell Euridice Hernández-Sánchez

    2015-09-01

    Full Text Available The cactus OpsDHN1 dehydrin belongs to a large family of disordered and highly hydrophilic proteins known as Late Embryogenesis Abundant (LEA proteins, which accumulate during the late stages of embryogenesis and in response to abiotic stresses. Herein, we present the in vivo OpsDHN1 subcellular localization by N-terminal GFP translational fusion; our results revealed a cytoplasmic and nuclear localization of the GFP::OpsDHN1 protein in Nicotiana benthamiana epidermal cells. In addition, dimer assembly of OpsDHN1 in planta using a Bimolecular Fluorescence Complementation (BiFC approach was demonstrated. In order to understand the in vivo role of the histidine-rich motif, the OpsDHN1-ΔHis version was produced and assayed for its subcellular localization and dimer capability by GFP fusion and BiFC assays, respectively. We found that deletion of the OpsDHN1 histidine-rich motif restricted its localization to cytoplasm, but did not affect dimer formation. In addition, the deletion of the S-segment in the OpsDHN1 protein affected its nuclear localization. Our data suggest that the deletion of histidine-rich motif and S-segment show similar effects, preventing OpsDHN1 from getting into the nucleus. Based on these results, the histidine rich motif is proposed as a targeting element for OpsDHN1 nuclear localization.

  19. Pluronic F127 nanomicelles engineered with nuclear localized functionality for targeted drug delivery

    International Nuclear Information System (INIS)

    PKKKRKV (Pro-Lys-Lys-Lys-Arg-Lys-Val, PV7), a seven amino acid peptide, has emerged as one of the primary nuclear localization signals that can be targeted into cell nucleus via the nuclear import machinery. Taking advantage of chemical diversity and biological activities of this short peptide sequence, in this study, Pluronic F127 nanomicelles engineered with nuclear localized functionality were successfully developed for intracellular drug delivery. These nanomicelles with the size ∼ 100 nm were self-assembled from F127 polymer that was flanked with two PV7 sequences at its both terminal ends. Hydrophobic anticancer drug doxorubicin (DOX) with inherent fluorescence was chosen as the model drug, which was found to be efficiently encapsulated into nanomicelles with the encapsulation efficiency at 72.68%. In comparison with the non-functionalized namomicelles, the microscopic observation reveals that PV7 functionalized nanomicelles display a higher cellular uptake, especially into the nucleus of HepG2 cells, due to the nuclear localization signal effects. Both cytotoxicity and apoptosis studies show that the DOX-loaded nanomicelles were more potent than drug nanomicelles without nuclear targeting functionality. It was thus concluded that PV7 functionalized nanomicelles could be a potentially alternative vehicle for nuclear targeting drug delivery. - Highlights: ► A new nuclear targeted drug delivery system based on micelles is developed. ► This micellar system features a core-shell structure with the size peaked at 100 nm. ► PV7, a short peptide sequence, is adopted as a nuclear targeting ligand. ► PV7 functionalized drug loaded micelles are more potent in killing tumor cells

  20. Stable replication of the EBNA1/OriP-mediated baculovirus vector and its application to anti-HCV gene therapy

    Directory of Open Access Journals (Sweden)

    Chang Myint OO

    2009-10-01

    Full Text Available Abstract Background Hepatitis C virus (HCV is one of the main causes of liver-related morbidity and mortality. Although combined interferon-α-ribavirin therapy is effective for about 50% of the patients with HCV, better therapies are needed and preventative vaccines have yet to be developed. Short-hairpin RNAs (shRNAs inhibit gene expression by RNA interference. The application of transient shRNA expression is limited, however, due to the inability of the shRNA to replicate in mammalian cells and its inefficient transduction. The duration of transgene (shRNA expression in mammalian cells can be significantly extended using baculovirus-based shRNA-expressing vectors that contain the latent viral protein Epstein-Barr nuclear antigen 1 (EBNA1 and the origin of latent viral DNA replication (OriP sequences. These recombinant vectors contain compatible promoters and are highly effective for infecting primary hepatocyte and hepatoma cell lines, making them very useful tools for studies of hepatitis B and hepatitis C viruses. Here, we report the use of these baculovirus-based vector-derived shRNAs to inhibit core-protein expression in full-length hepatitis C virus (HCV replicon cells. Results We constructed a long-term transgene shRNA expression vector that contains the EBV EBNA1 and OriP sequences. We also designed baculovirus vector-mediated shRNAs against the highly conserved core-protein region of HCV. HCV core protein expression was inhibited by the EBNA1/OriP baculovirus vector for at least 14 days, which was considerably longer than the 3 days of inhibition produced by the wild-type baculovirus vector. Conclusion These findings indicate that we successfully constructed a long-term transgene (shRNA expression vector (Ac-EP-shRNA452 using the EBNA1/OriP system, which was propagated in Escherichia coli and converted into mammalian cells. The potential anti-HCV activity of the long-term transgene (shRNA expression vector was evaluated with the view

  1. Identification of a functional nuclear localization signal within the human USP22 protein

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Jianjun [Key Laboratory of Jiangxi Province for the Systems Bio-Medicine, Jiujiang, Jiangxi Province 332000 (China); College of Basic Medical Science, Jiujiang University, Jiujiang, Jiangxi Province 332000 (China); Wang, Yaqin [Reproductive Medical Center, Renmin Hospital of Wuhan University, Wuhan, Hubei Province 430060 (China); Gong, Zhen [Key Laboratory of Jiangxi Province for the Systems Bio-Medicine, Jiujiang, Jiangxi Province 332000 (China); Liu, Jianyun [College of Basic Medical Science, Jiujiang University, Jiujiang, Jiangxi Province 332000 (China); Li, Weidong, E-mail: lwd626518@163.com [College of Basic Medical Science, Jiujiang University, Jiujiang, Jiangxi Province 332000 (China)

    2014-06-20

    Highlights: • USP22 was accumulated in nucleus. • We identified of a functional USP22 NLS. • The KRRK amino acid residues are indispensable in NLS. • The KRRK motif is conserved in USP22 homologues. - Abstract: Ubiquitin-specific processing enzyme 22 (USP22), a member of the deubiquitinase family, is over-expressed in most human cancers and has been implicated in tumorigenesis. Because it is an enzymatic subunit of the human SAGA transcriptional cofactor, USP22 deubiquitylates histone H2A and H2B in the nucleus, thus participating in gene regulation and cell-cycle progression. However, the mechanisms regulating its nuclear translocation have not yet been elucidated. It was here demonstrated that USP22 is imported into the nucleus through a mechanism mediated by nuclear localization signal (NLS). The bipartite NLS sequence KRELELLKHNPKRRKIT (aa152–168), was identified as the functional NLS for its nuclear localization. Furthermore, a short cluster of basic amino acid residues KRRK within this bipartite NLS plays the primary role in nuclear localization and is evolutionarily conserved in USP22 homologues. In the present study, a functional NLS and the minimal sequences required for the active targeting of USP22 to the nucleus were identified. These findings may provide a molecular basis for the mechanism underlying USP22 nuclear trafficking and function.

  2. Identification of a functional nuclear localization signal within the human USP22 protein

    International Nuclear Information System (INIS)

    Highlights: • USP22 was accumulated in nucleus. • We identified of a functional USP22 NLS. • The KRRK amino acid residues are indispensable in NLS. • The KRRK motif is conserved in USP22 homologues. - Abstract: Ubiquitin-specific processing enzyme 22 (USP22), a member of the deubiquitinase family, is over-expressed in most human cancers and has been implicated in tumorigenesis. Because it is an enzymatic subunit of the human SAGA transcriptional cofactor, USP22 deubiquitylates histone H2A and H2B in the nucleus, thus participating in gene regulation and cell-cycle progression. However, the mechanisms regulating its nuclear translocation have not yet been elucidated. It was here demonstrated that USP22 is imported into the nucleus through a mechanism mediated by nuclear localization signal (NLS). The bipartite NLS sequence KRELELLKHNPKRRKIT (aa152–168), was identified as the functional NLS for its nuclear localization. Furthermore, a short cluster of basic amino acid residues KRRK within this bipartite NLS plays the primary role in nuclear localization and is evolutionarily conserved in USP22 homologues. In the present study, a functional NLS and the minimal sequences required for the active targeting of USP22 to the nucleus were identified. These findings may provide a molecular basis for the mechanism underlying USP22 nuclear trafficking and function

  3. EpCAM nuclear localization identifies aggressive Thyroid Cancer and is a marker for poor prognosis

    Directory of Open Access Journals (Sweden)

    MacMillan Christina

    2010-06-01

    Full Text Available Abstract Background Proteolytic cleavage of the extracellular domain (EpEx of Epithelial cell adhesion molecule (EpCAM and nuclear signaling by its intracellular oncogenic domain Ep-ICD has recently been implicated in increased proliferation of cancer cells. The clinical significance of Ep-ICD in human tumors remains an enigma. Methods EpEx, Ep-ICD and β-catenin immunohistochemistry using specific antibodies was conducted on 58 archived thyroid cancer (TC tissue blocks from 34 patients and correlated with survival analysis of these patients for up to 17 years. Results The anaplastic (ATC and aggressive thyroid cancers showed loss of EpEx and increased nuclear and cytoplasmic accumulation of Ep-ICD. In contrast, the low grade papillary thyroid cancers (PTC showed membranous EpEx and no detectable nuclear Ep-ICD. The ATC also showed concomitant nuclear expression of Ep-ICD and β-catenin. Kaplan-Meier Survival analysis revealed reduced overall survival (OS for TC patients showing nuclear Ep-ICD expression or loss of membranous EpEx (p Conclusion We report reciprocal loss of membrane EpEx but increased nuclear and cytoplasmic accumulation of Ep-ICD in aggressive TC; nuclear Ep-ICD correlated with poor OS of TC patients. Thus nuclear Ep-ICD localization may serve as a useful biomarker for aggressive TC and may represent a novel diagnostic, prognostic and therapeutic target for aggressive TC.

  4. Nuclear binding energies: global collective structures and local shell-model correlations

    International Nuclear Information System (INIS)

    In this contribution the global behaviour of nuclear binding energies, or alternatively two-neutron separation energies, are studied. Recent high-precision mass measurements show local deviations from an overall macroscopic behaviour, and it is shown how a consistent and simultaneously description of both can be given within the Interacting Boson Model. (orig.)

  5. The N-terminal 33 amino acid domain of Siva-1 is sufficient for nuclear localization

    International Nuclear Information System (INIS)

    Siva-1 induces apoptosis in multiple pathological processes and plays an important role in the suppression of tumor metastasis, protein degradation, and other functions. Although many studies have demonstrated that Siva-1 functions in the cytoplasm, a few have found that Siva-1 can relocate to the nucleus. In this study, we found that the first 33 amino acid residues of Siva-1 are required for its nuclear localization. Further study demonstrated that the green fluorescent protein can be imported into the nucleus after fusion with these 33 amino acid residues. Other Siva-1 regions and domains showed less effect on Siva-1 nuclear localization. By site-mutagenesis of all of these 33 amino acid residues, we found that mutants of the first 1-18 amino acids affected Siva-1 nuclear compartmentalization but could not complete this localization independently. In summary, we demonstrated that the N-terminal 33 amino acid residues were sufficient for Siva-1 nuclear localization, but the mechanism of this translocation needs additional investigation

  6. The N-terminal 33 amino acid domain of Siva-1 is sufficient for nuclear localization

    Energy Technology Data Exchange (ETDEWEB)

    Chen, J.Y.; Yang, L.X. [Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou (China); Huang, Z.F. [Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou (China); Department of Biochemistry, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou (China); Key Laboratory of Tropical Diseases Control, Sun Yat-sen University, Ministry of Education in China, Guangzhou (China)

    2013-12-02

    Siva-1 induces apoptosis in multiple pathological processes and plays an important role in the suppression of tumor metastasis, protein degradation, and other functions. Although many studies have demonstrated that Siva-1 functions in the cytoplasm, a few have found that Siva-1 can relocate to the nucleus. In this study, we found that the first 33 amino acid residues of Siva-1 are required for its nuclear localization. Further study demonstrated that the green fluorescent protein can be imported into the nucleus after fusion with these 33 amino acid residues. Other Siva-1 regions and domains showed less effect on Siva-1 nuclear localization. By site-mutagenesis of all of these 33 amino acid residues, we found that mutants of the first 1-18 amino acids affected Siva-1 nuclear compartmentalization but could not complete this localization independently. In summary, we demonstrated that the N-terminal 33 amino acid residues were sufficient for Siva-1 nuclear localization, but the mechanism of this translocation needs additional investigation.

  7. Nuclear magnetic resonance of iron-57 nuclei in local fields in yttrium and iron garnets

    International Nuclear Information System (INIS)

    We have demonstrated the nuclear resonance of 57Fe nuclei in the local field of each of the two magnetic sub-lattices of yttrium and iron garnets. The resonance frequencies and the relaxation times have been measured as a function of the temperature. (author)

  8. Nuclear localization of human DNA mismatch repair protein exonuclease 1 (hEXO1)

    DEFF Research Database (Denmark)

    Knudsen, Nina Østergaard; Nielsen, Finn Cilius; Vinther, Lena;

    2007-01-01

    localization signals (NLSs) in hEXO1. Using fluorescent fusion proteins, we show that the sequence 418KRPR421, which exhibit strong homology to other monopartite NLS sequences, is responsible for correct nuclear localization of hEXO1. This NLS sequence is located in a region that is also required for hEXO1......Human exonuclease 1 (hEXO1) is implicated in DNA mismatch repair (MMR) and mutations in hEXO1 may be associated with hereditary nonpolyposis colorectal cancer (HNPCC). Since the subcellular localization of MMR proteins is essential for proper MMR function, we characterized possible nuclear...... interaction with hMLH1 and we show that defective nuclear localization of hEXO1 mutant proteins could be rescued by hMLH1 or hMSH2. Both hEXO1 and hMLH1 form complexes with the nuclear import factors importin beta/alpha1,3,7 whereas hMSH2 specifically recognizes importin beta/alpha3. Taken together, we infer...

  9. Characterization, subcellular localization and nuclear targeting of casein kinase 2 from Zea mays

    DEFF Research Database (Denmark)

    Peracchia, G; Jensen, A B; Culiáñez-Macià, F A;

    1999-01-01

    by using in-frame fusions of the maize CK2alpha subunit to the reporter gene encoding beta-glucuronidase (GUS) which were assayed in transiently transformed onion epidermal cells. Analysis of chimeric constructs identified one region containing a nuclear localization signal (NLS) that is highly conserved...

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

    International Nuclear Information System (INIS)

    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

  11. Do nuclear collisions create a locally equilibrated quark-gluon plasma?

    CERN Document Server

    Romatschke, Paul

    2016-01-01

    Experimental results on azimuthal correlations in high energy nuclear collisions (nucleus-nucleus, proton-nucleus and proton-proton) seem to be well described by viscous hydrodynamics. It is often argued that this agreement implies either local thermal equilibrium or at least local isotropy. In this note, I present arguments why this is not the case. Neither local near-equilibrium nor near-isotropy are required in order for hydrodynamics to offer a successful and accurate description of experimental results. However, I predict the breakdown of hydrodynamics at momenta of order twenty times the temperature, corresponding to a smallest possible QCD liquid drop size of 0.05 fm.

  12. INVESTIGATION OF LATENT NUCLEAR LOCALIZATION SEQUENCE(NLS) OF STAT3

    Institute of Scientific and Technical Information of China (English)

    杨镇珲; 李惠; 叶中德; 冯健男; 沈倍奋

    2004-01-01

    Objective: To investigate the latent nuclear localization sequence (NLS) OF STAT3. Methods: Clustal X (1.81) was used to alignment the DNA binding domain of the STAT family. According to structure characters, corresponding expression plasmids were constructed via oligonucleotides designed with gene tool software. Through in vitro transfection, images were observed with laser confocal microscopy. Results: homology positions of arginine and lysine were found in DNA binding domain of the stat family. The wild type-STAT3 proteins primarily localized in the cytoplasm and translocated into the nucleus after interleukin-6 stimulation. However, the truncated mutant of DSTAT3-GFP protein was exclusively expressed in the cytoplasm. Conclusion: The potential NLS in the DNA binding domain of STAT3 is exposed to nuclear importing receptor when cells are stinulated by cytokine, which promotes the translocation of STAT3 into nuclear.

  13. Rad52 multimerization is important for its nuclear localization in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Plate, Iben; Albertsen, Line; Lisby, Michael;

    2008-01-01

    Rad52 is essential for all homologous recombination and DNA double strand break repair events in Saccharomyces cerevisiae. This protein is multifunctional and contains several domains that allow it to interact with DNA as well as with different repair proteins. However, it has been unclear how Ra...... on the results we propose a model where the additive effect of multiple NLS(Rad52) sequences in a Rad52 ring-structure ensures efficient nuclear localization of Rad52.......52 enters the nucleus. In the present study, we have used a combination of mutagenesis and sequence analysis to show that Rad52 from S. cerevisiae contains a single functional pat7 type NLS essential for its nuclear localization. The region containing the NLS seems only to be involved in nuclear...

  14. Construction of recombinant baculoviruses expressing hemagglutinin of H5N1 avian influenza and research on the immunogenicity

    Science.gov (United States)

    Ge, Jingping; An, Qi; Gao, Dongni; Liu, Ying; Ping, Wenxiang

    2016-01-01

    Recombinant baculoviruses with different promoter and regulatory elements were constructed to enhance the expression of target protein and boost the efficacies of avian influenza vaccine. Hemagglutinin gene was cloned into the baculovirus transfer vectors driven by cytomegaloviru (CMV) and White spot syndrome virus immediate-early promoter one (WSSV ie1) promoter respectively, with different regulatory elements. The recombinant baculoviruses were directly used as vaccines to immunize specific pathogen-free chickens. The protein expression levels of recombinant baculoviruses BV-S-HA and BV-S-ITRs-HA were respectively 2.43 and 2.67 times than that of BV-S-con-HA, while the protein expression levels of BV-A-HA and BV-A-ITRs-HA were respectively 2.44 and 2.69 times than that of BV-S-con-HA. Immunoglobulin G (IgG) antibody levels induced by BV-A and BV-S series recombinant baculovirus were significantly higher than the commercialized vaccine group (P < 0.05). Among the groups with same promoter, the IgG antibody levels induced by the baculovirus containing regulatory elements were significantly higher than control group. Additionally, the immune effects induced by BV-A series recombinant baculoviruses with WSSV ie1 promoter were significantly stronger than the BV-S series recombinant baculoviruses with CMV promoter. The avian influenza vaccine prepared based on baculovirus vector can simultaneously stimulate the humoral and cellular immune responses. PMID:27063566

  15. Nuclear localization of Rad51B is independent of BRCA2

    Energy Technology Data Exchange (ETDEWEB)

    Miller, K A; Hinz, J M; Yamada, A; Thompson, L H; Albala, J S

    2005-06-28

    Human Rad51 is critical for the maintenance of genome stability through its role in the repair of DNA double-strand breaks. Rad51B (Rad51L1/hRec2) is one of the five known paralogs of human Rad51 found in a multi-protein complex with three other Rad51 paralogs, Rad51C, Rad51D and Xrcc2. Examination of EGFP-Rad51B fusion protein in HeLa S3 cells and immunofluorescence in several human cell lines confirms the nuclear localization of Rad51B. This is the first report to detail putative interactions of a Rad51 paralog protein with BRCA2. Utilization of a BRCA2 mutant cell line, CAPAN-1 suggests that Rad51B localizes to the nucleus independent of BRCA2. Although both Rad51B and BRCA2 are clearly involved in the homologous recombinational repair pathway, Rad51B and BRCA2 do not appear to associate directly. Furthermore, mutations in the KKLK motif of Rad51B, amino acid residues 4-7, mislocalizes Rad51B to the cytoplasm suggesting that this is the nuclear localization signal for the Rad51B protein. Examination of wild-type EGFP-Rad51B fusion protein in mammalian cells deficient in Rad51C showed that Rad51B localizes to the nucleus independent of Rad51C; further suggesting that Rad51B, like Rad51C, contains its own nuclear localization signal.

  16. Oscillator strengths, first-order properties, and nuclear gradients for local ADC(2)

    Energy Technology Data Exchange (ETDEWEB)

    Schütz, Martin, E-mail: martin.schuetz@chemie.uni-regensburg.de [Institute of Physical and Theoretical Chemistry, University of Regensburg, Universitätsstraße 31, D-93040 Regensburg (Germany)

    2015-06-07

    We describe theory and implementation of oscillator strengths, orbital-relaxed first-order properties, and nuclear gradients for the local algebraic diagrammatic construction scheme through second order. The formalism is derived via time-dependent linear response theory based on a second-order unitary coupled cluster model. The implementation presented here is a modification of our previously developed algorithms for Laplace transform based local time-dependent coupled cluster linear response (CC2LR); the local approximations thus are state specific and adaptive. The symmetry of the Jacobian leads to considerable simplifications relative to the local CC2LR method; as a result, a gradient evaluation is about four times less expensive. Test calculations show that in geometry optimizations, usually very similar geometries are obtained as with the local CC2LR method (provided that a second-order method is applicable). As an exemplary application, we performed geometry optimizations on the low-lying singlet states of chlorophyllide a.

  17. 1. European conference on local democracies and nuclear power plants. Conference proceedings

    International Nuclear Information System (INIS)

    The Group of European Municipalities with Nuclear Facilities (GMF) was constituted in 1999. Its basic objectives are to keep and strengthen the relations between municipalities with the aim of consolidating a European network of municipalities with common problems; and to participate in European forums on the future of nuclear energy, contributing and helping in decision making processes. The GMF wants to become a support element to the local authorities of nuclear areas in order to foster their participation in the decision making processes, working continuously in collaboration with other agents of the nuclear sector, especially with the authorities of the European Commission, assuming that its major contribution is the knowledge of the everyday territorial reality, as a fundamental element that should be born in mind in all defined policies, whether on european or national level. It is stated that the debate on the future of nuclear energy represents a new era for the municipalities with nuclear facilities across Europe and that international cooperation will open the way for a better use of nuclear energy in Europe

  18. Nuclear localization of platelet-activating factor receptor controls retinal neovascularization

    Science.gov (United States)

    K Bhosle, Vikrant; Rivera, José Carlos; Zhou, Tianwei (Ellen); Omri, Samy; Sanchez, Melanie; Hamel, David; Zhu, Tang; Rouget, Raphael; Rabea, Areej Al; Hou, Xin; Lahaie, Isabelle; Ribeiro-da-Silva, Alfredo; Chemtob, Sylvain

    2016-01-01

    Platelet-activating factor (PAF) is a pleiotropic phospholipid with proinflammatory, procoagulant and angiogenic actions on the vasculature. We and others have reported the presence of PAF receptor (Ptafr) at intracellular sites such as the nucleus. However, mechanisms of localization and physiologic functions of intracellular Ptafr remain poorly understood. We hereby identify the importance of C-terminal motif of the receptor and uncover novel roles of Rab11a GTPase and importin-5 in nuclear translocation of Ptafr in primary human retinal microvascular endothelial cells. Nuclear localization of Ptafr is independent of exogenous PAF stimulation as well as intracellular PAF biosynthesis. Moreover, nuclear Ptafr is responsible for the upregulation of unique set of growth factors, including vascular endothelial growth factor, in vitro and ex vivo. We further corroborate the intracrine PAF signaling, resulting in angiogenesis in vivo, using Ptafr antagonists with distinct plasma membrane permeability. Collectively, our findings show that nuclear Ptafr translocates in an agonist-independent manner, and distinctive functions of Ptafr based on its cellular localization point to another dimension needed for pharmacologic selectivity of drugs. PMID:27462464

  19. Regulation of RelA Subcellular Localization by a Putative Nuclear Export Signal and p50

    OpenAIRE

    Harhaj, Edward W; Sun, Shao-Cong

    1999-01-01

    Nuclear factor κB (NF-κB) represents a family of dimeric DNA binding proteins, the pleotropic form of which is a heterodimer composed of RelA and p50 subunits. The biological activity of NF-κB is controlled through its subcellular localization. Inactive NF-κB is sequestered in the cytoplasm by physical interaction with an inhibitor, IκBα. Signal-mediated IκBα degradation triggers the release and subsequent nuclear translocation of NF-κB. It remains unknown whether the NF-κB shuttling between ...

  20. EpCAM nuclear localization identifies aggressive Thyroid Cancer and is a marker for poor prognosis

    International Nuclear Information System (INIS)

    Proteolytic cleavage of the extracellular domain (EpEx) of Epithelial cell adhesion molecule (EpCAM) and nuclear signaling by its intracellular oncogenic domain Ep-ICD has recently been implicated in increased proliferation of cancer cells. The clinical significance of Ep-ICD in human tumors remains an enigma. EpEx, Ep-ICD and β-catenin immunohistochemistry using specific antibodies was conducted on 58 archived thyroid cancer (TC) tissue blocks from 34 patients and correlated with survival analysis of these patients for up to 17 years. The anaplastic (ATC) and aggressive thyroid cancers showed loss of EpEx and increased nuclear and cytoplasmic accumulation of Ep-ICD. In contrast, the low grade papillary thyroid cancers (PTC) showed membranous EpEx and no detectable nuclear Ep-ICD. The ATC also showed concomitant nuclear expression of Ep-ICD and β-catenin. Kaplan-Meier Survival analysis revealed reduced overall survival (OS) for TC patients showing nuclear Ep-ICD expression or loss of membranous EpEx (p < 0.0004), median OS = 5 months as compared to 198 months for patients who did not show nuclear Ep-ICD or demonstrated only membranous EpE. We report reciprocal loss of membrane EpEx but increased nuclear and cytoplasmic accumulation of Ep-ICD in aggressive TC; nuclear Ep-ICD correlated with poor OS of TC patients. Thus nuclear Ep-ICD localization may serve as a useful biomarker for aggressive TC and may represent a novel diagnostic, prognostic and therapeutic target for aggressive TC

  1. Phosphatidylinositol 3-kinase/Akt signaling enhances nuclear localization and transcriptional activity of BRCA1

    International Nuclear Information System (INIS)

    Signaling pathways involved in regulating nuclear-cytoplasmic distribution of BRCA1 have not been previously reported. Here, we provide evidence that heregulin β1-induced activation of the Akt pathway increases the nuclear content of BRCA1. First, treatment of T47D breast cancer cells with heregulin β1 results in a two-fold increase in nuclear BRCA1 as assessed by FACS analysis, immunoblotting and immunofluorescence. This heregulin-induced increase in nuclear BRCA1 is blocked by siRNA-mediated down-regulation of Akt. Second, mutation of threonine 509 in BRCA1, the site of Akt phosphorylation, to an alanine, attenuates the ability of heregulin to induce BRCA1 nuclear accumulation. These data suggest that Akt-catalyzed phosphorylation of BRCA1 is required for the heregulin-regulated nuclear concentration of BRCA1. Because most functions ascribed to BRCA1 occur within the nucleus, we postulated that phosphorylation-dependent nuclear accumulation of BRCA1 would result in enhanced nuclear activity, specifically transcriptional activity, of BRCA1. This postulate is affirmed by our observation that the ability of BRCA1 to transactivate GADD45 promoter constructs was enhanced in T47D cells treated with heregulin β1. Furthermore, the heterologous expression of BRCA1 in HCC1937 human breast cancer cells, which have constitutively active Akt, also induces GADD45 promoter activity, whereas the expression of BRCA1 in which threonine 509 has been mutated to an alanine is able to only minimally induce promoter activity. These findings implicate Akt in upstream events leading to BRCA1 nuclear localization and function

  2. Cloning, expression and nuclear localization of human NPM3, a member of the nucleophosmin/nucleoplasmin family of nuclear chaperones

    Directory of Open Access Journals (Sweden)

    Ganguly Amit

    2001-11-01

    Full Text Available Abstract Background Studies suggest that the related proteins nucleoplasmin and nucleophosmin (also called B23, NO38 or numatrin are nuclear chaperones that mediate the assembly of nucleosomes and ribosomes, respectively, and that these activities are accomplished through the binding of basic proteins via their acidic domains. Recently discovered and less well characterized members of this family of acidic phosphoproteins include mouse nucleophosmin/nucleoplasmin 3 (Npm3 and Xenopus NO29. Here we report the cloning and initial characterization of the human ortholog of Npm3. Results Human genomic and cDNA clones of NPM3 were isolated and sequenced. NPM3 lies 5.5 kb upstream of FGF8 and thus maps to chromosome 10q24-26. In addition to amino acid similarities, NPM3 shares many physical characteristics with the nucleophosmin/nucleoplasmin family, including an acidic domain, multiple potential phosphorylation sites and a putative nuclear localization signal. Comparative analyses of 14 members of this family from various metazoans suggest that Xenopus NO29 is a candidate ortholog of human and mouse NPM3, and they further group both proteins closer with the nucleoplasmins than with the nucleophosmins. Northern blot analysis revealed that NPM3 was strongly expressed in all 16 human tissues examined, with especially robust expression in pancreas and testis; lung displayed the lowest level of expression. An analysis of subcellular fractions of NIH3T3 cells expressing epitope-tagged NPM3 revealed that NPM3 protein was localized solely in the nucleus. Conclusions Human NPM3 is an abundant and widely expressed protein with primarily nuclear localization. These biological activities, together with its physical relationship to the chaparones nucleoplasmin and nucleophosmin, are consistent with the proposed function of NPM3 as a molecular chaperone functioning in the nucleus.

  3. Localized corrosion of a candidate container material for high-level nuclear waste disposal

    International Nuclear Information System (INIS)

    Localized corrosion is one of the important considerations in the design of metallic containers used for the geologic disposal of high-level nuclear waste. This paper addresses the effect of environmental factors on the localized corrosion behavior of alloy 825, one of the candidate alloys for containers in the Yucca Mountain repository site. A two-level, full factorial experimental design was used to examine the main effects and interactions of chloride, sulfate, nitrate, fluoride, and temperature. This was augmented by additional experiments involving chloride and temperature at several levels. Cyclic, potentiodynamic polarization tests were used to determine the relative susceptibility of the alloy to localized corrosion. Crevice corrosion was detected at chloride levels as low as 20 ppm, and both pitting and crevice corrosion were observed at higher chloride levels. Among the environmental factors, chloride and sulfate were found to be promoters of localized corrosion, while nitrate and fluoride were inhibitors of localized corrosion. The experiments indicated that the electrochemical parameters (e.g., pitting potential, repassivation potential, or the difference between them) were not sufficient indicators of localized corrosion. Instead, the visual observation and electrochemical parameters were combined into an index, termed localized corrosion index (LCI), to quantify the extent of localized corrosion

  4. State and local planning procedures dealing with social and economic impacts from nuclear power plants

    International Nuclear Information System (INIS)

    The roles of state and local agencies in planning for and managing social and economic impacts of nuclear power plants are studied. In order to be effective in these roles state and local agencies must work with each other as well as the NRC. A comparative case study approach is used which analyzes six sites in three West Coast states. The case studies included plants in operation, plants under construction, and plants still in the planning stages. In contrast to some states, all three of these states have moderately centralized procedures for siting power plants, and all have strong environmental laws

  5. Final disposal of nuclear waste. Local communities and the site selection process

    International Nuclear Information System (INIS)

    The purpose of this report is to give information about how the five local Swedish communities that have been involved in the plans for selecting a site for the Swedish repository for spent nuclear fuels, have planned their work. The information is primarily directed towards representatives for the local communities that in the future might get involved in the selection process. The report tries to answer questions like: Which decision stages can be foreseen? What possibilities - formal and real - exist for leaving the selection process at a later stage? What demands will arise for inquiries and for public information? What support can governmental agencies give?

  6. Expression, Delivery and Function of Insecticidal Proteins Expressed by Recombinant Baculoviruses

    Directory of Open Access Journals (Sweden)

    Jeremy A. Kroemer

    2015-01-01

    Full Text Available Since the development of methods for inserting and expressing genes in baculoviruses, a line of research has focused on developing recombinant baculoviruses that express insecticidal peptides and proteins. These recombinant viruses have been engineered with the goal of improving their pesticidal potential by shortening the time required for infection to kill or incapacitate insect pests and reducing the quantity of crop damage as a consequence. A wide variety of neurotoxic peptides, proteins that regulate insect physiology, degradative enzymes, and other potentially insecticidal proteins have been evaluated for their capacity to reduce the survival time of baculovirus-infected lepidopteran host larvae. Researchers have investigated the factors involved in the efficient expression and delivery of baculovirus-encoded insecticidal peptides and proteins, with much effort dedicated to identifying ideal promoters for driving transcription and signal peptides that mediate secretion of the expressed target protein. Other factors, particularly translational efficiency of transcripts derived from recombinant insecticidal genes and post-translational folding and processing of insecticidal proteins, remain relatively unexplored. The discovery of RNA interference as a gene-specific regulation mechanism offers a new approach for improvement of baculovirus biopesticidal efficacy through genetic modification.

  7. Assessment of national systems for obtaining local acceptance of nuclear waste management siting activities

    International Nuclear Information System (INIS)

    On behalf of the United States Department of Energy (DOE), International Energy Associates Limited (IEAL) of Washington, D.C. has conducted surveys and analyses of fourteen countries' plans and approaches for dealing with the problems of obtaining local siting acceptance for nuclear waste management facilities. It was determined that the following elements of the formal systems generally facilitate and/or expedite waste management siting decisions: (1) a clear-cut pro-nuclear power position on the part of the government; (2) a willingness on the part of the central government to exert (with prudence and restraint) its pre-emptive rights in nuclear matters; (3) political structures in which the heads of regional or provincial governments are appointed by the central government; (4) national laws that link reactor licensing with a detailed plan for waste management; (5) an established and stable policy with regard to reprocessing. In contrast, it was determined that the following elements of the formal system generally hinder waste management siting activities: (1) historically strong local land used veto laws; (2) the use of national referenda for making nuclear decisions; (3) requirements for public hearings. The informal approaches fall into the following five categories: (1) political: e.g. assertion of will by political leaders, activities to enlist support of local politicians, activities to broaden involvement in decision-making; (2) economic: e.g. emphasis on normal benefits, provision for additional economic benefits; (3) siting: e.g. at or near existing nuclear facilities, on government or utility property, at multiple locations to spread the political burden; (4) timing: e.g. decoupling drilling activities from ultimate repository site decision, deliberate deferral to (long-range) future; (5) education: e.g. creation of special government programmes, enlisting of media support

  8. Automated local bright feature image analysis of nuclear protein distribution identifies changes in tissue phenotype

    International Nuclear Information System (INIS)

    The organization of nuclear proteins is linked to cell and tissue phenotypes. When cells arrest proliferation, undergo apoptosis, or differentiate, the distribution of nuclear proteins changes. Conversely, forced alteration of the distribution of nuclear proteins modifies cell phenotype. Immunostaining and fluorescence microscopy have been critical for such findings. However, there is an increasing need for quantitative analysis of nuclear protein distribution to decipher epigenetic relationships between nuclear structure and cell phenotype, and to unravel the mechanisms linking nuclear structure and function. We have developed imaging methods to quantify the distribution of fluorescently-stained nuclear protein NuMA in different mammary phenotypes obtained using three-dimensional cell culture. Automated image segmentation of DAPI-stained nuclei was generated to isolate thousands of nuclei from three-dimensional confocal images. Prominent features of fluorescently-stained NuMA were detected using a novel local bright feature analysis technique, and their normalized spatial density calculated as a function of the distance from the nuclear perimeter to its center. The results revealed marked changes in the distribution of the density of NuMA bright features as non-neoplastic cells underwent phenotypically normal acinar morphogenesis. In contrast, we did not detect any reorganization of NuMA during the formation of tumor nodules by malignant cells. Importantly, the analysis also discriminated proliferating non-neoplastic cells from proliferating malignant cells, suggesting that these imaging methods are capable of identifying alterations linked not only to the proliferation status but also to the malignant character of cells. We believe that this quantitative analysis will have additional applications for classifying normal and pathological tissues

  9. Automated local bright feature image analysis of nuclear proteindistribution identifies changes in tissue phenotype

    Energy Technology Data Exchange (ETDEWEB)

    Knowles, David; Sudar, Damir; Bator, Carol; Bissell, Mina

    2006-02-01

    The organization of nuclear proteins is linked to cell and tissue phenotypes. When cells arrest proliferation, undergo apoptosis, or differentiate, the distribution of nuclear proteins changes. Conversely, forced alteration of the distribution of nuclear proteins modifies cell phenotype. Immunostaining and fluorescence microscopy have been critical for such findings. However, there is an increasing need for quantitative analysis of nuclear protein distribution to decipher epigenetic relationships between nuclear structure and cell phenotype, and to unravel the mechanisms linking nuclear structure and function. We have developed imaging methods to quantify the distribution of fluorescently-stained nuclear protein NuMA in different mammary phenotypes obtained using three-dimensional cell culture. Automated image segmentation of DAPI-stained nuclei was generated to isolate thousands of nuclei from three-dimensional confocal images. Prominent features of fluorescently-stained NuMA were detected using a novel local bright feature analysis technique, and their normalized spatial density calculated as a function of the distance from the nuclear perimeter to its center. The results revealed marked changes in the distribution of the density of NuMA bright features as non-neoplastic cells underwent phenotypically normal acinar morphogenesis. In contrast, we did not detect any reorganization of NuMA during the formation of tumor nodules by malignant cells. Importantly, the analysis also discriminated proliferating non-neoplastic cells from proliferating malignant cells, suggesting that these imaging methods are capable of identifying alterations linked not only to the proliferation status but also to the malignant character of cells. We believe that this quantitative analysis will have additional applications for classifying normal and pathological tissues.

  10. Attitudes towards risks and opinions on a repository for spent nuclear fuel in four local communities

    International Nuclear Information System (INIS)

    A questionnaire was mailed to 1500 persons in each of the four communities that are candidates for siting, or otherwise involved, of the nuclear waste repository. 80 of the non-respondents were interviewed over telephone. The questionnaire included questions about the inhabitants attitude towards locating a repository in their home community, about participating in a local referendum, about trust in experts and organisations and about knowledge in the radioactive waste issues

  11. A toolbox of endogenous and heterologous nuclear localization sequences for the methylotrophic yeast Pichia pastoris

    OpenAIRE

    Weninger, Astrid; Glieder, Anton; Vogl, Thomas

    2015-01-01

    Nuclear localization sequences (NLSs) are required for the import of proteins in the nucleus of eukaryotes. However many proteins from bacteria or bacteriophages are used for basic studies in molecular biology, to generate synthetic genetic circuits or for genome editing applications. Prokaryotic recombinases, CRISPR-associated proteins such as Cas9 or bacterial and viral polymerases require efficient NLSs to function in eukaryotes. The yeast Pichia pastoris is a widely used expression platfo...

  12. Nuclear charge-exchange excitations in localized covariant density functional theory

    International Nuclear Information System (INIS)

    The recent progress in the studies of nuclear charge-exchange excitations with localized covariant density functional theory is briefly presented, by taking the fine structure of spin-dipole excitations in 16O as an example. It is shown that the constraints introduced by the Fock terms of the relativistic Hartree-Fock scheme into the particle-hole residual interactions are straightforward and robust. (authors)

  13. Nuclear charge-exchange excitations in localized covariant density functional theory

    CERN Document Server

    Liang, H Z; Nakatsukasa, T; Niu, Z M; Ring, P; Roca-Maza, X; Van Giai, N; Zhao, P W

    2014-01-01

    The recent progress in the studies of nuclear charge-exchange excitations with localized covariant density functional theory is briefly presented, by taking the fine structure of spin-dipole excitations in 16O as an example. It is shown that the constraints introduced by the Fock terms of the relativistic Hartree-Fock scheme into the particle-hole residual interactions are straightforward and robust.

  14. Simulating the dynamics of the neutron flux in a nuclear reactor by locally recurrent neural networks

    International Nuclear Information System (INIS)

    In this paper, a locally recurrent neural network (LRNN) is employed for approximating the temporal evolution of a nonlinear dynamic system model of a simplified nuclear reactor. To this aim, an infinite impulse response multi-layer perceptron (IIR-MLP) is trained according to a recursive back-propagation (RBP) algorithm. The network nodes contain internal feedback paths and their connections are realized by means of IIR synaptic filters, which provide the LRNN with the necessary system state memory

  15. Method and device for calibration of measurement contrivances for local power distribution in nuclear reactors

    International Nuclear Information System (INIS)

    For calibration purposes a heating cable is installed inside a bundle of cables leading to thermocouples used together with a body that is heated by gamma irradiation (gamma thermometer), for measurement of local power distribution in a nuclear reactor. The signals from the thermocouples are measured during the calibration period for analysis of the signal reaction that is caused by induced heat into the body, sudden stop of electrical heating or by supply of sinus shaped heating current to the heating cable

  16. 'Nuclear emergency preparedness' for local residents. Support of on-site training of many kinds of places and people

    International Nuclear Information System (INIS)

    In order to support and ensure the nuclear emergency preparedness system and safety of residents in cities, towns and villages, NPO Nuclear Emergency Preparedness Support Center was established in May, 2003. 130 on-site training and education classes were held and above 2,000 participants attended to them for two years. Objects of the countermeasure of nuclear emergency preparedness in local area and residents, what is nuclear emergency for inhabitants, what is use of Table of International Nuclear Event Scale (INES)?, a use of INES, relation between INES level and the nuclear emergency preparedness system are discussed. (S.Y.)

  17. Process-based modeling of the control of beet armyworm, Spodoptera exigua, with baculoviruses in greenhouse chrysanthemum

    NARCIS (Netherlands)

    Bianchi, F.J.J.A.

    2001-01-01

    This thesis describes the development of a process-based simulation model for the population dynamics of beet armyworm, Spodoptera exigua , and baculoviruses in greenhouse chrysanthemum. The model (BACSIM) has been validated for two baculoviruses with clear differences in biological characteristics,

  18. Local fracture properties and dissimilar weld integrity in nuclear power plants

    Science.gov (United States)

    Wang, Guozhen; Wang, Haitao; Xuan, Fuzhen; Tu, Shantung; Liu, Changjun

    2013-09-01

    In this paper, the local fracture properties in a Alloy52M dissimilar metal welded joint (DMWJ) between A508 ferritic steel and 316 L stainless steel in nuclear power plants were investigated by using the single-edge notched bend (SENB) specimens, and their use in integrity assessment of DMWJ structures was analyzed. The results show that the local fracture resistance in the DMWJ is determined by local fracture mechanism, and which is mainly related to the microstructures and local strength mismatches of materials at the crack locations. The initial cracks always grow towards the materials with lower strength, and the crack path deviation is mainly controlled by the local strength mismatch. If the local fracture properties could not be used for cracks in the heat affected zones (HAZs), interface and near interface zones, the use of the fracture properties ( J-resistance curves) of base metals or weld metals following present codes will unavoidably produce non-conservative (unsafe) or excessive conservative assessment results. In most cases, the assessment results will be potentially unsafe. Therefore, it is recommended to obtain and use local mechanical and fracture properties in the integrity assessment of DMWJs.

  19. Nuclear matter properties from local chiral interactions with $\\Delta$ isobar intermediate states

    CERN Document Server

    Logoteta, Domenico; Kievsky, Alejandro

    2016-01-01

    Using two-nucleon and three-nucleon interactions derived in the framework of chiral perturbation theory (ChPT) with and without the explicit $\\Delta$ isobar contributions, we calculate the energy per particle of symmetric nuclear matter and pure neutron matter in the framework of the microscopic Brueckner-Hartree-Fock approach. In particular, we present for the first time nuclear matter calculations using the new fully local in coordinate-space two-nucleon interaction at the next-to-next-to-next-to-leading-order (N3LO) of ChPT with $\\Delta$ isobar intermediate states (N3LO$\\Delta$) recently developed by Piarulli et al. [arXiv:1606:06335]. We find that using this N3LO$\\Delta$ potential, supplemented with a local N2LO three-nucleon interaction with explicit $\\Delta$ isobar degrees of freedom, it is possible to obtain a satisfactory saturation point of symmetric nuclear matter. For this combination of two- and three-nucleon interactions we also calculate the nuclear symmetry energy and we compare our results wit...

  20. Transcription-dependent nuclear localization of DAZAP1 requires an N-terminal signal

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Yi-Tzu; Wen, Wan-Ching [Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan (China); Yen, Pauline H., E-mail: pyen@ibms.sinica.edu.tw [Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan (China)

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer DAZAP1 shuttles between the nucleus and the cytoplasm. Black-Right-Pointing-Pointer DAZAP1 accumulates in the cytoplasm when the nuclear transcription is inhibited. Black-Right-Pointing-Pointer DAZAP1's transcription-dependent nuclear localization requires N-terminal N42. Black-Right-Pointing-Pointer SLIRP binds to N42 and may be involved in the process. -- Abstract: Deleted in Azoospermia Associated Protein 1 (DAZAP1) is a ubiquitous hnRNP protein required for normal development and spermatogenesis. It resides predominantly in the nucleus and moves between the nucleus and the cytoplasm via a ZNS shuttling signal at its C-terminus. DAZAP1 accumulates in the cytoplasm when RNA polymerase II activity is inhibited by actinomycin D. Here we report the mapping of a 42-amino acid segment (N42) at the N-terminus of DAZAP1 that is both necessary and sufficient for its transcription-dependent nuclear localization. In addition, using a yeast two-hybrid system, we have identified SLIRP as a N42-binding protein which may regulate DAZAP1 subcellular localization.

  1. TRX-1 Regulates SKN-1 Nuclear Localization Cell Non-autonomously in Caenorhabditis elegans.

    Science.gov (United States)

    McCallum, Katie C; Liu, Bin; Fierro-González, Juan Carlos; Swoboda, Peter; Arur, Swathi; Miranda-Vizuete, Antonio; Garsin, Danielle A

    2016-05-01

    The Caenorhabditis elegans oxidative stress response transcription factor, SKN-1, is essential for the maintenance of redox homeostasis and is a functional ortholog of the Nrf family of transcription factors. The numerous levels of regulation that govern these transcription factors underscore their importance. Here, we add a thioredoxin, encoded by trx-1, to the expansive list of SKN-1 regulators. We report that loss of trx-1 promotes nuclear localization of intestinal SKN-1 in a redox-independent, cell non-autonomous fashion from the ASJ neurons. Furthermore, this regulation is not general to the thioredoxin family, as two other C. elegans thioredoxins, TRX-2 and TRX-3, do not play a role in this process. Moreover, TRX-1-dependent regulation requires signaling from the p38 MAPK-signaling pathway. However, while TRX-1 regulates SKN-1 nuclear localization, classical SKN-1 transcriptional activity associated with stress response remains largely unaffected. Interestingly, RNA-Seq analysis revealed that loss of trx-1 elicits a general, organism-wide down-regulation of several classes of genes; those encoding for collagens and lipid transport being most prevalent. Together, these results uncover a novel role for a thioredoxin in regulating intestinal SKN-1 nuclear localization in a cell non-autonomous manner, thereby contributing to the understanding of the processes involved in maintaining redox homeostasis throughout an organism. PMID:26920757

  2. Nuclear Localization of p38 MAPK in Response to DNA Damage

    Directory of Open Access Journals (Sweden)

    C. David Wood, Tina M. Thornton, Guadalupe Sabio, Roger A. Davis, Mercedes Rincon

    2009-01-01

    Full Text Available p38 MAP kinase (MAPK is activated in response to environmental stress, cytokines and DNA damage, and mediates death, cell differentiation and cell cycle checkpoints. The intracellular localization of p38 MAPK upon activation remains unclear, and may depend on the stimulus. We show here that activation of p38 MAPK by stimuli that induce DNA double strand breaks (DSBs, but not other stimuli, leads to its nuclear translocation. In addition, naturally occurring DSBs generated through V(DJ recombination in immature thymocytes also promote nuclear accumulation of p38 MAPK. Nuclear translocation of p38 MAPK does not require its catalytic activity, but is induced by a conformational change of p38 MAPK triggered by phosphorylation within the active site. The selective nuclear accumulation of p38 MAPK in response to DNA damage could be a mechanism to facilitate the phosphorylation of p38 MAPK nuclear targets for the induction of a G2/M cell cycle checkpoint and DNA repair.

  3. Relation between the local scale transformation approach and the nuclear energy density formalism

    International Nuclear Information System (INIS)

    An expression for the energy density functional is obtaiped. Local scale transformation method iS applied to the one-particle Dirac density matrix of a particular form. A nuclear system is described by the Hamiltonian with Skyrme forces. An additional term, widely known as Weizsecker's kinetic energy correction, appears as well as the term that is transformed into the Thomas-Fermi kinetic energy expression for slowly varying local density. The equation for the local density distribution is resolved by applying the core-spline method. Numerical calculations of the density distributions and mean-square radius rms of charge distributions are carried out for 12C, 16O, 32S, 40Ca, 48Ca, 66Zn, 140Ce, and 208Pb. It is seen from the results that the local scale transformation method can be successfully used for finding the explicit form of the energy density functional. The constructed functional, although derived from a nonrealistic model density matrix proves to be quite reasonable for the reproduction of nuclear characteristics

  4. Localization of latency-associated nuclear antigen (LANA) on mitotic chromosomes.

    Science.gov (United States)

    Rahayu, Retno; Ohsaki, Eriko; Omori, Hiroko; Ueda, Keiji

    2016-09-01

    In latent infection of Kaposi's sarcoma-associated herpesvirus (KSHV), viral gene expression is extremely limited and copy numbers of viral genomes remain constant. Latency-associated nuclear antigen (LANA) is known to have a role in maintaining viral genome copy numbers in growing cells. Several studies have shown that LANA is localized in particular regions on mitotic chromosomes, such as centromeres/pericentromeres. We independently examined the distinct localization of LANA on mitotic chromosomes during mitosis, using super-resolution laser confocal microscopy and correlative fluorescence microscopy-electron microscopy (FM-EM) analyses. We found that the majority of LANA were not localized at particular regions such as telomeres/peritelomeres, centromeres/pericentromeres, and cohesion sites, but at the bodies of condensed chromosomes. Thus, LANA may undergo various interactions with the host factors on the condensed chromosomes in order to tether the viral genome to mitotic chromosomes and realize faithful viral genome segregation during cell division. PMID:27254595

  5. The Drosophila nucleoporin DNup88 localizes DNup214 and CRM1 on the nuclear envelope and attenuates NES-mediated nuclear export

    OpenAIRE

    Roth, Peggy; Xylourgidis, Nikos; Sabri, Nafiseh; Uv, Anne; Fornerod, Maarten; Samakovlis, Christos

    2003-01-01

    Many cellular responses rely on the control of nucleocytoplasmic transport of transcriptional regulators. The Drosophila nucleoporin Nup88 is selectively required for nuclear accumulation of Rel proteins and full activation of the innate immune response. Here, we investigate the mechanisms underlying its role in nucleocytoplasmic transport. Nuclear import of an nuclear localization signal-enhanced green fluorescent protein (NLS-EGFP) reporter is not affected in DNup88 (members only; mbo) muta...

  6. Radiation doses to local populations near nuclear weapons test sites worldwide.

    Science.gov (United States)

    Simon, Steven L; Bouville, André

    2002-05-01

    Nuclear weapons testing was conducted in the atmosphere at numerous sites worldwide between 1946 and 1980, which resulted in exposures to local populations as a consequence of fallout of radioactive debris. The nuclear tests were conducted by five nations (United States, Soviet Union, United Kingdom, France, and China) primarily at 16 sites. The 16 testing sites, located in nine different countries on five continents (plus Oceania) contributed nearly all of the radioactive materials released to the environment by atmospheric testing; only small amounts were released at a fewother minor testing sites. The 16 sites discussed here are Nevada Test Site, USA (North American continent), Bikini and Enewetak, Marshall Islands (Oceania); Johnston Island, USA (Oceania), Christmas and Malden Island, Kiribati (Oceania); Emu Field, Maralinga, and Monte Bello Islands, Australia (Australian continent); Mururoa and Fangataufa, French Polynesia (Oceania), Reggane, Algeria (Africa), Novaya Zemlya and Kapustin Yar, Russia (Europe), Semipalatinsk, Kazakhstan (Asia), and Lop Nor, China (Asia). There were large differences in the numbers of tests conducted at each location and in the total explosive yields. Those factors, as well as differences in population density, lifestyle, environment, and climate at each site, led to large differences in the doses received by local populations. In general, the tests conducted earliest led to the highest individual and population exposures, although the amount of information available for a few of these sites is insufficient to provide any detailed evaluation of radiation exposures. The most comprehensive information for any site is for the Nevada Test Site. The disparities in available information add difficulty to determining the radiation exposures of local populations at each site. It is the goal of this paper to summarize the available information on external and internal doses received by the public living in the regions near each of the

  7. Radiation doses to local populations near nuclear weapons test sites worldwide

    International Nuclear Information System (INIS)

    Nuclear weapons testing was conducted in the atmosphere at numerous sites worldwide between 1946 and 1980, which resulted in exposures to local populations as a consequence of fallout of radioactive debris. The nuclear tests were conducted by five nations (United States, Soviet Union, United Kingdom, France, and China) primarily at 16 sites. The 16 testing sites, located in nine different countries on five continents (plus Oceania) contributed nearly all of the radioactive materials released to the environment by atmospheric testing; only small amounts were released at a few other minor testing sites. The 16 sites discussed here are Nevada Test Site, USA (North American continent), Bikini and Enewetak, Marshall Islands (Oceania); Johnston Island, USA (Oceania), Christmas and Malden Island, Kiribati (Oceania); Emu Field, Maralinga, and Monte Bello Islands, Australia (Australian continent); Mururoa and Fangataufa, French Polynesia (Oceania), Reggane, Algeria (Africa), Novaya Zemlya and Kapustin Yar, Russia (Europe), Semipalatinsk, Kazakhstan (Asia), and Lop Nor, China (Asia). There were large differences in the numbers of tests conducted at each location and in the total explosive yields. Those factors, as well as differences in population density, lifestyle, environment, and climate at each site, led to large differences in the doses received by local populations. In general, the tests conducted earliest led to the highest individual and population exposures, although the amount of information available for a few of these sites is insufficient to provide any detailed evaluation of radiation exposures. The most comprehensive information for any site is for the Nevada Test Site. The disparities in available information add difficulty to determining the radiation exposures of local populations at each site. It is the goal of this paper to summarize the available information on external and internal doses received by the public living in the regions near each of the

  8. Generating a host range-expanded recombinant baculovirus.

    Science.gov (United States)

    Wu, Chunfeng; Deng, Zihao; Long, Zhao; Cai, Yi; Ying, Zhongfu; Yin, Hanqi; Yuan, Meijin; Clem, Rollie J; Yang, Kai; Pang, Yi

    2016-01-01

    As baculoviruses usually have a narrow insecticidal spectrum, knowing the mechanisms by which they control the host-range is prerequisite for improvement of their applications as pesticides. In this study, from supernatant of culture cells transfected with DNAs of an Autographa californica multiple nucleopolyhedrovirus (AcMNPV) mutant lacking the antiapoptotic gene p35 (vAc(∆P35)) and a cosmid representing a fragment of Spodoptera exigua nucleopolyhedrovirus (SeMNPV), a viral strain was plaque-purified and named vAcRev. vAcRev had a broader host range than either vAc(∆P35) or SeMNPV parental virus, being able to infect not only the permissive hosts of its parental viruses but also a nonpermissive host (Spodoptera litura). Genome sequencing indicated that vAcRev comprises a mixture of two viruses with different circular dsDNA genomes. One virus contains a genome similar to vAc(∆P35), while in the other viral genome, a 24.4 kbp-fragment containing 10 essential genesis replaced with a 4 kbp-fragment containing three SeMNPV genes including a truncated Se-iap3 gene. RNA interference and ectopic expression assays found that Se-iap3 is responsible for the host range expansion of vAcRev, suggesting that Se-iap3 inhibits the progression of apoptosis initiated by viral infection and promotes viral propagation in hosts both permissive and non-permissive for AcMNPV and SeMNPV. PMID:27321273

  9. Identification and characterization of multiple conserved nuclear localization signals within adenovirus E1A

    Energy Technology Data Exchange (ETDEWEB)

    Marshall, Kris S.; Cohen, Michael J.; Fonseca, Greg J.; Todorovic, Biljana; King, Cason R. [Department of Microbiology and Immunology, Western University, London Regional Cancer Program, London, ON, Canada N6A 4L6 (Canada); Yousef, Ahmed F. [Department of Chemical and Environmental Engineering, Masdar Institute, Abu Dhabi (United Arab Emirates); Zhang, Zhiying [College of Animal Science and Technologies, Northwest A and F University, Yangling, Shaanxi 712100 (China); Mymryk, Joe S., E-mail: jmymryk@uwo.ca [Department of Microbiology and Immunology, Western University, London Regional Cancer Program, London, ON, Canada N6A 4L6 (Canada); Department of Oncology, Western University, London Regional Cancer Program, London, ON, Canada N6A 4L6 (Canada)

    2014-04-15

    The human adenovirus 5 (HAdV-5) E1A protein has a well defined canonical nuclear localization signal (NLS) located at its C-terminus. We used a genetic assay in the yeast Saccharomyces cerevisiae to demonstrate that the canonical NLS is present and functional in the E1A proteins of each of the six HAdV species. This assay also detects a previously described non-canonical NLS within conserved region 3 and a novel active NLS within the N-terminal/conserved region 1 portion of HAdV-5 E1A. These activities were also present in the E1A proteins of each of the other five HAdV species. These results demonstrate that, despite substantial differences in primary sequence, HAdV E1A proteins are remarkably consistent in that they contain one canonical and two non-canonical NLSs. By utilizing independent mechanisms, these multiple NLSs ensure nuclear localization of E1A in the infected cell. - Highlights: • HAdV E1A uses multiple mechanisms for nuclear import. • We identified an additional non-canonical NLS in the N-terminal/CR1 portion of E1A. • The new NLS does not contact importin-alpha directly. • All NLSs are functionally conserved in the E1A proteins of all 6 HAdV species.

  10. A functional nuclear localization sequence in the C. elegans TRPV channel OCR-2.

    Directory of Open Access Journals (Sweden)

    Meredith J Ezak

    Full Text Available The ability to modulate gene expression in response to sensory experience is critical to the normal development and function of the nervous system. Calcium is a key activator of the signal transduction cascades that mediate the process of translating a cellular stimulus into transcriptional changes. With the recent discovery that the mammalian Ca(v1.2 calcium channel can be cleaved, enter the nucleus and act as a transcription factor to control neuronal gene expression, a more direct role for the calcium channels themselves in regulating transcription has begun to be appreciated. Here we report the identification of a nuclear localization sequence (NLS in the C. elegans transient receptor potential vanilloid (TRPV cation channel OCR-2. TRPV channels have previously been implicated in transcriptional regulation of neuronal genes in the nematode, although the precise mechanism remains unclear. We show that the NLS in OCR-2 is functional, being able to direct nuclear accumulation of a synthetic cargo protein as well as the carboxy-terminal cytosolic tail of OCR-2 where it is endogenously found. Furthermore, we discovered that a carboxy-terminal portion of the full-length channel can localize to the nucleus of neuronal cells. These results suggest that the OCR-2 TRPV cation channel may have a direct nuclear function in neuronal cells that was not previously appreciated.

  11. The BRCA1-binding protein BRAP2 can act as a cytoplasmic retention factor for nuclear and nuclear envelope-localizing testicular proteins.

    Science.gov (United States)

    Davies, Rebecca G; Wagstaff, Kylie M; McLaughlin, Eileen A; Loveland, Kate L; Jans, David A

    2013-12-01

    Regulation of nuclear protein import is central to many cellular processes such as development, with a key mechanism being factors that retain cargoes in the cytoplasm that normally localize in the nucleus. The breast cancer antigen BRCA1-binding protein BRAP2 has been reported as a novel negative regulator of nuclear import of various nuclear localization signal (NLS)-containing viral and cellular proteins, but although implicated in differentiation pathways and highly expressed in tissues including testis, the gamut of targets for BRAP2 action in a developmental context is unknown. As a first step towards defining the BRAP2 interactome, we performed a yeast-2-hybrid screen to identify binding partners of BRAP2 in human testis. Here we report characterization for the first time of three of these: the high mobility group (HMG)-box-domain-containing chromatin component HMG20A, nuclear mitotic apparatus protein NuMA1 and synaptic nuclear envelope protein SYNE2. Co-immunoprecipitation experiments indicate association of BRAP2 with HMG20A, NuMA1, and SYNE2 in testis, underlining the physiological relevance of the interactions, with immunohistochemistry showing that where BRAP2 is co-expressed with HMG20A and NuMA1, both are present in the cytoplasm, in contrast to their nuclear localization in other testicular cell types. Importantly, quantitative confocal microscopic analysis of cultured cells indicates that ectopic expression of BRAP2 inhibits nuclear localization of HMG20A and NuMA1, and prevents nuclear envelope accumulation of SYNE2, the first report of BRAP2 altering localization of a non-nuclear protein. These results imply for the first time that BRAP2 may have an important role in modulating subcellular localization during testicular development. PMID:23707952

  12. Identification of a nuclear localization sequence in the polyomavirus capsid protein VP2

    Science.gov (United States)

    Chang, D.; Haynes, J. I. 2nd; Brady, J. N.; Consigli, R. A.; Spooner, B. S. (Principal Investigator)

    1992-01-01

    A nuclear localization signal (NLS) has been identified in the C-terminal (Glu307-Glu-Asp-Gly-Pro-Gln-Lys-Lys-Lys-Arg-Arg-Leu318) amino acid sequence of the polyomavirus minor capsid protein VP2. The importance of this amino acid sequence for nuclear transport of newly synthesized VP2 was demonstrated by a genetic "subtractive" study using the constructs pSG5VP2 (expressing full-length VP2) and pSG5 delta 3VP2 (expressing truncated VP2, lacking amino acids Glu307-Leu318). These constructs were transfected into COS-7 cells, and the intracellular localization of the VP2 protein was determined by indirect immunofluorescence. These studies revealed that the full-length VP2 was localized in the nucleus, while the truncated VP2 protein was localized in the cytoplasm and not transported to the nucleus. A biochemical "additive" approach was also used to determine whether this sequence could target nonnuclear proteins to the nucleus. A synthetic peptide identical to VP2 amino acids Glu307-Leu318 was cross-linked to the nonnuclear proteins bovine serum albumin (BSA) or immunoglobulin G (IgG). The conjugates were then labeled with fluorescein isothiocyanate and microinjected into the cytoplasm of NIH 3T6 cells. Both conjugates localized in the nucleus of the microinjected cells, whereas unconjugated BSA and IgG remained in the cytoplasm. Taken together, these genetic subtractive and biochemical additive approaches have identified the C-terminal sequence of polyoma-virus VP2 (containing amino acids Glu307-Leu318) as the NLS of this protein.

  13. Characterization of Local Wind Patterns around the Kori Nuclear Power Plant using Cluster Analysis and WRF meteorological modeling

    International Nuclear Information System (INIS)

    To accurately predict the atmospheric diffusion of radioactive effluent, detailed analysis of local wind patterns nearby nuclear power plants are necessary. In this study, the characteristics of typical local winds around the Kori Nuclear Power Plant (Kori NPP) were investigated using the cluster analysis and Weather Research and Forecasting (WRF) meteorological modeling. In this study, the local wind characteristics around the Kori NPP were analyzed using cluster analysis and WRF meteorological modeling. As a result of the cluster analysis, four wind patterns around the Kori NPP were selected. The model study indicated the possibility that the local winds in the target area can largely contribute to the atmospheric diffusion of radioactive effluents

  14. High sensitivity localization of parathyroid tumors by nuclear imaging in 50 proven cases

    International Nuclear Information System (INIS)

    Fifty patients with surgically and biochemically proven hyperparathyroidism had imaging studies of the neck performed with Thallium-201 chloride (2-3mCi) and Tc-99m pertechnetate, a gamma camera and pinhole collimator, and acquired and processed with a DEC PDP-11 computer with color coding for differences in localization of Tl-201 (in thyroid and parathyroid tumor) compared with pertechnetate (in the thyroid alone). Thirty-eight of 43 single parathyroid adenomas (88%) were successfully localized, 6 of 9 multiple adenomas in 3 such patients, and 11 of 12 hyperplastic glands in 3 patients with secondary hyperparathyroidism, and 1 of 1 with cancer. With regard to tumor size, 92% of tumors 1.0 cm. or larger were found, but only 50% of those smaller than 0.7 cm. One substernal adenoma 0.6 cm. in size was missed, but 5 of 6 adenomas previously missed at surgery were found successfully. No normal or functionally suppressed glands were identified, and no false positive studies were seen, even in patients with abnormal thyroid glands. Nuclear imaging studies with computerized comparison are the most efficacious single method for preoperative localization of parathyroid tumors yet reported, and distinguish single versus multiple disease with high reliability. All 3 intrathyroid parathyroid adenomas were identified as to site with this method. Advantages of preoperative localization of parathyroid tumors include planning of surgical approach, shortening of operations, and localization of thyroidotomy for intrathyroid tumors

  15. Decision making on nuclear waste in the local community. Three northern communities

    International Nuclear Information System (INIS)

    The main objective of the project is to examine how citizens, politicians, and other actors in the three Swedish municipalities of Storuman, Malaa and Oeverkalix reacted politically on the idea that the Swedish Nuclear Fuel and Waste Management Co was to perform a pre-study in the municipality with the purpose of producing a safer basis for decision so as to later on be able to decide if the company should proceed with examinations and possibly build a repository for spent nuclear fuel there. As a second objective we would like to examine the spent nuclear fuel issue as a public policy problem. This is done by applying the theory of collective goods on the disposal issue. The third objective is to develop the democratic and scientific dimensions of the nuclear waste problem. Should the issue be determined at the national, county or local level? Should the question about a geographic and geologic place for the disposal be determined by research, and how should the balance between research and democracy be effected?

  16. Intense deuterium nuclear fusion of pycnodeuterium-lumps coagulated locally within highly deuterated atom clusters

    International Nuclear Information System (INIS)

    Embedded nano-Pd particles of 5 nm in size instantly abundant D-atoms more than 250% in the atomic ratio against Pd-atoms at room temperature when they are kept in D2 gas pressurized to less than 10 atm. In such ultrahigh densities, 2-4 D-atoms can be coagulated inside each octahedral space of Pd lattice (pycnodeuterium-lump). When a stimulation energy such as latticequake causing by ultrasonic wave was supplied to those highly deuterated Pd particles, intense deuterium nuclear fusion (''solid fusion'') was generated there and both excess heat and 4He gas were abundantly produced. Naturally, these facts can not be realized at all in bulk Pd. The results show that the nuclear fusion occurs without any hazardous rays in pycnodeuterium-lumps coagulated locally inside the each cell of the host metal lattice. These unit cells correspond to minimum unit of the solid fusion reactor as a ''Lattice Reactor''. (author)

  17. Intense deuterium nuclear fusion of pycnodeuterium-lumps coagulated locally within highly deuterated atom clusters

    CERN Document Server

    Yoshiaki, A; Zhang, Y C

    2002-01-01

    Embedded nano-Pd particles of 5 nm in size instantly abundant D-atoms more than 250% in the atomic ratio against Pd-atoms at room temperature when they are kept in D sub 2 gas pressurized to less than 10 atm. In such ultrahigh densities, 2-4 D-atoms can be coagulated inside each octahedral space of Pd lattice (pycnodeuterium-lump). When a stimulation energy such as latticequake causing by ultrasonic wave was supplied to those highly deuterated Pd particles, intense deuterium nuclear fusion (''solid fusion'') was generated there and both excess heat and sup 4 He gas were abundantly produced. Naturally, these facts can not be realized at all in bulk Pd. The results show that the nuclear fusion occurs without any hazardous rays in pycnodeuterium-lumps coagulated locally inside the each cell of the host metal lattice. These unit cells correspond to minimum unit of the solid fusion reactor as a ''Lattice Reactor''. (author)

  18. Stellar and Dust Properties of Local Elliptical Galaxies: Clues to the Onset of Nuclear Activity

    CERN Document Server

    Zhang, Yu; Ho, Luis C

    2008-01-01

    We study the stellar and dust properties of a well-defined sample of local elliptical galaxies to investigate the relationship between host galaxy properties and nuclear activity. We select a complete sample of 45 ellipticals from the Palomar spectroscopic survey of nearby galaxies, which includes 20 low-luminosity active galactic nuclei classified as LINERs and 25 inactive galaxies. Using a stellar population synthesis method, we compare the derived stellar population properties of the LINER versus the inactive subsamples. We also study the dust and stellar surface brightness distributions of the central regions of these galaxies using high-resolution images obtained with the {\\it Hubble Space Telescope}. Relative to the inactive subsample, ellipticals hosting LINERs share similar total optical and near-infrared luminosity, central stellar velocity dispersions, and nuclear stellar populations as judged from their luminosity-weighted ages and metallicities. LINERs, on the other hand, have a larger fraction of...

  19. Review study 1995. Localization of the repository for spent nuclear fuel

    International Nuclear Information System (INIS)

    This report gives an overview of the studies performed by SKB pertinent to selection of a site for the Swedish repository for spent nuclear fuels, and is written for both experts in the various fields involved, decision-makers and the interested general public. The review can not comprise all detailed factors necessary for deciding the localization, but deals mainly with conditions on the land surface and can point out areas which are not well suited or less interesting as a site. It also treats several scientific, technical and social bases in different parts of the country. 120 refs, 53 figs

  20. Distinct effects of nuclear membrane localization on gene transcription silencing in Drosophila S2 cells and germ cells

    Institute of Scientific and Technical Information of China (English)

    Lu Sui; Yanhong Yang

    2011-01-01

    Nuclear envelope proteins have important roles in chromatin organization and signal-dependent transcriptional regulation. A previous study reported that the inner nuclear membrane protein, Otefin (Ote), was essential for germline stem cell (GSC) maintenance via interaction with Smad complex. The interaction of Otc with the Smad complex recruits the bam locus to the nuclear periphery and subsequently results in bam transcriptional silencing, revealing that nuclear peripheral localization is essential for bam gene regulation. However, it remains unknown whether the nuclear peripheral localization is sufficient for bam silencing. To address this issue, we have established a tethering system, in which the Gal4 DNA binding domain (DBD) of the Flag:Gal4 DBD:Ote △ LEM fusion protein physically interacts with the Gal4 binding sites upstream of bamP-gfp to artificially recruit the reporter gene gfp to the nuclear membrane. Our data demonstrated that the nuclear peripheral localization seemed to affect the expression of the target naked gene in S2 cells. By contrast, in Drosophila germ cells, the nuclear membrane localization was not sufficient for gene silencing.

  1. Baculovirus as a PRRSV and PCV2 bivalent vaccine vector: baculovirus virions displaying simultaneously GP5 glycoprotein of PRRSV and capsid protein of PCV2.

    Science.gov (United States)

    Xu, Xin-Gang; Wang, Zhi-Sheng; Zhang, Qi; Li, Zhao-Cai; Ding, Li; Li, Wei; Wu, Hung-Yi; Chang, Ching-Dong; Lee, Long-Huw; Tong, De-Wen; Liu, Hung-Jen

    2012-02-01

    The GP5 glycoprotein of PRRSV is the main target for inducing neutralizing antibodies and protective immunity in the natural host. The capsid (Cap) protein is the major immunogenic protein and associated with the production of PCV2-specific neutralizing antibodies. In the present study, one genetic recombinant baculovirus BacSC-Dual-GP5-Cap was constructed. This virus displays simultaneously histidine-tagged GP5 and Cap proteins with the baculovirus glycoprotein gp64 TM and CTD on the virion surface as well as the surface of the virus-infected cells. After infection, the GP5 and Cap proteins were expressed and anchored simultaneously on the plasma membrane of Sf-9 cells, as revealed by Western blot and confocal microscopy. This report demonstrated first that both GP5 and Cap proteins were displayed successfully on the viral surface, revealed by immunogold electron microscopy. Vaccination of swine with recombinant baculovirus BacSC-Dual-GP5-Cap elicited significantly higher GP5 and Cap ELISA antibody titers in swine than the control groups. Virus neutralization test also showed that serum from the BacSC-Dual-GP5-Cap treated swine had significant levels of virus neutralization titers. Lymphocyte proliferation responses could be induced in swine immunized with BacSC-Dual-GP5-Cap than the control groups. These findings demonstrate that the BacSC-Dual-GP5-Cap bivalent subunit vaccine can be a potential vaccine against PRRSV and PCV2 infections. PMID:22172969

  2. The effect of intermittent operation on local fission rate in the McMaster Nuclear Reactor

    International Nuclear Information System (INIS)

    The McMaster Nuclear Reactor operates on a 16-hr/day/5-day/week schedule causing cyclic loading of Xenon in the core and requiring compensation by the control systems to maintain operations. The constant control rod interaction affects local fission rates. This paper confirms the relationship between Xenon load and control rod movement and studies the relationship between local fission rate and control rod insertion. The results provide information related to analysis approximations used in depletion calculations. In addition, comparisons are made between the current MNR operation cycle and a proposed continuous operational approach. The results are further discussed in the context of proposed Molybdenum-99 production at MNR. (author)

  3. Baculovirus virions displaying Plasmodium berghei circumsporozoite protein protect mice against malaria sporozoite infection

    International Nuclear Information System (INIS)

    The display of foreign proteins on the surface of baculovirus virions has provided a tool for the analysis of protein-protein interactions and for cell-specific targeting in gene transfer applications. To evaluate the baculovirus display system as a vaccine vehicle, we have generated a recombinant baculovirus (AcNPV-CSPsurf) that displays rodent malaria Plasmodium berghei circumsporozoite protein (PbCSP) on the virion surface as a fusion protein with the major baculovirus envelope glycoprotein gp64. The PbCSP-gp64 fusion protein was incorporated and oligomerized on the virion surface and led to a 12-fold increase in the binding activity of AcNPV-CSPsurf virions to HepG2 cells. Immunization with adjuvant-free AcNPV-CSPsurf virions induced high levels of antibodies and gamma interferon-secreting cells against PbCSP and protected 60% of mice against sporozoite challenge. These data demonstrate that AcNPV-CSPsurf displays sporozoite-like PbCSP on the virion surface and possesses dual potentials as a malaria vaccine candidate and a liver-directed gene delivery vehicle

  4. Over-expression and characterization of active recombinant rat liver carnitine palmitoyltransferase II using baculovirus.

    Science.gov (United States)

    Johnson, T M; Mann, W R; Dragland, C J; Anderson, R C; Nemecek, G M; Bell, P A

    1995-01-01

    The cDNA encoding rat liver carnitine palmitoyltransferase II (CPT-II) was heterologously expressed using a recombinant baculovirus/insect cell system. Unlike Escherichia coli, the baculovirus-infected insect cells expressed mostly soluble active recombinant CPT-II (rCPT-II). CPT activity from crude lysates of recombinant baculovirus-infected insect cells was maximal between 50 and 72 h post-infection, with a peak specific activity of 100-200 times that found in the mock- or wild-type-infected control lysates. Milligram quantities (up to 1.8 mg/l of culture) of active rCPT-II were chromatographically purified from large-scale cultures of insect cells infected with the recombinant baculovirus. The rCPT-II was found to be: (1) similar in size to the native rat liver enzyme (approximately 70 kDa) as judged by SDS/PAGE; (2) immunoreactive with a polyclonal serum raised against rat liver CPT-II; and (3) not glycosylated. Kinetic analysis of soluble rCPT-II revealed Km values for carnitine and palmitoyl-CoA of 950 +/- 27 microM and 34 +/- 5.6 microM respectively. Images Figure 1 Figure 2 Figure 4 PMID:7626037

  5. CHARACTERIZATION OF THE DNA OF A NONOCCLUDED BACULOVIRUS, HZ-1V

    Science.gov (United States)

    The DNA of the nonoccluded baculovirus (Hz-1V) obtained from the IMC-Hz-1 cell line was characterized by physicochemical and restriction endonuclease techniques. Hz-1V DNA isolated from purified virus had buoyant densities of 1.58 and 1.54 g/ml in CsC1-ethidium bromide density gr...

  6. Iron levels change in larval Heliothis virescens tissues following baculovirus infection

    Science.gov (United States)

    Inductively-coupled plasma mass spectrometry (ICP-MS) and 59Fe radiotracers were used to investigate changes in levels of iron (Fe) in the tissues of Heliothis virescens following baculovirus infection. Fe concentrations were determined by ICP-MS in hemolymph collected from 4th instar larvae infect...

  7. Changes in trace metals in hemolymph of baculovirus infected noctuid larvae

    Science.gov (United States)

    We studied how biologically relevant trace metals (i.e., micronutrients) in the plasma of larvae of Heliothis virescens and Helicoverpa zea (Lepidoptera: Noctuidae) changed in response to per os baculovirus infection, larval development, and injection of heat-killed bacteria. Concentrations of plas...

  8. Method to characterize local meteorology at nuclear facilities for application to emergency response needs

    International Nuclear Information System (INIS)

    Effluent dispersion is evaluated using computer codes that require various meteorological parameters such as wind and stability data. These data will be based on current conditions at the site in question, and on forecasts of the expected local meteorology for the time period to be simulated. To assist NRC personnel in preparing these forecasts, a weather-typing model was implemented to analyze the characteristic behavior of local meteorology as it responds to various synoptic-scale weather features (e.g., warm fronts, cold fronts, high pressure systems). Historical observations acquired by instrumented towers at several nuclear power plants were analyzed as a function of the prevailing synoptic weather feature, synoptic-scale pressure gradient, and time of year. This study focused on sites located in shoreline and complex terrain environments because of the occurrence of mesoscale circulations, which are the sea/lake-land breeze and valley wind systems. Such circulations produce diurnally changing wind and stability conditions that cannot be readily identified by synoptic-scale weather forecasts. The advantage in analyzing the climatological behavior of local meteorology as it responds to various synoptic weather systems is that certain weather systems will control the local meteorology and produce persistent conditions

  9. Nuclear Fragile X Mental Retardation Protein is localized to Cajal bodies.

    Directory of Open Access Journals (Sweden)

    Alain Y Dury

    2013-10-01

    Full Text Available Fragile X syndrome is caused by loss of function of a single gene encoding the Fragile X Mental Retardation Protein (FMRP. This RNA-binding protein, widely expressed in mammalian tissues, is particularly abundant in neurons and is a component of messenger ribonucleoprotein (mRNP complexes present within the translational apparatus. The absence of FMRP in neurons is believed to cause translation dysregulation and defects in mRNA transport essential for local protein synthesis and for synaptic development and maturation. A prevalent model posits that FMRP is a nucleocytoplasmic shuttling protein that transports its mRNA targets from the nucleus to the translation machinery. However, it is not known which of the multiple FMRP isoforms, resulting from the numerous alternatively spliced FMR1 transcripts variants, would be involved in such a process. Using a new generation of anti-FMRP antibodies and recombinant expression, we show here that the most commonly expressed human FMRP isoforms (ISO1 and 7 do not localize to the nucleus. Instead, specific FMRP isoforms 6 and 12 (ISO6 and 12, containing a novel C-terminal domain, were the only isoforms that localized to the nuclei in cultured human cells. These isoforms localized to specific p80-coilin and SMN positive structures that were identified as Cajal bodies. The Cajal body localization signal was confined to a 17 amino acid stretch in the C-terminus of human ISO6 and is lacking in a mouse Iso6 variant. As FMRP is an RNA-binding protein, its presence in Cajal bodies suggests additional functions in nuclear post-transcriptional RNA metabolism. Supporting this hypothesis, a missense mutation (I304N, known to alter the KH2-mediated RNA binding properties of FMRP, abolishes the localization of human FMRP ISO6 to Cajal bodies. These findings open unexplored avenues in search for new insights into the pathophysiology of Fragile X Syndrome.

  10. Application of the counterfactual method to assess of the local economic impact of a nuclear power station

    International Nuclear Information System (INIS)

    Several studies have examined the local economic effects of nuclear power stations. However, their heterogeneity within countries and between countries indicate that one must be very cautious in drawing general conclusions about the positive and negative local economic effects of nuclear power stations over time. Using information about a specific nuclear facility in Spain and its zone of influence, we investigate the local economic impact of a nuclear facility by applying an infrequently used methodology called the “counterfactual” method. The purpose of this application of the counterfactual method is to establish what would have happened if the nuclear power plant had not been built where it was. This method permits observation of the impact of the nuclear power plant on the evolution of a set of variables whose influence is extremely important for the local area, such as the population, unemployment level, per capita income, and municipal governments’ revenues. Generalization of this method could offer the possibility of providing comparative results. However, the method must be complemented with other short-term approaches to provide a more specific analysis of the economic effects on the local actors involved and to offer incentives to policymakers to design and develop policies aimed at boosting economic activity in the area. - Highlights: • The Trillo Nuclear Power Plant plays a key role in stimulating its local environment. • The counterfactual method is estimated based on the performance of control group. • At least at the beginning of its operation the population dynamism was clear. • There is not strong evidence about the rise of employment and local public revenues. • The expenditure flow assesses the local turnover, jobs and municipal revenues

  11. 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.

  12. 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)

    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

  13. Local nuclear slope and curvature in high-energy pp and anti-pp elastic scattering

    International Nuclear Information System (INIS)

    The local nuclear slope B(s, t) = (d / dt) (ln (dσn (s, t) /dt)) is reconstructed from the experimental angular distributions with a procedure that uses overlapping t-bins, for an energy that ranges from the ISR to the S anti-pp S and the Tevraton. Predictions of several models of (p, p) and (anti-p, p) elastic scattering at high energy are tested in B(s, t) at small vertical stoke t vertical stroke. Only a model with two-components pomeron and odderon gives a satisfactory agreement with the (non-fitted) slope data, in particular for the evolution of B(s, t) with s as a function of t in anti-pp scattering. This modell predicts a similar behavior for pp and anti-pp scattering at small vertical stroke t vertical stroke. A detailed confirmation for pp collisions would be expected from RHIC. The extreme sensitivity of the local nuclear curvature C(s, t) = (1 / 2) (d / dt) (B (s, t)) with the choise for a pomeron model is emphasized. The present model predicts a change of sign. The ideal place to search for an eventual confirmation of this prediction would be LHC:

  14. Protein Sub-Nuclear Localization Based on Effective Fusion Representations and Dimension Reduction Algorithm LDA

    Directory of Open Access Journals (Sweden)

    Shunfang Wang

    2015-12-01

    Full Text Available An effective representation of a protein sequence plays a crucial role in protein sub-nuclear localization. The existing representations, such as dipeptide composition (DipC, pseudo-amino acid composition (PseAAC and position specific scoring matrix (PSSM, are insufficient to represent protein sequence due to their single perspectives. Thus, this paper proposes two fusion feature representations of DipPSSM and PseAAPSSM to integrate PSSM with DipC and PseAAC, respectively. When constructing each fusion representation, we introduce the balance factors to value the importance of its components. The optimal values of the balance factors are sought by genetic algorithm. Due to the high dimensionality of the proposed representations, linear discriminant analysis (LDA is used to find its important low dimensional structure, which is essential for classification and location prediction. The numerical experiments on two public datasets with KNN classifier and cross-validation tests showed that in terms of the common indexes of sensitivity, specificity, accuracy and MCC, the proposed fusing representations outperform the traditional representations in protein sub-nuclear localization, and the representation treated by LDA outperforms the untreated one.

  15. A proline-tyrosine nuclear localization signal (PY-NLS) is required for the nuclear import of fission yeast PAB2, but not of human PABPN1.

    Science.gov (United States)

    Mallet, Pierre-Luc; Bachand, François

    2013-03-01

    Nuclear poly(A)-binding proteins (PABPs) are evolutionarily conserved proteins that play key roles in eukaryotic gene expression. In the fission yeast Schizosaccharomyces pombe, the major nuclear PABP, Pab2, functions in the maturation of small nucleolar RNAs as well as in nuclear RNA decay. Despite knowledge about its nuclear functions, nothing is known about how Pab2 is imported into the nucleus. Here, we show that Pab2 contains a proline-tyrosine nuclear localization signal (PY-NLS) that is necessary and sufficient for its nuclear localization and function. Consistent with the role of karyopherin β2 (Kapβ2)-type receptors in the import of PY-NLS cargoes, we show that the fission yeast ortholog of human Kapβ2, Kap104, binds to recombinant Pab2 and is required for Pab2 nuclear localization. The absence of arginine methylation in a basic region N-terminal to the PY-core motif of Pab2 did not affect its nuclear localization. However, in the context of a sub-optimal PY-NLS, we found that Pab2 was more efficiently targeted to the nucleus in the absence of arginine methylation, suggesting that this modification can affect the import kinetics of a PY-NLS cargo. Although a sequence resembling a PY-NLS motif can be found in the human Pab2 ortholog, PABPN1, our results indicate that neither a functional PY-NLS nor Kapβ2 activity are required to promote entry of PABPN1 into the nucleus of human cells. Our findings describe the mechanism by which Pab2 is imported into the nucleus, providing the first example of a PY-NLS import system in fission yeast. In addition, this study suggests the existence of alternative or redundant nuclear import pathways for human PABPN1. PMID:23279110

  16. Subcellular localization of calcium and Ca-ATPase activity during nuclear maturation in Bufo arenarum oocytes.

    Science.gov (United States)

    Ramos, Inés; Cisint, Susana B; Crespo, Claudia A; Medina, Marcela F; Fernández, Silvia N

    2009-08-01

    The localization of calcium and Ca-ATPase activity in Bufo arenarum oocytes was investigated by ultracytochemical techniques during progesterone-induced nuclear maturation, under in vitro conditions. No Ca2+ deposits were detected in either control oocytes or progesterone-treated ones for 1-2 h. At the time when nuclear migration started, electron dense deposits of Ca2+ were visible in vesicles, endoplasmic reticulum cisternae and in the space between the annulate lamellae membranes. Furthermore, Ca-ATPase activity was also detected in these membrane structures. As maturation progressed, the cation deposits were observed in the cytomembrane structures, which underwent an important reorganization and redistribution. Thus, they moved from the subcortex and became located predominantly in the oocyte cortex area when nuclear maturation ended. Ca2+ stores were observed in vesicles surrounding or between the cortical granules, which are aligned close to the plasma membrane. The positive Ca-ATPase reaction in these membrane structures could indicate that the calcium deposit is an ATP-dependent process. Our results suggest that during oocyte maturation calcium would be stored in membrane structures where it remains available for release at the time of fertilization. Data obtained under our experimental conditions indicate that calcium from the extracellular medium would be important for the oocyte maturation process. PMID:19397840

  17. Nuclear localization of Lyn tyrosine kinase mediated by inhibition of its kinase activity

    International Nuclear Information System (INIS)

    Src-family kinases, cytoplasmic enzymes that participate in various signaling events, are found at not only the plasma membrane but also subcellular compartments, such as the nucleus, the Golgi apparatus and late endosomes/lysosomes. Lyn, a member of the Src-family kinases, is known to play a role in DNA damage response and cell cycle control in the nucleus. However, it is still unclear how the localization of Lyn to the nucleus is regulated. Here, we investigated the mechanism of the distribution of Lyn between the cytoplasm and the nucleus in epitheloid HeLa cells and hematopoietic THP-1 cells. Lyn was definitely detected in purified nuclei by immunofluorescence and immunoblotting analyses. Nuclear accumulation of Lyn was enhanced upon treatment of cells with leptomycin B (LMB), an inhibitor of Crm1-mediated nuclear export. Moreover, Lyn mutants lacking the sites for lipid modification were highly accumulated in the nucleus upon LMB treatment. Intriguingly, inhibition of the kinase activity of Lyn by SU6656, Csk overexpression, or point mutation in the ATP-binding site induced an increase in nuclear Lyn levels. These results suggest that Lyn being imported into and rapidly exported from the nucleus preferentially accumulates in the nucleus by inhibition of the kinase activity and lipid modification

  18. Improved Production Efficiency of Virus-Like Particles by the Baculovirus Expression Vector System.

    Directory of Open Access Journals (Sweden)

    Javier López-Vidal

    Full Text Available Vaccines based on virus-like particles (VLPs have proven effective in humans and animals. In this regard, the baculovirus expression vector system (BEVS is one of the technologies of choice to generate such highly immunogenic vaccines. The extended use of these vaccines for human and animal populations is constrained because of high production costs, therefore a significant improvement in productivity is crucial to ensure their commercial viability. Here we describe the use of the previously described baculovirus expression cassette, called TB, to model the production of two VLP-forming vaccine antigens in insect cells. Capsid proteins from porcine circovirus type 2 (PCV2 Cap and from the calicivirus that causes rabbit hemorrhagic disease (RHDV VP60 were expressed in insect cells using baculoviruses genetically engineered with the TB expression cassette. Productivity was compared to that obtained using standard counterpart vectors expressing the same proteins under the control of the polyhedrin promoter. Our results demonstrate that the use of the TB expression cassette increased the production yields of these vaccine antigens by around 300% with respect to the standard vectors. The recombinant proteins produced by TB-modified vectors were fully functional, forming VLPs identical in size and shape to those generated by the standard baculoviruses, as determined by electron microscopy analysis. The use of the TB expression cassette implies a simple modification of the baculovirus vectors that significantly improves the cost efficiency of VLP-based vaccine production, thereby facilitating the commercial viability and broad application of these vaccines for human and animal health.

  19. Improved Production Efficiency of Virus-Like Particles by the Baculovirus Expression Vector System.

    Science.gov (United States)

    López-Vidal, Javier; Gómez-Sebastián, Silvia; Bárcena, Juan; Nuñez, Maria del Carmen; Martínez-Alonso, Diego; Dudognon, Benoit; Guijarro, Eva; Escribano, José M

    2015-01-01

    Vaccines based on virus-like particles (VLPs) have proven effective in humans and animals. In this regard, the baculovirus expression vector system (BEVS) is one of the technologies of choice to generate such highly immunogenic vaccines. The extended use of these vaccines for human and animal populations is constrained because of high production costs, therefore a significant improvement in productivity is crucial to ensure their commercial viability. Here we describe the use of the previously described baculovirus expression cassette, called TB, to model the production of two VLP-forming vaccine antigens in insect cells. Capsid proteins from porcine circovirus type 2 (PCV2 Cap) and from the calicivirus that causes rabbit hemorrhagic disease (RHDV VP60) were expressed in insect cells using baculoviruses genetically engineered with the TB expression cassette. Productivity was compared to that obtained using standard counterpart vectors expressing the same proteins under the control of the polyhedrin promoter. Our results demonstrate that the use of the TB expression cassette increased the production yields of these vaccine antigens by around 300% with respect to the standard vectors. The recombinant proteins produced by TB-modified vectors were fully functional, forming VLPs identical in size and shape to those generated by the standard baculoviruses, as determined by electron microscopy analysis. The use of the TB expression cassette implies a simple modification of the baculovirus vectors that significantly improves the cost efficiency of VLP-based vaccine production, thereby facilitating the commercial viability and broad application of these vaccines for human and animal health. PMID:26458221

  20. Feasibility of baculovirus-mediated reporter gene delivery for efficient monitoring of islet transplantation in vivo

    International Nuclear Information System (INIS)

    Objective: The objective of this study was to explore the feasibility of baculovirus vector-mediated sodium iodide symporter (NIS) gene delivery to monitor islet transplantation. Methods: Baculovirus vectors expressing green fluorescent protein (GFP) or NIS (Bac-GFP and Bac-NIS) were established using the Bac-to-Bac baculovirus expression system. The GFP expression of Bac-GFP-infected rat islets was observed in vitro by fluorescence microscopy. Iodine uptake and inhibition of iodine uptake by NaClO4 in Bac-NIS-infected islets were dynamically monitored in vitro. Bac-GFP- or Bac-NIS-infected islets were implanted into the left axillary cavity of NOD-SCID mice, and fluorescence imaging and 125I NanoSPECT/CT imaging were subsequently performed in vivo. Results: Bac-GFP efficiently infected rat islets (over 95% infected at MOI = 40), and the expression of GFP lasted approximately two weeks. NaClO4 could inhibit iodine uptake by Bac-NIS-infected islets. In vivo imaging revealed that the fluorescence intensity of the transplant sites in Bac-GFP-infected groups was significantly higher than in the non-infected group. Grafts could be clearly observed by 125I NanoSPECT/CT imaging for up to 8 h. Conclusion: Baculovirus vectors are powerful vehicles for studying rat islets in gene delivery. It is feasible to use a baculovirus vector to delivery an NIS gene for non-invasive monitoring transplanted islets in vivo by the expression of the target gene

  1. Application of local computer networks to nuclear power plant problems at the level of automated control of the production process

    International Nuclear Information System (INIS)

    The requirements placed on the hardware and software for the automated system of production control at a nuclear power plant are summarized. In the design of the Temelin nuclear power plant, this system is local network-based, with two-way communication between the local stations. The control functions are distributed among 6 local stations, viz. the file server, interactive terminal, information terminal, data storage terminal, calculation terminal and communication terminal. A brief characterization is given of the necessary software, including both the system software and the basic and applications user software. (Z.M.)

  2. A study on the feasibility and a possible form of local residents participation in nuclear safety regulation

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Sang Hoon; Son, Moon Kyu [Korea Association for Nuclear Technology, Taejon (Korea, Republic of); Kim, Seong Sook [Shila Univ., Busan (Korea, Republic of)

    2003-02-15

    It is expected that the results of this study will help policy makers in the government and nuclear industry enhance the confidence of local residents and citizens in the safety of nuclear energy. It is also believed that the citizen confidence on the nuclear energy will help them implement nuclear policies more efficiently and more safely. Considering that the newly sworn-in government is to involve citizens in wide variety of policy areas, this study fits with the policy directions of the new government. Most importantly, desirable citizen participation in public decision-making is to contribute to the enhancement of the working democracy.

  3. Evaluation of the basic concepts of approaches for the coexistence of nuclear energy and people/local community

    International Nuclear Information System (INIS)

    In November 2007, the Policy Evaluation Committee compiled the report, which evaluated the basic concepts of approaches to the coexistence of nuclear energy and people/local community, specified in the Framework for Nuclear Energy Policy. The report states that the 'concerned administrative bodies are carrying out measures related to the coexistence of nuclear energy and people/local communities in line with these basic concept' and summarizes fifteen proposals conductive to the betterment and improvement of these measures, which were classified as 1) secure transparency and promotion of mutual understanding with the public, 2) development and enrichment of learning opportunities and public participation, 3) relationship between the government and local governments and 4) coexistence with local residents. The Japan Atomic Energy Commission (JAEC) considers this report to be reasonable. This article presented an overview of this activity. (T. Tanaka)

  4. Interaction between a plasma membrane-localized ankyrin-repeat protein ITN1 and a nuclear protein RTV1

    International Nuclear Information System (INIS)

    Highlights: ► ITN1, a plasma membrane ankyrin protein, interacts with a nuclear DNA-binding protein RTV1. ► The nuclear transport of RTV1 is partially inhibited by interaction with ITN1. ► RTV1 can promote the nuclear localization of ITN1. ► Both overexpression of RTV1 and the lack of ITN1 increase salicylic acids sensitivity in plants. -- Abstract: The increased tolerance to NaCl 1 (ITN1) protein is a plasma membrane (PM)-localized protein involved in responses to NaCl stress in Arabidopsis. The predicted structure of ITN1 is composed of multiple transmembrane regions and an ankyrin-repeat domain that is known to mediate protein–protein interactions. To elucidate the molecular functions of ITN1, we searched for interacting partners using a yeast two-hybrid assay, and a nuclear-localized DNA-binding protein, RTV1, was identified as a candidate. Bimolecular fluorescence complementation analysis revealed that RTV1 interacted with ITN1 at the PM and nuclei in vivo. RTV1 tagged with red fluorescent protein localized to nuclei and ITN1 tagged with green fluorescent protein localized to PM; however, both proteins localized to both nuclei and the PM when co-expressed. These findings suggest that RTV1 and ITN1 regulate the subcellular localization of each other.

  5. Interaction between a plasma membrane-localized ankyrin-repeat protein ITN1 and a nuclear protein RTV1

    Energy Technology Data Exchange (ETDEWEB)

    Sakamoto, Hikaru [Department of Bioproduction, Faculty of Bioindustry, Tokyo University of Agriculture, 196 Yasaka, Abashiri-shi, Hokkaido 093-2422 (Japan); Sakata, Keiko; Kusumi, Kensuke [Department of Biology, Faculty of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Kojima, Mikiko; Sakakibara, Hitoshi [RIKEN Plant Science Center, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045 (Japan); Iba, Koh, E-mail: koibascb@kyushu-u.org [Department of Biology, Faculty of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan)

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer ITN1, a plasma membrane ankyrin protein, interacts with a nuclear DNA-binding protein RTV1. Black-Right-Pointing-Pointer The nuclear transport of RTV1 is partially inhibited by interaction with ITN1. Black-Right-Pointing-Pointer RTV1 can promote the nuclear localization of ITN1. Black-Right-Pointing-Pointer Both overexpression of RTV1 and the lack of ITN1 increase salicylic acids sensitivity in plants. -- Abstract: The increased tolerance to NaCl 1 (ITN1) protein is a plasma membrane (PM)-localized protein involved in responses to NaCl stress in Arabidopsis. The predicted structure of ITN1 is composed of multiple transmembrane regions and an ankyrin-repeat domain that is known to mediate protein-protein interactions. To elucidate the molecular functions of ITN1, we searched for interacting partners using a yeast two-hybrid assay, and a nuclear-localized DNA-binding protein, RTV1, was identified as a candidate. Bimolecular fluorescence complementation analysis revealed that RTV1 interacted with ITN1 at the PM and nuclei in vivo. RTV1 tagged with red fluorescent protein localized to nuclei and ITN1 tagged with green fluorescent protein localized to PM; however, both proteins localized to both nuclei and the PM when co-expressed. These findings suggest that RTV1 and ITN1 regulate the subcellular localization of each other.

  6. CHARACTERIZATION OF THE GLYCOSYLATED ECDYSTEROIDS IN THE HEMOLYMPH OF BACULOVIRUS-INFECTED GYPSY MOTH LARVAE AND CELLS IN CULTURE

    Science.gov (United States)

    Fourth-instar gypsy moth (Lymantria dispar; Lepidoptera: Lymantriidae) larvae, infected with the gypsy moth baculovirus (LdNPV), show an elevated and prolonged extension of the hemolymph ecdysteroid titer peak associated with molting. The ecdysteroid immunoreactivity associated w...

  7. Rabies-virus-glycoprotein-pseudotyped recombinant baculovirus vaccine confers complete protection against lethal rabies virus challenge in a mouse model.

    Science.gov (United States)

    Wu, Qunfeng; Yu, Fulai; Xu, Jinfang; Li, Yang; Chen, Huanchun; Xiao, Shaobo; Fu, Zhen F; Fang, Liurong

    2014-06-25

    Rabies virus has been an ongoing threat to humans and animals. Here, we developed a new strategy to generate a rabies virus vaccine based on a pseudotyped baculovirus. The recombinant baculovirus (BV-RVG/RVG) was pseudotyped with the rabies virus glycoprotein (RVG) and also simultaneously expressed another RVG under the control of the immediate early CMV promoter. In vitro, this RVG-pseudotyped baculovirus vector induced syncytium formation in insect cells and displayed more efficient gene delivery into mammalian cells. Mice immunized with BV-RVG/RVG developed higher levels of virus-neutralizing antibodies, and conferred 100% protection against rabies viral challenge. These data indicate that the RVG-pseudotyped baculovirus BV-RVG/RVG can be used as an alternative strategy to develop a safe and efficacious vaccine against the rabies virus. PMID:24793501

  8. Experimental Approaches to Study Mitochondrial Localization and Function of a Nuclear Cell Cycle Kinase, Cdk1.

    Science.gov (United States)

    Candas, Demet; Qin, Lili; Fan, Ming; Li, Jian-Jian

    2016-01-01

    Although mitochondria possess their own transcriptional machinery, merely 1% of mitochondrial proteins are synthesized inside the organelle. The nuclear-encoded proteins are transported into mitochondria guided by their mitochondria targeting sequences (MTS); however, a majority of mitochondrial localized proteins lack an identifiable MTS. Nevertheless, the fact that MTS can instruct proteins to go into the mitochondria provides a valuable tool for studying mitochondrial functions of normally nuclear and/or cytoplasmic proteins. We have recently identified the cell cycle kinase CyclinB1/Cdk1 complex in the mitochondria. To specifically study the mitochondrial functions of this complex, mitochondrial overexpression and knock-down of this complex without interfering with its nuclear or cytoplasmic functions were essential. By tagging CyclinB1/Cdk1 with MTS, we were able to achieve mitochondrial overexpression of this complex to study its mitochondrial targets as well as functions. Via tagging dominant-negative Cdk1 with MTS, inhibition of Cdk1 activity was accomplished particularly in the mitochondria. Potential mitochondrial targets of CyclinB1/Cdk1 complex were identified using a gel-based proteomics approach. Unlike traditional 2D gel analysis, we employed 2-dimensional difference gel electrophoresis (2D-DIGE) technology followed by phosphoprotein staining to fluorescently label differentially phosphorylated proteins in mitochondrial Cdk1 expressing cells. Identification of phosphoprotein spots that were altered in wild type versus dominant negative Cdk1 bearing mitochondria revealed the identity of mitochondrial targets of Cdk1. Finally, to determine the effect of CyclinB1/Cdk1 mitochondrial localization in cell cycle progression, a cell proliferation assay using a synthetic thymidine analogue EdU (5-ethynyl-2'-deoxyuridine) was used to monitor the cells as they go through the cell cycle and replicate their DNA. Altogether, we demonstrated a variety of approaches

  9. Efficient, low-cost protein factories: expression of human adenosine deaminase in baculovirus-infected insect larvae.

    OpenAIRE

    Medin, J A; Hunt, L; Gathy, K; Evans, R K; Coleman, M S

    1990-01-01

    Human adenosine deaminase (EC 3.5.4.4), a key purine salvage enzyme essential for immune competence, has been overproduced in Spodoptera frugiperda cells and in Trichoplusia ni (cabbage looper) larvae infected with recombinant baculovirus. The coding sequence of human adenosine deaminase was recombined into a baculovirus immediately downstream from the strong polyhedrin gene promoter. Approximately 60 hr after infection of insect cells with the recombinant virus, maximal levels of intracellul...

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Science.gov (United States)

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

    2016-01-01

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

  12. GD3 nuclear localization after apoptosis induction in HUT-78 cells

    International Nuclear Information System (INIS)

    Glycosphingolipids are essential components of plasma membrane and act as antigens, mediators of cell adhesion, and modulators of signal transduction. Following activation of the Fas receptor, gangliosides are recuited in various intracellular compartments. We have evaluated whether the pro-apoptotic anti-CD95 antibody induces a nuclear localization of GD3 in HUT-78 cells. Our data shows that GD3 translocation from cytosol to nuclei is strongly correlated to concomitant rapid phosphorylation of histone H1 shortly after induction of apoptosis. This work advances the hypothesis that GD3 induces a post-translational modification of histone H1 thus influencing the apoptosis process through transcriptional activation/repression of specific genes

  13. Monodisperse magnetite nanoparticles coupled with nuclear localization signal peptide for cell-nucleus targeting.

    Science.gov (United States)

    Xu, Chenjie; Xie, Jin; Kohler, Nathan; Walsh, Edward G; Chin, Y Eugene; Sun, Shouheng

    2008-03-01

    Functionalization of monodisperse superparamagnetic magnetite (Fe(3)O(4)) nanoparticles for cell specific targeting is crucial for cancer diagnostics and therapeutics. Targeted magnetic nanoparticles can be used to enhance the tissue contrast in magnetic resonance imaging (MRI), to improve the efficiency in anticancer drug delivery, and to eliminate tumor cells by magnetic fluid hyperthermia. Herein we report the nucleus-targeting Fe(3)O(4) nanoparticles functionalized with protein and nuclear localization signal (NLS) peptide. These NLS-coated nanoparticles were introduced into the HeLa cell cytoplasm and nucleus, where the particles were monodispersed and non-aggregated. The success of labeling was examined and identified by fluorescence microscopy and MRI. The work demonstrates that monodisperse magnetic nanoparticles can be readily functionalized and stabilized for potential diagnostic and therapeutic applications. PMID:18080259

  14. [GnRH analogues containing SV-40 virus T-antigen nuclear localization sequence].

    Science.gov (United States)

    Burov, S V; Iablokova, T V; Dorosh, M Iu; Kriviziuk, E V; Efremov, A M; Orlov, S V

    2010-01-01

    To improve the efficiency of anticancer drugs due to their delivery to intracellular targets a set of GnRH analogues containing nuclear localization signal (NLS) of SV-40 virus large T-antigen have been synthesized. NLS was attached to the parent molecule via ε-amino group of D-Lysine in position 1 or 6 of peptide sequence using orthogonal protection strategy. The biological activity studies revealed that incorporation of NLS moiety significantly increases cytotoxic activity of palmitoyl-containing GnRH analogues in vitro. The influence of tested peptides on tumor cells does not accompanied by the destruction of cell membrane, as confirmed in experiments with normal fibroblasts, used as a control. PMID:21063449

  15. Effect of nuclear mass on carrier-envelope-phase-controlled electron localization in dissociating molecules

    Science.gov (United States)

    Xu, Han; Xu, Tian-Yu; He, Feng; Kielpinski, D.; Sang, R. T.; Litvinyuk, I. V.

    2014-04-01

    We explore the effect of nuclear mass on the laser-driven electron localization process. We dissociate a mixed H2 and D2 target with intense, carrier-envelope-phase (CEP) stable 6 fs laser pulses and detect the products in a reaction microscope. We observe a very strong CEP-dependent asymmetry in proton and deuteron emission for low dissociation energy channels. This asymmetry is stronger for H2 than for D2. We also observe a large CEP offset between the asymmetry spectra for H2 and D2. Our theoretical simulations, based on a one-dimensional two-channel model, agree very well with the asymmetry spectra, but fail to account properly for the phase difference between the two isotopes.

  16. Recombinant baculovirus mediates dsRNA specific to rr2 delivery and its protective efficacy against WSSV infection.

    Science.gov (United States)

    Rattanarojpong, Triwit; Khankaew, Suthiwat; Khunrae, Pongsak; Vanichviriyakit, Rapeepun; Poomputsa, Kanokwan

    2016-07-10

    White spot syndrome virus (WSSV) is a major causative agent in shrimp farming. Consequently, RNAi technology is an effective strategy to prevent WSSV infection in shrimp especially dsRNA targeting to rr2 of WSSV. In an effort to develop dsRNA expression in shrimp for control of WSSV infection, we developed a recombinant baculovirus expressing recombinant VP28 as the gene delivery system to carry a gene encoding dsRNA specific to rr2 for triggering the RNAi process in shrimp. The results showed that the recombinant baculovirus harboring VP28 was able to express VP28 indicated by Western blot with polyclonal antibody specific to VP28. VP28 transcript was detected in shrimp hemocytes after co-culture hemocytes with the recombinant baculovirus displaying VP28. In addition, we found that shrimp injected with the recombinant baculovirus displaying VP28 and encoding dsRNA synthetic gene specific to rr2 (Bac-VP28-dsrr2) showed the lowest cumulative mortality (33%) at 14days post infection (dpi) when compared to shrimp injected with baculovirus displaying VP28 (Bac-VP28) (64% cumulative mortality) (pBac-VP28-dsrr2 also showed significantly lower WSSV copies than shrimp injected with Bac-VP28 (pBac-VP28-dsrr2 was effective in prevention of WSSV infection. Therefore, the results obtained here can be applied to the prevention of WSSV infection by mixing the recombinant baculovirus with shrimp feed in the future. PMID:27164257

  17. The RelA nuclear localization signal folds upon binding to IκBα

    Science.gov (United States)

    Cervantes, Carla F.; Bergqvist, Simon; Kjaergaard, Magnus; Kroon, Gerard; Sue, Shih-Che; Dyson, H. Jane; Komives, Elizabeth A.

    2010-01-01

    Summary The nuclear localization signal (NLS) polypeptide of RelA, the canonical NFκB family member, is responsible for regulating the nuclear localization of RelA-containing NFκB dimers. The RelA NLS polypeptide also plays a crucial role in mediating the high affinity and specificity of the interaction of RelA-containing dimers with the inhibitor IκBα, forming two helical motifs according to the published X-ray crystal structure. In order to define the nature of the interaction between the RelA NLS and IκBα under solution conditions, we conducted NMR and isothermal titration calorimetry (ITC) studies using a truncated form of IκBα containing residues 67-206 and a peptide spanning residues 293-321 of RelA. The NLS peptide, though largely unfolded, has a weak tendency toward helical structure when free in solution. Upon addition of the labeled peptide to unlabeled IκBα, the resonance dispersion in the NMR spectrum is significantly greater, providing definitive evidence that the RelA NLS polypeptide folds upon binding IκBα. ITC studies of single-point mutants reveal that residue F309, which is located in the middle of the more C-terminal of the two helices (helix 4) in the IκBα-bound RelA NLS polypeptide, is critical for the binding of the RelA NLS polypeptide to IκBα. These results help to explain the role of helix 4 in mediating the high affinity of RelA for IκBα. PMID:21094161

  18. Colloquium / Preparation for nuclear post-accident. Citizens and local authorities involvement in major risks governance - proceedings

    International Nuclear Information System (INIS)

    During 40 years, France has chosen nuclear energy as main energy source for power generation. Today, nuclear energy covers 80% of the French electricity needs. For this reason, in France, each inhabitant lives at less than 200 km of a nuclear power plant. The September 11, 2001 terror attack has led to reconsider the nuclear risk in terms of security. In 2005, the French nuclear safety authority (ASN) has been entrusted with the preparation of a nuclear post-accident management study. This study has been mainly based on the experience feedback of the Chernobyl accident and on the dialogue with different actors of the French territory: the local authorities, the habitants, the associations and the health, environment and education actors. This colloquium represents the opening act of the joint elaboration of the management of a potential nuclear accident at the region scale. The colloquium was organized around 5 round tables: 1 - Did the nuclear safety transparency law change the situation?; 2 - Examples of pluralistic dialogues: the long-term management of a nuclear accident; 3 - How to encourage the necessary skills development of citizens? 4 - Regional development strategies in terms of nuclear risks management (incidents, effluents, wastes, transports..); 5 - New territory liabilities and citizens' legitimate aspiration in terms of health/environment: building something together. This document is the proceedings of this colloquium. It reports the exchanges between the participants

  19. Nuclear binding energies: Global collective structure and local shell-model correlations

    International Nuclear Information System (INIS)

    Nuclear binding energies and two-neutron separation energies are analysed starting from the liquid-drop model and the nuclear shell model in order to describe the global trends of the above observables. We subsequently concentrate on the Interacting Boson Model (IBM) and discuss a new method in order to provide a consistent description of both, ground-state and excited-state properties. We address the artefacts that appear when crossing mid-shell using the IBM formulation and perform detailed numerical calculations for nuclei situated in the 50-82 shell. We also concentrate on local deviations from the above global trends in binding energy and two-neutron separation energies that appear in the neutron-deficient Pb region. We address possible effects on the binding energy, caused by mixing of low-lying 0+ intruder states into the ground state, using configuration mixing in the IBM framework. We also study ground-state properties using a macroscopic-microscopic model. Detailed comparisons with recent experimental data in the Pb region are amply discussed

  20. ALS mutant FUS disrupts nuclear localization and sequesters wild-type FUS within cytoplasmic stress granules.

    Science.gov (United States)

    Vance, Caroline; Scotter, Emma L; Nishimura, Agnes L; Troakes, Claire; Mitchell, Jacqueline C; Kathe, Claudia; Urwin, Hazel; Manser, Catherine; Miller, Christopher C; Hortobágyi, Tibor; Dragunow, Mike; Rogelj, Boris; Shaw, Christopher E

    2013-07-01

    Mutations in the gene encoding Fused in Sarcoma (FUS) cause amyotrophic lateral sclerosis (ALS), a fatal neurodegenerative disorder. FUS is a predominantly nuclear DNA- and RNA-binding protein that is involved in RNA processing. Large FUS-immunoreactive inclusions fill the perikaryon of surviving motor neurons of ALS patients carrying mutations at post-mortem. This sequestration of FUS is predicted to disrupt RNA processing and initiate neurodegeneration. Here, we demonstrate that C-terminal ALS mutations disrupt the nuclear localizing signal (NLS) of FUS resulting in cytoplasmic accumulation in transfected cells and patient fibroblasts. FUS mislocalization is rescued by the addition of the wild-type FUS NLS to mutant proteins. We also show that oxidative stress recruits mutant FUS to cytoplasmic stress granules where it is able to bind and sequester wild-type FUS. While FUS interacts with itself directly by protein-protein interaction, the recruitment of FUS to stress granules and interaction with PABP are RNA dependent. These findings support a two-hit hypothesis, whereby cytoplasmic mislocalization of FUS protein, followed by cellular stress, contributes to the formation of cytoplasmic aggregates that may sequester FUS, disrupt RNA processing and initiate motor neuron degeneration. PMID:23474818

  1. Local government's view on Chuetsu-oki earthquake and nuclear power plant facilities

    International Nuclear Information System (INIS)

    Niigataken Chuetsu-oki earthquake occurred in July 2007 and damaged individual's housing and properties in local residents and Kashiwazaki-Kariwa nuclear power plant facilities. The earthquake caused very strong ground motions far exceeding the assumed values in the design. The impacts were a fire breakout of the Unit 3 transformer and a release of small amount of radioactive materials to the environment at Unit 6 and 7. A joint of the driving shaft of the overhead crane of the Unit 6 reactor building was damaged. The fire of the Unit 3 transformer developed safety concerns among the public because of insufficient effectiveness of the private fire brigade, unavailability of fire control systems, and delayed notification to the external fire station, and as its consequence a lot of time needed to bring the fire under control. Delayed information dissemination gave deep concerns about the possible radioactive contamination in the environment. Based on New Regulatory Guide for Reviewing Seismic Design of Nuclear Power Reactor Facilities revised in September 2006, detail understanding of shakes and additional investigation of geological faults around the plant site were underway. Precise explanations on in-situ investigation and evaluation results to the residents would be necessary to answer their seismic safety concerns. (T. Tanaka)

  2. Activation of AMP-activated protein kinase stimulates the nuclear localization of glyceraldehyde 3-phosphate dehydrogenase in human diploid fibroblasts

    OpenAIRE

    Kwon, Hyun Jin; Rhim, Ji Heon; Jang, Ik-Soon; Kim, Go-Eun; Park, Sang Chul; Yeo, Eui-Ju

    2010-01-01

    In addition to its well-known glycolytic activity, GAPDH displays multiple functions, such as nuclear RNA export, DNA replication and repair, and apoptotic cell death. This functional diversity depends on its intracellular localization. In this study, we explored the signal transduction pathways involved in the nuclear translocation of GAPDH using confocal laser scanning microscopy of immunostained human diploid fibroblasts (HDFs). GAPDH was present mainly in the cytoplasm when cultured wi...

  3. Centrin 2 Localizes to the Vertebrate Nuclear Pore and Plays a Role in mRNA and Protein Export▿ †

    OpenAIRE

    Resendes, Karen K.; Rasala, Beth A.; Forbes, Douglass J.

    2008-01-01

    Centrins in vertebrates have traditionally been associated with microtubule-nucleating centers such as the centrosome. Unexpectedly, we found centrin 2 to associate biochemically with nucleoporins, including the Xenopus laevis Nup107-160 complex, a critical subunit of the vertebrate nuclear pore in interphase and of the kinetochores and spindle poles in mitosis. Immunofluorescence of Xenopus cells and in vitro reconstituted nuclei indeed revealed centrin 2 localized at the nuclear pores. Use ...

  4. Consumption of locally grown foods by the populations residing near nuclear power stations

    International Nuclear Information System (INIS)

    The studies carried out by the Geographical Institute of Aix-en-Provence and the B.E.G.E.A. are in three forms: the first studies the local rural production (farming, animal rearing and fishing); the second is concerned with the food intake of the population within a radius of 5 to 10 km around the site; the third consists in working out a soil utilization map covering an area of about 2500 hectares around the site in order to determine the agricultural and urban areas as well as the open spaces, and the exact sort of cultivation carried out, plot by plot. The map also represents all the phenomena concerning hydrography, irrigation, watering and the supply of potable water. All these studies concern only a few parishes and enable an accurate knowledge of the environment to be acquired. The food studies seek an order of magnitude of the amount of local produce consumed by the populations of the nuclear power station sites with a view to determining radioactive contamination levels. The results show the specific food consumption features of a population bearing the deep stamp of its environment and its rural origin

  5. Expression and localization of heterogeneous nuclear ribonucleoprotein K in mouse ovaries and preimplantation embryos.

    Science.gov (United States)

    Zhang, Ping; Wang, Ningling; Lin, Xianhua; Jin, Li; Xu, Hong; Li, Rong; Huang, Hefeng

    2016-02-26

    Heterogeneous nuclear ribonucleoprotein K (hnRNP K), an evolutionarily conserved protein, is involved in several important cellular processes that are relevant to cell proliferation, differentiation, apoptosis, and cancer development. However, details of hnRNP K expression during mammalian oogenesis and preimplantation embryo development are lacking. The present study investigates the expression and cellular localization of K protein in the mouse ovaries and preimplantation embryos using immunostaining. We demonstrate, for the first time, that hnRNP K is abundantly expressed in the nuclei of mouse oocytes in primordial, primary and secondary follicles. In germ vesicle (GV)-stage oocytes, hnRNP K accumulates in the germinal vesicle in a spot distribution manner. After germinal vesicle breakdown, speckled hnRNP K is diffusely distributed in the cytoplasm. However, after fertilization, the K protein relocates into the female and male pronucleus and persists in the blastomere nuclei. Localization of K protein in the human ovary and ovarian granulosa cell tumor (GCT) was also investigated. Overall, this study provides important morphological evidence to better understand the possible roles of hnRNP K in mammalian oogenesis and early embryo development. PMID:26850853

  6. A Mathematical Model of Baculovirus Infection on Insect Cells at Low Multiplicity of Infection

    Institute of Scientific and Technical Information of China (English)

    You-Hong ZHANG; Josée C. MERCHUK

    2004-01-01

    The expression efficiency of the insect cells-baculovirus system used for insecticidal virus production and the expression of medically useful foreign genes is closely related with the dynamics of infection. The present studies develop a model of the dynamic process of insect cell infection with baculovirus at low multiplicity of infection (MOI), which is based on the multi-infection cycles of insect cell infection at low MOI. A mathematical model for the amount of viruses released from primary infected cells and the amount of free viruses before secondary infected cells release viruses has been developed. Comparison of the simulation results with the experimental data confirms qualitatively that this model is highly reasonable before secondary infected cells release viruses. This model is considered as a base for further modeling the entire complicated infection process.

  7. Functional analysis of the C-terminal region of human adenovirus E1A reveals a misidentified nuclear localization signal

    Energy Technology Data Exchange (ETDEWEB)

    Cohen, Michael J.; King, Cason R.; Dikeakos, Jimmy D. [Department of Microbiology and Immunology, The University of Western Ontario, A4-833 London Regional Cancer Centre, 800 Commissioners Road E., London, Ontario, N6A 4L6 Canada (Canada); Mymryk, Joe S., E-mail: jmymryk@uwo.ca [Department of Microbiology and Immunology, The University of Western Ontario, A4-833 London Regional Cancer Centre, 800 Commissioners Road E., London, Ontario, N6A 4L6 Canada (Canada); Department of Oncology, The University of Western Ontario, London Regional Cancer Centre, Ontario (Canada)

    2014-11-15

    The immortalizing function of the human adenovirus 5 E1A oncoprotein requires efficient localization to the nucleus. In 1987, a consensus monopartite nuclear localization sequence (NLS) was identified at the C-terminus of E1A. Since that time, various experiments have suggested that other regions of E1A influence nuclear import. In addition, a novel bipartite NLS was recently predicted at the C-terminal region of E1A in silico. In this study, we used immunofluorescence microscopy and co-immunoprecipitation analysis with importin-α to verify that full nuclear localization of E1A requires the well characterized NLS spanning residues 285–289, as well as a second basic patch situated between residues 258 and 263 ({sup 258}RVGGRRQAVECIEDLLNEPGQPLDLSCKRPRP{sup 289}). Thus, the originally described NLS located at the C-terminus of E1A is actually a bipartite signal, which had been misidentified in the existing literature as a monopartite signal, altering our understanding of one of the oldest documented NLSs. - Highlights: • Human adenovirus E1A is localized to the nucleus. • The C-terminus of E1A contains a bipartite nuclear localization signal (NLS). • This signal was previously misidentified to be a monopartite NLS. • Key basic amino acid residues within this sequence are highly conserved.

  8. A Dual Mechanism Controls Nuclear Localization in the Atypical Basic-Helix-Loop-Helix Protein PAR1 of Arabidopsis thaliana

    Institute of Scientific and Technical Information of China (English)

    Anahit Galstyan; Jordi Bou-Torrent; Irma Roig-Villanova; Jaime F. Martínez-García

    2012-01-01

    PAR1 is an atypical basic-helix-loop-helix (bHLH) protein that negatively regulates the shade avoidance syndrome in Arabidopsis thaliana acting as a transcriptional cofactor.Consistently with this function,PAR1 has to be in the nucleus to display biological activity.Previous structure-function analyses revealed that the N-terminal region of PAR1 drives the protein to the nucleus.However,truncated forms of PAR1 lacking this region still display biological activity,implying that PAR1 has additional mechanisms to localize into the nucleus.In this work,we compared the primary structure of PAR1 and various related and unrelated plant bHLH proteins,which led us to suggest that PAR1 contains a non-canonical nuclear localization signal (NLS) in the N-terminal region.By overexpressing truncated and mutated derivatives of PAR1,we have also investigated the importance of other regions of PAR1,such as the acidic and the extended HLH dimerization domains,for its nuclear localization.We found that,in the absence of the N-terminal region,a functional HLH domain is required for nuclear localization.Our results suggest the existence of a dual mechanism for PAR1 nuclear localization:(1) one mediated by the N-terminal non-consensus NLS and (2) a second one that involves interaction with other proteins via the dimerization domain.

  9. Development of the local stress analysis methodology for nuclear class 1 piping welded to the seal plate

    International Nuclear Information System (INIS)

    Lugs, brackets, stiffners and other attachments may be welded, bolted and studded to the outside or inside of piping and the local stress arise because of the radial thermal expansion of the piping, the dilatation of the piping due to its internal pressure, the circumferential contraction of the pipe as a results of an axial tensile force (Poisson's effect), etc., being constrained by those. So the evaluation of the local stress for the piping constrained by those. So the evaluation of the local stress for the piping constrained by the attachments in accordance with the ASME Section III NB-3651.3 is required for the nuclear class 1 piping. In this report the local stress analysis procedure for the nuclear class 1 piping welded to the seal plate was established. SInce the stress analysis for the nuclear class 1 piping require the fatigue analysis procedure was established using the commercial code ANSYS 5.1 for the structure analysis in this report. The procedure which was developed in this report can be used very effectively for the design for the seal plate and the local stress analysis of the nuclear class 1 piping welded to the seal plate. 14 figs., 5 refs. (Author) .new

  10. Localization of nuclear cathepsin L and its association with disease progression and poor outcome in colorectal cancer.

    LENUS (Irish Health Repository)

    Sullivan, Shane

    2012-02-01

    Previous in vitro studies have identified a nuclear isoform of Cathepsin L. The aim of this study was to examine if nuclear Cathepsin L exists in vivo and examine its association with clinical, pathological and patient outcome data. Cellular localization (nuclear and cytoplasmic) and expression levels v of Cathespin L in 186 colorectal cancer cases using immunohistochemistry. The molecular weight and activity of nuclear and cytoplasmic Cathepsin L in vivo and in vitro were assessed by Western blotting and ELISA, respectively. Epithelial nuclear staining percentage (p = 0.04) and intensity (p = 0.006) increased with advancing tumor stage, whereas stromal cytoplasmic staining decreased (p = 0.02). Using multivariate statistical analysis, survival was inversely associated with staining intensity in the epithelial cytoplasm (p = 0.01) and stromal nuclei (p = 0.007). In different colorectal cell lines and in vivo tumors, pro- and active Cathepsin L isoforms were present in both the cytoplasm and nuclear samples, with pro-Cathepsin L at 50 kDa and active Cathepsin L at 25 kDa. Purified nuclear and cytoplasmic fractions from cell lines and tumors showed active Cathepsin L activity. The identification of nuclear Cathepsin L may play an important prognostic role in colorectal disease progression and patient outcome. Moreover, these findings suggest that altering active nuclear Cathepsin L may significantly influence disease progression.

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

    Energy Technology Data Exchange (ETDEWEB)

    Patterson, Edward I. [Charles Sturt University, School of Animal and Veterinary Sciences, Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); EH Graham Centre for Agricultural Innovation (NSW Department of Primary Industries and Charles Sturt University), Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); Dombrovski, Andrew K. [Charles Sturt University, School of Biomedical Sciences, Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); Swarbrick, Crystall M.D. [Charles Sturt University, School of Biomedical Sciences, Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); EH Graham Centre for Agricultural Innovation (NSW Department of Primary Industries and Charles Sturt University), Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); Raidal, Shane R. [Charles Sturt University, School of Animal and Veterinary Sciences, Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); EH Graham Centre for Agricultural Innovation (NSW Department of Primary Industries and Charles Sturt University), Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); Forwood, Jade K., E-mail: jforwood@csu.edu.au [Charles Sturt University, School of Biomedical Sciences, Boorooma St., Wagga Wagga, New South Wales 2678 (Australia); EH Graham Centre for Agricultural Innovation (NSW Department of Primary Industries and Charles Sturt University), Boorooma St., Wagga Wagga, New South Wales 2678 (Australia)

    2013-09-06

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

  12. Baculovirus Envelope Protein ODV-E66 Is a Novel Chondroitinase with Distinct Substrate Specificity*

    OpenAIRE

    Sugiura, Nobuo; Setoyama, Yuka; Chiba, Mie; Kimata, Koji; Watanabe, Hideto

    2011-01-01

    Chondroitin sulfate is a linear polysaccharide of alternating d-glucuronic acid and N-acetyl-d-galactosamine residues with sulfate groups at various positions of the sugars. It interacts with and regulates cytokine and growth factor signal transduction, thus influencing development, organ morphogenesis, inflammation, and infection. We found chondroitinase activity in medium conditioned by baculovirus-infected insect cells and identified a novel chondroitinase. Sequence analysis revealed that ...

  13. Enhanced enterovirus 71 virus-like particle yield from a new baculovirus design.

    Science.gov (United States)

    Lin, Shih-Yeh; Yeh, Chia-Tsui; Li, Wan-Hua; Yu, Cheng-Ping; Lin, Wen-Chin; Yang, Jyh-Yuan; Wu, Hsueh-Ling; Hu, Yu-Chen

    2015-10-01

    Enterovirus 71 (EV71) is responsible for the outbreaks of hand-foot-and-mouth disease in the Asia-Pacific region. To produce the virus-like particle (VLP) vaccine, we previously constructed recombinant baculoviruses to co-express EV71 P1 polypeptide and 3CD protease using the Bac-to-Bac(®) vector system. The recombinant baculoviruses resulted in P1 cleavage by 3CD and subsequent VLP assembly in infected insect cells, but caused either low VLP yield or excessive VLP degradation. To tackle the problems, here we explored various expression cassette designs and flashBAC GOLD™ vector system which was deficient in v-cath and chiA genes. We found that the recombinant baculovirus constructed using the flashBAC GOLD™ system was insufficient to improve the EV71 VLP yield. Nonetheless, BacF-P1-C3CD, a recombinant baculovirus constructed using the flashBAC GOLD(TM) system to express P1 under the polh promoter and 3CD under the CMV promoter, dramatically improved the VLP yield while alleviating the VLP degradation. Infection of High Five(TM) cells with BacF-P1-C3CD enhanced the total and extracellular VLP yield to ≈268 and ≈171 mg/L, respectively, which enabled the release of abundant VLP into the supernatant and simplified the downstream purification. Intramuscular immunization of mice with 5 μg purified VLP induced cross-protective humoral responses and conferred protection against lethal virus challenge. Given the significantly improved extracellular VLP yield (≈171 mg/L) and the potent immunogenicity conferred by 5 μg VLP, one liter High Five(TM) culture produced ≈12,000 doses of purified vaccine, thus rendering the EV71 VLP vaccine economically viable and able to compete with inactivated virus vaccines. PMID:25997678

  14. Improved Production Efficiency of Virus-Like Particles by the Baculovirus Expression Vector System

    OpenAIRE

    López-Vidal, Javier; Gómez-Sebastián, Silvia; Bárcena, Juan; Nuñez, Maria del Carmen; Martínez-Alonso, Diego; Dudognon, Benoit; Guijarro, Eva; José M Escribano

    2015-01-01

    Vaccines based on virus-like particles (VLPs) have proven effective in humans and animals. In this regard, the baculovirus expression vector system (BEVS) is one of the technologies of choice to generate such highly immunogenic vaccines. The extended use of these vaccines for human and animal populations is constrained because of high production costs, therefore a significant improvement in productivity is crucial to ensure their commercial viability. Here we describe the use of the previousl...

  15. Expression and Characterization of Recombinant Serratia liquefaciens Nucleases Produced with Baculovirus-mediated Silkworm Expression System.

    Science.gov (United States)

    Iiyama, Kazuhiro; Lee, Jae Man; Tatsuke, Tuneyuki; Mon, Hiroaki; Kusakabe, Takahiro

    2016-06-01

    Baculovirus-Bombyx mori protein expression system has mainly been used for translation of eukaryotic proteins. In contrast, information pertaining to bacterial protein expression using this system is not sufficient. Therefore, recombinant nucleases from Serratia liquefaciens (rSlNucAs) were expressed in a Baculovirus-B. mori protein expression system. rSlNucAs containing the native signal peptide (rSlNucA-NSP) or silkworm 30-K signal peptide (rSlNucA-30K) at the NH2-terminus were constructed to enable secretion into the extracellular fraction. Both rSlNucA-30K and rSlNucA-NSP were successfully secreted into hemolymph of B. mori larvae. Affinity-purified rSlNucAs showed high nuclease activity. Optimum pH was 7.5 and half of maximum activity was maintained between pH 7.0 and 9.5. Optimum temperature was 35 °C. rSlNucAs showed sufficient activity in twofold-diluted radioimmunoprecipitation assay buffer and undiluted, mild lysis buffer. Genomic DNA of Escherichia coli was efficiently digested by rSlNucAs in the bacterial lysate. The results in this study suggest that rSlNucAs expressed by the Baculovirus-B. mori protein expression system will be a useful tool in molecular biology. Functional recombinant protein of bacteria was produced by Baculovirus-B. mori protein expression system. This system may be highly suitable for bacterial extracellular protein secreted via Sec pathway. PMID:27059494

  16. Ultra Deep Sequencing of a Baculovirus Population Reveals Widespread Genomic Variations

    Directory of Open Access Journals (Sweden)

    Aurélien Chateigner

    2015-07-01

    Full Text Available Viruses rely on widespread genetic variation and large population size for adaptation. Large DNA virus populations are thought to harbor little variation though natural populations may be polymorphic. To measure the genetic variation present in a dsDNA virus population, we deep sequenced a natural strain of the baculovirus Autographa californica multiple nucleopolyhedrovirus. With 124,221X average genome coverage of our 133,926 bp long consensus, we could detect low frequency mutations (0.025%. K-means clustering was used to classify the mutations in four categories according to their frequency in the population. We found 60 high frequency non-synonymous mutations under balancing selection distributed in all functional classes. These mutants could alter viral adaptation dynamics, either through competitive or synergistic processes. Lastly, we developed a technique for the delimitation of large deletions in next generation sequencing data. We found that large deletions occur along the entire viral genome, with hotspots located in homologous repeat regions (hrs. Present in 25.4% of the genomes, these deletion mutants presumably require functional complementation to complete their infection cycle. They might thus have a large impact on the fitness of the baculovirus population. Altogether, we found a wide breadth of genomic variation in the baculovirus population, suggesting it has high adaptive potential.

  17. Infection, transfection, and co-transfection of baculoviruses by microprojectile bombardment of larvae.

    Science.gov (United States)

    Obregón-Barboza, Verónica; Del Rincón-Castro, Ma Cristina; Cabrera-Ponce, José L; Ibarra, Jorge E

    2007-03-01

    The use of baculoviruses as expression vectors for heterologous proteins has been practically limited to the use of the Autographa californica multiple nucleopolyhedrovirus (AcMNPV). In this work, infection, transfection and co-transfection events with the baculoviruses AcMNPV and Trichoplusia ni granulovirus (TnGV) were accomplished by bombardment of T. ni first-instar larvae with microprojectiles coated with virions, viral DNA, and viral DNA and a transfer vector, respectively. A series of shooting conditions were tested until positive results were obtained. The use of 1.6 microm gold particles at 900 psi shooting pressure, 400 Torr vacuum, 7 cm distance to target, on sets of 20 first-instar larvae held in a 16 mm diameter container, proved to be the best shooting conditions. Typical infection symptoms were shown by larvae when shot with viruses or viral DNA from AcMNPV or TnGV. Co-transfected recombinant AcMNPV and TnGV were identified by the formation of occlusion bodies and GFP, respectively, in bombarded larvae. This technique opens a wide range of possibilities, not only to use an extensive number of baculoviruses as expression vectors for heterologous proteins, but also be used to infect, transfect or co-transfect a wide variety of viruses into animal cells. PMID:17184851

  18. Conservation of a proteinase cleavage site between an insect retrovirus (gypsy) Env protein and a baculovirus envelope fusion protein

    International Nuclear Information System (INIS)

    The predicted Env protein of insect retroviruses (errantiviruses) is related to the envelope fusion protein of a major division of the Baculoviridae. The highest degree of homology is found in a region that contains a furin cleavage site in the baculovirus proteins and an adjacent sequence that has the properties of a fusion peptide. In this investigation, the homologous region in the Env protein of the gypsy retrovirus of Drosophila melanogaster (DmegypV) was investigated. Alteration of the predicted DmegypV Env proteinase cleavage site from RIAR to AIAR significantly reduced cleavage of Env in both Spodoptera frugiperda (Sf-9) and D. melanogaster (S2) cell lines. When the predicted DmegypV Env cleavage site RIAR was substituted for the cleavage sequence RRKR in the Lymantria dispar nucleopolyhedrovirus fusion protein (LD130) sequence, cleavage of the hybrid LD130 molecules still occurred, although at a reduced level. The conserved 21-amino acid sequence just downstream of the cleavage site, which is thought to be the fusion peptide in LD130, was also characterized. When this sequence from DmegypV Env was substituted for the homologous sequence in LD130, cleavage still occurred, but no fusion was observed in either cell type. In addition, although a DmegypV-Env-green fluorescent protein construct localized to cell membranes, no cell fusion was observed

  19. Nuclear F-actin enhances the transcriptional activity of β-catenin by increasing its nuclear localization and binding to chromatin.

    Science.gov (United States)

    Yamazaki, Shota; Yamamoto, Koji; de Lanerolle, Primal; Harata, Masahiko

    2016-04-01

    Actin plays multiple roles both in the cytoplasm and in the nucleus. Cytoplasmic actin, in addition to its structural role in the cytoskeleton, also contributes to the subcellular localization of transcription factors by interacting with them or their partners. The transcriptional cofactor β-catenin, which acts as an intracellular transducer of canonical Wnt signaling, indirectly associates with the cytoplasmic filamentous actin (F-actin). Recently, it has been observed that F-actin is transiently formed within the nucleus in response to serum stimulation and integrin signaling, and also during gene reprogramming. Despite these earlier observations, information about the function of nuclear F-actin is poorly defined. Here, by facilitating the accumulation of nuclear actin artificially, we demonstrate that polymerizing nuclear actin enhanced the nuclear accumulation and transcriptional function of β-catenin. Our results also show that the nuclear F-actin colocalizes with β-catenin and enhances the binding of β-catenin to the downstream target genes of the Wnt/β-catenin signaling pathway, including the genes for the cell cycle regulators c-myc and cyclin D, and the OCT4 gene. Nuclear F-actin itself also associated with these genes. Since Wnt/β-catenin signaling has important roles in cell differentiation and pluripotency, our observations suggest that nuclear F-actin formed during these biological processes is involved in regulating Wnt/β-catenin signaling. PMID:26900020

  20. EGFR Signaling Regulates Maspin/SerpinB5 Phosphorylation and Nuclear Localization in Mammary Epithelial Cells

    Science.gov (United States)

    Reina, Jeffrey; Morais Freitas, Vanessa

    2016-01-01

    Maspin (SerpinB5) is a non-inhibitory serpin (serine protease inhibitor) with very diverse biological activities including regulation of cell adhesion, migration, death, control of gene expression and oxidative stress response. Initially described as a tumor and metastasis suppressor, clinical data brought controversies to the field, as some studies reported no correlation between SerpinB5 expression and prognosis value. These data underscore the importance of understanding SerpinB5 function in a normal physiological context and the molecular mechanism involved. Several SerpinB5 phosphoforms have been detected in different cell lines, but the signaling pathways involved and the biological significance of this post-translational modification in vivo remains to be explored. In this study we investigated SerpinB5 expression, subcellular localization and phosphorylation in different stages of the mouse mammary gland development and the signaling pathway involved. Here we show that SerpinB5 is first detected in late pregnancy, reaches its highest levels in lactation and remains at constant levels during post-lactational regression (involution). Using high resolution isoelectric focusing followed but immunoblot, we found at least 8 different phosphoforms of SerpinB5 during lactation, which decreases steadily at the onset of involution. In order to investigate the signaling pathway involved in SerpinB5 phosphorylation, we took advantage of the non-transformed MCF-10A model system, as we have previously observed SerpinB5 phosphorylation in these cells. We detected basal levels of SerpinB5 phosphorylation in serum- and growth factor-starved cells, which is due to amphiregulin autocrine activity on MCF-10A cells. EGF and TGF alpha, two other EGFR ligands, promote important SerpinB5 phosphorylation. Interestingly, EGF treatment is followed by SerpinB5 nuclear accumulation. Altogether, these data indicate that SerpinB5 expression and phosphorylation are developmentally

  1. A Study of Community Leaders in a Nuclear Host Community: Local Issues, Expectations and Support and Opposition.

    Science.gov (United States)

    Bronfman, B. H.

    As part of a continuing effort to assess the social impacts on communities of energy facility planning, construction, operation, and decommissioning, a May 1977 survey of 37 community leaders in Hartsville, Tennessee (site of a nuclear power plant) establishes major local issues (past, present, and future) which leaders feel are important to…

  2. Analytic approach to constructing effective local potentials in nuclear reaction theory

    International Nuclear Information System (INIS)

    Full text: Recently a method of constructing effective local potentials between strongly interacting composite particles was suggested [1]. The approach is based on the requirement of the proper structure of the nearest to the physical region singularities of scattering amplitudes as well as on the methods of the inverse scattering problem. In the given work the method is generalized to the case of the presence of the long-range Coulomb interaction which drastically changes the analytic structure of the S matrix. The effective potential is defined as the local operator which, being inserted into the Lippmann-Schwinger equation, generates required discontinuities of partial-wave scattering amplitudes. Its strong part is written in the form V(r)=∫μ0C(α)e-ar dα, where μ is determined by the position of the nearest to the physical region dynamical singularity. For all the processes considered, these singularities correspond to pole Feynman diagrams describing the elastic transfer mechanism. The C(α) function is found as a solution of the inverse scattering problem equations, the kernels of which are determined by the discontinuities at the nearest dynamical cuts. The Coulomb interaction is treated by introducing reduced Coulomb-nuclear scattering amplitudes and, in addition, by taking into account Coulomb corrections in the three-particle intermediate states and in the vertex functions of the pole diagrams. The processes of nd, pd, p3He, nα, pα and 3Heα scattering were considered. To calculate the required discontinuities, the information on the corresponding vertex constants and binding energies was used. Effective potentials, scattering lengths and low-energy phase shifts for the processes under consideration were obtained. The work was supported by the Russian Foundation for Basic Research (grant No.04-02-16602) and by the 'Russian Universities' program (grant No. 02.02.027)

  3. ANCLI's white book about the local governance of nuclear activities; Livre blanc de l'ANCLI sur la gouvernance locale des activites nucleaires

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2005-05-15

    This white book is the result of the work carried out by the local commissions of information about nuclear activities (CLI) with the objective of making a common status and recommendations about 4 main topics: the clear definition of the structure and juridical status of CLI's and the acknowledgement of their competence at the local scale, the role and participation of CLI's, and of the national association of CLI's (ANCLI), in the decision processes at the national and international scale, the access of local CLI actors to expertise, and the consideration of ANCLI's opinion in the solutions proposed for the management of radioactive wastes. This last point is taken as an illustration of the previous 3 topics. (J.S.)

  4. Cytomegalovirus Assembly Protein Precursor and Proteinase Precursor Contain Two Nuclear Localization Signals That Mediate Their Own Nuclear Translocation and That of the Major Capsid Protein

    OpenAIRE

    Plafker, Scott M.; Gibson, Wade

    1998-01-01

    The cytomegalovirus (CMV) assembly protein precursor (pAP) interacts with the major capsid protein (MCP), and this interaction is required for nuclear translocation of the MCP, which otherwise remains in the cytoplasm of transfected cells (L. J. Wood et al., J. Virol. 71:179–190, 1997). We have interpreted this finding to indicate that the CMV MCP lacks its own nuclear localization signal (NLS) and utilizes the pAP as an NLS-bearing escort into the nucleus. The CMV pAP amino acid sequence has...

  5. Basic amino acid residues located in the N-terminal region of BEND3 are essential for its nuclear localization

    International Nuclear Information System (INIS)

    BEN domain-containing protein 3 (BEND3) has recently been reported to function as a heterochromatin-associated protein in transcriptional repression in the nucleus. BEND3 should have nuclear localization signals (NLSs) to localize to the nucleus in light of its molecular weight, which is higher than that allowed to pass through nuclear pore complexes. We here analyzed the subcellular localization of deletion/site-directed mutants of human BEND3 by an immunofluorescence assay in an attempt to identify the amino acids essential for its nuclear localization. We found that three basic amino acid residues located in the N-terminal region of BEND3 (BEND356–58, KRK) are essential, suggesting that these residues play a role as a functional NLS. These results provide valuable information for progressing research on BEND3. - Highlights: • BEND3 localizes to the nucleus. • The N-terminal 60 amino acids region of BEND3 contains NLS. • Amino acids located between 56 and 58 of BEND3 (KRK) are part of NLS. • KRK motif is highly conserved among BEND3 homologs

  6. Basic amino acid residues located in the N-terminal region of BEND3 are essential for its nuclear localization

    Energy Technology Data Exchange (ETDEWEB)

    Shiheido, Hirokazu, E-mail: shiheido@ak.med.kyoto-u.ac.jp; Shimizu, Jun

    2015-02-20

    BEN domain-containing protein 3 (BEND3) has recently been reported to function as a heterochromatin-associated protein in transcriptional repression in the nucleus. BEND3 should have nuclear localization signals (NLSs) to localize to the nucleus in light of its molecular weight, which is higher than that allowed to pass through nuclear pore complexes. We here analyzed the subcellular localization of deletion/site-directed mutants of human BEND3 by an immunofluorescence assay in an attempt to identify the amino acids essential for its nuclear localization. We found that three basic amino acid residues located in the N-terminal region of BEND3 (BEND3{sub 56–58}, KRK) are essential, suggesting that these residues play a role as a functional NLS. These results provide valuable information for progressing research on BEND3. - Highlights: • BEND3 localizes to the nucleus. • The N-terminal 60 amino acids region of BEND3 contains NLS. • Amino acids located between 56 and 58 of BEND3 (KRK) are part of NLS. • KRK motif is highly conserved among BEND3 homologs.

  7. An analysis of inhabitants opinion on energy issues. Comparison between localities of nuclear installation sites and of urban communities

    International Nuclear Information System (INIS)

    For the purpose of comparing the mental attitudes toward nuclear power generation held by the public between localities containing nuclear installation sites and those of urban communities, a survey has been conducted by mail and by direct interviews. The survey conversed Fukushima, Niigata and Fukui Prefectures, representing the former group of localities, whereas Tokyo and Osaka, representing the latter. The mail survey revealed differences between the two groups of localities in their image of power and in their perception of its present status, as well as in their sense of values in daily life. The interview survey indicated that the factors behind the differences included regional characteristics such as temperaments of resident population, and effects on the private life style brought by nuclear installation. Narrowing down this gap of mental attitude between the two groups of localities should call indispensably for better mutual understanding. In bringing this about, consideration should be given to particular local characteristics, including popular mentality and personal view of life, as well as image and knowledge of energy sources held by the individuals. (author)

  8. Localization of hydrophobic ions in phospholipid bilayers using 1H nuclear Overhauser effect spectroscopy

    International Nuclear Information System (INIS)

    The binding location for the hydrophobic ions tetraphenylphosphonium (TPP+) and tetraphenylboron (TPB-) was studied in sonicated phosphatidylcholine (PC) vesicles by measuring time-dependent and steady-state intermolecular 1H nuclear Overhauser effects (NOE's). Intermolecular cross-relaxation was also investigated by two-dimensional NOE spectroscopy. Information on the distance and order parameter dependence of the NOE's was obtained from a simple simulation of the NOE's in the alkyl chain region. Taken together, the NOE data and the simulation provide strong evidence that TPB- and TPP+, at low concentrations (≤ 10 mol %), are localized in the alkyl chain region of the bilayer. At these lower concentrations of TPP+ or TPB-, no significant effect on lipid 13C T1 or T2 relaxation rates is detected. The proposed location is consistent with the expected free energy profiles for hydrophobic ions and with the carbonyl oxygens or interfacial water as the source of the membrane dipole potential. At higher ion/lipid ratios (≥ 20 mol %), TPB-/lipid NOE's increase. This results from a specific association of TPB- with the choline head group

  9. Importance weighting of local flux measurements to improve reactivity predictions in nuclear systems

    Energy Technology Data Exchange (ETDEWEB)

    Dulla, Sandra; Hoh, Siew Sin; Nervo, Marta; Ravetto, Piero [Politecnico di Torino, Dipt. Energia (Italy)

    2015-07-15

    The reactivity monitoring is a key aspect for the safe operation of nuclear reactors, especially for subcritical source-driven systems. Various methods are available for both, off-line and on-line reactivity determination from direct measurements carried out on the reactor. Usually the methods are based on the inverse point kinetic model applied to signals from neutron detectors and results may be severely affected by space and spectral effects. Such effects need to be compensated and correction procedures have to be applied. In this work, a new approach is proposed, by using the full information from different local measurements to generate a global signal through a proper weighting of the signals provided by single neutron detectors. A weighting techique based on the use of the adjoint flux proves to be efficient in improving the prediction capability of inverse techniques. The idea is applied to the recently developed algorithm, named MAρTA, that can be used in both off-line and online modes.

  10. Improved radiological/nuclear source localization in variable NORM background: An MLEM approach with segmentation data

    Science.gov (United States)

    Penny, Robert D.; Crowley, Tanya M.; Gardner, Barbara M.; Mandell, Myron J.; Guo, Yanlin; Haas, Eric B.; Knize, Duane J.; Kuharski, Robert A.; Ranta, Dale; Shyffer, Ryan; Labov, Simon; Nelson, Karl; Seilhan, Brandon; Valentine, John D.

    2015-06-01

    A novel approach and algorithm have been developed to rapidly detect and localize both moving and static radiological/nuclear (R/N) sources from an airborne platform. Current aerial systems with radiological sensors are limited in their ability to compensate for variable naturally occurring radioactive material (NORM) background. The proposed approach suppresses the effects of NORM background by incorporating additional information to segment the survey area into regions over which the background is likely to be uniform. The method produces pixelated Source Activity Maps (SAMs) of both target and background radionuclide activity over the survey area. The task of producing the SAMs requires (1) the development of a forward model which describes the transformation of radionuclide activity to detector measurements and (2) the solution of the associated inverse problem. The inverse problem is ill-posed as there are typically fewer measurements than unknowns. In addition the measurements are subject to Poisson statistical noise. The Maximum-Likelihood Expectation-Maximization (MLEM) algorithm is used to solve the inverse problem as it is well suited for under-determined problems corrupted by Poisson noise. A priori terrain information is incorporated to segment the reconstruction space into regions within which we constrain NORM background activity to be uniform. Descriptions of the algorithm and examples of performance with and without segmentation on simulated data are presented.

  11. Alteration of 'R7T7' type nuclear glasses: statistical approach, experimental validation, local evolution model

    International Nuclear Information System (INIS)

    The aim of this work is to propose an evolution of nuclear (R7T7-type) glass alteration modeling. The first part of this thesis is about development and validation of the 'r(t)' model. This model which predicts the decrease of alteration rates in confined conditions is based upon a coupling between a first-order dissolution law and a diffusion barrier effect of the alteration gel layer. The values and the uncertainties regarding the main adjustable parameters of the model (α, Dg and C*) have been determined from a systematic study of the available experimental data. A program called INVERSION has been written for this purpose. This work lead to characterize the validity domain of the 'r(t)' model and to parametrize it. Validation experiments have been undertaken, confirming the validity of the parametrization over 200 days. A new model is proposed in the second part of this thesis. It is based on an inhibition of glass dissolution reaction by silicon coupled with a local description of silicon retention in the alteration gel layer. This model predicts the evolutions of boron and silicon concentrations in solution as well as the concentrations and retention profiles in the gel layer. These predictions have been compared to measurements of retention profiles by the secondary ion mass spectrometry (SIMS) method. The model has been validated on fractions of gel layer which reactivity present low or moderate disparities. (author)

  12. Improved radiological/nuclear source localization in variable NORM background: An MLEM approach with segmentation data

    International Nuclear Information System (INIS)

    A novel approach and algorithm have been developed to rapidly detect and localize both moving and static radiological/nuclear (R/N) sources from an airborne platform. Current aerial systems with radiological sensors are limited in their ability to compensate for variable naturally occurring radioactive material (NORM) background. The proposed approach suppresses the effects of NORM background by incorporating additional information to segment the survey area into regions over which the background is likely to be uniform. The method produces pixelated Source Activity Maps (SAMs) of both target and background radionuclide activity over the survey area. The task of producing the SAMs requires (1) the development of a forward model which describes the transformation of radionuclide activity to detector measurements and (2) the solution of the associated inverse problem. The inverse problem is ill-posed as there are typically fewer measurements than unknowns. In addition the measurements are subject to Poisson statistical noise. The Maximum-Likelihood Expectation-Maximization (MLEM) algorithm is used to solve the inverse problem as it is well suited for under-determined problems corrupted by Poisson noise. A priori terrain information is incorporated to segment the reconstruction space into regions within which we constrain NORM background activity to be uniform. Descriptions of the algorithm and examples of performance with and without segmentation on simulated data are presented

  13. Local power peaking factor estimation in nuclear fuel by artificial neural networks

    International Nuclear Information System (INIS)

    This paper presents the training of an artificial neural network (ANN) to accurately predict, in very short time, a physical parameter used in nuclear fuel reactor optimization: the local power peaking factor (LPPF) in a typical boiling water reactor (BWR) fuel lattice. The ANN training patterns are distribution of fissile and burnable poison materials in the fuel lattice and their associated LPPF. These data were obtained by modeling the fuel lattices with a neutronic simulator: the HELIOS transport code. The combination of the pin U235 enrichment and the Gd2O3 (gadolinia) concentration, inside the 10 x 10 fuel lattice array, was encoded by three different methods. However, the only encoding method that was able to give a good prediction of the LPPF was the method which added the U235 enrichment and the gadolinia concentration. The results show that the relative error in the estimation of the LPPF, obtained by the trained ANN, ranged from 0.022% to 0.045%, with respect to the HELIOS results

  14. Local seismic network for monitoring of a potential nuclear power plant area

    Science.gov (United States)

    Tiira, Timo; Uski, Marja; Kortström, Jari; Kaisko, Outi; Korja, Annakaisa

    2016-04-01

    This study presents a plan for seismic monitoring of a region around a potential nuclear power plant. Seismic monitoring is needed to evaluate seismic risk. The International Atomic Energy Agency has set guidelines on seismic hazard evaluation and monitoring of such areas. According to these guidelines, we have made a plan for a local network of seismic stations to collect data for seismic source characterization and seismotectonic interpretations, as well as to monitor seismic activity and natural hazards. The detection and location capability of the network were simulated using different station configurations by computing spatial azimuthal coverages and detection threshold magnitudes. Background noise conditions around Pyhäjoki were analyzed by comparing data from different stations. The annual number of microearthquakes that should be detected with a dense local network centered around Pyhäjoki was estimated. The network should be dense enough to fulfill the requirements of azimuthal coverage better than 180° and automatic event location capability down to ML ˜ 0 within a distance of 25 km from the site. A network of 10 stations should be enough to reach these goals. With this setup, the detection threshold magnitudes are estimated to be ML = -0.1 and ML = 0.1 within a radius of 25 and 50 km from Pyhäjoki, respectively. The annual number of earthquakes detected by the network is estimated to be 2 (ML ≥ ˜ -0.1) within 25 km radius and 5 (ML ≥ ˜-0.1 to ˜0.1) within 50 km radius. The location accuracy within 25 km radius is estimated to be 1-2 and 4 km for horizontal coordinates and depth, respectively. Thus, the network is dense enough to map out capable faults with horizontal accuracy of 1-2 km within 25 km radius of the site. The estimation is based on the location accuracies of five existing networks in northern Europe. Local factors, such as seismic noise sources, geology and infrastructure might limit the station configuration and detection and

  15. 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 effects on the production of silk. In nature these

  16. Exact electronic and nuclear time-dependent potential energy surfaces for attosecond electron localization in the dissociation of H2+

    International Nuclear Information System (INIS)

    We study the electron localization dynamics in the dissociation of H2+ using a 1D model Hamiltonian. To this end we calculate the exact time-dependent potential energy surfaces (TDPES) both for the electron and for the nuclei. The exact electronic TDPES shows that the final electron localization is determined when the interatomic barrier becomes large and prohibits electron tunneling. The exact nuclear TDPES shows the mechanism of slowdown of the dissociation. It is found that the nuclear potential cannot be approximated by the weighted average of the quasi-static state potential energy surfaces, but can be approximated well by the transition between them. We show these two time-dependent potentials are the exact potential functionals of the time-dependent multicomponent density functional theory and can reproduce the whole phenomena of electron localization dynamics.

  17. Interleaved localized 1H/31P nuclear magnetic resonance spectroscopy of skeletal muscle

    International Nuclear Information System (INIS)

    Nuclear magnetic resonance (NMR) has been used as a spectroscopic method in physics and chemistry before it was developed to become a diagnostic imaging tool in medicine. When NMR spectroscopy is applied to human tissue, metabolism can be studied in normal physiological and pathological states in vivo. Metabolite concentrations and rates can be monitored dynamically and with localization of a defined region of interest. The 'window' which is opened for observation, i.e. which quantities are measured, depends on the nucleus used for RF excitation. Mechanisms of adenosine tri-phosphate (ATP) resynthesis, as a direct source of energy for muscle contraction, are phosphocreatine (PCr) splitting, glycolysis, beta-oxidation and, finally, oxidative phosphorylation. Whilst the dependency of these processes' fractional contribution to muscular energy supply on exercise type and duration is well known, quantitative models of the regulating mechanisms involved are still subject of current research. A large fraction of the established knowledge about metabolism is based on biochemical analysis of tissue acquired invasively (e.g. microdialysis and open-flow microperfusion) or representing averaged metabolic concentrations for the whole body (via serum metabolites or gas exchange analysis). Localized NMR spectroscopy, however, is capable of non-invasively acquiring time-resolved data from a defined volume of interest, in vivo. In contrast to the vast majority of MRS studies investigating metabolism, where spectra of a single nucleus (commonly 1H, 31P or 13C) were acquired or several MR spectra with different nuclei were measured in separate experiments, this work opens an additional 'window' on muscle metabolism by interleaved localized acquisition of 1H and 31P NMR spectra from human calf muscle in vivo, during rest, exercise and recovery, in a single experiment. Using this technique, the time courses of the concentrations of phosphocreatine, inorganic phosphate (Pi), ATP, total

  18. New nuclear data service at CNEA: retrieval of the update libraries from a local Web-Server

    International Nuclear Information System (INIS)

    A new On-line Nuclear Data Service was implemented at National Atomic Energy Commission (CNEA) Web-Site. The information usually issued by the Nuclear Data Section of IAEA (NDS-IAEA) on CD-ROM, as well as complementary libraries periodically downloaded from the a mirror server of NDS-IAEA Service located at IPEN, Brazil are available on the new CNEA Web page. In the site, users can find numerical data on neutron, charged-particle, and photonuclear reactions, nuclear structure, and decay data, with related bibliographic information. This data server is permanently maintained and updated by CNEA staff members. This crew also offers assistance on the use and retrieval of nuclear data to local users. (author)

  19. Allocation of responsibilities between central and local authorities concerning nuclear power plant licensing

    International Nuclear Information System (INIS)

    This paper examines Belgian regulations on licences to construct and operate nuclear power plants in the context of implementation of the 1980 Act concerning regionalisation. It also reviews the relevant nuclear legislation in certain other countries. (NEA)

  20. Identification of a nuclear localization motif in the serine/arginine protein kinase PSRPK of physarum polycephalum

    Directory of Open Access Journals (Sweden)

    Tian Shengli

    2009-08-01

    Full Text Available Abstract Background Serine/arginine (SR protein-specific kinases (SRPKs are conserved in a wide range of organisms, from humans to yeast. Studies showed that SRPKs can regulate the nuclear import of SR proteins in cytoplasm, and regulate the sub-localization of SR proteins in the nucleus. But no nuclear localization signal (NLS of SRPKs was found. We isolated an SRPK-like protein PSRPK (GenBank accession No. DQ140379 from Physarum polycephalum previously, and identified a NLS of PSRPK in this study. Results We carried out a thorough molecular dissection of the different domains of the PSRPK protein involved in its nuclear localization. By truncation of PSRPK protein, deletion of and single amino acid substitution in a putative NLS and transfection of mammalian cells, we observed the distribution of PSRPK fluorescent fusion protein in mammalian cells using confocal microscopy and found that the protein was mainly accumulated in the nucleus; this indicated that the motif contained a nuclear localization signal (NLS. Further investigation with truncated PSPRK peptides showed that the NLS (318PKKGDKYDKTD328 was localized in the alkaline Ω-loop of a helix-loop-helix motif (HLHM of the C-terminal conserved domain. If the 318PKKGDK322 sequence was deleted from the loop or K320 was mutated to T320, the PSRPK fluorescent fusion protein could not enter and accumulate in the nucleus. Conclusion This study demonstrated that the 318PKKGDKYDKTD328 peptides localized in the C-terminal conserved domain of PSRPK with the Ω-loop structure could play a crucial role in the NLS function of PSRPK.

  1. Baculovirus-mediated Expression of p35 Confers Resistance to Apoptosis in Human Embryo Kidney 293 cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Baculovirus has many advantages as vectors for gene transfer. We demonstrated that recombinant baculovirus vectors expressing p35 (Ac-CMV-p35) and eGFP (Ac-CMV-GFP) could be transduced into human kidney 293 cells efficiently. The level of transgene expression was viral dose dependent and high-level expression of the target gene could be achieved under the heterogonous promoter. MTT assay suggested that both Ac-CMV-p35 and Ac-CMV-GFP did not have cytotoxic effect on human embryo kidney 293 cells. Cell growth curve showed the Ac-CMV-p35 and Ac- CMV-GFP transduced and non-transduced cells had similar proliferation rate, so baculovirus-mediated p35expression had no adverse effect on cell proliferation. In addition, baculovirus-mediated p35 gene expression protected human embryo kidney 293 cells against apoptosis induced by various apoptosis inducers such as Actinomycin D, UV or serum-free media. These results suggested that the baculovirus vector mediated p35 gene expression was functional and it could be widely used in molecular research and even gene therapy.

  2. A novel bipartite nuclear localization signal guides BPM1 protein to nucleolus suggesting its Cullin3 independent function.

    Directory of Open Access Journals (Sweden)

    Dunja Leljak Levanić

    Full Text Available BPM1 belongs to the MATH-BTB family of proteins, which act as substrate-binding adaptors for the Cullin3-based E3 ubiquitin ligase. MATH-BTB proteins associate with Cullin3 via the BTB domain and with the substrate protein via the MATH domain. Few BPM1-interacting proteins with different functions are recognized, however, specific roles of BPM1, depending on its cellular localization have not been studied so far. Here, we found a novel bipartite nuclear localization signal at the C-terminus of the BPM1 protein, responsible for its nuclear and nucleolar localization and sufficient to drive the green fluorescent protein and cytoplasmic BPM4 protein into the nucleus. Co-localization analysis in live Nicotiana tabacum BY2 cells indicates a Cullin3 independent function since BPM1 localization is predominantly nucleolar and thus devoid of Cullin3. Treatment of BY2 cells with the proteasome inhibitor MG132 blocks BPM1 and Cullin3 degradation, suggesting turnover of both proteins through the ubiquitin-proteasome pathway. Possible roles of BPM1 in relation to its in vivo localization are discussed.

  3. Nuclear localization signal deletion mutants of lamin A and progerin reveal insights into lamin A processing and emerin targeting

    Science.gov (United States)

    Wu, Di; Flannery, Andrew R; Cai, Helen; Ko, Eunae; Cao, Kan

    2014-01-01

    Lamin A is a major component of the lamina, which creates a dynamic network underneath the nuclear envelope. Mutations in the lamin A gene (LMNA) cause severe genetic disorders, one of which is Hutchinson-Gilford progeria syndrome (HGPS), a disease triggered by a dominant mutant named progerin. Unlike the wild-type lamin A, whose farnesylated C-terminus is excised during post-translational processing, progerin retains its farnesyl tail and accumulates on the nuclear membrane, resulting in abnormal nuclear morphology during interphase. In addition, membrane-associated progerin forms visible cytoplasmic aggregates in mitosis. To examine the potential effects of cytoplasmic progerin, nuclear localization signal (NLS) deleted progerin and lamin A (PGΔNLS and LAΔNLS, respectively) have been constructed. We find that both ΔNLS mutants are farnesylated in the cytosol and associate with a sub-domain of the ER via their farnesyl tails. While the farnesylation on LAΔNLS can be gradually removed, which leads to its subsequent release from the ER into the cytoplasm, PGΔNLS remains permanently farnesylated and membrane-bounded. Moreover, both ΔNLS mutants dominantly affect emerin’s nuclear localization. These results reveal new insights into lamin A biogenesis and lamin A-emerin interaction. PMID:24637396

  4. Threats and benefits updated information on local opinions regarding the spent nuclear fuel repository in Finland - 16128

    International Nuclear Information System (INIS)

    The aim of the paper is to provide updated information on local opinion regarding the siting of a spent nuclear fuel repository in Finland. The main question is how the residents of the municipality perceive the threats and benefits of the repository. In accordance with the Decision in Principle by the Council of State passed in 2000, the Olkiluoto area in Municipality of Eurajoki was chosen as the location for the repository to accommodate spent nuclear fuel produced in Finland. Updated information on local opinions is needed as the siting process is approaching the next phase, the application for a construction license by 2012. The nuclear waste management company Posiva, owned by the utilities Teollisuuden Voima and Fortum Power and Heat, has also applied for a new Decision in Principle (DiP) for expansion of the repository. The data provided in this paper is based on a survey carried out in June 2008. The respondents were selected from the residents of the municipality of Eurajoki and the neighbouring municipalities using stratified random sampling (N=3000). The response rate of the survey was 20% (N=606). The paper is part of a joint research project between the University of Jyvaeskylae and the University of Tampere. The research project 'Follow-up research regarding socio-economic effects and communication of final disposal facility of spent nuclear fuel in Eurajoki and its neighbouring municipalities' is funded by the Finnish Research Programme on Nuclear Waste Management (KYT2010). (authors)

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

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

    International Nuclear Information System (INIS)

    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

  7. Identification and Characterization of Nuclear Localization Signals within the Nucleocapsid Protein VP15 of White Spot Syndrome Virus

    Institute of Scientific and Technical Information of China (English)

    Li-juan LI; Hua-jun ZHANG; Cong ZHANG; Zheng-li SHI

    2009-01-01

    The nucleocapsid protein VP15 of white spot syndrome virus (WSSV) is a basic DNA-binding protein. Three canonical bipartite nuclear localization signals (NLSs), called NLS1 (aa 11-27), NLS2 (aa 33-49) and NLS3 (44-60), have been detected in this protein, using the ScanProsite computer program. To determine the nuclear localization sequence of VP15, the full-length open reading frame, or the sequence of one of the three NLSs, was fused to the green fluorescent protein (GFP) gene, and transiently expressed in insect Sf9 cells. Transfection with full-length VP15 resulted in GFP fluorescence being distributed exclusively in the nucleus. NLS 1 alone could also direct GFP to the nucleus, but less efficiently. Neither of the other two NLSs (NLS2 and 3) was functional when expressed alone, but exhibited similar activity to NLS1 when they were expressed as a fusion peptide. Furthermore, a mutated VP15, in which the two basic amino acids (11RR12) of NLSI were changed to two alanines (11AA12), caused GFP to be localized only in the cytoplasm of Sf9 cells. These results demonstrated that VP15, as a nuclear localization protein, needs cooperation between its three NLSs, and that the two residues (11RR12) of NLS1 play a key role in transporting the protein to the nucleus.

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

    Directory of Open Access Journals (Sweden)

    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.

  9. Nuclear Quadrupole Resonance Study of the Nitrogen Mustards and Local Anesthetics.

    Science.gov (United States)

    Buess, Michael Lee

    The density matrix description of pulsed nitrogen -14 nuclear quadrupole resonance (NQR) spin-echoes is presented. The parallel between this problem, when formulated in terms of the fictitious spin- 1/2 operators, and that of spin - 1/2 NMR spin-echoes in liquids is discussed along with the complications which arise in multiple-pulse NQR experiments in powders due to the random orientation of the electric field gradient tensors. The equipment and procedures involved in searching for, detecting and identifying NQR resonances using pulsed techniques are described. The ('14)N NQR spectra of several nitrogen mustard compounds in the solid state are reported and analyzed in the framework of the Townes and Dailey theory. For the aniline derivatives, a correlation exists between l -(sigma), l being the nitrogen lone-pair electron density and (sigma) the average N-C sigma bond electron density, and the enhanced Hammett sigma constant (sigma)('-). An improved correlation is obtained between l-(sigma) and (sigma)(,R)('-), which emphasizes the importance of resonance effects in determining l-(sigma). The increase of hydrolysis and alkylation rates with increasing values of l-(sigma) is in agreement with the identification of the cyclic immonium ion as the intermediate in the hydrolysis and alkylation processes of the aromatic nitrogen mustards. A possible correlation is noted between the ('35)Cl NQR spectra for some of the mustards and measures of toxic and antitumor activity. ('14)N NQR spectra for several local anesthetics in the solid state are also reported and analyzed using the Townes and Dailey approach. The changes in the electron distributions at various nitrogen sites, produced by protonating the tertiary amino nitrogen, are discussed and shown to be in general agreement with expectations bases on the increased electrophilic character of the protonated amino group.

  10. The prevention of the local nuclear accidents in the Republic of Moldova

    International Nuclear Information System (INIS)

    Although there are no nuclear reactors in Moldova, there are numerous radiation sources situated in a few waste disposal sites as well as the sources applied in medicine, science, education, industry, agriculture, which demand serious concern from safety and radiation protection point of view. Under cooperation with IAEA national Regulatory Control in the Field of Nuclear Protection and Safety was established since 1993. A number of governmental regulatory bodies supervise the following activities: radiation standardization; radiological supervision of sources storage, exploitation and disposal; radiological monitoring of radioactive substances, food products, building materials; supervision of personnel exposure and environmental exposure; investigation of radiological accidents; etc. In 1998, Moldova has joined The International Convention for early Notification of Nuclear Accidents; The Convention on Nuclear Safety; The Convention on Assistance in Case of Nuclear Accident of Radiological Emergency and The Convention on the Physical protection of Nuclear Material

  11. Nuclear β-catenin localization supports homology of feathers, avian scutate scales, and alligator scales in early development.

    Science.gov (United States)

    Musser, Jacob M; Wagner, Günter P; Prum, Richard O

    2015-01-01

    Feathers are an evolutionary novelty found in all extant birds. Despite recent progress investigating feather development and a revolution in dinosaur paleontology, the relationship of feathers to other amniote skin appendages, particularly reptile scales, remains unclear. Disagreement arises primarily from the observation that feathers and avian scutate scales exhibit an anatomical placode-defined as an epidermal thickening-in early development, whereas alligator and other avian scales do not. To investigate the homology of feathers and archosaur scales we examined patterns of nuclear β-catenin localization during early development of feathers and different bird and alligator scales. In birds, nuclear β-catenin is first localized to the feather placode, and then exhibits a dynamic pattern of localization in both epidermis and dermis of the feather bud. We found that asymmetric avian scutate scales and alligator scales share similar patterns of nuclear β-catenin localization with feathers. This supports the hypothesis that feathers, scutate scales, and alligator scales are homologous during early developmental stages, and are derived from early developmental stages of an asymmetric scale present in the archosaur ancestor. Furthermore, given that the earliest stage of β-catenin localization in feathers and archosaur scales is also found in placodes of several mammalian skin appendages, including hair and mammary glands, we hypothesize that a common skin appendage placode originated in the common ancestor of all amniotes. We suggest a skin placode should not be defined by anatomical features, but as a local, organized molecular signaling center from which an epidermal appendage develops. PMID:25963196

  12. Construction and immunogenicity of recombinant pseudotype baculovirus expressing the capsid protein of porcine circovirus type 2 in mice.

    Science.gov (United States)

    Fan, Huiying; Pan, Yongfei; Fang, Liurong; Wang, Dang; Wang, Shengping; Jiang, Yunbo; Chen, Huanchun; Xiao, Shaobo

    2008-06-01

    Baculovirus has emerged recently as a novel and attractive gene delivery vehicle for mammalian cells. Porcine circovirus type 2 (PCV2) is known to be associated with post-weaning multisystemic wasting syndrome (PMWS), an emerging swine disease which results in tremendous economic losses. In this study, baculovirus pseudotyped with vesicular stomatitis virus glycoprotein (VSV-G) was used as a vector to express capsid (Cap) protein, the most important immunogen of PCV2, under the transcriptional control of cytomegalovirus immediate early (CMV-IE) enhancer/promoter. The resultant recombinant baculovirus (BV-G-ORF2) efficiently transduced and expressed the Cap protein in mammalian cells, as demonstrated by Western blot and flow cytometric analyses. After direct vaccination with 1x10(8) or 1x10(9)plaque forming units (PFU)/mouse of BV-G-ORF2, significant PCV2-specific ELISA antibodies, neutralizing antibodies, as well as cellular immune responses could be induced in mice. BV-G-ORF2 exhibited better immunogenicity than a DNA vaccine encoding the Cap protein, even at a dose of 1x10(8)PFU/mouse. Taken together, the improved immunogenicity of BV-G-ORF2, together with the unique advantages of pseudotype baculovirus, including easy manipulation, simple scale-up, lack of toxicity, and no pre-existing antibody against baculovirus in the hosts, indicate that pseudotype baculovirus-mediated gene delivery can be utilized as an alternative strategy to develop a new generation of vaccines against PCV2 infection. PMID:18394722

  13. Construction of recombinant baculovirus Ac-CMV-hSox9 for gene therapy of intervertebral disc degeneration

    Institute of Scientific and Technical Information of China (English)

    LIU Xiao-yun; YANG Shu-hua; LIANG Chang-yong; SONG Jian-hua; LI Kang-hua; CHEN Xin-wen

    2007-01-01

    Objective: To construct the recombinant baculovirus Ac-cytomegalovirus (CMV)-hSox9 for gene therapy of intervertebral disc degeneration. Methods: Bac-to-Bac system was used for the construction of baculovirus Ac-CMV-hSox9. The cDNA of hSox9 was first cloned into a plasmid vector under the control of CMV promotor to generate the donor plasmid pFastBacDul-green fluorescene protein (GFP)-CMV (pFGC)-hSox9.The resultant plasmid was transformed into DH10Bac cells and then the transformation mixture was spread on Luria-Bertani (LB) agarose culture medium containing isopropyl-β-D-thiogalactoside (IPTG), X-gal, gentamicin, kanamycin and tetracycline.The white colonies were selected and cultured for amplification, and the hSox9Bacmid DNA was extracted. After verification, recombinant baculovirus Ac-CMV-hSox9 was obtained through transfecting Sf 21 cells.The expression of hSox9 gene in the intervertebral disc cells in rabbits was determined by Western blotting and immunohistochemical staining.Results: Polymerase chain reaction (PCR) confirmed the presence of hSox9 gene in the recombinant baculovirus and the Sf 21 cells transfected by the baculovirus showed the expression of fluorescence protein.Western blotting and immunohistochemical staining analysis indicated that exogenous hSox9 gene was expressed in the disc cells.Conclusions: The successful construction of the recombinant baculovirus Ac-CMV-hSox9 and the confirmation of the target gene expression provides a novel expression vector system for basic research and clinical treatment of intervertebral degenerative disc disease.

  14. Interstitial tissue-specific gene expression in mouse testis by intra-tunica albuguineal injection of recombinant baculovirus

    Institute of Scientific and Technical Information of China (English)

    Hyun Jung Park; Won Young Lee; Jin Hoi Kim; Jae Hwan Kim; Hun Jong Jung; Nam Hyung Kim; Bo Kyung Kim; Hyuk Song

    2009-01-01

    The purpose of this study is to establish a gene delivery system for interstitial tissue-specific protein expression in mice testes using modified recombinant baculovirus. Green fluorescent protein (GFP)-expressing recombinant bacuiovirus (GFP-baculovirus), in which the insect cell-specific polyhedron promoter was replaced by the cytomegalovirus (CMV)-IE promoter, was used to transfect testicular cells in vitro, and for intra-tunica albuguineai injection of the interstitial tissue of the testis. GFP expression was monitored in frozen testes sections by fluorescence microscopy. Expression of GFP in testicular tissues was also assessed by reverse transcription polymerase chain reaction (RT-PCR), and protein expression was assessed by Western blot. Testicular cells in vitro were infected efficiently by modified recombinant GFP-baculovirus. Intra-tunica albuguineal injection of GFP-baculovirus into the mouse testis resulted in a high level of GFP expression in the interstitial tissues. RT-PCR analysis clearly showed GFP gene expression in the testis, particularly interstitial tissues. Intra-tunica albuguineal injection of a modified baculovirus that encoded recombinant rat insulin-like growth factor binding protein (IGFBP)-5 resulted in an increase in IGFBP-5 in testis and semen. In conclusion, we have developed an efficient delivery system for gene expression in vivo in testicular cells, particularly cells of the interstitial tissue using intra-tunica albuguineal injection of a modified recombinant baculovirus. This method will be particularly relevant for application that requires gene delivery and protein expression in the testicular cells of the outer seminiferous tubule of the testis.

  15. Baculovirus Coinfection Strategy for Improved Galactosylation of Recombinant Glycoprotein Produced by Insect Cell Culture

    Science.gov (United States)

    Ney, Yap Wei; Rahman, Badarulhisam Abdul; Aziz, Azila Abdul

    Baculovirus Expression Vector System (BEVS) is widely used for the production of recombinant glycoproteins, but it is not ideal for pharmaceutical glycoprotein production due to incomplete glycosylation. The factors that ensure successful glycosylation are the presence of sufficient amount of glycosyltransferases, sugar nucleotides as the substrate donor and the recombinant protein as the substrate acceptor. In this study, we analyzed the galactosylation process by the introduction of ß-1,4galactosyltransferase (ß-1,4GalT) as the glycosyltransferase of interest and uridine-5`-diphosphogalactose (UDP-Gal) as the substrate donor. Recombinant human transferrin (rhTf) as a model protein was used as the substrate acceptor. Insect cell lines have been reported to produce a small amount of ß-1,4GalT and thus insufficient for effective galactosylation. In this study, we developed a method to produce galactosylated rhTf and optimized the expression of rhTf with better N-glycan quality. Recombinant ß-1,4GalT was introduced during protein expression by the coinfection of the BEVS with baculovirus carrying bovine ß-1,4GalT. To evaluate the extent of galactosylation by the coinfection strategy, a binding assay was established. In this binding assay, glycoprotein acceptor was absorbed onto ELISA plate surface. A lectin known as Ricinus communis agglutinin-I (RCA-I) labeled with peroxidase, was added and allowed to recognize Gal ß1>4GlcNAc group on the N-glycan of the glycoprotein, followed by appropriate color reaction measurements. Coexpression between rhTf and ß-1,4GalT did not show encouraging results due to the reduction of UDP-Gal upon baculovirus infection. This interesting finding suggested that the introduction of ß-1,4GalT alone was not sufficient for successful galactosylation. Alternatively, post harvest glycosylation method strategy seems to be a promising technique in the improvement of glycoprotein quality.

  16. Proteomic analyses of baculovirus Anticarsia gemmatalis multiple nucleopolyhedrovirus budded and occluded virus.

    Science.gov (United States)

    Braconi, Carla Torres; Ardisson-Araújo, Daniel Mendes Pereira; Paes Leme, Adriana Franco; Oliveira, Juliana Velasco de Castro; Pauletti, Bianca Alves; Garcia-Maruniak, Alejandra; Ribeiro, Bergmann Morais; Maruniak, James E; Zanotto, Paolo Marinho de Andrade

    2014-04-01

    Baculoviruses infect insects, producing two distinct phenotypes during the viral life cycle: the budded virus (BV) and the occlusion-derived virus (ODV) for intra- and inter-host spread, respectively. Since the 1980s, several countries have been using Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) as a biological control agent against the velvet bean caterpillar, A. gemmatalis. The genome of AgMNPV isolate 2D (AgMNPV-2D) carries at least 152 potential genes, with 24 that possibly code for structural proteins. Proteomic studies have been carried out on a few baculoviruses, with six ODV and two BV proteomes completed so far. Moreover, there are limited data on virion proteins carried by AgMNPV-2D. Therefore, structural proteins of AgMNPV-2D were analysed by MALDI- quadrupole-TOF and liquid chromatography MS/MS. A total of 44 proteins were associated with the ODV and 33 with the BV of AgMNPV-2D. Although 38 structural proteins were already known, we found six new proteins in the ODV and seven new proteins carried by the AgMNPV-2D BV. Eleven cellular proteins that were found on several other enveloped viruses were also identified, which are possibly carried with the virion. These findings may provide novel insights into baculovirus biology and their host interaction. Moreover, our data may be helpful in subsequent applied studies aiming to improve AgMNPV use as a biopesticide and a biotechnology tool for gene expression or delivery. PMID:24443474

  17. Construction of Recombinant Baculovirus Surface-Displayed the Capsid Protein of Porcine Circovirus Type 2%表面展示猪圆环病毒2型衣壳蛋白的重组杆状病毒的构建

    Institute of Scientific and Technical Information of China (English)

    程晓亮; 林文耀; 叶煜; 陈筱薇; 严常燕; 廖明; 樊惠英

    2011-01-01

    利用PCR方法扩增猪圆环病毒2型核定位信号区缺失的Cap蛋白基因,将其亚克隆入杆状病毒表面展示质粒pBACsurf-1的gp64信号肽和gp64成熟蛋白之间.将此融合片段亚克隆到质粒pcDNA3.1(+),获得重组质粒pcDNA3.1-gp64-dCap.然后将含有CMV启动子,gp64及dCap的基因片段克隆到杆状病毒转移质粒pFastBac-V,得到重组质粒pFastBac-dCap-V.然后将此转化DH10Bac感受态细胞,获得的重组穿梭质粒Bacmid-dCap-V,经脂质体转染St9细胞,获得重组杆状病毒Ac-dCap-V.该重组病毒感染Sf9细胞后可以产生典型的细胞病变,转导哺乳动物细胞后经间接免疫荧光试验证明,该重组杆状病毒成功转导哺乳动物细胞并表达dCap蛋白.%The open capsid(Cap)protein gene without nuclear localization signal (NLS) of porcine circo-virus 2(PCV2)was amplified by PCR and sub-cloned into pBACsurf-1 between the upstream gp64 signal sequence and downstresm gp64 mature domain. The fusion gene containing dCap and gp64 was inserted into pcDNA3.1 ( + ) to construct the recombinant plasmid pcDNA3. L-gp64-dCap. A fragment of the CMV promoter-gp64-dCap fusion gene cassette was excised from pcDNA3. L-gp64-dCap and inserted into the baculovirus transfer vector pFastBac-V to construct the recombinant plasmid pFastBac-dCap-V. It was followed by the transformation of plasmid into Escherichia coli DHlOBac competent cells to obtain the recombinant shuttle vector Bacmid-dCap-V. Eventually, the recombinant Bacmid was transfected into Sf9 cells to produce the recombinant baculovirus Ac-dCap-V using the LipofectamineTM 2000. The Sf9 cells produced cytopathic after infection of recombinant baculoviruses Ac-dCap-V. Indirect immunofluoresent assay demonstrated that the Ac-dCap-V efficiently transducted the mammalian cells in vitro and the dCap protein was expressed successfully.

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

    International Nuclear Information System (INIS)

    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

  19. Molecular characterization and baculovirus expression of the glycoprotein B of a seal herpesvirus (phocid herpesvirus-1).

    Science.gov (United States)

    Harder, T C; Osterhaus, A D

    1997-01-20

    A glycoprotein B (gB) gene homologue was identified in a 5.4-kb BamHl genomic fragment of the phocid herpesvirus type-1 (PhHV-1) which represents a widespread and important pathogen of pinnipeds. Sequence analysis revealed a gB-specific open-reading frame comprising 881 amino acids. Phylogenetic analysis gave evidence for a close evolutionary relationship between PhHV-1 and members of the Varicellovirus genus of the alpha-Herpesvirinae and canid herpesvirus in particular. In PhHV-1-infected Crandell feline kidney cells gB is expressed as a 113-kDa glycosylated molecule which is proteolytically cleaved into at least two fragments of 67 and 53-59 kDa apparently forming disulfide-linked heterodimers of 140 kDa. Cell surface expression of PhHV-1 gB was confirmed by FACS analysis. Thus, synthesis and processing of the gB protein of PhHV-1 follows a pattern also observed in other Varicelloviruses. Since the gB protein of herpesviruses, expressed in the baculovirus system, has been shown to be a suitable target for vaccine design, we used this system for expression of PhHV-1 gB. Recombinant (rec) baculovirus-expressed gB was identified as a 105-kDa glycosylated molecule. Proteolytic cleavage into fragments of 62 and 52 kDa was markedly delayed compared to wild-type (wt) gB. Wt and rec gB harbored endoglycosidase H (precursor)- as well as N-glycosidase F-sensitive N-glycans (proteolytic fragments). Baculovirus-expressed gB appeared to be antigenically authentic, since it was recognized in radioimmunoprecipitation and immune peroxidase monolayer assays by PhHV-1-neutralizing seal sera and by gB-specific neutralizing murine monoclonal antibodies. Furthermore, PhHV-1-neutralizing antibodies were induced in mice following immunization with baculovirus-expressed gB, indicating its suitability for incorporation in a candidate vaccine for seals. PMID:9018133

  20. High-yield production of canine parvovirus virus-like particles in a baculovirus expression system.

    Science.gov (United States)

    Jin, Hongli; Xia, Xiaohong; Liu, Bing; Fu, Yu; Chen, Xianping; Wang, Huihui; Xia, Zhenqiang

    2016-03-01

    An optimized VP2 gene from the current prevalent CPV strain (new CPV-2a) in China was expressed in a baculovirus expression system. It was found that the VP2 proteins assembled into virus-like particles (VLPs) with antigenic properties similar to those of natural CPV and with an especially high hemagglutination (HA) titer (1:2(20)). Dogs intramuscularly or orally immunized with VLPs produced antibodies against CPV with >1:80 hemagglutination inhibition (HI) units for at least 3 months. The CPV VLPs could be considered for use as a vaccine against CPV or as a platform for research on chimeric VLP vaccines against other diseases. PMID:26666439

  1. Multi-region fuzzy logic controller with local PID controllers for U-tube steam generator in nuclear power plant

    OpenAIRE

    Puchalski Bartosz; Duzinkiewicz Kazimierz; Rutkowski Tomasz

    2015-01-01

    In the paper, analysis of multi-region fuzzy logic controller with local PID controllers for steam generator of pressurized water reactor (PWR) working in wide range of thermal power changes is presented. The U-tube steam generator has a nonlinear dynamics depending on thermal power transferred from coolant of the primary loop of the PWR plant. Control of water level in the steam generator conducted by a traditional PID controller which is designed for nominal power level of the nuclear react...

  2. Identification and characterization of a nuclear localization signal of TRIM28 that overlaps with the HP1 box

    Energy Technology Data Exchange (ETDEWEB)

    Moriyama, Tetsuji; Sangel, Percival [Laboratory of Nuclear Transport Dynamics, National Institutes of Biomedical Innovation, Health and Nutrition (NIBIOHN), Osaka 567-0085 (Japan); Yamaguchi, Hiroki [School of Medicine, Osaka University, Osaka 565-0871 (Japan); Obuse, Chikashi [Graduate School of Life Science, Hokkaido University, Sapporo 001-0021 (Japan); Miyamoto, Yoichi [Laboratory of Nuclear Transport Dynamics, National Institutes of Biomedical Innovation, Health and Nutrition (NIBIOHN), Osaka 567-0085 (Japan); Oka, Masahiro, E-mail: moka@nibiohn.go.jp [Laboratory of Nuclear Transport Dynamics, National Institutes of Biomedical Innovation, Health and Nutrition (NIBIOHN), Osaka 567-0085 (Japan); Laboratory of Biomedical Innovation, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan); Yoneda, Yoshihiro, E-mail: y-yoneda@nibiohn.go.jp [National Institutes of Biomedical Innovation, Health and Nutrition (NIBIOHN), Osaka 567-0085 (Japan); Laboratory of Biomedical Innovation, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871 (Japan)

    2015-07-03

    Tripartite motif-containing 28 (TRIM28) is a transcription regulator, which forms a repressor complex containing heterochromatin protein 1 (HP1). Here, we report identification of a nuclear localization signal (NLS) within the 462-494 amino acid region of TRIM28 that overlaps with its HP1 binding site, HP1 box. GST-pulldown experiments revealed the interaction of the arginine-rich TRIM28 NLS with various importin α subtypes (α1, α2 and α4). In vitro transport assay demonstrated that nuclear localization of GFP-TRIM28 NLS is mediated by importin αs, in conjunction with importin β1 and Ran. Further, we demonstrated that HP1 and importin αs compete for binding to TRIM28. Together, our findings suggest that importin α has an essential role in the nuclear delivery and preferential HP1 interaction of TRIM28. - Highlights: • TRIM28 contains an NLS within the 462-494 amino acid region. • The nuclear import of TRIM28 is mediated by importin α/importin β1. • TRIM28 NLS overlaps with HP1 Box. • HP1 and importin α compete for binding to TRIM28.

  3. Identification and characterization of a nuclear localization signal of TRIM28 that overlaps with the HP1 box

    International Nuclear Information System (INIS)

    Tripartite motif-containing 28 (TRIM28) is a transcription regulator, which forms a repressor complex containing heterochromatin protein 1 (HP1). Here, we report identification of a nuclear localization signal (NLS) within the 462-494 amino acid region of TRIM28 that overlaps with its HP1 binding site, HP1 box. GST-pulldown experiments revealed the interaction of the arginine-rich TRIM28 NLS with various importin α subtypes (α1, α2 and α4). In vitro transport assay demonstrated that nuclear localization of GFP-TRIM28 NLS is mediated by importin αs, in conjunction with importin β1 and Ran. Further, we demonstrated that HP1 and importin αs compete for binding to TRIM28. Together, our findings suggest that importin α has an essential role in the nuclear delivery and preferential HP1 interaction of TRIM28. - Highlights: • TRIM28 contains an NLS within the 462-494 amino acid region. • The nuclear import of TRIM28 is mediated by importin α/importin β1. • TRIM28 NLS overlaps with HP1 Box. • HP1 and importin α compete for binding to TRIM28

  4. Nuclear and nucleolar localization signals and their targeting function in phosphatidylinositol 4-kinase PI4K230

    International Nuclear Information System (INIS)

    PI4K230, an isoform of phosphatidylinositol 4-kinase, known primarily as a cytoplasmic membrane-bound enzyme, was detected recently also in the nucleolus of several cells. Here we provide mechanistic insight on the targeting function of its putative nuclear localization signal (NLS) sequences using molecular modeling, digitonin-permeabilized HeLa cells and binding to various importins. The synthetic sequence 916NFNHIHKRIRRVADKYLSG934 comprising a putative monopartite NLS (NLS1), targeted covalently bound fluorescent BSA to the nucleoplasm via classical importin α/β mechanism employing importins α1 and α3 but not α5. This transport was inhibited by wheat germ agglutinin and GTPγS. The sequence 1414SKKTNRGSQLHKYYMKRRTL1433, a putative bipartite NLS (NLS2) proved ineffective in nuclear targeting if conjugated to fluorescently labeled BSA. Nonetheless, NLS2 or either of its basic clusters directed to the nucleolus soybean trypsin inhibitor that can pass the nuclear pore complex passively; moreover, an expressed 58 kDa fragment of PI4K230 (AA1166-1667) comprising NLS2 was also imported into the nucleus by import factors of reticulocyte lysate or by importin α1/β or α3/β complexes and localized to the nucleolus. We conclude that the putative bipartite NLS itself is a nucleolar targeting signal, and for nuclear import PI4K230 requires a larger sequence around it or, alternatively, the monopartite NLS

  5. Importance of nuclear localization for the apoptosis-induced activity of a fungal galectin AAL (Agrocybe aegerita lectin)

    International Nuclear Information System (INIS)

    Agrocybe aegerita lectin (AAL) was identified previously in our group as a novel galectin from medicinal fungi Agrocybe aegerita, and has been shown to effectively induce cancer cell cycle arrest and apoptosis in vitro and tumor regression in vivo. Here, AAL was observed to translocate into the HeLa cell nucleus and induce cell apoptosis when it was predominantly in the nucleus. The N-terminus and C-terminus of AAL were required for nuclear localization. Site mutated proteins were generated based on AAL structure. Dimer interface mutant I25G, carbohydrate recognition domain (CRD) mutant R63H, and loop region mutant L33A could not enter the nucleus and lost the ability to induce apoptosis. CRD mutant H59Q and loop region mutant I144G maintained nuclear localization activity, and H59Q retained residual bioability but I144G had no activity, indicating that nuclear localization is important but not sufficient for AAL to become apoptotically active. Our findings provide a novel antitumor mechanism of fungal galectin.

  6. Down-regulation of β-catenin Nuclear Localization by Aspirin Correlates with Growth Inhibition of Jurkat Cell Line

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    In this study, we examined the effects of aspirin on the growth rates, subcellar distribution of β-catenin protein, the expression of β-catenin/TCF signaling pathway target gene cyclinD1 mRNA,and cell cycle of Jurkat cell line (Human T-acute lymphoblastic leukemia). Our results showed that the treatment with aspirin inhibited the growth of Jurkat cell line. Jurkat cells treated with 3 mmol/L of aspirin could significantly decrease nuclear localization of β-catenin, and at 5 mmol/L of aspirin,the nuclear localization of β-catenin was undetectable. QRT-PCR showed that the target gene cyclinD1 mRNA expression was gradually decreased with the dosage of aspirin. Aspirin induced G0/G1cell cycle arrest in Jurkat cells. We are led to conclude that aspirin acts through β-catenin-independent mechanisms. The effects of aspirin include down-regulation of β-catenin nuclear localization and G0/G1 cell cycle arrest, which might serve as a means of growth inhibition in aspirin-treated human Jurkat cell line.

  7. Elevated expression of fatty acid synthase and nuclear localization of carnitine palmitoyltransferase 1C are common among human gliomas.

    Science.gov (United States)

    Wakamiya, Tomihiro; Suzuki, Satoshi O; Hamasaki, Hideomi; Honda, Hiroyuki; Mizoguchi, Masahiro; Yoshimoto, Koji; Iwaki, Toru

    2014-10-01

    Fatty acid synthase (FASN) and carnitine palmitoyltransferase 1C (CPT1C), a brain-specific isoform of the CPT1 family, are upregulated in certain types of cancers, including gliomas. Acetyl-CoA carboxylase (ACC) catalyzes the carboxylation of acetyl-CoA to malonyl-CoA, the rate-limiting step in fatty acid synthesis, and its phosphorylated form inhibits lipid synthesis. We examined the expression and subcellular localization of these fatty acid metabolism-related molecules in human gliomas. We performed immunostaining of two glioma cell lines (U373MG and U87MG) and 41 surgical specimens of diffuse gliomas with various histological grades (21 with the isocitrate dehydrogenase 1(IDH1) R132H mutation and 20 without the mutation). In the cultured glioma cells, CPT1C and phosphorylated ACC (p-ACC) were mainly localized to the nuclei, whereas FASN localized to the cytoplasm. In the surgical specimens, most glioma tissues showed nuclear staining for CPT1C and p-ACC, and cytoplasmic staining for FASN, regardless of the genetic status of IDH1 and the histological grade. Therefore, elevated cytoplasmic expression of FASN and nuclear localization of CPT1C are common among human diffuse gliomas, which may be regulated by the differential phosphorylation status of ACC in the cellular compartment. PMID:24984811

  8. Transcription-dependent nucleolar cap localization and possible nuclear function of DExH RNA helicase RHAU

    International Nuclear Information System (INIS)

    RHAU (RNA helicase associated with AU-rich element) is a DExH protein originally identified as a factor accelerating AU-rich element-mediated mRNA degradation. The discovery that RHAU is predominantly localized in the nucleus, despite mRNA degradation occurring in the cytoplasm, prompted us to consider the nuclear functions of RHAU. In HeLa cells, RHAU was found to be localized throughout the nucleoplasm with some concentrated in nuclear speckles. Transcriptional arrest altered the localization to nucleolar caps, where RHAU is closely localized with RNA helicases p68 and p72, suggesting that RHAU is involved in transcription-related RNA metabolism in the nucleus. To see whether RHAU affects global gene expression transcriptionally or posttranscriptionally, we performed microarray analysis using total RNA from RHAU-depleted HeLa cell lines, measuring both steady-state mRNA levels and mRNA half-lives by actinomycin D chase. There was no change in the half-lives of most transcripts whose steady-state levels were affected by RHAU knockdown, suggesting that these transcripts are subjected to transcriptional regulation. We propose that RHAU has a dual function, being involved in both the synthesis and degradation of mRNA in different subcellular compartments

  9. Issues associated with assessing nuclear localization of N-terminally unphosphorylated β-catenin with monoclonal antibody 8E7

    Directory of Open Access Journals (Sweden)

    Russell Susan

    2009-02-01

    Full Text Available Abstract Background β-catenin is a dual function adhesion/transcriptional co-activator protein, and both functions are critical for normal tissue homeostasis. Since the transcriptional functions of β-catenin are more often implicated in various disease processes, there is much interest in the development and use of reagents to interrogate spatial and temporal evidence of β-catenin nuclear signaling in cells and tissues. An important study demonstrated that the signaling form of β-catenin is specifically unphosphorylated at residues S37 and T41, and suggested that this form exhibits a propensity for cytosolic/nuclear accumulation relative to the total pool of β-catenin. Results We show that monoclonal antibody, 8E7, which recognizes the signaling form of β-catenin specifically unphosphorylated at S37 and T41 (Active B-Catenin, ABC, also cross-reacts with a widely expressed, variably accessible nuclear antigen that is not β-catenin. In cell types commonly used to study Wnt activation, this non-specific nuclear staining can be robust, obscuring the ABC signal. Definitive detection of nuclear localized ABC can be confirmed through an ability of classical cadherins to sequester ABC to cell junctions. In tissues, milder antigen retrieval methods can reduce the accessibility of mAb 8E7 to this cross-reacting nuclear antigen. Conclusion These findings reveal that interpretation of nuclear, signaling active β-catenin using monoclonal antibody 8E7 should be considered judiciously, and in conjunction with independent methods. Reviewers This article was reviewed by Frank J. T. Staal (nominated by Rachel Gerstein, Jyoti M. Sen (nominated by Avinash Bhandoola and Manabu Sugai.

  10. Experiences from two local processes of debate and referenda on the issue of siting high level nuclear waste

    International Nuclear Information System (INIS)

    Full text of publication follows: the paper summarizes results from two interview studies conducted in the communities of Storuman (1995) and Malaa (1997) in northern Sweden regarding whether to continue investigations of the areas for siting of a deep level repository for high level nuclear waste. Active, local participants in the work and discussions preceding each local referendum on the issue were asked to reflect on reasons and considerations related to their opinions, as well as the overall outcome for achieving a deeper understanding of the local processes. The first referendum (1995) yielded a strongly voiced rejection of continuing local investigations (72%), whereas the second (1997) referendum resulted in a marginally negative response (54%). A comparison of the results of the interview studies showed e.g. that the decision processes differed across communities, regarding both time interval and content, and that the local strategies and tactics related to the campaigns preceding the referenda differed. Among the similarities were the types of questions which remained unclear, often related to a long term perspective, e.g. risks and uncertainties regarding material reliability, access to and future safety of the repository, concern for future generations, national and international long-term decision procedures, and roles of responsibility. The discussion focuses on considerations around the issue of local vs. centralized political decisions and the tool provided by the referendum, and touches upon some issues which appeared rather paradoxical. (author)

  11. On the local economy of nuclear power and the construction of Jiangxi

    International Nuclear Information System (INIS)

    Therefore, the development of nuclear power both for Jiangxi provide a stable economic development and protect the environment backed energy, reduce environmental pollution, thereby promoting the development of Jiangxi's economy by leaps and bounds. (authors)

  12. Tropomyosin is localized in the nuclear matrix and chromosome scaffold of physarum polycephalum

    Institute of Scientific and Technical Information of China (English)

    ZENGXIANLU; XIAOGUANGWANG; 等

    1999-01-01

    The nuclei and chromosomes were isolated from plasmodia of Physarum polycephalum.The nuclear matrix and chromosome scaffold were obtained after the DNA and most of the proteins were extracted with DNase I and 2 M NaCl.SD-PAGE analyses revealed that the nuclear matrix and chromosome scaffold contained a 37 kD polypeptide which is equivalent to tropomyosin in molecular weight.Immunofluorescence observations upon slide preparations labeled with anti-tropomyosin antibody showed that the nuclear matrix and chromosome scaffold emanated bright fluorescence,suggesting the presence of the antigen in them.Immunodotting results confirmed the presence of tropomyosin in the nuclear matrix and chromosome scaffold.Immunoelectron microscopic observations further demonstrated that tropomyosin was dispersively distributed in the interphase nuclei and metaphase chromosomes.

  13. An efficient method of constructing homologous recom binant baculovirus with PCR-amplified fragments

    Institute of Scientific and Technical Information of China (English)

    HOU; Songwang; (侯松旺); CHEN; Xinwen; (陈新文); WANG; Hanzhong; (王汉中); HU; Zhihong; (胡志红)

    2003-01-01

    This paper describes a rapid method of constructing homologous recombinant baculovirus in E. coli with PCR-amplified fragments. By using this method, the traditional steps of constructing transfer vector are omitted. The method is based on phage λ red system which can promote the recombination between the homologous fragments with the length above 36 bp. Taking HaSNPV as an example, this paper describes the rapid recombination process by using chloramphenicol resistance gene (CmR) to replace orf135 in HaSNPV genome. A pair of primers with length of 60 bp was synthesized, in which 40 bp was homologous to the each end sequence of orf135, and the rest 20 bp was homologous to the each end sequence of CmR. By using these primers, a linear fragment containing the complete CmR gene between 40 bp of homologous arms of orf135 was generated by PCR with the plasmid pKD3 which contains CmR as the template. By transforming the linear fragment into the E. coli containing the bacterial artificial chromosome of HaSNPV and with the help of a plasmid expressing λ recombinase, the recombinants on which the homologue replacement had taken place were selected by chloramphenicol resistance. This method greatly shortens the process of constructing recombinant baculovirus since the process was performed in E. coli and does not need to construct transfer vectors. It can be further used for gene replacement and gene deletion of other large viral genomes.

  14. The construction and preliminary analysis of a Tn5 transposon based random mutant library of baculovirus

    Institute of Scientific and Technical Information of China (English)

    Li Hui; Zhao Minglei; Yin Juan; Zhong Jiang

    2006-01-01

    A transposon-based random mutation library of AcMNPV,the type species of baculovirus,was constructed using a Tn5 transposon.The green fluorescence protein gene under the control of the Drosophila hsp70 promoter was inserted into the transposon for easy tracking in insect cells.In vitro transposition was carried out using the transposon and AcMNPV genomic DNA to allow the random insertion of the transposon into the virus genome.The transposed genome was then used to transfect Sf21 insect cells,and a library of mutant viruses capable of expressing green fluorescence protein was obtained.Two mutant viruses,B9F and Li6A were isolated,and the sites of transposon insertion were determined to be within the coding regions of the 94k and p10 genes,respectively.Both genes were determined to be nonessential in viral replication and infection.This technique will be very useful in the functional study of baculovirus genes.

  15. Crystallization and preliminary crystallographic studies of the metalloglycoprotein esterase A4 using a baculovirus expression system

    International Nuclear Information System (INIS)

    Esterase A4 (EA4) is a timer protein found in diapause eggs of the silkworm Bombyx mori. The gene for this metalloglycoprotein was cloned from B. mori eggs and expressed using a baculovirus expression system in silkworm pupae. Crystals of the purified protein have been grown that diffract to beyond 2.1 Å resolution at 100 K using synchrotron radiation. Esterase A4 (EA4) is a timer protein found in diapause eggs of the silkworm Bombyx mori. The gene for this metalloglycoprotein was cloned from B. mori eggs and expressed using a baculovirus expression system in silkworm pupae. Crystals of the purified protein have been grown that diffract to beyond 2.1 Å resolution at 100 K using synchrotron radiation. The protein crystals belong to space group P21, with unit-cell parameters a = 47.1, b = 73.9, c = 47.4 Å, β = 104.1°. With one dimer per asymmetric unit, the crystal volume per unit protein weight (VM) is 2.3 Å3 Da−1 and the solvent content is 47%

  16. Sunlight stability and rain-fastness of formulations of Baculovirus heliothis

    International Nuclear Information System (INIS)

    Sunlight-Ultraviolet, with an activity spectrum from 290 to 400 nm, is the most destructive factor affecting the persistence of baculoviruses. Benzopurpurin (a disazo dye) and carbon provided the best protection when polyhedral inclusion bodies (PIB) of Baculovirus heliothis were exposed to an artificial spectrum simulating sunlight-UV (UV). Greater than 75% of the original PIB activity was still present after 48 h of sunlight-UV. When sprayed on soybeans and exposed to natural sunlight, only formulations with carbon provided significant protection of PIB. The half-life of formulations were PIB-only 4.9 +/- 1.4 h (mean +/- SE), PIB + polymer (pyrrolidone-based sticker) 3.3 +/- 0.6 h, PIB + polymer + benzopurpurin 3.4 +/- 0.7 h, and PIB + polymer + carbon 27.7 +/- 5.2 h. PIB of B. heliothis tenaciously adhere to soybean, Glycine max (L.) Merrill, leaflets after spraying and drying. Less than 6% of the PIB activity of nonformulated PIB was lost after a drenching, simulated rainfall. More than 97% of the original PIB activity of carbon formulations was still present on soybean leaflets after 10 h of exposure to sunlight-UV. In contrast, 20% was present for formulations without carbon

  17. Analysis of nucleolar morphology and protein localization as an indicator of nuclear reprogramming

    DEFF Research Database (Denmark)

    Østrup, Olga; Pedersen, Hanne Skovsgaard; Holm, Hanne M.;

    2015-01-01

    the nucleolus are summarized in this developmental context, but also as they occur in assisted reproductive technologies such as in vitro fertilization and somatic cell nuclear transfer. Moreover, detailed protocols for monitoring the nucleolar changes by transmission electron microscopy and...... cloning by somatic cell nuclear transfer. However, when cells are reprogrammed by less fundamental means, as for example treatment by Xenopus extract or expression of pluripotency genes, more subtle nucleolar modulations can also be noted. The monitoring and understanding of the reprogramming...

  18. Continuity equation and local gauge invariance for the N3LO nuclear Energy Density Functionals

    OpenAIRE

    Raimondi, F.; Carlsson, B G; Dobaczewski, J.; Toivanen, J.

    2011-01-01

    Background: The next-to-next-to-next-to-leading order (N3LO) nuclear energy density functional extends the standard Skyrme functional with new terms depending on higher-order derivatives of densities, introduced to gain better precision in the nuclear many-body calculations. A thorough study of the transformation properties of the functional with respect to different symmetries is required, as a step preliminary to the adjustment of the coupling constants. Purpose: Determine to which extent t...

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

    Science.gov (United States)

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

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    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.

  1. Characterization of the recombinant proteins of porcine circovirus type2 field isolate expressed in the baculovirus system.

    Science.gov (United States)

    Kim, Yuna; Kim, Jinhyun; Kang, Kyoungsoo; Lyoo, Young S

    2002-03-01

    Porcine circovirus (PCV) type2 was isolated using primary porcine kidney cells from lymph node of piglets with typical PMWS. The presence of the virus was identified by PCR using primers specific to PCV type2. The ORFs 1 and 2 were amplified by PCR using primers corresponding to the target genes of the PCV type 2. Cloned genes were inserted into the baculovirus expression vector and PCV recombinant proteins were expressed using baculovirus expression system. Recombinant protein expression was determined by indirect immunofluorescent assay (IFA) and immunoblotting using polyclonal antiserum to PCV. ORF1 gene expressed two proteins with approximately 17 kDa and 31 kDa proteins in the baculovirus system. Recombinant protein of the ORF2 was similar to that of the native virus except minor bands with different molecular weight were detected. Recombinant protein expressed in the baculovirus system showed at least two glycosylation sites based on the tunicamycin treatment. Recombinant protein of the ORF2 assembled virus-like particle in recombinant virus infected insect cells. PMID:14614268

  2. Expression of the hemagglutinin HA1 subunit of the equine influenza virus using a baculovirus expression system.

    Science.gov (United States)

    Sguazza, Guillermo H; Fuentealba, Nadia A; Tizzano, Marco A; Galosi, Cecilia M; Pecoraro, Marcelo R

    2013-01-01

    Equine influenza virus is a leading cause of respiratory disease in horses worldwide. Disease prevention is by vaccination with inactivated whole virus vaccines. Most current influenza vaccines are generated in embryonated hens' eggs. Virions are harvested from allantoic fluid and chemically inactivated. Although this system has served well over the years, the use of eggs as the substrate for vaccine production has several well-recognized disadvantages (cost, egg supply, waste disposal and yield in eggs). The aim of this study was to evaluate a baculovirus system as a potential method for producing recombinant equine influenza hemagglutinin to be used as a vaccine. The hemagglutinin ectodomain (HA1 subunit) was cloned and expressed using a baculovirus expression vector. The expression was determined by SDS-PAGE and immunoblotting. A high yield, 20μg/ml of viral protein, was obtained from recombinant baculovirus-infected cells. The immune response in BALB/c mice was examined following rHA1 inoculation. Preliminary results show that recombinant hemagglutinin expressed from baculovirus elicits a strong antibody response in mice; therefore it could be used as an antigen for subunit vaccines and diagnostic tests. PMID:24401775

  3. Behaviour of wild-type and genetically modified baculoviruses in the Helicoverpa armigera - cotton system: a simulation approach

    NARCIS (Netherlands)

    Sun, X.

    2005-01-01

    Keywords:   Helicoverpa armigera , baculovirus, genetic modification, cotton,transmissionLocal panels and maintainability human factors assessment for AP1000 nuclear power plant

    International Nuclear Information System (INIS)

    A document entitled 'AP1000 Local Panels and Maintainability Human Factors Design Guidelines' was produced to aid the designers to specifically include human factors (HF) considerations in the design, operation, and maintenance of local control stations and plant equipment. To ensure that the applicable HF design guidelines are appropriately applied to the design of local panels and maintenance activities, and identify any HF improvement opportunities that can readily be implemented at the design stage, a HF assessment of maintenance activities and local plant operations is underway. This assessment gives priority to local control stations and equipment which have been identified as having a potential impact on safety. This includes risk-significant systems, structures and components (SSCs) identified through the probabilistic risk assessment (PRA), and local operator actions as required by the Emergency Operating Procedures (EOPs). Local actions, maintenance activities and associated operator interfaces are reviewed against the relevant HF guidelines. The results of the assessment include a description of the component, associated local actions and/or required maintenance activities, good design features and/or potential issues, and recommendations for change or improvement. These results are communicated to responsible design engineers who evaluate the impact to plant design and implement design changes, if deemed necessary. (author)

  4. The feasibility of using a baculovirus vector to deliver the sodium-iodide symporter gene as a reporter

    Energy Technology Data Exchange (ETDEWEB)

    Zhou Xiang; Li Biao; Wang Jun; Yin Hongyan [Department of Nuclear Medicine, Ruijin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200025 (China); Zhang Yifan [Department of Nuclear Medicine, Ruijin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200025 (China)], E-mail: zhangyifan1992@yahoo.com.cn

    2010-04-15

    Purpose: To evaluate the efficiency of baculovirus vectors in transducing FTC-133 cells and to examine the feasibility of using baculovirus vectors for the delivery of the sodium-iodide symporter (NIS) gene as a reporter through co-transduction to monitor the expression of the target gene. Method: Two recombinant baculoviruses were constructed to express NIS and green fluorescent protein (GFP) respectively. FTC-133, 8050C, SW1116, A549 cells, were infected with Bac-GFP. The infection efficiency of Bac-GFP and the intensity of fluorescence, in either the presence or absence of sodium butyrate, were monitored by flow cytometry. The iodine uptake by FTC-133 cells infected with Bac-NIS was measured using a {gamma} counter. FTC-133 cells were infected with a mixture of equal amounts of Bac-NIS and Bac-GFP at different setting of multiplicity of infection (MOI). The changes of GFP fluorescence intensity and iodine uptake were monitored 24 h after infection in the coinfected cells. Results: We have successfully constructed recombinant baculoviruses carrying NIS and GFP under the control of the cytomegalovirus IE-1 promoter. We found that transduced efficiency of baculovirus in 8505C, SW1116, A549 cells are low in absence of sodium butyrate. Yet Bac-GFP infects FTC-133 cells at a high efficiency, 77.67%, 85.57% and 93.23% with MOI of 100, 200 and 400, respectively. The fluorescence intensity of the Bac-GFP infected tumor cells correlated positively with the MOI of the virus. Sodium butyrate induction increased both the infection efficiency and the fluorescence intensity, but increase of infection efficiency was insignificant in FTC-133 cells. Reporter gene (GFP) expression in FTC-133 is stable within 7 days after infection. The radioactivity incorporated by the tumor cells infected with Bac-NIS correlated positively with the MOI of Bac-NIS as well. In tumor cells co-infected with Bac-NIS and Bac-GFP, the amount of radioactivity incorporated significantly correlated with

  5. The feasibility of using a baculovirus vector to deliver the sodium-iodide symporter gene as a reporter

    International Nuclear Information System (INIS)

    Purpose: To evaluate the efficiency of baculovirus vectors in transducing FTC-133 cells and to examine the feasibility of using baculovirus vectors for the delivery of the sodium-iodide symporter (NIS) gene as a reporter through co-transduction to monitor the expression of the target gene. Method: Two recombinant baculoviruses were constructed to express NIS and green fluorescent protein (GFP) respectively. FTC-133, 8050C, SW1116, A549 cells, were infected with Bac-GFP. The infection efficiency of Bac-GFP and the intensity of fluorescence, in either the presence or absence of sodium butyrate, were monitored by flow cytometry. The iodine uptake by FTC-133 cells infected with Bac-NIS was measured using a γ counter. FTC-133 cells were infected with a mixture of equal amounts of Bac-NIS and Bac-GFP at different setting of multiplicity of infection (MOI). The changes of GFP fluorescence intensity and iodine uptake were monitored 24 h after infection in the coinfected cells. Results: We have successfully constructed recombinant baculoviruses carrying NIS and GFP under the control of the cytomegalovirus IE-1 promoter. We found that transduced efficiency of baculovirus in 8505C, SW1116, A549 cells are low in absence of sodium butyrate. Yet Bac-GFP infects FTC-133 cells at a high efficiency, 77.67%, 85.57% and 93.23% with MOI of 100, 200 and 400, respectively. The fluorescence intensity of the Bac-GFP infected tumor cells correlated positively with the MOI of the virus. Sodium butyrate induction increased both the infection efficiency and the fluorescence intensity, but increase of infection efficiency was insignificant in FTC-133 cells. Reporter gene (GFP) expression in FTC-133 is stable within 7 days after infection. The radioactivity incorporated by the tumor cells infected with Bac-NIS correlated positively with the MOI of Bac-NIS as well. In tumor cells co-infected with Bac-NIS and Bac-GFP, the amount of radioactivity incorporated significantly correlated with the GFP

  6. Dynamic nuclear polarization enhanced nuclear magnetic resonance and electron spin resonance studies of hydration and local water dynamics in micelle and vesicle assemblies.

    Science.gov (United States)

    McCarney, Evan R; Armstrong, Brandon D; Kausik, Ravinath; Han, Songi

    2008-09-16

    We present a unique analysis tool for the selective detection of local water inside soft molecular assemblies (hydrophobic cores, vesicular bilayers, and micellar structures) suspended in bulk water. Through the use of dynamic nuclear polarization (DNP), the (1)H NMR signal of water is amplified, as it interacts with stable radicals that possess approximately 658 times higher spin polarization. We utilized stable nitroxide radicals covalently attached along the hydrophobic tail of stearic acid molecules that incorporate themselves into surfactant-based micelle or vesicle structures. Here, we present a study of local water content and fluid viscosity inside oleate micelles and vesicles and Triton X-100 micelles to serve as model systems for soft molecular assemblies. This approach is unique because the amplification of the NMR signal is performed in bulk solution and under ambient conditions with site-specific spin labels that only detect the water that is directly interacting with the localized spin labels. Continuous wave (cw) electron spin resonance (ESR) analysis provides rotational dynamics of the spin-labeled molecular chain segments and local polarity parameters that can be related to hydration properties, whereas we show that DNP-enhanced (1)H NMR analysis of fluid samples directly provides translational water dynamics and permeability of the local environment probed by the spin label. Our technique therefore has the potential to become a powerful analysis tool, complementary to cw ESR, to study hydration characteristics of surfactant assemblies, lipid bilayers, or protein aggregates, where water dynamics is a key parameter of their structure and function. In this study, we find that there is significant penetration of water inside the oleate micelles with a higher average local water viscosity (approximately 1.8 cP) than in bulk water, and Triton X-100 micelles and oleate vesicle bilayers mostly exclude water while allowing for considerable surfactant chain

  7. Multi-region fuzzy logic controller with local PID controllers for U-tube steam generator in nuclear power plant

    Directory of Open Access Journals (Sweden)

    Puchalski Bartosz

    2015-12-01

    Full Text Available In the paper, analysis of multi-region fuzzy logic controller with local PID controllers for steam generator of pressurized water reactor (PWR working in wide range of thermal power changes is presented. The U-tube steam generator has a nonlinear dynamics depending on thermal power transferred from coolant of the primary loop of the PWR plant. Control of water level in the steam generator conducted by a traditional PID controller which is designed for nominal power level of the nuclear reactor operates insufficiently well in wide range of operational conditions, especially at the low thermal power level. Thus the steam generator is often controlled manually by operators. Incorrect water level in the steam generator may lead to accidental shutdown of the nuclear reactor and consequently financial losses. In the paper a comparison of proposed multi region fuzzy logic controller and traditional PID controllers designed only for nominal condition is presented. The gains of the local PID controllers have been derived by solving appropriate optimization tasks with the cost function in a form of integrated squared error (ISE criterion. In both cases, a model of steam generator which is readily available in literature was used for control algorithms synthesis purposes. The proposed multi-region fuzzy logic controller and traditional PID controller were subjected to broad-based simulation tests in rapid prototyping software - Matlab/Simulink. These tests proved the advantage of multi-region fuzzy logic controller with local PID controllers over its traditional counterpart.

  8. The estrogen receptor alpha nuclear localization sequence is critical for fulvestrant-induced degradation of the receptor.

    Science.gov (United States)

    Casa, Angelo J; Hochbaum, Daniel; Sreekumar, Sreeja; Oesterreich, Steffi; Lee, Adrian V

    2015-11-01

    Fulvestrant, a selective estrogen receptor down-regulator (SERD) is a pure competitive antagonist of estrogen receptor alpha (ERα). Fulvestrant binds ERα and reduces the receptor's half-life by increasing protein turnover, however, its mechanism of action is not fully understood. In this study, we show that removal of the ERα nuclear localization sequence (ERΔNLS) resulted in a predominantly cytoplasmic ERα that was degraded in response to 17-β-estradiol (E2) but was resistant to degradation by fulvestrant. ERΔNLS bound the ligands and exhibited receptor interaction similar to ERα, indicating that the lack of degradation was not due to disruption of these processes. Forcing ERΔNLS into the nucleus with a heterologous SV40-NLS did not restore degradation, suggesting that the NLS domain itself, and not merely receptor localization, is critical for fulvestrant-induced ERα degradation. Indeed, cloning of the endogenous ERα NLS onto the N-terminus of ERΔNLS significantly restored both its nuclear localization and turnover in response to fulvestrant. Moreover, mutation of the sumoylation targets K266 and K268 within the NLS impaired fulvestrant-induced ERα degradation. In conclusion, our study provides evidence for the unique role of the ERα NLS in fulvestrant-induced degradation of the receptor. PMID:26272024

  9. Growing from a local into an international nuclear services provider - challenges and opportunities

    International Nuclear Information System (INIS)

    Apart from nuclear power plants' (NPPs) internally staffed expert resources, the support of qualified external nuclear maintenance and modification services providers (NSPs) is extremely important. The postponement of the nuclear power renaissance along with an aging workforce is pushing the industry to search for different approaches to maintain and improve the required level of expertise and quality. Operating in a domestic market with only one NPP unit, the leading Slovenian NSP has been partnering with complimentary companies to jointly satisfy the needs of the domestic, US and EU customers. By remaining competitive in quality and price and sharing resources, global experience and good practices, results in added value to all of the interested parties is achieved. In addition to supporting peak outage season shortages of qualified resources, the concept also aids in load-levelling the resource needs throughout the year. The challenges and the opportunities related to the concept are discussed in the paper. (authors)

  10. Localization of nuclear retained mRNAs in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Thomsen, Rune; Libri, Domenico; Boulay, Jocelyne;

    2003-01-01

    In the yeast Saccharomyces cerevisiae, a common conditional phenotype associated with deletion or mutation of genes encoding mRNA export factors is the rapid accumulation of mRNAs in intranuclear foci, suggested to be near transcription sites. The nuclear RNA exosome has been implicated in retain......In the yeast Saccharomyces cerevisiae, a common conditional phenotype associated with deletion or mutation of genes encoding mRNA export factors is the rapid accumulation of mRNAs in intranuclear foci, suggested to be near transcription sites. The nuclear RNA exosome has been implicated...... in retaining RNAs in these foci; on deletion of the exosome component Rrp6p, the RNA is released. To determine the exact nuclear location of retained as well as released mRNAs, we have used mRNA export mutant strains to analyze the spatial relationship between newly synthesized heat shock mRNA, the chromosomal...

  11. Prolonged, brain-wide expression of nuclear-localized GCaMP3 for functional circuit mapping

    Directory of Open Access Journals (Sweden)

    Christina Kay Kim

    2014-11-01

    Full Text Available Larval zebrafish offer the potential for large-scale optical imaging of neural activity throughout the central nervous system; however, several barriers challenge their utility. First, ~panneuronal probe expression has to date only been demonstrated at early larval stages up to 7 days post-fertilization (dpf, precluding imaging at later time points when circuits are more mature. Second, nuclear exclusion of genetically-encoded calcium indicators (GECIs limits the resolution of functional fluorescence signals collected during imaging. Here, we report the creation of transgenic zebrafish strains exhibiting robust, nuclearly targeted expression of GCaMP3 across the brain up to at least 14 dpf utilizing a previously described optimized Gal4-UAS system. We confirmed both nuclear targeting and functionality of the modified probe in vitro and measured its kinetics in response to action potentials. We then demonstrated in vivo functionality of nuclear-localized GCaMP3 in transgenic zebrafish strains by identifying eye position-sensitive fluorescence fluctuations in caudal hindbrain neurons during spontaneous eye movements. Our methodological approach will facilitate studies of larval zebrafish circuitry by both improving resolution of functional Ca2+ signals and by allowing brain-wide expression of improved GECIs, or potentially any probe, further into development.

  12. Self-reliance and innovation to improve the localization ability of nuclear power construction

    International Nuclear Information System (INIS)

    Construction is a crucial link in the course of design, site selection, construction and operation for a nuclear power plant. The quality during construction directly affects the nuclear safety. And the construction quality involves various aspects including quality culture, feasibility of construction technique, quality assurance, and quality control. Through technical management innovation, great achievements have been made in the civil construction of Unit 3 of Qinshan II Extension Project. As a result, the construction period was shortened and the concrete quality reached the fairfaced concrete standard. (authors)

  13. Localization of phosphorylated TrkA in carrier vesicles involved in its nuclear translocation in U251 cell line

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    A number of transmembrane receptors are targeted to the nucleus and convincingly localized therein. However, what remains a conundrum is how these cell-surface receptors end up in the nucleus. In this study, we reported that the transmembrane receptor phosphorylated TrkA was located in a series of carrier vesicles, including ring-like vesicles near the plasma membrane, large core vesicles and small dense core vesicles around the nuclei, as well as in the nucleus in human glioma cell line U251 using immunocytochemistry and immunofluorescence staining. Meanwhile, we also showed that small dense core vesicles budded from large core vesicles, and interacted with the nuclear envelope. Accordingly, our results suggested that such a series of membrane compartments might be involved in the pathway of nuclear translocation of the transmembrane receptor TrkA.

  14. Localization of phosphorylated TrkA in carrier vesicles involved in its nuclear translocation in U251 cell line

    Institute of Scientific and Technical Information of China (English)

    GONG AiHua; ZHANG ZhiJian; XIAO DeSheng; YANG Yong; WANG YongZhong; CHEN YongChang

    2007-01-01

    A number of transmembrane receptors are targeted to the nucleus and convincingly localized therein.However, what remains a conundrum is how these cell-surface receptors end up in the nucleus. In this study, we reported that the transmembrane receptor phosphorylated TrkA was located in a series of carrier vesicles, including ring-like vesicles near the plasma membrane, large core vesicles and small dense core vesicles around the nuclei, as well as in the nucleus in human glioma cell line U251 using immunocytochemistry and immunofluorescence staining. Meanwhile, we also showed that small dense core vesicles budded from large core vesicles, and interacted with the nuclear envelope. Accordingly,our results suggested that such a series of membrane compartments might be involved in the pathway of nuclear translocation of the transmembrane receptor TrkA.

  15. Localized Beampipe Heating due to $e^-$ Capture and Nuclear Excitation in Heavy Ion Colliders

    OpenAIRE

    Klein, Spencer R.

    2000-01-01

    At heavy ion colliders, two major sources of beam loss are expected to be $e^+e^-$ production, where the $e^-$ is bound to one of the nuclei, and photonuclear excitation and decay via neutron emission. Both processes alter the ions charged to mass ratio by well defined amounts, creating beams of particles with altered magnetic rigidity. These beams will deposit their energy in a localized region of the accelerator, causing localized heating, The size of the target region depends on the collid...

  16. Phenomenon of Local Natural Circulation in a Circuit of Nuclear Power Plant

    International Nuclear Information System (INIS)

    The report presents results of experimental research performed at the BN-350 reactor facility, which have allowed identifying a local natural circulation phenomenon arising in some sections of heat removal loops. The nature of this phenomenon and features of its interaction with total-circuit natural circulation is described. In particular, specific features and degree of possible influence of local natural circulation on operational modes of reactor facility are analyzed. (author)

  17. Participation of local citizens groups in the Swedish nuclear waste process

    International Nuclear Information System (INIS)

    The Waste Network's conclusions and views on the nuclear waste issue are summarised in the following points: The management of the nuclear waste is not solved. In order to minimise the amount of waste, the further operating of nuclear reactors should be questioned. The choice of method should be made before the siting. The deadlock to the KBS method must come to an end. The choice of method must be based on clearly expressed functional conditions formulated in advance. The siting must be based on considerations to environment and security, not political acceptance. The pilot studies in municipalities must cease and should be replaced by a clear and understandable sieving process at a national scale. An independent authority must control and supervise the EIA process instead of the nuclear industry. A well performed EIA process is the necessary condition to give the choice of method and site enough legitimacy and acceptance. Environmental organisations being the representatives of the public must be given reasonable conditions and resources to take part in the EIA process to handle the question

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

    International Nuclear Information System (INIS)

    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

  19. Nuclear localization of Src-family tyrosine kinases is required for growth factor-induced euchromatinization

    International Nuclear Information System (INIS)

    Src-family kinases (SFKs), which participate in various signaling events, are found at not only the plasma membrane but also several subcellular compartments, including the nucleus. Nuclear structural changes are frequently observed during transcription, cell differentiation, senescence, tumorigenesis, and cell cycle. However, little is known about signal transduction in the alteration of chromatin texture. Here, we develop a pixel imaging method for quantitatively evaluating chromatin structural changes. Growth factor stimulation increases euchromatic hypocondensation and concomitant heterochromatic hypercondensation in G1 phase, and the levels reach a plateau by 30 min, sustain for at least 5 h and return to the basal levels after 24 h. Serum-activated SFKs in the nucleus were more frequently detected in the euchromatin areas than the heterochromatin areas. Nuclear expression of kinase-active SFKs, but not unrelated Syk kinase, drastically increases both euchromatinization and heterochromatinization in a manner dependent on the levels of nuclear tyrosine phosphorylation. However, growth factor stimulation does not induce chromatin structural changes in SYF cells lacking SFKs, and reintroduction of one SFK member into SYF cells can, albeit insufficiently, induce chromatin structural changes. These results suggest that nuclear tyrosine phosphorylation by SFKs plays an important role in chromatin structural changes upon growth factor stimulation.

  1. Design of a small single-purpose nuclear desalination plant compatible with the local resources in the Middle East

    International Nuclear Information System (INIS)

    In this paper results of the preliminary stages of the design of a 2.6 million gallons per day (1000 cubic meters/day) single-purpose 40 MW thermal MSF-heavy water nuclear plant are reported. Design objectives and philosophy are reviewed. A description is given of the neutronic, thermal, control and mechanical design parameters of the plant. The design is based on utilization of available local material, technology and manpower. The feasibility of production of heavy water in the fertilizer production plants and the extraction of natural uranium from the phosphate mines on the Red Sea coast are considered pointing out further stages of development and future plans

  2. WNT Protein-independent Constitutive Nuclear Localization of β-Catenin Protein and Its Low Degradation Rate in Thalamic Neurons*

    OpenAIRE

    Misztal, Katarzyna; Wisniewska, Marta B.; Ambrozkiewicz, Mateusz; Nagalski, Andrzej; Kuznicki, Jacek

    2011-01-01

    Nuclear localization of β-catenin is a hallmark of canonical Wnt signaling, a pathway that plays a crucial role in brain development and the neurogenesis of the adult brain. We recently showed that β-catenin accumulates specifically in mature thalamic neurons, where it regulates the expression of the Cav3.1 voltage-gated calcium channel gene. Here, we investigated the mechanisms underlying β-catenin accumulation in thalamic neurons. We report that a lack of soluble factors produced either by ...

  3. Nuclear-localized Calcineurin Homologous Protein CHP1 Interacts with Upstream Binding Factor and Inhibits Ribosomal RNA Synthesis*

    OpenAIRE

    Jiménez-Vidal, Maite; Srivastava, Jyoti; Putney, Luanna K; Barber, Diane L.

    2010-01-01

    Calcineurin homologous protein 1 (CHP1) is a widely expressed, 22-kDa myristoylated EF-hand Ca2+-binding protein that shares a high degree of similarity with the regulatory B subunit of calcineurin (65%) and with calmodulin (59%). CHP1 localizes to the plasma membrane, the Golgi apparatus, and the nucleus and functions to regulate trafficking of early secretory vesicles, activation of T cells, and expression and transport of the Na-H exchanger NHE1. Although CHP1 contains nuclear export signa...

  4. The local heat treatment technology for not easily accessible welded joints of sodium-conducting pipes in nuclear power stations

    International Nuclear Information System (INIS)

    Experimental works results in local heat treatment of not easily accessible welded joints for the nuclear power stations sodium-conducting pipes using flexible wired resistance heaters are described. Vertical welded joints 820x13, 630x13, 219x10 and 245x11 were studied. It was found out that for the tubes welded joints 219x10 and 245x11mm the normal process of austenization without reflectors inside the welded joint is achieved under heating by means of one electric heater. It was suggested that the electric heater capacity should be increased for tubes welded joints 820x13 and 630x13 mm

  5. Review study 1995. Localization of the repository for spent nuclear fuel; Oeversiktsstudie 95. Lokalisering av djupfoervar foer anvaent kaernbraensle

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1995-10-01

    This report gives an overview of the studies performed by SKB pertinent to selection of a site for the Swedish repository for spent nuclear fuels, and is written for both experts in the various fields involved, decision-makers and the interested general public. The review can not comprise all detailed factors necessary for deciding the localization, but deals mainly with conditions on the land surface and can point out areas which are not well suited or less interesting as a site. It also treats several scientific, technical and social bases in different parts of the country. 120 refs, 53 figs.

  6. Characterization of the immune responses elicited by baculovirus-based vector vaccines against influenza virus hemagglutinin

    Institute of Scientific and Technical Information of China (English)

    Zhi-peng HU; Juan YIN; Yuan-yuan ZHANG; Shu-ya JIA; Zuo-jia CHEN; Jiang ZHONG

    2012-01-01

    Aim:To compare the specific immune responses elicited by different baculovirus vectors in immunized mice.Methods:We constructed and characterized two recombinant baculoviruses carrying the expression cassette for the H5N1 influenza virus hemagglutinin (HA) gene driven by either an insect cell promoter (vAc-HA) or a dual-promoter active both in insect and mammalian cells (vAc-HA-DUAL).Virus without the HA gene (vAc-EGFP) was used as a control.These viruses were used to immunize mice subcutaneously and intraperitoneally.The production of total and specific antibodies was determined by ELISA and competitive ELISA.Cytokine production by the spleen cells of immunized mice was studied using the ELISPOT assay.Results:Both the vAc-HA and vAc-HA-DUAL vectors expressed HA proteins in insect Sf9 cells,and HA antigen was present in progeny virions.The vAc-HA-DUAL vector also mediated HA expression in virus-transduced mammalian cell lines (BHK and A547).Both vAo-HA and vAc-HA-DUAL exhibited higher transduction efficiencies than vAc-EGFP in mammalian cells,as shown by the expression of the reporter gene egfp.Additionally,both vAc-HA and vAc-HA-DUAL induced high levels of HA-specific antibody production in immunized mice; vAc-HA-DUAL was more efficient in inducing IFN-Y and IL-2 upon stimulation with specific antigen,whereas vAc-HA was more efficient in inducing IL-4 and IL-6.Conclusion:Baculovirus vectors elicited efficient,specific immune responses in immunized mice.The vector displaying the HA antigen on the virion surface (vAc-HA) elicited a Th2-biased immune response,whereas the vector displaying HA and mediating HA expression in the cell (vAc-HA-DUAL) elicited a Th1-biased immune response.

  7. Does nuclear power-related facility siting always lower the local property Values? Comparative analysis among the sites in Japan

    International Nuclear Information System (INIS)

    Yamane et al. (2011) carried out hedonic house rent analyses for several nuclear power plant sites in Japan, compared the result for each plant, and obtained some empirical results indicating that the local residents' marginal willingness-to-pay (MWTP) for living near the plant was not necessarily positive and that the MWTP was correlated with operation years of the plants and some of the host communities' attributes (i.e., population density, financial condition and public service improvement). However, these results may suffer from biases and inefficiency in estimating hedonic functions, caused by spatial dependency: spatial autoregression and spatial autocorrelation. In this paper, we introduce spatial econometric techniques to settle this problem. As a revised result, it is indicated that the local residents' MWTP is correlated with past accidents in the plants, education service improvement in the host communities and so on. (author)

  8. Local stakeholder involvement in the perspective of nuclear waste management: lessons form the Cowam network

    International Nuclear Information System (INIS)

    The management of high level radioactive waste is nowadays recognised as a complex decision-making process entailing technical, environmental, ethical, social, political and economic dimensions where no solution can be reached solely on the basis of technical considerations. While this issue is acknowledged as a problem for the community as a whole, waste management remains a global problem looking for a local solution. Starting from this view, COWAM network (Community Waste Management), developed under the Fifth Framework Programme of the European Commission, addressed the following objectives: 1) To empower local actors through a networking process; 2) To gather and discuss the available experiences of decision-making processes at the local level within their national context in Europe; 3) To set up an arena for balanced exchanges between local actors, NGOs, regulators and implementers; 4) To promote new approaches to decision-making in national contexts in Europe. COWAM network comprises 230 delegates from 10 European countries, involving in priority local communities and NGOs. The emphasis put on the local participation enabled members of COWAM network to overcome distrust and to build common lessons and views beyond usual stakeholder positions. Through the analysis of case studies different issues were identified, among them two relate more specifically to: 1) Expertise what is the purpose of expertise on environmental impact in the decision-making process? How is this expertise linked with other scientific and non scientific issues? What is the role of stakeholders in expertise? 2) Environmental quality in the long term and sustainable development how is the impact of radioactive waste management facilities on the environment in the long term taken into account? how is this associated with the sustainable development of the hosting community? How are local stakeholders involved in these issues and what is the expected benefit from their participation? (author)

  9. Characterization of amino acid residues within the N-terminal region of Ubc9 that play a role in Ubc9 nuclear localization

    International Nuclear Information System (INIS)

    As the sole E2 enzyme for SUMOylation, Ubc9 is predominantly nuclear. However, the underlying mechanisms of Ubc9 nuclear localization are still not well understood. Here we show that RNAi-depletion of Imp13, an importin known to mediate Ubc9 nuclear import, reduces both Ubc9 nuclear accumulation and global SUMOylation. Furthermore, Ubc9-R13A or Ubc9-H20D mutation previously shown to interrupt the interaction of Ubc9 with nucleus-enriched SUMOs reduces the nuclear enrichment of Ubc9, suggesting that the interaction of Ubc9 with the nuclear SUMOs may enhance Ubc9 nuclear retention. Moreover, Ubc9-R17E mutation, which is known to disrupt the interaction of Ubc9 with both SUMOs and Imp13, causes a greater decrease in Ubc9 nuclear accumulation than Ubc9-R13A or Ubc9-H20D mutation. Lastly, Ubc9-K74A/S89D mutations that perturb the interaction of Ubc9 with nucleus-enriched SUMOylation-consensus motifs has no effect on Ubc9 nuclear localization. Altogether, our results have elucidated that the amino acid residues within the N-terminal region of Ubc9 play a pivotal role in regulation of Ubc9 nuclear localization. - Highlights: • Imp13-mediated nuclear import of Ubc9 is critical for global SUMOylation. • Ubc9 mutations disrupting Ubc9-SUMO interaction decrease Ubc9 nuclear accumulation. • N-terminal amino acid residues of Ubc9 are critical for Ubc9 nuclear enrichment

  10. Adipogenesis stimulates the nuclear localization of EWS with an increase in its O-GlcNAc glycosylation in 3T3-L1 cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Qiang; Kamemura, Kazuo, E-mail: k_kamemura@nagahama-i-bio.ac.jp

    2014-07-18

    Highlights: • The majority of EWS localizes stably in the cytosol in 3T3-L1 preadipocytes. • Adipogenic stimuli induce the nuclear localization of EWS. • Adipogenesis promotes O-GlcNAcylation of EWS. • O-GlcNAcylation stimulates the recruitment of EWS to the nuclear periphery. - Abstract: Although the Ewing sarcoma (EWS) proto-oncoprotein is found in the nucleus and cytosol and is associated with the cell membrane, the regulatory mechanisms of its subcellular localization are still unclear. Here we found that adipogenic stimuli induce the nuclear localization of EWS in 3T3-L1 cells. Tyrosine phosphorylation in the C-terminal PY-nuclear localization signal of EWS was negative throughout adipogenesis. Instead, an adipogenesis-dependent increase in O-linked β-N-acetylglucosamine (O-GlcNAc) glycosylation of EWS was observed. Pharmacological inactivation of O-GlcNAcase in preadipocytes promoted perinuclear localization of EWS. Our findings suggest that the nuclear localization of EWS is partly regulated by the glycosylation.

  11. Pre-operative localization of parathyroid adenomas: A comparison of power and colour Doppler ultrasonography with nuclear medicine scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Scheiner, Jac D.; Dupuy, Damian E.; Monchik, John M.; Noto, Richard B.; Cronan, John J

    2001-12-01

    AIM: To compare power and colour Doppler ultrasonography (US) with nuclear medicine scintigraphy (NM) in the preoperative localization of parathyroid adenomas in patients with primary hyperparathyroidism (PHPT). MATERIALS AND METHODS: Thirty-one patients with biochemical evidence of PHPT underwent pre-operative US and NM for parathyroid adenoma localization. Both studies were interpreted independently without prior knowledge of the other study's findings. All patients had surgical removal of the parathyroid adenoma utilizing standard neck exploration or minimally invasive unilateral surgical techniques with rapid serum assay of circulating parathyroid hormone levels. RESULTS: All patients had single parathyroid adenomas at surgery. Prospective sensitivities for US, NM and both studies combined were 65%, 68%, and 74%, respectively, with a positive predictive value of 100% each. The adenoma was localized by only one imaging modality in 16% of cases. CONCLUSIONS: US and NM provide complementary roles in the pre-operative localization of parathyroid adenomas in patients with PHPT. Scheiner, J.D.et al. (2001)

  12. Control room, emergency control system and local control panels in nuclear power plants

    International Nuclear Information System (INIS)

    The requirements on planning and construction of control boards including ergonomic-technical designing are specified in this rule. The specifications put the requirements on the design of place, process and environment of work, which are mentioned in the sections 90 and 91 of the labor-management relations act, into more concrete terms for the safety-relevant control panels as work places in a nuclear power station. The work places at control panels are not considered as video workstations in the sense of the 'Safety Rules for Video Workstations in the Office Sector' published by the General Association of the Industrial Trade Associations. The requirements are based on the operation and information technology realized at present in control panels of stationary nuclear power plants. (orig./HP)

  13. Localized form of Fock terms in nuclear covariant density functional theory

    CERN Document Server

    Liang, Haozhao; Ring, Peter; Roca-Maza, Xavier; Meng, Jie

    2012-01-01

    In most of the successful versions of covariant density functional theory in nuclei, the Fock terms are not included explicitly, which leads to local functionals and forms the basis of their widespread applicability at present. However, it has serious consequences for the description of Gamow-Teller resonances (GTR) and spin-dipole resonances (SDR) which can only be cured by adding further phenomenological parameters. Relativistic Hartree-Fock models do not suffer from these problems. They can successfully describe the GTR and SDR as well as the isovector part of the Dirac effective mass without any additional parameters. However, they are non-local and require considerable numerical efforts. By the zero-range reduction and the Fierz transformation, a new method is proposed to take into account the Fock terms in local functionals, which retains the simplicity of conventional models and provides proper descriptions of the spin-isospin channels and the Dirac masses.

  14. Monodisperse Magnetite Nanoparticles Coupled with Nuclear Localization Signal Peptide for Cell-Nucleus Targeting

    OpenAIRE

    Xu, Chenjie; Xie, Jin; Kohler, Nathan; Walsh, Edward G; Chin, Y. Eugene; Sun, Shouheng

    2008-01-01

    Functionalization of monodisperse superparamagnetic magnetite (Fe3O4) nanoparticles for cell specific targeting is crucial for cancer diagnostics and therapeutics. Targeted magnetic nanoparticles can be used to enhance the tissue contrast in magnetic resonance imaging (MRI), to improve the efficiency in anticancer drug delivery, and to eliminate tumor cells by magnetic fluid hyperthermia. Herein we report the nucleus-targeting Fe3O4 nanoparticles functionalized with protein and nuclear locali...

  15. On-site storage of high level nuclear waste: Attitudes and perceptions of local residents

    International Nuclear Information System (INIS)

    No public policy issue has been as difficult as high-level nuclear waste. Debates continue regarding Yucca Mountain as a disposal site, and - more generally - the appropriateness of geologic disposal and the need to act quickly. Previous research has focused on possible social, political, and economic consequences of a facility in Nevada. Impacts have been predicted to be potentially large and to emanate mainly from stigmatization of the region due to increased perceptions of risk. Analogous impacts from leaving waste at power plants have been either ignored or assumed to be negligible. This paper presents survey results on attitudes of residents in three countries where nuclear waste is currently stored. Topics include perceived risk, knowledge of nuclear waste and radiation, and impacts on jobs, tourism, and housing values from leaving waste on site. Results are similar to what has been reported for Nevada; the public is concerned about possible adverse effects from on-site storage of waste. 24 refs., 7 figs., 5 tabs

  16. Construction and characteristics of a transformed lepidopteran cell clone expressing baculovirus p35

    Institute of Scientific and Technical Information of China (English)

    ZHENG Guiling; LI Changyou; LI Guoxun; WANG Ping; Robert R. Granados

    2005-01-01

    A transformed cell line was constructed from Mythimna separata cells Ms7311 by lipofection method. TMs7311 cells were generated using a double selection technique involving a selection in the antibiotic Zeocin, followed by a second round of selection by exhibiting cell characterization. A cell clone expressing p35 was obtained with high level of AcMNPV and recombinant proteins. Compared with wild type Ms7311 cells, the cell clone showed increased resistance to Actinamycin D-induced apoptosis and a profound resistance to nutrient development (PBS). When the cell clone was infected with recombinant baculoviruses expressing secreted alkaline phosphatase (SEAP) and β-galactosi- dase, expression of the recombinant proteins from TMs7311 cells exceeded that from parental Ms7311 cells. Production of budded virus and occlusion body was significantly higher than that from parental cells Ms7311.

  17. Production of CCHF Virus-Like Particle by a Baculovirus-Insect Cell Expression System

    Institute of Scientific and Technical Information of China (English)

    Zhao-rui Zhou; Man-li Wang; Fei Deng; Tian-xian Li; Zhi-hong Hu; Hua-fin Wang

    2011-01-01

    Crimean-Congo Haemorrhagic Fever Virus(CCHFV)is a tick-born virus of the Nairovirus genus within the Bunyaviridae family,which is widespread and causes,high fatality. The nucleocapsid of CCHFV is comprised of N proteins that are encoded by the S segment. In this research,the N protein of CCHFV was expressed in insect cells using a recombinant baculovirus. Under an electron microscope,Virus-Like Particles (VLPs)with various size and morphology were observed in cytoplasmic vesicles in the infected cells.Sucrose-gradient purification of the cell lysate indicated that the VLPs were mainly located in the upper fraction after ultracentrifugation,which was confirmed by Western blot analysis and immuno-electron microscopy(IEM).

  18. Efficient production of type 2 porcine circovirus-like particles by a recombinant baculovirus.

    Science.gov (United States)

    Liu, Lan-Jun; Suzuki, Takako; Tsunemitsu, Hiroshi; Kataoka, Michiyo; Ngata, Noriyo; Takeda, Naokazu; Wakita, Takaji; Miyamura, Tatsuo; Li, Tian-Cheng

    2008-01-01

    The capsid protein of PCV2 was expressed by using a recombinant baculovirus with insect Tn5 cells. A large amount of 28-kDa protein was released into the culture medium and self-assembled into PCV2-like particles (PCV2-LPs) with a buoyant density of 1.365 g/cm(3) and a diameter of 20 nm. PCV2-LPs were efficiently expressed, yielding 1 mg of purified particles per 10(7) Tn5 cells. The PCV2-LPs have antigenicity similar to that of authentic PCV2 particles, allowing us to develop a method for sensitively detecting PCV2-specific IgG antibodies. In addition, the PCV2-LPs appeared to be the most promising PCV2 vaccine candidate, by virtue of their potent immunogenicity. PMID:18998045

  19. Identification of a single-nucleocapsid baculovirus isolated from Clanis bilineata tsingtauica (Lepidoptera: Sphingidae).

    Science.gov (United States)

    Wang, Liqun; Yi, Jianping; Zhu, Shanying; Li, Bing; Chen, Yan; Shen, Weide; Wang, Wenbing

    2008-01-01

    A nucleopolyhedrovirus isolated from infected larvae of Clanis bilineata tsingtauica was characterized. Electron microscopical studies on the ultrastructure of C. bilineata nucleopolyhedrovirus (ClbiSNPV) occlusion bodies (OBs) showed several virions (up to 16) with a single nucleocapsid packaged within a single viral envelope. The diameter of the OBs was 0.77-1.7 mum with a mean of 1.13 +/- 0.19 mum. The complete sequence of the ClbiSNPV polyhedrin (polh) gene contained 741 nucleotides, predicting a protein of 246 amino acids. Phylogenetic analyses using the complete sequence of the polh genes indicated that ClbiSNPV clusters with Group II NPVs. This is the first record of a baculovirus from C. bilineata. PMID:18584114

  20. Construction of occluded recombinant baculoviruses containing the full-length cry1Ab and cry1Ac genes from Bacillus thuringiensis

    OpenAIRE

    Ribeiro, B M; Crook, N. E.

    1998-01-01

    The administration of baculoviruses to insects for bioassay purposes is carried out, in most cases, by contamination of food surfaces with a known amount of occlusion bodies (OBs). Since per os infection is the natural route of infection, occluded recombinant viruses containing crystal protein genes (cry1Ab and cry1Ac) from Bacillus thuringiensis were constructed for comparison with the baculovirus prototype Autographa californica nucleopolyhedrovirus (AcNPV). The transfer vector pAcUW2B was ...

  1. Hypercholesterolemia Increases the Production of Leukotriene B4 in Neutrophils by Enhancing the Nuclear Localization of 5-Lipoxygenase

    Directory of Open Access Journals (Sweden)

    Xiao-Feng Lai

    2014-11-01

    Full Text Available Aims: Neutrophils can synthesize leukotriene B4 (LTB4 by activating the 5-lipoxygenase (5-LOsignaling pathway. LTB4 is a pro-inflammatory mediator associated with the etiology and progression of atherosclerosis. It can increase function and number of neutrophils in an autocrine manner. Since hypercholesterolemia is associated with an increase in the number and function of neutrophils, we hypothesized that this effect could be mediated through increased production of LTB4 in neutrophils. Methods/Results: Hypercholesterolemia was modeled in Wistar rats by feeding them with a high cholesterol diet. The induction of hypercholesterolemia caused an increase in the plasma levels of LTB4, following lipopolysaccharide stimulation. This effect was recapitulated in vitro, both in the presence and absence of stimulation with the activator of 5-LO, A23187. Neutrophils in hypercholesterolemia rats expressed similar total levels of 5-LO as control rats, but displayed increased nuclear localization of 5-LO, as well as elevated levels of phosphorylated 5-LO and ERK1/2. In vitro, MβCD/cholesterol complexes enriched cholesterol in neutrophils, resulted in similar changes in 5-LO/LTB4. In addition, these alterations could be inhibited with the ERK inhibitor PD98059. Conclusion: Hypercholesterolemia increases LTB4 production in neutrophils by increasing the nuclear localization of 5-LO, which is the result of its phosphorylation by activated ERK1/2.

  2. Localization of an irregular power flux increase source in the core of a nuclear reactor with the use of a special parameter

    International Nuclear Information System (INIS)

    An algorithm of localization of a local neutron flux increase source in the core of a nuclear reactor is described. The algorithm allows determining the position of the neutron flux source using the signals of the three or more detectors. The detectors are fission chambers distributed under the bottom of the core

  3. Construction of recombinant baculovirus vaccines for Newcastle disease virus and an assessment of their immunogenicity.

    Science.gov (United States)

    Ge, Jingping; Liu, Ying; Jin, Liying; Gao, Dongni; Bai, Chengle; Ping, Wenxiang

    2016-08-10

    Newcastle disease (ND) is a lethal avian infectious disease caused by Newcastle disease virus (NDV) which poses a substantial threat to China's poultry industry. Conventional live vaccines against NDV are available, but they can revert to virulent strains and do not protect against mutant strains of the virus. Therefore, there is a critical unmet need for a novel vaccine that is safe, efficacious, and cost effective. Here, we designed novel recombinant baculovirus vaccines expressing the NDV F or HN genes. To optimize antigen expression, we tested the incorporation of multiple regulatory elements including: (1) truncated vesicular stomatitis virus G protein (VSV-GED), (2) woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), (3) inverted terminal repeats (ITRs) of adeno-associated virus (AAV Serotype II), and (4) the cytomegalovirus (CMV) promoter. To test the in vivo efficacy of the viruses, we vaccinated chickens with each construct and characterized the cellular and humoral immune response to challenge with virulent NDV (F48E9). All of the vaccine constructs provided some level of protection (62.5-100% protection). The F-series of vaccines provided a greater degree of protection (87.5-100%) than the HN-series (62.5-87.5%). While all of the vaccines elicited a robust cellular and humoral response subtle differences in efficacy were observed. The combination of the WPRE and VSV-GED regulatory elements enhanced the immune response and increased antigen expression. The ITRs effectively increased the length of time IFN-γ, IL-2, and IL-4 were expressed in the plasma. The F-series elicited higher titers of neutralizing antibody and NDV-specific IgG. The baculovirus system is a promising platform for NDV vaccine development that combines the immunostimulatory benefits of a recombinant virus vector with the non-replicating benefits of a DNA vaccine. PMID:27015979

  4. Baculovirus-mediated GCRV vp7 and vp6 genes expression in silkworm and grass carp.

    Science.gov (United States)

    Liu, Bo; Gong, Yongchang; Li, Zhen; Hu, Xiaolong; Cao, Guangli; Xue, Renyu; Gong, Chengliang

    2016-06-01

    Grass carp hemorrhagic disease is a common fish disease and often results in significant economic losses in grass carp aquaculture in China. This study was aimed to develop a novel oral vaccine against grass carp reovirus (GCRV). GCRV vp6 and vp7 genes with β-actin promoter of Megalobrama amblycephala and polyhedrin promoter (Ph10) of baculovirus, respectively, were cloned into plasmid pFast™-Dual to construct a vector pFast-PHVP7-AVP6, which was used to generate a recombinant baculovirus BacFish-vp6/vp7 via Bac-to-Bac system. The VP7 expression was analyzed from freeze-dried powder of the BacFish-vp6/vp7-infected silkworm pupae by western blotting, and VP6 expression was analyzed from orally vaccinated fish with the freeze-dried powder by RT-PCR. The VP6 expression was also analyzed from both CIK cells transduced with BacFish-vp6/vp7 and tissues of vaccinated fish by immunofluorescence analysis. Recombinant VP7 could be detected from the BacFish-vp6/vp7-infected silkworm pupae. Pathological changes were not observed in CIK cells transduced with BacFish-vp6/vp7, and VP6 expression was found in CIK cells. When the grass carps were orally administrated with the freeze-dried powder, vp6 gene transcription was found in blood of the vaccinated fishes and VP6 protein was observed in liver and kidney of the vaccinated fish by immunofluorescence analysis. These results indicated that vp7 gene was expressed in the BacFish-vp6/vp7-infected silkworm and vp6 gene was expressed in orally vaccinated fish with freeze-dried powder of the BacFish-vp6/vp7-infected silkworm pupae, suggesting the possibility to use the powder as an orally administrated vaccine. PMID:27085857

  5. Nuclear localization and cytosolic overexpression of LASP-1 correlates with tumor size and nodal-positivity of human breast carcinoma

    Directory of Open Access Journals (Sweden)

    Dietl Johannes

    2007-10-01

    Full Text Available Abstract Background LIM and SH3 protein 1 (LASP-1, initially identified from human breast cancer, is a specific focal adhesion protein involved in cell proliferation and migration, which was reported to be overexpressed in 8–12 % of human breast cancers and thought to be exclusively located in cytoplasm. Methods In the present work we analyzed the cellular and histological expression pattern of LASP-1 and its involvement in biological behavior of human breast cancer through correlation with standard clinicopathological parameters and expression of c-erbB2 (HER-2/neu, estrogen- (ER and progesterone-receptors (PR. For this purpose immunohistochemical staining intensity and percentage of stained cells were semi-quantitatively rated to define a LASP-1 immunoreactive score (LASP-1-IRS. LASP-1-IRS was determined in 83 cases of invasive ductal breast carcinomas, 25 ductal carcinomas in situ (DCIS and 18 fibroadenomas. Cellular LASP-1 distribution and expression pattern was visualized by immunofluorescence and confocal microscopy and assessed through separate Western blots of nuclear and cytosol preparations of BT-20, MCF-7, MDA-MB231, and ZR-75/1 breast cancer cells. Results Statistical analysis revealed that the resulting LASP-1-IRS was significantly higher in invasive carcinomas compared to fibroadenomas (p = 0.0176. Strong cytoplasmatic expression of LASP-1 was detected in 55.4 % of the invasive carcinomas, which correlated significantly with nuclear LASP-1-positivity (p = 0.0014, increased tumor size (p = 0.0159 and rate of nodal-positivity (p = 0.0066. However, levels of LASP-1 expression did not correlate with average age at time point of diagnosis, histological tumor grading, c-erbB2-, ER- or PR-expression. Increased nuclear localization and cytosolic expression of LASP-1 was found in breast cancer with higher tumor stage as well as in rapidly proliferating epidermal basal cells. Confocal microscopy and separate Western blots of cytosolic and

  6. Nuclear localization and cytosolic overexpression of LASP-1 correlates with tumor size and nodal-positivity of human breast carcinoma

    International Nuclear Information System (INIS)

    LIM and SH3 protein 1 (LASP-1), initially identified from human breast cancer, is a specific focal adhesion protein involved in cell proliferation and migration, which was reported to be overexpressed in 8–12 % of human breast cancers and thought to be exclusively located in cytoplasm. In the present work we analyzed the cellular and histological expression pattern of LASP-1 and its involvement in biological behavior of human breast cancer through correlation with standard clinicopathological parameters and expression of c-erbB2 (HER-2/neu), estrogen- (ER) and progesterone-receptors (PR). For this purpose immunohistochemical staining intensity and percentage of stained cells were semi-quantitatively rated to define a LASP-1 immunoreactive score (LASP-1-IRS). LASP-1-IRS was determined in 83 cases of invasive ductal breast carcinomas, 25 ductal carcinomas in situ (DCIS) and 18 fibroadenomas. Cellular LASP-1 distribution and expression pattern was visualized by immunofluorescence and confocal microscopy and assessed through separate Western blots of nuclear and cytosol preparations of BT-20, MCF-7, MDA-MB231, and ZR-75/1 breast cancer cells. Statistical analysis revealed that the resulting LASP-1-IRS was significantly higher in invasive carcinomas compared to fibroadenomas (p = 0.0176). Strong cytoplasmatic expression of LASP-1 was detected in 55.4 % of the invasive carcinomas, which correlated significantly with nuclear LASP-1-positivity (p = 0.0014), increased tumor size (p = 0.0159) and rate of nodal-positivity (p = 0.0066). However, levels of LASP-1 expression did not correlate with average age at time point of diagnosis, histological tumor grading, c-erbB2-, ER- or PR-expression. Increased nuclear localization and cytosolic expression of LASP-1 was found in breast cancer with higher tumor stage as well as in rapidly proliferating epidermal basal cells. Confocal microscopy and separate Western blots of cytosolic and nuclear preparations confirmed nuclear

  7. Internal ribosome entry site of Rhopalosiphum padi virus is functional in mammalian cells and has cryptic promoter activity in baculovirus-in fected Sf21 cells

    Institute of Scientific and Technical Information of China (English)

    Yi-jane WU; Chao-yi TENG; Yu-jie CHEN; Seng-chi CHEN; Ying-ju CHEN; Yi-ting LIN; Tzong-yuan WU

    2008-01-01

    Aim: To substantiate the in vitro translational studies of a cross-kingdom, inter- nal ribosome entry site (IRES), the 5"untranslated region of the Rhopalosiphum padi virus (RhPV), can function in mammalian cells and act as a shuttle IRES between insect cells and mammalian cells. Methods: Cytomegalovirus (CMV) promoter-based bicistronic mammalian cell expression vectors, either in plasmids or baculovirus vectors, were generated. Plasmid transient transfection and baculovirus transduction assays were performed to test whether the RhPV IRES can mediate translation activity in versatile mammalian cell lines. Results: Both plasmids and recombinant baculoviruses containing the CMV promoter and the RhPV IRES can mediate bicistronic gene expression in mammalian cells. However, in the CMV promoter containing recombinant baculovirus-infected insect Sf21 cells, only the second cistron gene expression was observed. Northern blot analysis and a promoterless assay demonstrated that the RhPV IRES exhibited cryptic promoter activity in baculovirus-infected insect cells. Conclusion: RhPV IRES can mediate gene expression in both insect cells and mammalian cells, and this characteristic of the RhPV IRES will facilitate the development of a bicistronic baculovirus gene therapy vectors.

  8. Nuclear localization of phosphorylated c-Myc protein in human tumor cells.

    Directory of Open Access Journals (Sweden)

    C. Soldani

    2010-05-01

    Full Text Available Using immunocytochemical techniques at light and electron microscopy, we analysed the distribution of phosphorylated c-Myc in actively proliferating human HeLa cells. The distribution pattern of c-Myc was also compared with those of other ribonucleoprotein (RNP-containing components (PANA, hnRNP-core proteins, fibrillarin or RNP-associated nuclear proteins (SC-35 splicing factor. Our results provide the first evidence that phosphorylated c-Myc accumulates in the nucleus of tumor cells, where it colocalizes with fibrillarin, both in the nucleolus and in extranucleolar structures.

  9. Sizewell nuclear power station: investigation of radiation exposure pathways from liquid effluents. Local habits survey 1981

    International Nuclear Information System (INIS)

    A habits and consumption survey to review radiation exposure pathways due to liquid effluents released from the CEGB Sizewell site is described. It is relevant to both the Sizewell A and proposed Sizewell B nuclear power stations. The main objectives are to provide input data to a radiological assessment by means of identifying critical groups and to provide data for guidance in a review of environmental monitoring programmes. The way in which data for the different pathways should be combined in order to aid the subsequent radiological assessment is discussed. Recommendations are made for adjustments to the present monitoring programmes. (U.K.)

  10. A localized navigation algorithm for radiation evasion for nuclear facilities: Optimizing the “Radiation Evasion” criterion: Part I

    International Nuclear Information System (INIS)

    Highlights: ► A new navigation algorithm for radiation evasion around nuclear facilities. ► An optimization criteria minimized under algorithm operation. ► A man-borne device guiding the occupational worker towards paths that warrant least radiation × time products. ► Benefits of using localized navigation as opposed to global navigation schemas. ► A path discrimination function for finding the navigational paths exhibiting the least amounts of radiation. -- Abstract: In this paper, we introduce a navigation algorithm having general utility for occupational workers at nuclear facilities and places where radiation poses serious health hazards. This novel algorithm leverages the use of localized information for its operation. Therefore, the need for central processing and decision resources is avoided, since information processing and the ensuing decision-making are done aboard a man-borne device. To acquire the information needed for path planning in radiation avoidance, a well-designed and distributed wireless sensory infrastructure is needed. This will automatically benefit from the most recent trends in technology developments in both sensor networks and wireless communication. When used to navigate based on local radiation information, the algorithm will behave more reliably when accidents happen, since no long-haul communication links are required for information exchange. In essence, the proposed algorithm is designed to leverage nearest neighbor information coming in through the sensory network overhead, to compute successful navigational paths from one point to another. The proposed algorithm is tested under the “Radiation Evasion” criterion. It is also tested for the case when more information, beyond nearest neighbors, is made available; here, we test its operation for different numbers of step look-ahead. We verify algorithm performance by means of simulations, whereby navigational paths are calculated for different radiation fields

  11. Nuclear localization of Vpr is crucial for the efficient replication of HIV-1 in primary CD4+ T cells

    International Nuclear Information System (INIS)

    The human immunodeficiency virus type 1 (HIV-1) accessory protein Vpr appears to make a substantial contribution to the replication of HIV-1 in established T cell lines when HIV-1 is present at very low multiplicities of infection. However, the role of Vpr in viral replication in primary CD4+ T cells remains to be clarified. In this study, we generated a panel of viruses that encoded mutant forms of Vpr that lacked either the ability to accumulate in the nucleus and induce G2 arrest or the ability to induce apoptosis, which has been shown to occur independently of G2 arrest of the cell cycle. We demonstrate here that the nuclear localization of Vpr and consequent G2 arrest but not the induction of apoptosis by Vpr are important for viral replication in primary CD4+ T cells at both high and low multiplicities of infection. Viruses that encoded mutant forms of Vpr that failed to be imported into the nucleus in the presence of cytoplasmic extracts from primary CD4+ T cells in an in vitro nuclear import assay replicated at drastically reduced rates. Thus, Vpr might be a key regulator of the viral nuclear import process during infection in primary CD4+ T cells. By contrast, a mutant form of Vpr that exhibited diffuse cytosolic staining exclusively in an immunofluorescence assay of HeLa cells and was not imported into nucleus by the cytosol from HeLa cells was effectively imported into the nucleus by cytosol from primary CD4+ T cells. This Vpr mutant virus replicated well in primary CD4+ T cells, indicating that cellular factors in primary CD4+ T cells are indispensable for the accumulation of Vpr in the nucleus and, thus, for viral replication. Our results suggest that the nuclear import of Vpr might be a good target in efforts to block the early stages of replication of HIV-1

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

    Directory of Open Access Journals (Sweden)

    Aviad Levin

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

  13. Nuclear localization of Obox4 is dependent on its homeobox domain

    OpenAIRE

    Park, Geon Tae; Lee, Kyung-Ah

    2013-01-01

    Objective Oocyte-specific homeobox 4 (Obox4) is preferentially expressed in oocytes and plays an important role in the completion of meiosis of oocytes. However, the Obox4 expression pattern has not been reported yet. In this study, we investigated the subcellular localization of Obox4 using a green fluorescent protein (GFP) fusion expression system. Methods Three regions of Obox4 were divided and fused to the GFP expression vector. The partly deleted homeodomain (HD) regions of Obox4 were al...

  14. Effect of local and large-scale environments on nuclear activity and star formation

    CERN Document Server

    Argudo-Fernández, M; Sabater, J; Puertas, S Duarte; Verley, S; Yang, X

    2016-01-01

    Active galactic nuclei (AGN) is one of the main drivers for transition from star-forming disk to passive spheroidal galaxies. However, the role of large-scale environment versus one-on-one interactions in triggering different types of AGN is still uncertain. We present a statistical study of the prevalence of the nuclear activity in isolated galaxies and physically bound isolated pairs. For the purpose of this study we considered optically and radio selected nuclear activity types. We aim to assess the effect of one-on-one interaction on the fraction of AGN and the role of their large-scale environment. To study the effect of one-on-one interaction on the fraction of AGN in isolated galaxy pairs, we compare with a sample of isolated galaxies homogeneously selected under the same isolation criterion. We examine the effect of the large-scale environment by comparing with control samples of single galaxies and galaxy pairs. In general we found no difference in the prevalence of optical AGN for the considered sam...

  15. Lysine 271 but not lysine 210 of XRCC4 is required for the nuclear localization of XRCC4 and DNA ligase IV

    Energy Technology Data Exchange (ETDEWEB)

    Fukuchi, Mikoto; Wanotayan, Rujira; Liu, Sicheng; Imamichi, Shoji; Sharma, Mukesh Kumar; Matsumoto, Yoshihisa, E-mail: yoshim@nr.titech.ac.jp

    2015-06-12

    XRCC4 and DNA Ligase IV (LIG4) cooperate to join two DNA ends at the final step of DNA double-strand break (DSB) repair through non-homologous end-joining (NHEJ). However, it is not fully understood how these proteins are localized to the nucleus. Here we created XRCC4{sup K271R} mutant, as Lys271 lies within the putative nuclear localization signal (NLS), and XRCC4{sup K210R} mutant, as Lys210 was reported to undergo SUMOylation, implicated in the nuclear localization of XRCC4. Wild-type and mutated XRCC4 with EGFP tag were introduced into HeLa cell, in which endogenous XRCC4 had been knocked down using siRNA directed to 3′-untranslated region, and tested for the nuclear localization function by fluorescence microscopy. XRCC4{sup K271R} was defective in the nuclear localization of itself and LIG4, whereas XRCC4{sup K210R} was competent for the nuclear localization with LIG4. To examine DSB repair function, wild-type and mutated XRCC4 were introduced into XRCC4-deficient M10. M10-XRCC4{sup K271R}, but not M10-XRCC4{sup K210R}, showed significantly reduced surviving fraction after 2 Gy γ-ray irradiation as compared to M10-XRCC4{sup WT}. The number of γ-H2AX foci remaining 2 h after 2 Gy γ-ray irradiation was significantly greater in M10-XRCC4{sup K271R} than in M10-XRCC4{sup WT}, whereas it was only marginally increased in M10-XRCC4{sup K210R} as compared to M10-XRCC4{sup WT}. The present results collectively indicated that Lys271, but not Lys210, of XRCC4 is required for the nuclear localization of XRCC4 and LIG4 and that the nuclear localizing ability is essential for DSB repair function of XRCC4. - Highlights: • XRCC4{sup K271R} is defective in the nuclear localization of itself and LIG4. • XRCC4{sup K210R} is competent for the nuclear localization of itself and LIG4. • XRCC4{sup K271R} is deficient in DSB repair function. • XRCC4{sup K210R} is mostly normal in DSB repair function.

  16. Lysine 271 but not lysine 210 of XRCC4 is required for the nuclear localization of XRCC4 and DNA ligase IV

    International Nuclear Information System (INIS)

    XRCC4 and DNA Ligase IV (LIG4) cooperate to join two DNA ends at the final step of DNA double-strand break (DSB) repair through non-homologous end-joining (NHEJ). However, it is not fully understood how these proteins are localized to the nucleus. Here we created XRCC4K271R mutant, as Lys271 lies within the putative nuclear localization signal (NLS), and XRCC4K210R mutant, as Lys210 was reported to undergo SUMOylation, implicated in the nuclear localization of XRCC4. Wild-type and mutated XRCC4 with EGFP tag were introduced into HeLa cell, in which endogenous XRCC4 had been knocked down using siRNA directed to 3′-untranslated region, and tested for the nuclear localization function by fluorescence microscopy. XRCC4K271R was defective in the nuclear localization of itself and LIG4, whereas XRCC4K210R was competent for the nuclear localization with LIG4. To examine DSB repair function, wild-type and mutated XRCC4 were introduced into XRCC4-deficient M10. M10-XRCC4K271R, but not M10-XRCC4K210R, showed significantly reduced surviving fraction after 2 Gy γ-ray irradiation as compared to M10-XRCC4WT. The number of γ-H2AX foci remaining 2 h after 2 Gy γ-ray irradiation was significantly greater in M10-XRCC4K271R than in M10-XRCC4WT, whereas it was only marginally increased in M10-XRCC4K210R as compared to M10-XRCC4WT. The present results collectively indicated that Lys271, but not Lys210, of XRCC4 is required for the nuclear localization of XRCC4 and LIG4 and that the nuclear localizing ability is essential for DSB repair function of XRCC4. - Highlights: • XRCC4K271R is defective in the nuclear localization of itself and LIG4. • XRCC4K210R is competent for the nuclear localization of itself and LIG4. • XRCC4K271R is deficient in DSB repair function. • XRCC4K210R is mostly normal in DSB repair function

  17. Understanding of the characteristics of the local newspapers providing media coverage on the matters of nuclear energy in the regions where nuclear facilities are located. Based on analysis of the media reports and interviews with journalists

    International Nuclear Information System (INIS)

    Taking into consideration the influence of the media coverage, this research aims to analyze the characteristics of the local newspapers that cover diverse events relevant to nuclear energy in regional areas where nuclear facilities are located (hereinafter called the 'region'). According to the previous surveys, local residents in the region are more interested in the nuclear energy matters than those who live in urban areas. Plus, the local newspapers turn out to report more events of nuclear energy from a variety of angles. Through interviews with executives and journalists of the local newspaper companies in the regions, it is revealed that the local newspapers tend not to report news sensationally, but they would rather take a supportive stance toward the development in their regions. The interviewees hope that various activities of the nuclear industry will promote education, employment and cooperation among government, industry and academia. They also desire that the industry's activities will help to increase benefits in their regions. It appears that the interviewees' awareness reflects articles of the local newspapers. As a result of the surveys conducted for this research, it is considered that the journalists expect that their region will make particularly qualitative progress in the future. (author)

  18. Compatibility of localized wave packets and unrestricted single particle dynamics for cluster formation in nuclear collisions

    International Nuclear Information System (INIS)

    Anti-symmetrized molecular dynamics with quantum branching is generalized so as to allow finite time duration of the unrestricted coherent mean field propagation which is followed by the decoherence into wave packets. In this new model, the wave packet shrinking by the mean field propagation is respected as well as the diffusion, so that it predicts a one-body dynamics similar to that in mean field models. The shrinking effect is expected to change the diffusion property of nucleons in nuclear matter and the global one-body dynamics. The central 129Xe + Sn collisions at 50 MeV/nucleon are calculated by the models with and without shrinking, and it is shown that the inclusion of the wave packet shrinking has a large effect on the multifragmentation in a big expanding system with a moderate expansion velocity. (author)

  19. The human papillomavirus (HPV) E6 oncoproteins promotes nuclear localization of active caspase 8

    International Nuclear Information System (INIS)

    The HPV-16 E6 and E6⁎ proteins have been shown previously to be capable of regulating caspase 8 activity. We now show that the capacity of E6 to interact with caspase 8 is common to diverse HPV types, being also seen with HPV-11 E6, HPV-18 E6 and HPV-18 E6⁎. Unlike most E6-interacting partners, caspase 8 does not appear to be a major proteasomal target of E6, but instead E6 appears able to stimulate caspase 8 activation, without affecting the overall apoptotic activity. This would appear to be mediated in part by the ability of the HPV E6 oncoproteins to recruit active caspase 8 to the nucleus. - Highlights: • Multiple HPV E6 oncoproteins interact with the caspase 8 DED domain. • HPV E6 stimulates activation of caspase 8. • HPV E6 promotes nuclear accumulation of caspase 8

  20. Study of community leaders in a nuclear host community: local issues, expectations and support and opposition

    International Nuclear Information System (INIS)

    A study of community leaders was undertaken in Hartsville, Tennessee, site of the TVA Hartsville Nuclear Power Plant currently under construction. Leaders were found to be extremely supportive of the plant and of TVA's efforts to mitigate impacts expected to result from construction. Like their citizen counterparts, leaders expect economic benefits and some growth-related disruption to occur as a result of the plant, while environmental impacts are seen as extremely unlikely to occur. Plant-related issues, such as housing availability and traffic congestion, dominate leaders' thinking about current issues. These issues are expected to continue to be important in the future, and new issues dealing with growth and planning, employment and taxation are expected to arise

  1. The human papillomavirus (HPV) E6 oncoproteins promotes nuclear localization of active caspase 8

    Energy Technology Data Exchange (ETDEWEB)

    Manzo-Merino, Joaquin [Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México. Av. San Fernando No. 22, Col. Sección XVI, Tlalpan 14080 (Mexico); Massimi, Paola [International Centre for Genetic Engineering and Biotechnology, Padriciano 99, I-34149 Trieste (Italy); Lizano, Marcela, E-mail: lizanosoberon@gmail.com [Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México. Av. San Fernando No. 22, Col. Sección XVI, Tlalpan 14080 (Mexico); Banks, Lawrence, E-mail: banks@icgeb.org [International Centre for Genetic Engineering and Biotechnology, Padriciano 99, I-34149 Trieste (Italy)

    2014-02-15

    The HPV-16 E6 and E6{sup ⁎} proteins have been shown previously to be capable of regulating caspase 8 activity. We now show that the capacity of E6 to interact with caspase 8 is common to diverse HPV types, being also seen with HPV-11 E6, HPV-18 E6 and HPV-18 E6{sup ⁎}. Unlike most E6-interacting partners, caspase 8 does not appear to be a major proteasomal target of E6, but instead E6 appears able to stimulate caspase 8 activation, without affecting the overall apoptotic activity. This would appear to be mediated in part by the ability of the HPV E6 oncoproteins to recruit active caspase 8 to the nucleus. - Highlights: • Multiple HPV E6 oncoproteins interact with the caspase 8 DED domain. • HPV E6 stimulates activation of caspase 8. • HPV E6 promotes nuclear accumulation of caspase 8.

  2. Local decision-making facing issues of national interest experiences from the swedish siting process for a spent nuclear fuel repository

    International Nuclear Information System (INIS)

    It is common knowledge that there are difficulties in convincing the general public and their democratically elected representatives that final disposal of spent nuclear fuel can be made in safe way. Special problems for the decision-makers are created by the demands put on today's generations to make a responsible risk assessment in a area with genuine uncertainties and characterised by any expressions of lack of confidence in social institutions. The current Swedish process for siting a deep repository for spent nuclear fuel has evolved during a period of many years, through inputs by the industry, Government, regulatory authorities and concerned municipalities. It is clear that the nuclear industry, represented by the Swedish Nuclear Fuel and Waste Management CO (SKB), has the full responsibility to find a solution to the waste management problem and to implement the solution - and to for this under the supervision of Government and regulating authorities. But, given the strong tradition of local self-government, the concerned municipalities, the local population in this process. this is simply the following fact: For people who have engaged themselves in local politics - and are prepared to take their responsibility for the well-being and development of their local community - the issue of a possible nuclear repository in the neighbourhood is difficult to handle. A relevant question is: Why should the nation as a whole expect these locally elected representatives to feel a responsibility for an issue of national importance? (author)

  3. Novel strategies to exploit existing natural infections: synergisms between baculoviruses and other toxins. Research Project Final Report

    OpenAIRE

    Hesketh, Helen; Hails, Rosemary

    2007-01-01

    EXECUTIVE SUMMARY Baculoviruses are a well studied group of insect viruses which have known advantages as biological control agents of insect pests in agriculture and forestry; strains used are very specific to the insect species they can infect, they can be formulated and sprayed conventionally and they are easily broken down by UV thereby leaving minimal non-toxic residues. They are the major group of viruses which infect insects but they do not replicate in vertebrates, plants or other ...

  4. Structured modeling of recombinant protein production in batch and fed-batch culture of baculovirus-infected insect cells

    OpenAIRE

    Jang, J. D.; Sanderson, C.S.; Chan, L. C. L.; Barford, J. P.; Reid, S

    2000-01-01

    The infection of insect cells with baculovirus was described in a mathematical model as a part of the structured dynamic model describing whole animal cell metabolism. The model presented here is capable of simulating cell population dynamics, the concentrations of extracellular and intracellularviral components, and the heterologous product titers. The model describes the whole processes of viral infection and theeffect of the infection on the host cell metabolism. Dynamic simulation of the ...

  5. Pathogen Persistence in the Environment and Insect-Baculovirus Interactions: Disease-Density Thresholds, Epidemic Burnout and Insect Outbreaks

    OpenAIRE

    Fuller, Emma; Elderd, Bret D.; Dwyer, Greg

    2012-01-01

    Classical epidemic theory focuses on directly transmitted pathogens, but many pathogens are instead transmitted when hosts encounter infectious particles. Theory has shown that for such diseases pathogen persistence time in the environment can strongly affect disease dynamics, but estimates of persistence time, and consequently tests of the theory, are extremely rare. We consider the consequences of persistence time for the dynamics of the gypsy moth baculovirus, a pathogen transmitted when l...

  6. Improvement of traditional local rice varieties through induced mutations using nuclear techniques

    International Nuclear Information System (INIS)

    'Improvement of local rice varieties for high yield, resistance to disease and insect pests (brown plant hopper and rice blast) and export quality through induced mutations for the Mekong Delta' started in 1993. After six years, it showed effecting on the field in the MD as well as at the south of Vietnam. TNDB-100 manifest very wide adaptation and yield stable variety. THDB is suitable for deepwater rice region, coastal area, where rice cultivation effected by acid sulphate and salinity conditions. Both varieties are good example for the method. Thank to good Co-operation from extension center from provinces, hundred classes of extension were organized to recommend to the farmers. And thank to the strongly supporting from IAEA so that nearly 400,000 ha of TNDB-100 occupied at the south of Vietnam as well as nearly 15,000 ha of THDB grown in the coastal as well as rainfed lowland rice areas at the South of Vietnam. To continue the rice improvement by this technique, seeds of six traditional local varieties were exposed under different dose of gamma rays to create new mutants. At present day hundred improved breeding lines were selected, a dozen of uniform lines were isolated and entranced the yield trail as well as regional testing program. From these improved varieties would be selected to contribute to the rice cultivation at the south of Vietnam in the next years. (author)

  7. Measurement of local dose rates in controlled areas of nuclear power plants. Version 6/85

    International Nuclear Information System (INIS)

    Local dose rate measurements according to this regulation, serve the following purposes: Development of preventive measures for radiation protection with respect to maintenance and modifications in controlled areas, e.g. determination of individual measures for personnel protection, development of flow charts and dose rate estimates; monitoring of the personnel's location of activity within the controlled area, including determination of time-dependence of local dose rates and determination of immediate radiation protection measures required; limitation and identification of controlled areas. Such measurements are performed to ensure that the principles of radiation protection as stated in Art. 28 (1) of the Radiation Protection Ordinance with respect to external radiation exposure of the personnel, and the requirements of Art. 61 (1) of the Radiation Protection Ordinance relating to controlled areas are met and the limitation and identification of controlled areas as prescribed in Art 57 (1) of the Radiation Protection Ordinance can be performed. They are not used to determine body doses as per Art. 63 of the Radiation Protection Ordinance. (orig./HP)

  8. An apoptosis-inhibiting gene from a nuclear polyhedrosis virus encoding a polypeptide with Cys/His sequence motifs.

    OpenAIRE

    Birnbaum, M. J.; Clem, R J; Miller, L K

    1994-01-01

    Two different baculovirus genes are known to be able to block apoptosis triggered upon infection of Spodoptera frugiperda cells with p35 mutants of the insect baculovirus Autographa californica nuclear polyhedrosis virus (AcMNPV):p35 (P35-encoding gene) of AcMNPV (R. J. Clem, M. Fechheimer, and L. K. Miller, Science 254:1388-1390, 1991) and iap (inhibitor of apoptosis gene) of Cydia pomonella granulosis virus (CpGV) (N. E. Crook, R. J. Clem, and L. K. Miller, J. Virol. 67:2168-2174, 1993). Us...

  9. IRES mediated expression of viral 3C protease for enhancing the yield of FMDV empty capsids using baculovirus system.

    Science.gov (United States)

    Vivek Srinivas, V M; Basagoudanavar, Suresh H; Hosamani, Madhusudan

    2016-03-01

    For expression of FMDV empty capsids, high protease activity associated with 3C co-expressed with P1 polyprotein has been reported to adversely affect the yields of capsids. Limiting the levels of 3Cpro relative to P1-2A polypeptide is thus critical to enhance the yields. In this study, FMDV internal ribosome entry site (IRES) sequence which serves as an alternative to the CAP-dependent translation initiation mechanism, was used for controlled translation of 3C protease. Baculovirus expressing bicistronic cDNA cassette containing two open reading frames-FMDV capsid gene (P1-2A) and 3Cpro intervened by IRES was prepared. Analysis of the expression in insect cells infected with baculovirus showed increased accumulation of processed capsids. Recombinant capsids showed higher immunoreactivity similar to the whole virus antigen, when reacted with polyclonal antibodies against the purified whole virus 146S particles. Thus, inclusion of the IRES upstream of 3Cpro facilitated reduced expression of the protease in baculovirus expression system, without causing significant proteolysis, thereby contributing to improved yields of the processed capsid antigens. PMID:26775685

  10. Introduction of temperature-sensitive helper and donor plasmids into Bac-to-Bac baculovirus expression systems

    Institute of Scientific and Technical Information of China (English)

    Zhihong; Huang; Ao; Li; Mengjia; Pan; Wenbi; Wu; Meijin; Yuan; Kai; Yang

    2015-01-01

    In the baculovirus shuttle vector(bacmid) system, a helper plasmid and a donor plasmid are employed to insert heterologous genes into a cloned baculovirus genome via Tn7 transposition in Escherichia coli. The helper and donor plasmids are usually cotransfected with constructed bacmids into insect cells, which will lead to integration of these plasmids into the viral genome,and hence to the production of defective virions. In this study, to facilitate the preparation of plasmid-free recombinant bacmids, we modified a set of helper and donor plasmids by replacing their replication origins with that of a temperature-sensitive(ts) plasmid, p SIM6. Using the resulting ts helper plasmid p MON7124 ts and the ts donor plasmid p FB1ts-PH-GFP, a recombinant bacmid,b Ac WT-PG(-), was constructed, and the transposition efficiency was found to be 33.1%. The plasmids were then removed by culturing at 37 °C. For b Ac WT-PG(-), the infectious progeny virus titer and the protein expression level under the control of the polyhedrin promoter were similar to those of a bacmid constructed with unmodified helper and donor plasmids. These ts plasmids will be useful for obtaining plasmid-free bacmids for both heterologous protein production and fundamental studies of baculovirus biology.

  11. The conserved baculovirus protein p33 (Ac92) is a flavin adenine dinucleotide-linked sulfhydryl oxidase

    International Nuclear Information System (INIS)

    Open reading frame 92 of the Autographa californica baculovirus (Ac92) is one of about 30 core genes present in all sequenced baculovirus genomes. Computer analyses predicted that the Ac92 encoded protein (called p33) and several of its baculovirus orthologs were related to a family of flavin adenine dinucleotide (FAD)-linked sulfhydryl oxidases. Alignment of these proteins indicated that, although they were highly diverse, a number of amino acids in common with the Erv1p/Alrp family of sulfhydryl oxidases are present. Some of these conserved amino acids are predicted to stack against the isoalloxazine and adenine components of FAD, whereas others are involved in electron transfer. To investigate this relationship, Ac92 was expressed in bacteria as a His-tagged fusion protein, purified, and characterized both spectrophotometrically and for its enzymatic activity. The purified protein was found to have the color (yellow) and absorption spectrum consistent with it being a FAD-containing protein. Furthermore, it was demonstrated to have sulfhydryl oxidase activity using dithiothreitol and thioredoxin as substrates.

  12. Hydrogeological modelling of the eastern region of Areco river locally detailed on Atucha I and II nuclear power plants area

    International Nuclear Information System (INIS)

    Water flow behaviour of Pampeano aquifer was modeled using Visual Mod-flow software Package 2.8.1 with the assumption of a free aquifer, within the region of the Areco river and extending to the rivers of 'Canada Honda' and 'de la Cruz'. Steady state regime was simulated and grid refinement allows obtaining locally detailed calculation in the area of Atucha I and II Nuclear power plants, in order to compute unsteady situations as the consequence of water flow variations from and to the aquifer, enabling the model to study the movement of possible contaminant particles in the hydrogeologic system. In this work the effects of rivers action, the recharge conditions and the flow lines are analyzed, taking always into account the range of reliability of obtained results, considering the incidence of uncertainties introduced by data input system, the estimates and interpolation of parameters used. (author)

  13. Age determination of the nuclear stellar population of Active Galactic Nuclei using Locally Weighted Regression

    CERN Document Server

    Estrada-Piedra, T; Terlevich, R J; Fuentes, O; Terlevich, E; Estrada-Piedra, Trilce; Torres-Papaqui, Juan Pablo; Terlevich, Roberto; Fuentes, Olac; Terlevich, Elena

    2003-01-01

    We present a new technique to segregate old and young stellar populations in galactic spectra using machine learning methods. We used an ensemble of classifiers, each classifier in the ensemble specializes in young or old populations and was trained with locally weighted regression and tested using ten-fold cross-validation. Since the relevant information concentrates in certain regions of the spectra we used the method of sequential floating backward selection offline for feature selection. The application to Seyfert galaxies proved that this technique is very insensitive to the dilution by the Active Galactic Nucleus (AGN) continuum. Comparing with exhaustive search we concluded that both methods are similar in terms of accuracy but the machine learning method is faster by about two orders of magnitude.

  14. Local structure of LiB3O5 single crystal from 7Li nuclear magnetic resonance

    International Nuclear Information System (INIS)

    The local structure of LiB3O5 single crystal was investigated with 7Li (I=3/2) nuclear magnetic resonance measurements. We observed four different spectra, which could be divided into two groups corresponding to two kinds of lithium atoms, LiA and LiB, lying at crystallographically equivalent sites and magnetically inequivalent sites. From these results, the quadrupole coupling constants and the asymmetry parameters were determined at room temperature and are e2qQ/h=143±1 kHz and η=0.6±0.1 for 7Li. The directions of the principal axes of the electric field gradient tensors were also determined. The spectra for the two groups have the same principal values of the electric field gradient tensor, but different orientations, and originate from magnetically inequivalent sites. Also, the 7Li spin-lattice relaxation rate was measured, and the measured relaxation rate was found to be proportional to the temperature. The temperature-dependent single phonon process is considered to be more effective than the Raman process for nuclear quadrupole relaxation. In addition, we discuss the correlation between the asymmetry parameter and the largest nonlinear optical coefficient

  15. Structure of Importin-α from a Filamentous Fungus in Complex with a Classical Nuclear Localization Signal.

    Directory of Open Access Journals (Sweden)

    Natalia E Bernardes

    Full Text Available Neurospora crassa is a filamentous fungus that has been extensively studied as a model organism for eukaryotic biology, providing fundamental insights into cellular processes such as cell signaling, growth and differentiation. To advance in the study of this multicellular organism, an understanding of the specific mechanisms for protein transport into the cell nucleus is essential. Importin-α (Imp-α is the receptor for cargo proteins that contain specific nuclear localization signals (NLSs that play a key role in the classical nuclear import pathway. Structures of Imp-α from different organisms (yeast, rice, mouse, and human have been determined, revealing that this receptor possesses a conserved structural scaffold. However, recent studies have demonstrated that the Impα mechanism of action may vary significantly for different organisms or for different isoforms from the same organism. Therefore, structural, functional, and biophysical characterization of different Impα proteins is necessary to understand the selectivity of nuclear transport. Here, we determined the first crystal structure of an Impα from a filamentous fungus which is also the highest resolution Impα structure already solved to date (1.75 Å. In addition, we performed calorimetric analysis to determine the affinity and thermodynamic parameters of the interaction between Imp-α and the classical SV40 NLS peptide. The comparison of these data with previous studies on Impα proteins led us to demonstrate that N. crassa Imp-α possess specific features that are distinct from mammalian Imp-α but exhibit important similarities to rice Imp-α, particularly at the minor NLS binding site.

  16. Long isoform of ErbB3 binding protein, p48, mediates protein kinase B/Akt-dependent HDM2 stabilization and nuclear localization

    International Nuclear Information System (INIS)

    p48 is a long isoform of the ErbB3 binding protein that has oncogenic functions including promotion of carcinogenesis and induction of malignant transformation through negative regulation of tumor suppressor p53. Here, we show that high level of p48 protein expression leads to enhance HDM2 phosphorylation by Akt and inhibits the self-ubiquitination of HDM2 by up-regulation of Akt activity, thereby promoting its protein stability. Moreover, p48 expression leads to accumulated nuclear localization of HDM2, whereas p48 depletion disturbs its nuclear localization. Hence, higher expression of p48 in cancer cells reduces p53 levels through modulation of HDM2 nuclear localization and protein stability via regulation of its Akt-mediated phosphorylation.

  17. Introduction of the anti-apoptotic baculovirus p35 gene in passion fruit induces herbicide tolerance, reduced bacterial lesions, but does not inhibits passion fruit woodiness disease progress induced by cowpea aphid-borne mosaic virus (CABMV).

    Science.gov (United States)

    de Freitas, Daniele Scandiucci; Coelho, Marly C Felipe; Souza, Manoel T; Marques, Abi; Ribeiro, E Bergmann Morais

    2007-01-01

    The introduction of anti-apoptotic genes into plants leads to resistance to environmental stress and broad-spectrum disease resistance. The anti-apoptotic gene (p35) from a baculovirus was introduced into the genome of passion fruit plants by biobalistics. Eleven regenerated plants showed the presence of the p35 gene by PCR and/or dot blot hybridization. Transcriptional analysis of regenerated plants showed the presence of specific p35 transcripts in 9 of them. Regenerated plants containing the p35 gene were inoculated with the cowpea aphid-borne mosaic virus (CABMV), the bacterium Xanthomonas axonopodis pv passiflorae, and the herbicide, glufosinate, (Syngenta). None of the plants showed resistance to CABMV. Regenerated plants (p35+) showed less than half of local lesions showed by non-transgenic plants when inoculated with X. axonopodis and some p35+ plants showed increased tolerance to the glufosinate herbicide when compared to non-transgenic plants. PMID:17016672

  18. Impact of the Fukushima Daiichi nuclear power plant accident on local community and healthcare services

    International Nuclear Information System (INIS)

    The Soso region of Japan, located in the northern part of the Pacific side of Fukushima Prefecture, has suffered tremendously from widespread damage caused by the earthquake, tsunami and the Fukushima Dai-ichi nuclear power plant accident. Immediately after the disaster it even seemed that the restoration of this region itself would not be possible. However after six months, the Indoor Restriction Order and evacuation orders within a 20 to 30 km zone from the plant, such as the Evacuation-Prepared Area in Case of Emergency, were lifted and all of those restrictions for the planned evaluation zone has been eased in line with the actual conditions of each region. A year and a month after the earthquake, the Caution Zone, which was declared to prohibit the entry to the zone within the 20 km radius from the plant, was lifted. Thus people can now enter as close as 10 km from the plant. Minami-Soma city is an administrative district which had the largest population (approximately 71,500 residents) within the 20 to 30 km zone prior to the earthquake. It was, also, the only district where the evacuation was not conducted by the municipality. The city is now called the Genpatsu frontline district as it is the closest city to the plant where people have continued to live. Due to the damage caused by the earthquake and Tsunami, the city has suffered both from destruction by the tsunami and radiation, and people are still facing numerous problems despite the fact that the city appears to have been restored on its surface. It is very unfortunate that much of the medical data from the region was lost in the confusion after the Great East Japan Earthquake. In this paper various facts after the disaster based on the data left in the So-so region, Minamisoma city, and Minamisoma municipal general hospital are reported. (author)

  19. Reconstruction of local fallout composition and gamma-ray exposure in a village contaminated by the first USSR nuclear test in the Semipalatinsk nuclear test site in Kazakhstan.

    Science.gov (United States)

    Imanaka, Tetsuji; Yamamoto, Masayoshi; Kawai, Kenta; Sakaguchi, Aya; Hoshi, Masaharu; Chaizhunusova, Nailya; Apsalikov, Kazbek

    2010-11-01

    After the disintegration of the USSR in end of 1991, it became possible for foreign scientists to visit Kazakhstan, in order to investigate the radiological consequences of nuclear explosions that had been conducted at the Semipalatinsk nuclear test site (SNTS). Since the first visit in 1994, our group has been continuing expeditions for soil sampling at various areas around SNTS. The current level of local fallout at SNTS was studied through γ-spectrometry for (137)Cs as well as α-spectrometry for (239,240)Pu. Average values of soil inventory from wide areas around SNTS were 3,500 and 3,700 Bq m(-2) for (137)Cs and (239,240)Pu, respectively, as of January 1, 2000. The average level of (137)Cs is comparable to that in Japan due to global fallout, while the level of (239,240)Pu is several tens of times larger than that in Japan. Areas of strong contamination were found along the trajectories of radioactive fallout, information on which was declassified after the collapse of the USSR. Our recent efforts of soil sampling were concentrated on the area around the Dolon village heavily affected by the radioactive plume from the first USSR atomic bomb test in 1949 and located 110 km east from ground zero of the explosion. Using soil inventory data, retrospective dosimetry was attempted by reconstructing γ-ray exposure from fission product nuclides deposited on the ground. Adopting representative parameters for the initial (137)Cs deposition (13 kBq m(-2)), the refractory/volatile deposition ratio (3.8) and the plume arrival time after explosion (2.5 h), an absorbed dose in air of 600 mGy was obtained for the 1-year cumulative dose in Dolon village, due to the first bomb test in 1949. Considering possible ranges of the parameters, 350 and 910 mGy were estimated for high and low cases of γ-ray dose in air, respectively. It was encouraging that the deduced value was consistent with other estimations using thermal luminescence and archived monitoring data. The present

  20. 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)

    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 R199H200 and R221R222R223 that likely function together as a bipartite nuclear localization signal. In this study, we report that mutating R221R222R223, but not R199H200, relocates Bel1 from the nucleus to the cytoplasm, suggesting an essential role for R221R222R223 in the nuclear localization of Bel1. Although not affecting the nuclear localization of Bel1, mutating R199H200 disables Bel1 from transactivating PFV promoters. Results of EMSA reveal that the R199H200 residues are vital for the binding of Bel1 to viral promoter DNA. Moreover, mutating R199H200 in Bel1 impairs PFV replication to a much greater extent than mutating R221R222R223. Collectively, our findings suggest that R199H200 directly participate in Bel1 binding to viral promoter DNA and are indispensible for Bel1 transactivation activity. - Highlights: • The R221R222R223 residues are essential for the nuclear localization of Bel1. • Although not affecting the nuclear localization of Bel1, mutating R199H200 disables Bel1 from transactivating PFV promoters. • The R199H200 residues directly participate in Bel1 binding to viral promoter DNA. • Mutating R199H200 in Bel1 impairs PFV replication to a much greater extent than mutating R221R222R223

  1. Local fracture behavior in an alloy 52M dissimilar metal welded joint in nuclear power plant

    International Nuclear Information System (INIS)

    Background: Dissimilar metal welded joints (DMWJs) were indicated to be vulnerable components in the primary systems, an accurate structural integrity assessment for such DMWJ is needed. Purpose: Understand the fracture performance of the welded joint containing defects, determine if appropriate to assess the defects in the weld joint with the present codes. Methods: The fracture tests of the initial crack located in the different positions of Alloy 52M DMWJ specimens were carried out by using single-edge notched bend (SENB) method. Results: Different crack resistance and crack propagation path were obtained for the SENB specimens, and the cracks always deviate to the materials with lower strength. Conclusions: The crack path deviation is mainly controlled by strength mismatch, rather than toughness mismatch. The J-resistance curves with larger crack path deviation only reflect the apparent fracture resistance along the crack growth region, rather than the intrinsic fracture resistance of the initial crack-tip region material., Without considering the local fracture resistance properties of heat affected zone (HAZ), interface region and internal affected region (IAZ), following present assessment codes will unavoidably produce non-conservative or excessive conservative assessment results. In most cases, the assessment results will be potentially non-conservative. (authors)

  2. Nuclear

    International Nuclear Information System (INIS)

    This document proposes a presentation and discussion of the main notions, issues, principles, or characteristics related to nuclear energy: radioactivity (presence in the environment, explanation, measurement, periods and activities, low doses, applications), fuel cycle (front end, mining and ore concentration, refining and conversion, fuel fabrication, in the reactor, back end with reprocessing and recycling, transport), the future of the thorium-based fuel cycle (motivations, benefits and drawbacks), nuclear reactors (principles of fission reactors, reactor types, PWR reactors, BWR, heavy-water reactor, high temperature reactor of HTR, future reactors), nuclear wastes (classification, packaging and storage, legal aspects, vitrification, choice of a deep storage option, quantities and costs, foreign practices), radioactive releases of nuclear installations (main released radio-elements, radioactive releases by nuclear reactors and by La Hague plant, gaseous and liquid effluents, impact of releases, regulation), the OSPAR Convention, management and safety of nuclear activities (from control to quality insurance, to quality management and to sustainable development), national safety bodies (mission, means, organisation and activities of ASN, IRSN, HCTISN), international bodies, nuclear and medicine (applications of radioactivity, medical imagery, radiotherapy, doses in nuclear medicine, implementation, the accident in Epinal), nuclear and R and D (past R and D programmes and expenses, main actors in France and present funding, main R and D axis, international cooperation)

  3. 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)

    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

  4. 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.

  5. Creation and development of the Local Information Computer Network of the Institute of High Energy Physics and Nuclear Physics NSC KIPT

    International Nuclear Information System (INIS)

    Results and the basic stages of creation and development of local information computer network at Institute of High Energy Physics and Nuclear Physics NSC KIPT for last five years are submitted. The purpose of work was increase of an overall performance of researchers due to rational use of resources and creation on base of local information computer network of shared system. In article obtained experience and specificity of realization of such systems is reflected

  6. Expression and purification of the matrix protein of Nipah virus in baculovirus insect cell system.

    Science.gov (United States)

    Masoomi Dezfooli, Seyedehsara; Tan, Wen Siang; Tey, Beng Ti; Ooi, Chien Wei; Hussain, Siti Aslina

    2016-01-01

    Nipah virus (NiV) causes fatal respiratory illness and encephalitis in humans and animals. The matrix (M) protein of NiV plays an important role in the viral assembly and budding process. Thus, an access to the NiV M protein is vital to the design of viral antigens as diagnostic reagents. In this study, recombinant DNA technology was successfully adopted in the cloning and expression of NiV M protein. A recombinant expression cassette (baculovirus expression vector) was used to encode an N-terminally His-tagged NiV M protein in insect cells. A time-course study demonstrated that the highest yield of recombinant M protein (400-500 μg) was expressed from 107 infected cells 3 days after infection. A single-step purification method based on metal ion affinity chromatography was established to purify the NiV M protein, which successfully yielded a purity level of 95.67% and a purification factor of 3.39. The Western blotting and enzyme-linked immunosorbent assay (ELISA) showed that the purified recombinant M protein (48 kDa) was antigenic and reacted strongly with the serum of a NiV infected pig. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 32:171-177, 2016. PMID:26519022

  7. Production and characterization of recombinant lignin peroxidase isozyme H2 from Phanerochaete chrysosporium using recombinant baculovirus.

    Science.gov (United States)

    Johnson, T M; Pease, E A; Li, J K; Tien, M

    1992-08-01

    Recombinant Phanerochaete chrysosporium lignin peroxidase isozyme H2 (pI 4.4) was produced in insect cells infected with a genetically engineered baculovirus containing a copy of the cDNA clone lambda ML-6. The recombinant enzyme was purified to near homogeneity and is capable of oxidizing veratryl alcohol, iodide, and, to a lesser extent, guaiacol. The Km of the recombinant enzyme for veratryl alcohol and H2O2 is similar to that of the fungal enzyme. The guaiacol oxidation activity or any other activity is not dependent upon Mn2+. The purified recombinant peroxidase is glycosylated with N-linked carbohydrate(s). The recombinant lignin peroxidase eluted from an anion exchange resin similar to that of native isozyme H1 rather than H2. However, the pI of the recombinant enzymes is different from both H1 and H2 isozymes. Further characterization of native isozymes H1 and H2 from the fungal cultures revealed identical N-terminus residues. This indicates that isozymes H1 and H2 differ in post-translational modification. PMID:1632652

  8. Analysis of expression and glycosylation of avian metapneumovirus attachment glycoprotein from recombinant baculoviruses.

    Science.gov (United States)

    Luo, Lizhong; Nishi, Krista; MacLeod, Erin; Sabara, Marta I; Li, Yan

    2010-11-01

    Recently, we reported the expression and glycosylation of avian metapneumovirus attachment glycoprotein (AMPV/C G protein) in eukaryotic cell lines by a transient-expression method. In the present study, we investigated the biosynthesis and O-linked glycosylation of the AMPV/C G protein in a baculovirus expression system. The results showed that the insect cell-produced G protein migrated more rapidly in SDS-PAGE as compared to LLC-MK2 cell-derived G proteins owing to glycosylation differences. The fully processed, mature form of G protein migrated between 78 and 86 kDa, which is smaller than the 110 kDa mature form of G expressed in LLC-MK2 cells. In addition, several immature G gene products migrating at 40-48 and 60-70 kDa were also detected by SDS-PAGE and represented glycosylated intermediates. The addition of the antibiotic tunicamycin, which blocks early steps of glycosylation, to insect cell culture resulted in the disappearance of two glycosylated forms of the G protein and identified a 38 kDa unglycosylated precursor. The maturation of the G protein was completely blocked by monensin, suggesting that the O-linked glycosylation of G initiated in the trans-Golgi compartment. The presence of O-linked sugars on the mature protein was further confirmed by lectin Arachis hypogaea binding assay. Furthermore, antigenic features of the G protein expressed in insect cells were evaluated by ELISA. PMID:20713098

  9. Cervix carcinoma is associated with an up-regulation and nuclear localization of the dual-specificity protein phosphatase VHR

    International Nuclear Information System (INIS)

    The 21-kDa Vaccinia virus VH1-related (VHR) dual-specific protein phosphatase (encoded by the DUSP3 gene) plays a critical role in cell cycle progression and is itself regulated during the cell cycle. We have previously demonstrated using RNA interference that cells lacking VHR arrest in the G1 and G2 phases of the cell cycle and show signs of beginning of cell senescence. In this report, we evaluated successfully the expression levels of VHR protein in 62 hysterectomy or conization specimens showing the various (pre) neoplastic cervical epithelial lesions and 35 additional cases of hysterectomy performed for non-cervical pathologies, from patients under 50 years of age. We used a tissue microarray and IHC technique to evaluate the expression of the VHR phosphatase. Immunofluorescence staining under confocal microscopy, Western blotting and RT-PCR methods were used to investigate the localization and expression levels of VHR. We report that VHR is upregulated in (pre) neoplastic lesions (squamous intraepithelial lesions; SILs) of the uterine cervix mainly in high grade SIL (H-SIL) compared to normal exocervix. In the invasive cancer, VHR is also highly expressed with nuclear localization in the majority of cells compared to normal tissue where VHR is always in the cytoplasm. We also report that this phosphatase is highly expressed in several cervix cancer cell lines such as HeLa, SiHa, CaSki, C33 and HT3 compared to primary keratinocytes. The immunofluorescence technique under confocal microscopy shows that VHR has a cytoplasmic localization in primary keratinocytes, while it localizes in both cytoplasm and nucleus of the cancer cell lines investigated. We report that the up-regulation of this phosphatase is mainly due to its post-translational stabilization in the cancer cell lines compared to primary keratinocytes rather than increases in the transcription of DUSP3 locus. These results together suggest that VHR can be considered as a new marker for cancer

  10. Troponin T3 regulates nuclear localization of the calcium channel Cavβ1a subunit in skeletal muscle

    International Nuclear Information System (INIS)

    The voltage-gated calcium channel (Cav) β1a subunit (Cavβ1a) plays an important role in excitation–contraction coupling (ECC), a process in the myoplasm that leads to muscle-force generation. Recently, we discovered that the Cavβ1a subunit travels to the nucleus of skeletal muscle cells where it helps to regulate gene transcription. To determine how it travels to the nucleus, we performed a yeast two-hybrid screening of the mouse fast skeletal muscle cDNA library and identified an interaction with troponin T3 (TnT3), which we subsequently confirmed by co-immunoprecipitation and co-localization assays in mouse skeletal muscle in vivo and in cultured C2C12 muscle cells. Interacting domains were mapped to the leucine zipper domain in TnT3 COOH-terminus (160–244 aa) and Cavβ1a NH2-terminus (1–99 aa), respectively. The double fluorescence assay in C2C12 cells co-expressing TnT3/DsRed and Cavβ1a/YFP shows that TnT3 facilitates Cavβ1a nuclear recruitment, suggesting that the two proteins play a heretofore unknown role during early muscle differentiation in addition to their classical role in ECC regulation. - Highlights: • Previously, we demonstrated that Cavβ1a is a gene transcription regulator. • Here, we show that TnT3 interacts with Cavβ1a. • We mapped TnT3 and Cavβ1a interaction domain. • TnT3 facilitates Cavβ1a nuclear enrichment. • The two proteins play a heretofore unknown role during early muscle differentiation

  11. The LSD1-type zinc finger motifs of Pisum sativa LSD1 are a novel nuclear localization signal and interact with importin alpha.

    Directory of Open Access Journals (Sweden)

    Shanping He

    Full Text Available BACKGROUND: Genetic studies of the Arabidopsis mutant lsd1 highlight the important role of LSD1 in the negative regulation of plant programmed cell death (PCD. Arabidopsis thaliana LSD1 (AtLSD1 contains three LSD1-type zinc finger motifs, which are involved in the protein-protein interaction. METHODOLOGY/PRINCIPAL FINDINGS: To further understand the function of LSD1, we have analyzed cellular localization and functional localization domains of Pisum sativa LSD1 (PsLSD1, which is a homolog of AtLSD1. Subcellular localization analysis of green fluorescent protein (GFP-tagged PsLSD1 indicates that PsLSD1 is localized in the nucleus. Using a series of GFP-tagged PsLSD1 deletion mutants, we found that the three LSD1-type zinc finger motifs of PsLSD1 alone can target GFP to the nucleus, whereas deletion of the three zinc finger motifs or any individual zinc finger motif causes PsLSD1 to lose its nuclear localization, indicating that the three zinc finger motifs are necessary and sufficient for its nuclear localization. Moreover, site-directed mutagenesis analysis of GFP-tagged PsLSD1 indicates that tertiary structure and basic amino acids of each zinc finger motif are necessary for PsLSD1 nuclear localization. In addition, yeast two-hybrid, pull-down, and BiFC assays demonstrate that the three zinc finger motifs of PsLSD1 directly bind to importin α in vitro and in vivo. CONCLUSIONS/SIGNIFICANCE: Our data demonstrate that the LSD1-type zinc finger motifs of PsLSD1 are a novel nuclear localization signal and directly bind to importin α, and suggest that the nuclear import of LSD1 may rely on the interaction between its zinc finger motifs and importin α. Moreover, the nuclear localization of PsLSD1 suggests that LSD1 may function as a transcription regulator involved in negatively regulating PCD.

  12. Nuclear envelope-localized EGF family protein amphiregulin activates breast cancer cell migration in an EGF-like domain independent manner

    International Nuclear Information System (INIS)

    Highlights: ► Nuclear envelope-localized proAREG activates cancer cell migration via its cytoplasmic domain. ► The induction of cell migration does not require the EGF-like domain or EGR function. ► Nuclear envelope-localized proAREG suppresses breast cancer cell growth without EGFR function. ► This study revealed a novel function mediated by the intracellular domain of proAREG. -- Abstract: Amphiregulin (AREG), an EGF family protein, is synthesized as a type I transmembrane precursor (proAREG) and expressed on the cell surface with an extracellular EGF-like domain and an intracellular short cytoplasmic tail. The ectodomain shedding yields a soluble EGF receptor ligand (soluble AREG) which binds to EGF receptor (EGFR) and concomitantly induces migration of unshed proAREG from the plasma membrane to the nuclear envelope (NE). AREG is known to play a potential role in breast cancer and has been intensively investigated as an EGF receptor ligand, while the function of the NE-localized proAREG remains unknown. In this study we used a truncated mutant that mimics NE-localized proAREG without shedding stimuli to discriminate between the functions of NE-localized and plasma membrane-localized proAREG and demonstrate that NE-localized proAREG activates breast cancer cell migration, but suppresses cell growth. Moreover, the present study shows that induction of cell migration by NE-localized proAREG does not require the extracellular growth factor domain or EGF receptor function. Collectively these data demonstrate a novel function mediated by the intracellular domain of proAREG and suggest a significant role for NE-localized proAREG in driving human breast cancer progression.

  13. Results of calculations of external gamma radiation exposure rates from local fallout and the related radionuclide compositions of two hypothetical 1-MT nuclear bursts. Final report

    International Nuclear Information System (INIS)

    This report presents data on calculated gamma radiation exposure rates and local surface deposition of related radionuclides resulting from two hypothetical 1-Mt nuclear bursts. Calculations are made of the debris from two types of bombs: one containing 235U as a fissionable material (designated oralloy), the other containing 238U (designated tuballoy). 4 references

  14. Results of calculations of external gamma radiation exposure rates from local fallout and the related radionuclide compositions of two hypothetical 1-MT nuclear bursts. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Hicks, H.

    1984-12-01

    This report presents data on calculated gamma radiation exposure rates and local surface deposition of related radionuclides resulting from two hypothetical 1-Mt nuclear bursts. Calculations are made of the debris from two types of bombs: one containing /sup 235/U as a fissionable material (designated oralloy), the other containing /sup 238/U (designated tuballoy). 4 references.

  15. A localized navigation algorithm for Radiation Evasion for nuclear facilities. Part II: Optimizing the “Nearest Exit” Criterion

    International Nuclear Information System (INIS)

    Highlights: ► A new navigation algorithm for Radiation Evasion around nuclear facilities. ► An optimization criteria minimized under algorithm operation. ► A man-borne device guiding the occupational worker towards paths that warrant least radiation × time products. ► Benefits of using localized navigation as opposed to global navigation schemas. ► A path discrimination function for finding the navigational paths exhibiting the least amounts of radiation. -- Abstract: In this extension from part I (Khasawneh et al., in press), we modify the navigation algorithm which was presented with the objective of optimizing the “Radiation Evasion” Criterion so that navigation would optimize the criterion of “Nearest Exit”. Under this modification, algorithm would yield navigation paths that would guide occupational workers towards Nearest Exit points. Again, under this optimization criterion, algorithm leverages the use of localized information acquired through a well designed and distributed wireless sensor network, as it averts the need for any long-haul communication links or centralized decision and monitoring facility thereby achieving a more reliable performance under dynamic environments. As was done in part I, the proposed algorithm under the “Nearest Exit” Criterion is designed to leverage nearest neighbor information coming in through the sensory network overhead, in computing successful navigational paths from one point to another. For comparison purposes, the proposed algorithm is tested under the two optimization criteria: “Radiation Evasion” and “Nearest Exit”, for different numbers of step look-ahead. We verify the performance of the algorithm by means of simulations, whereby navigational paths are calculated for different radiation fields. We, via simulations, also, verify the performance of the algorithm in comparison with a well-known global navigation algorithm upon which we draw our conclusions

  16. AKT activation drives the nuclear localization of CSE1L and a pro-oncogenic transcriptional activation in ovarian cancer cells.

    Science.gov (United States)

    Lorenzato, Annalisa; Biolatti, Marta; Delogu, Giuseppe; Capobianco, Giampiero; Farace, Cristiano; Dessole, Salvatore; Cossu, Antonio; Tanda, Francesco; Madeddu, Roberto; Olivero, Martina; Di Renzo, Maria Flavia

    2013-10-15

    The human homolog of the yeast cse1 gene (CSE1L) is over-expressed in ovarian cancer. CSE1L forms complex with Ran and importin-α and has roles in nucleocytoplasmic traffic and gene expression. CSE1L accumulated in the nucleus of ovarian cancer cell lines, while it was localized also in the cytoplasm of other cancer cell lines. Nuclear localization depended on AKT, which was constitutively active in ovarian cancer cells, as the CSE1L protein translocated to the cytoplasm when AKT was inactivated. Moreover, the expression of a constitutively active AKT forced the translocation of CSE1L from the cytoplasm to the nucleus in other cancer cells. Nuclear accrual of CSE1L was associated to the nuclear accumulation of the phosphorylated Ran Binding protein 3 (RanBP3), which depended on AKT as well. Also in samples of human ovarian cancer, AKT activation was associated to nuclear accumulation of CSE1L and phosphorylation of RanBP3. Expression profiling of ovarian cancer cells after CSE1L silencing showed that CSE1L was required for the expression of genes promoting invasion and metastasis. In agreement, CSE1L silencing impaired motility and invasiveness of ovarian cancer cells. Altogether these data show that in ovarian cancer cells activated AKT by affecting RanBP3 phosphorylation determines the nuclear accumulation of CSE1L and likely the nuclear concentration of transcription factors conveying pro-oncogenic signals. PMID:23948303

  17. H II regions, infrared dark molecular clouds and the local geometry of the Milky Way's nuclear star-forming ring

    CERN Document Server

    Liszt, H S

    2009-01-01

    To interpret the galactic center H II region complexes as constituents of a barred galaxy's nuclear star-forming ring, we compare 18cm VLA radiocontinuumm, $8-22\\mu$ MSX IR and 2.6mm BTL and ARO12m CO emission in the inner few hundred pc. Galactic center H II regions are comparable in their IR appearance, luminosity and SED to M17 or N!0, but the IR light distribution is strongly modified by extinction at 8-22$\\mu$, locally and overall. In Sgr B2 at $l > 0.6$\\degr strong radio H II regions are invisible in the IR. In two favorable cases, extinction from individual galactic center molecular clouds is shown to have $\\tau \\ga 1$ at 8-22$\\mu$ independent of wavelength. The gas kinematics are mostly rotational but with systematic $\\pm 30-50$ \\kms non-circular motion. Sgr B and C both show the same shell and high-velocity cap structure. The H II regions lie in a slightly-inclined ring of radius $\\approx$ 180 pc (1.2\\degr) whose near side appears at higher latitude and lower velocity and contains Sgr B. Sgr C is on ...

  18. The methylation of the C-terminal region of hnRNPQ (NSAP1) is important for its nuclear localization

    International Nuclear Information System (INIS)

    Protein arginine methylation is an irreversible post-translational protein modification catalyzed by a family of at least nine different enzymes entitled PRMTs (protein arginine methyl transferases). Although PRMT1 is responsible for 85% of the protein methylation in human cells, its substrate spectrum has not yet been fully characterized nor are the functional consequences of methylation for the protein substrates well understood. Therefore, we set out to employ the yeast two-hybrid system in order to identify new substrate proteins for human PRMT1. We were able to identify nine different PRMT1 interacting proteins involved in different aspects of RNA metabolism, five of which had been previously described either as substrates for PRMT1 or as functionally associated with PRMT1. Among the four new identified possible protein substrates was hnRNPQ3 (NSAP1), a protein whose function has been implicated in diverse steps of mRNA maturation, including splicing, editing, and degradation. By in vitro methylation assays we were able to show that hnRNPQ3 is a substrate for PRMT1 and that its C-terminal RGG box domain is the sole target for methylation. By further studies with the inhibitor of methylation Adox we provide evidence that hnRNPQ1-3 are methylated in vivo. Finally, we demonstrate by immunofluorescence analysis of HeLa cells that the methylation of hnRNPQ is important for its nuclear localization, since Adox treatment causes its re-distribution from the nucleus to the cytoplasm

  19. Processivity factor of KSHV contains a nuclear localization signal and binding domains for transporting viral DNA polymerase into the nucleus

    International Nuclear Information System (INIS)

    Kaposi's sarcoma-associated human herpesvirus (KSHV) encodes a processivity factor (PF-8, ORF59) that forms homodimers and binds to viral DNA polymerase (Pol-8, ORF9). PF-8 is essential for stabilizing Pol-8 on template DNA so that Pol-8 can incorporate nucleotides continuously. Here, the intracellular interaction of these two viral proteins was examined by confocal immunofluorescence microscopy. When individually expressed, PF-8 was observed exclusively in the nucleus, whereas Pol-8 was found only in the cytoplasm. However, when co-expressed, Pol-8 was co-translocated with PF-8 into the nucleus. Mutational analysis revealed that PF-8 contains a nuclear localization signal (NLS) as well as domains located at the N-terminus and the C-proximal regions that are required for Pol-8 binding. This study suggests that the mechanism that enables PF-8 to transport Pol-8 into the nucleus is the first critical step required for Pol-8 and PF-8 to function processively in KSHV DNA synthesis

  20. A localized corrosion assessment of 6% molybdenum stainless steel condenser tubing at the Calvert Cliffs Nuclear Power Plant

    International Nuclear Information System (INIS)

    A condition assessment was performed on the Calvert Cliffs Nuclear Power Plant Unit 1 Main Steam Condenser 6% molybdenum stainless steel tubing. This work consisted of eddy current (EC) examination, in-situ visual examination of tubes, and destructive analysis of pulled tubes. The assessment objectives were as follows: to determine the tubing degradation mechanism, to render the condenser leak-tight, to identify methods of improving condenser life management, and to develop a basis for projecting the remaining condenser life. Once the corrosion mechanism was identified, seawater corrosion tests were performed on UNS N08366 and N08367 tubing using vinyl sleeves, Type 316 stainless compression fittings, and epoxy paint as crevice formers. Based on eddy current examination results, greater than 95% of the 49,500 tubes in the condenser had no detectable degradation. Over 1,200 tubes were plugged with either eddy current indications measuring greater than 60% wall loss or with obstructions. One waterbox (11A) contained 52% of all eddy current indications with over half of these in one section (B). Destructive analysis of pulled tubes revealed crevice corrosion, a form of localized corrosion that initiated under deposits. This type of corrosion occurred most often and had the deepest penetration in the weld seam. This was attributed to molybdenum and chromium segregation in the as-welded tubes and to deleterious secondary phases formed during weld solidification

  1. Nuclear localization of CPI-17, a protein phosphatase-1 inhibitor protein, affects histone H3 phosphorylation and corresponds to proliferation of cancer and smooth muscle cells

    International Nuclear Information System (INIS)

    Highlights: •Non-canonical roles of the myosin phosphatase inhibitor (CPI-17) were studied. •CPI-17 is localized in the nucleus of hyperplastic cancer and smooth muscle cells. •CPI-17 Ser12 phosphorylation may regulate the nuclear import. •CPI-17 regulates histone H3 phosphorylation and cell proliferation. •The nuclear CPI-17-PP1 axis plays a proliferative role in cells. -- Abstract: CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17 kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear import. CPI-17 phosphorylated at Ser12 was not localized at nuclei, suggesting a suppressive role of Ser12 phosphorylation in the nuclear import. Activated CPI-17 bound to all three isoforms of PP1 catalytic subunit in Panc1 nuclear extracts. CPI-17 knockdown in Panc1 resulted in dephosphorylation of histone H3 at Thr3, Ser10 and Thr11, whereas it had no effects on the phosphorylation of myosin light chain and merlin, the known targets of MLCP. In parallel, CPI-17 knockdown suppressed Panc1 proliferation. We propose that CPI-17 accumulated in the nucleus through the N-terminal tail targets multiple PP1 signaling pathways regulating cell proliferation

  2. Nuclear localization of CPI-17, a protein phosphatase-1 inhibitor protein, affects histone H3 phosphorylation and corresponds to proliferation of cancer and smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Eto, Masumi, E-mail: masumi.eto@jefferson.edu [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kirkbride, Jason A.; Chugh, Rishika; Karikari, Nana Kofi [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kim, Jee In [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Cardiovascular Research Institute, Kyungpook National University School of Medicine, Daegu 700-422 (Korea, Republic of)

    2013-04-26

    Highlights: •Non-canonical roles of the myosin phosphatase inhibitor (CPI-17) were studied. •CPI-17 is localized in the nucleus of hyperplastic cancer and smooth muscle cells. •CPI-17 Ser12 phosphorylation may regulate the nuclear import. •CPI-17 regulates histone H3 phosphorylation and cell proliferation. •The nuclear CPI-17-PP1 axis plays a proliferative role in cells. -- Abstract: CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17 kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear import. CPI-17 phosphorylated at Ser12 was not localized at nuclei, suggesting a suppressive role of Ser12 phosphorylation in the nuclear import. Activated CPI-17 bound to all three isoforms of PP1 catalytic subunit in Panc1 nuclear extracts. CPI-17 knockdown in Panc1 resulted in dephosphorylation of histone H3 at Thr3, Ser10 and Thr11, whereas it had no effects on the phosphorylation of myosin light chain and merlin, the known targets of MLCP. In parallel, CPI-17 knockdown suppressed Panc1 proliferation. We propose that CPI-17 accumulated in the nucleus through the N-terminal tail targets multiple PP1 signaling pathways regulating cell proliferation.

  3. A unique sequence in the N-terminal regulatory region controls the nuclear localization of KLF8 by cooperating with the C-terminal zinc-fingers

    Institute of Scientific and Technical Information of China (English)

    Tina S Mehta; Heng Lu; Xianhui Wang; Alison M Urvalek; Kim-Hang H Nguyen; Farah Monzur; Jojo D Hammond; Jameson Q Ma; Jihe Zhao

    2009-01-01

    Kruppel-like factor 8 (KLF8) transcription factor plays a critical role in cell cycle progression, oncogenic trans-formation, epithelial to mesenchymal transition and invasion. However, its nuclear localization signal(s) (NLS) has not been identified. KLF8 shares with other KLFs monopartite NLSs (mNLS) and C2H2 zinc fingers (ZFs), both of which have been shown to be the NLSs for some other KLFs. In this report, using PCR-directed mutagenesis and immunofluorescent microscopy, we show that disruption of the mNLSs, deletion of any single ZF, or mutation of the Zn2+-binding or DNA-contacting motifs did not affect the nuclear localization of KLF8. Deletion of>1.5 ZFs from C-terminus, however, caused cytoplasmic accumulation of KLF8. Surprisingly, deletion of amino acid (aa) 151-200 re-gion almost eliminated KLF8 from the nucleus. S165A, K171E or K171R mutation, or treatment with PKC inhibitor led to partial cytoplasmic accumulation. Co-immunoprecipitation demonstrated that KLF8 interacted with importin-β and this interaction required the ZF motif. Deletion of aa 1-150 or 201-261 region alone did not alter the nuclear lo-calization. BrdU incorporation and cyclin D1 promoter luciferase assays showed that the KLF8 mutants defective in nuclear localization could not promote DNA synthesis or cyclin D1 promoter activation as the wild-type KLF8 did. Taken together, these results suggest that KLF8 has two NLSs, one surrounding S165 and K171 and the other being two tandem ZFs, which are critical for the regulation of KLF8 nuclear localization and its cellular functions.

  4. Exact electronic and nuclear time-dependent potential energy surfaces for attosecond electron localization in the dissociation of H{sub 2}{sup +}

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Yasumitsu; Abedi, Ali; Gross, Eberhard K.U. [Max Planck Institute of Microstructure Physics, Weinberg 2, D-06120 Halle (Germany); Maitra, Neepa T. [Department of Physics and Astronomy, Hunter College and the City University of New York, 695 Park Avenue, New York, New York 10065 (United States); Yamashita, Koichi [Department of Chemical System Engineering, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656 (Japan)

    2013-07-01

    We study the electron localization dynamics in the dissociation of H2+ using a 1D model Hamiltonian. To this end we calculate the exact time-dependent potential energy surfaces (TDPES) both for the electron and for the nuclei. The exact electronic TDPES shows that the final electron localization is determined when the interatomic barrier becomes large and prohibits electron tunneling. The exact nuclear TDPES shows the mechanism of slowdown of the dissociation. It is found that the nuclear potential cannot be approximated by the weighted average of the quasi-static state potential energy surfaces, but can be approximated well by the transition between them. We show these two time-dependent potentials are the exact potential functionals of the time-dependent multicomponent density functional theory and can reproduce the whole phenomena of electron localization dynamics.

  5. The use of additive and subtractive approaches to examine the nuclear localization sequence of the polyomavirus major capsid protein VP1

    Science.gov (United States)

    Chang, D.; Haynes, J. I. 2nd; Brady, J. N.; Consigli, R. A.; Spooner, B. S. (Principal Investigator)

    1992-01-01

    A nuclear localization signal (NLS) has been identified in the N-terminal (Ala1-Pro-Lys-Arg-Lys-Ser-Gly-Val-Ser-Lys-Cys11) amino acid sequence of the polyomavirus major capsid protein VP1. The importance of this amino acid sequence for nuclear transport of VP1 protein was demonstrated by a genetic "subtractive" study using the constructs pSG5VP1 (full-length VP1) and pSG5 delta 5'VP1 (truncated VP1, lacking amino acids Ala1-Cys11). These constructs were used to transfect COS-7 cells, and expression and intracellular localization of the VP1 protein was visualized by indirect immunofluorescence. These studies revealed that the full-length VP1 was expressed and localized in the nucleus, while the truncated VP1 protein was localized in the cytoplasm and not transported to the nucleus. These findings were substantiated by an "additive" approach using FITC-labeled conjugates of synthetic peptides homologous to the NLS of VP1 cross-linked to bovine serum albumin or immunoglobulin G. Both conjugates localized in the nucleus after microinjection into the cytoplasm of 3T6 cells. The importance of individual amino acids found in the basic sequence (Lys3-Arg-Lys5) of the NLS was also investigated. This was accomplished by synthesizing three additional peptides in which lysine-3 was substituted with threonine, arginine-4 was substituted with threonine, or lysine-5 was substituted with threonine. It was found that lysine-3 was crucial for nuclear transport, since substitution of this amino acid with threonine prevented nuclear localization of the microinjected, FITC-labeled conjugate.

  6. Beyond defense-in-depth: cost and funding of state and local government radiological emergency response plans and preparedness in support of commercial nuclear power stations

    International Nuclear Information System (INIS)

    Inadequate, sporadic, uncertain and frustrating are words local, state and Federal officials use to describe the current hodgepodge funding approach to State and local government radiological emergency response plans and preparedeness in support of commercial nuclear power stations. The creation of a Radiological Emergency Response Plans and Preparedness Fund for State and Local Government is offered as a preferred solution. Monies for the Fund could be derived from a one time Fee of $1 million levied on the operator of each nuclear power station. Every five years, adjustments could be made in the Fee to assure full recovery of costs because of inflation, revised criteria and other cost related factors. Any surplus would be refunded to the utilities. Any state that has obtained NRC concurrence or is in the process could be reimbursed for previous expenditures up to two years prior to NRC concurrence. Concurrence in all state and local government plans is the objective of the funding program. The Fund should be administered by the Nuclear Regulatory Commission. The report also discusses actions by Federal and state agencies and points to long range considerations, such as a training institute, including transportation and non-commercial and other fixed nuclear facilities, where preparedness could be enhanced by a coherent funding mechanism. All recommendations are based on an inquiry by the Office of state Programs, NRC, into the historical and future costs and funding of radiological emergency response plans and preparedness at the state and local government levels and are derived from discussions with many local, State and Federal officials

  7. Production of mink enteritis parvovirus empty capsids by expression in a baculovirus vector system: a recombinant vaccine for mink enteritis parvovirus in mink

    DEFF Research Database (Denmark)

    Christensen, J; Alexandersen, Søren; Bloch, B.;

    1994-01-01

    The VP-2 gene of mink enteritis parvovirus (MEV) was amplified by the polymerase chain reaction using MEV DNA isolated from the faeces of a naturally infected mink. Subsequently the VP-2 gene was cloned into a baculovirus expression vector. Recombinant baculo-viruses were isolated and the MEV VP-2......, the VP-2 gene encoded a valine and a tyrosine at amino acid positions 232 and 234, identical to the situation found in MEV type 1, but at position 300 there was a valine which is a determinant of MEV type 2. Immunization of mink with approximately 40000 haemagglutinating units of recombinant MEV VP-2...

  8. Prorenin processing enzyme (PPE) produced by Baculovirus-infected Sf-9 insect cells: PPE is the cysteine protease encoded in the acMNPV gene.

    Science.gov (United States)

    Gotoh, Takeshi; Awa, Hirono; Kikuchi, Ken-Ichi; Nirasawa, Satoru; Takahashi, Saori

    2010-01-01

    In infection cultures of Spodoptera frugiperda (Sf-9) insect cells with a recombinant baculovirus, vhpR, carrying human preprorenin cDNA in the polyhedrin locus of Autographa californica multiple nuclear polyhedrosis virus (AcMNPV), the expressed inactive recombinant human (rh)-prorenin is reported to be proteolytically processed to yield active rh-renin in the very late phase of culture (Takahashi et al., Biosci. Biotechnol. Biochem., 71, 2610-2613 (2007)). To identify the enzyme that catalyzes the processing of rh-prorenin, referred to as prorenin processing enzyme (PPE), we purified potential PPE from virus-infected Sf-9 culture supernatant by the use of an internally quenched fluorescent (IQF) substrate for PPE. The 32-kDa protein band agreed well with PPE activity on the final Mono Q FPLC. By N-terminal amino acid sequence analysis, the protein was revealed to be a cysteine protease encoded by the AcMNPV gene. Enzyme activity was inhibited by cysteine protease inhibitors but not by other protease inhibitors. When the purified rh-prorenin was incubated with the 32-kDa protein, renin activity appeared concomitant with the disappearance of rh-prorenin. The N-terminal amino acid sequence of the activated product was identical to that of the rh-renin that had accumulated in the infection cultures. These results indicate that the 32-kDa cysteine protease derived from the AcMNPV gene is the enzyme PPE of virus-infected Sf-9 cells. PMID:20139610

  9. Baculovirus expression of erythrovirus V9 capsids and screening by ELISA: serologic cross-reactivity with erythrovirus B19

    DEFF Research Database (Denmark)

    Heegaard, Erik D; Qvortrup, Klaus; Christensen, Jesper

    2002-01-01

    categorize V9 as an acute B19-like infection. Sequencing, combined with PCR studies, have since demonstrated the need for specific and differentiated techniques when examining samples for possible B19 or V9 viremia. The antigenic properties of the V9 capsid proteins have not been characterized previously. To...... address this question, V9 VP1 and VP2 open reading frames were cloned and expressed in insect cells using a baculovirus vector. Large quantities of purified recombinant V9 capsid protein were produced and electron micrographs revealed self-assembly of V9 VP1/VP2 and VP2 capsids into empty icosahedral...

  10. Development of cell lines from the cactophagous insect: Cactoblastis cactorum (Lepidoptera: Pyralidae) and their susceptibility to three baculoviruses.

    Science.gov (United States)

    Grasela, James J; McIntosh, Arthur H; Ringbauer, Joseph; Goodman, Cynthia L; Carpenter, James E; Popham, Holly J R

    2012-05-01

    The unintentional introduction of the cactus moth, Cactoblastis cactorum, a successful biological control agent formerly employed in the control of invasive prickly pear cactus species (Opuntia spp.) in Australia, Hawaii, South Africa, and various Caribbean islands, has posed great concern as to the possible threat to native, endangered species of cactus in the southeastern USA as well as with the potential to cause a major infestation of commercial and agricultural cactus crops in Mexico. A number of control measures have been investigated with varying degrees of success including, field exploration for cactus moth-specific parasitoids, insecticides, fungal, bacterial, and nematode agents. Current tactics used by the USA-Mexico binational program to eradicate cactus moth from Mexico and mitigate its westward movement in the USA include host plant removal, the manual removal and destruction of egg sticks and infected cacti stems, and the Sterile Insect Technique. One other approach not taken until now is the development of a cactus moth cell line as a tool to facilitate the investigation of baculoviruses as an alternative biocontrol method for the cactus moth. Consequently, we established C. cactorum cell lines derived from adult ovarian tissue designated as BCIRL-Cc-AM and BCIRL-Cc-JG. The mean cell population doubling time was 204.3 and 112 h for BCIRL-Cc-AM and BCIRL-Cc-JG, respectively, with weekly medium change, while the doubling time was 176.6 and 192.6 h for BCIRL-Cc-AM and BCIRL-Cc-JG, respectively, with a daily change of medium. In addition, the daily versus weekly change in medium was reflected in the percentage viability with both cell lines showing higher levels with a daily medium change. Of the three baculoviruses tested, only the recombinant AcMNPV-hsp70Red and GmMNPV at a multiplicity of infection (MOI) of 1.0 were able to demonstrate significant production of extracellular virus (ECV) in each of the cell lines, whereas both cell lines were

  11. Production and purification of VP2 protein of porcine parvovirus expressed in an insect-baculovirus cell system

    OpenAIRE

    Cui Shangjin; Yao Guizhe; Zhou Hongchao

    2010-01-01

    Abstract The porcine parvovirus (PPV) VP2 protein was expressed in an insect-baculovirus cell system and was purified using Ni-NTA affinity column chromatography. The recombinant 6-His-tagged VP2 protein with molecular mass (Mr) of about 64 kDa was detected by anti-his antibody and anti-PPV serum. Electron microscopy showed that the purified VP2 protein assembled into spherical particles with diameters ranging from 20 to 22 nm. The expressed VP2 was antigenically similar to the native capsid ...

  12. The scaffolding protein IQGAP1 co-localizes with actin at the cytoplasmic face of the nuclear envelope: implications for cytoskeletal regulation

    OpenAIRE

    Johnson, Michael A.; Henderson, Beric R.

    2012-01-01

    IQGAP1 is an important cytoskeletal regulator, known to act at the plasma membrane to bundle and cap actin filaments, and to tether the cortical actin meshwork to microtubules via plus-end binding proteins. Here we describe the novel subcellular localization of IQGAP1 at the cytoplasmic face of the nuclear envelope, where it co-located with F-actin. The IQGAP1 and F-actin staining overlapped that of microtubules at the nuclear envelope, revealing a pattern strikingly similar to that observed ...

  13. G-actin guides p53 nuclear transport: potential contribution of monomeric actin in altered localization of mutant p53

    Science.gov (United States)

    Saha, Taniya; Guha, Deblina; Manna, Argha; Panda, Abir Kumar; Bhat, Jyotsna; Chatterjee, Subhrangsu; Sa, Gaurisankar

    2016-01-01

    p53 preserves genomic integrity by restricting anomaly at the gene level. Till date, limited information is available for cytosol to nuclear shuttling of p53; except microtubule-based trafficking route, which utilizes minus-end directed motor dynein. The present study suggests that monomeric actin (G-actin) guides p53 traffic towards the nucleus. Histidine-tag pull-down assay using purified p53(1–393)-His and G-actin confirms direct physical association between p53 and monomeric G-actin. Co-immunoprecipitation data supports the same. Confocal imaging explores intense perinuclear colocalization between p53 and G-actin. To address atomistic details of the complex, constraint-based docked model of p53:G-actin complex was generated based on crystal structures. MD simulation reveals that p53 DNA-binding domain arrests very well the G-actin protein. Docking benchmark studies have been carried out for a known crystal structure, 1YCS (complex between p53DBD and BP2), which validates the docking protocol we adopted. Co-immunoprecipitation study using “hot-spot” p53 mutants suggested reduced G-actin association with cancer-associated p53 conformational mutants (R175H and R249S). Considering these findings, we hypothesized that point mutation in p53 structure, which diminishes p53:G-actin complexation results in mutant p53 altered subcellular localization. Our model suggests p53Arg249 form polar-contact with Arg357 of G-actin, which upon mutation, destabilizes p53:G-actin interaction and results in cytoplasmic retention of p53R249S. PMID:27601274

  14. G-actin guides p53 nuclear transport: potential contribution of monomeric actin in altered localization of mutant p53.

    Science.gov (United States)

    Saha, Taniya; Guha, Deblina; Manna, Argha; Panda, Abir Kumar; Bhat, Jyotsna; Chatterjee, Subhrangsu; Sa, Gaurisankar

    2016-01-01

    p53 preserves genomic integrity by restricting anomaly at the gene level. Till date, limited information is available for cytosol to nuclear shuttling of p53; except microtubule-based trafficking route, which utilizes minus-end directed motor dynein. The present study suggests that monomeric actin (G-actin) guides p53 traffic towards the nucleus. Histidine-tag pull-down assay using purified p53(1-393)-His and G-actin confirms direct physical association between p53 and monomeric G-actin. Co-immunoprecipitation data supports the same. Confocal imaging explores intense perinuclear colocalization between p53 and G-actin. To address atomistic details of the complex, constraint-based docked model of p53:G-actin complex was generated based on crystal structures. MD simulation reveals that p53 DNA-binding domain arrests very well the G-actin protein. Docking benchmark studies have been carried out for a known crystal structure, 1YCS (complex between p53DBD and BP2), which validates the docking protocol we adopted. Co-immunoprecipitation study using "hot-spot" p53 mutants suggested reduced G-actin association with cancer-associated p53 conformational mutants (R175H and R249S). Considering these findings, we hypothesized that point mutation in p53 structure, which diminishes p53:G-actin complexation results in mutant p53 altered subcellular localization. Our model suggests p53Arg249 form polar-contact with Arg357 of G-actin, which upon mutation, destabilizes p53:G-actin interaction and results in cytoplasmic retention of p53R249S. PMID:27601274

  15. AKT activation drives the nuclear localization of CSE1L and a pro-oncogenic transcriptional activation in ovarian cancer cells

    International Nuclear Information System (INIS)

    The human homolog of the yeast cse1 gene (CSE1L) is over-expressed in ovarian cancer. CSE1L forms complex with Ran and importin-α and has roles in nucleocytoplasmic traffic and gene expression. CSE1L accumulated in the nucleus of ovarian cancer cell lines, while it was localized also in the cytoplasm of other cancer cell lines. Nuclear localization depended on AKT, which was constitutively active in ovarian cancer cells, as the CSE1L protein translocated to the cytoplasm when AKT was inactivated. Moreover, the expression of a constitutively active AKT forced the translocation of CSE1L from the cytoplasm to the nucleus in other cancer cells. Nuclear accrual of CSE1L was associated to the nuclear accumulation of the phosphorylated Ran Binding protein 3 (RanBP3), which depended on AKT as well. Also in samples of human ovarian cancer, AKT activation was associated to nuclear accumulation of CSE1L and phosphorylation of RanBP3. Expression profiling of ovarian cancer cells after CSE1L silencing showed that CSE1L was required for the expression of genes promoting invasion and metastasis. In agreement, CSE1L silencing impaired motility and invasiveness of ovarian cancer cells. Altogether these data show that in ovarian cancer cells activated AKT by affecting RanBP3 phosphorylation determines the nuclear accumulation of CSE1L and likely the nuclear concentration of transcription factors conveying pro-oncogenic signals. - highlights: • CSE1L is a key player in nucleocytoplasmic traffic by forming complex with Ran. • AKT phosphorylates RanBP3 that regulates the nucleocytoplasmic gradient of Ran. • The activated oncogenic AKT drives the nuclear accumulation of CSE1L. • CSE1L in the nucleus up-regulates genes conveying pro-oncogenic signals. • CSE1L might contribute to tumor progression driven by the activated oncogenic AKT

  16. AKT activation drives the nuclear localization of CSE1L and a pro-oncogenic transcriptional activation in ovarian cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lorenzato, Annalisa; Biolatti, Marta [Department of Oncology, University of Torino School of Medicine, Torino (Italy); Institute for Cancer Research at Candiolo, Candiolo, Torino (Italy); Delogu, Giuseppe [Department of Biomedical Sciences-Histology, University of Sassari, Sassari (Italy); Capobianco, Giampiero [Department of Surgical, Microsurgical and Medical Sciences, University of Sassari, Sassari (Italy); Farace, Cristiano [Department of Biomedical Sciences-Histology, University of Sassari, Sassari (Italy); Dessole, Salvatore; Cossu, Antonio; Tanda, Francesco [Department of Surgical, Microsurgical and Medical Sciences, University of Sassari, Sassari (Italy); Madeddu, Roberto [Department of Biomedical Sciences-Histology, University of Sassari, Sassari (Italy); National Institute of Biostructures and Biosystems, Rome (Italy); Olivero, Martina [Department of Oncology, University of Torino School of Medicine, Torino (Italy); Institute for Cancer Research at Candiolo, Candiolo, Torino (Italy); Di Renzo, Maria Flavia, E-mail: mariaflavia.direnzo@unito.it [Department of Oncology, University of Torino School of Medicine, Torino (Italy); Institute for Cancer Research at Candiolo, Candiolo, Torino (Italy)

    2013-10-15

    The human homolog of the yeast cse1 gene (CSE1L) is over-expressed in ovarian cancer. CSE1L forms complex with Ran and importin-α and has roles in nucleocytoplasmic traffic and gene expression. CSE1L accumulated in the nucleus of ovarian cancer cell lines, while it was localized also in the cytoplasm of other cancer cell lines. Nuclear localization depended on AKT, which was constitutively active in ovarian cancer cells, as the CSE1L protein translocated to the cytoplasm when AKT was inactivated. Moreover, the expression of a constitutively active AKT forced the translocation of CSE1L from the cytoplasm to the nucleus in other cancer cells. Nuclear accrual of CSE1L was associated to the nuclear accumulation of the phosphorylated Ran Binding protein 3 (RanBP3), which depended on AKT as well. Also in samples of human ovarian cancer, AKT activation was associated to nuclear accumulation of CSE1L and phosphorylation of RanBP3. Expression profiling of ovarian cancer cells after CSE1L silencing showed that CSE1L was required for the expression of genes promoting invasion and metastasis. In agreement, CSE1L silencing impaired motility and invasiveness of ovarian cancer cells. Altogether these data show that in ovarian cancer cells activated AKT by affecting RanBP3 phosphorylation determines the nuclear accumulation of CSE1L and likely the nuclear concentration of transcription factors conveying pro-oncogenic signals. - highlights: • CSE1L is a key player in nucleocytoplasmic traffic by forming complex with Ran. • AKT phosphorylates RanBP3 that regulates the nucleocytoplasmic gradient of Ran. • The activated oncogenic AKT drives the nuclear accumulation of CSE1L. • CSE1L in the nucleus up-regulates genes conveying pro-oncogenic signals. • CSE1L might contribute to tumor progression driven by the activated oncogenic AKT.

  17. The complete genome of a baculovirus isolated from an insect of medical interest: Lonomia obliqua (Lepidoptera: Saturniidae).

    Science.gov (United States)

    Aragão-Silva, C W; Andrade, M S; Ardisson-Araújo, D M P; Fernandes, J E A; Morgado, F S; Báo, S N; Moraes, R H P; Wolff, J L C; Melo, F L; Ribeiro, B M

    2016-01-01

    Lonomia obliqua (Lepidoptera: Saturniidae) is a species of medical importance due to the severity of reactions caused by accidental contact with the caterpillar bristles. Several natural pathogens have been identified in L. obliqua, and among them the baculovirus Lonomia obliqua multiple nucleopolyhedrovirus (LoobMNPV). The complete genome of LoobMNPV was sequenced and shown to have 120,022 bp long with 134 putative open reading frames (ORFs). Phylogenetic analysis of the LoobMNPV genome showed that it belongs to Alphabaculovirus group I (lepidopteran-infective NPV). A total of 12 unique ORFs were identified with no homologs in other sequenced baculovirus genomes. One of these, the predicted protein encoded by loob035, showed significant identity to an eukaryotic transcription terminator factor (TTF2) from the Lepidoptera Danaus plexippus, suggesting an independent acquisition through horizontal gene transfer. Homologs of cathepsin and chitinase genes, which are involved in host integument liquefaction and viral spread, were not found in this genome. As L. obliqua presents a gregarious behavior during the larvae stage the impact of this deletion might be neglectable. PMID:27282807

  18. Baculovirus expression of beak and feather disease virus (BFDV) capsid protein capable of self-assembly and haemagglutination.

    Science.gov (United States)

    Stewart, Meredith E; Bonne, Nicolai; Shearer, Patrick; Khalesi, Bahman; Sharp, Margaret; Raidal, Shane

    2007-05-01

    Beak and feather disease virus (BFDV) is a common avian circovirus infection of wild Psittaciformes and is a recognised threat to endangered psittacine species. Currently, there is a requirement to develop BFDV antigen for diagnostic purposes and since efforts to propagate BFDV in vitro have so far been unsuccessful the entire coding region of BFDV ORF C1 was expressed in Sf9 insect cells using a baculovirus expression system. The entire coding region of BFDV ORF C1, the presumptive capsid, was expressed in Sf9 insect cells using baculovirus expression system. Electron microscopic examination of negatively stained material demonstrated that the recombinant protein self-assembled to produce virus-like particles (VLPs) thus confirming that ORF C1 is likely to be the sole determinant for capsid construction in vivo. BFDV VLPs also possessed haemagglutinating activity which provides further evidence that self-assembled BFDV VLPs retain receptor mediated biological activity and that the determinants for BFDV haemagglutination activity rely solely on the capsid protein. The recombinant protein reacted with anti-BFDV sera from naturally immune parrots and cockatoo and from chickens experimentally inoculated with native BFDV in both Western blots and haemagglutination inhibition (HI) assay. BFDV VLPs were also a suitable replacement antigen for serological detection of BFDV antibody by HI. PMID:17218022

  19. Functional display of an alpha2 integrin-specific motif (RKK) on the surface of baculovirus particles.

    Science.gov (United States)

    Riikonen, Reetta; Matilainen, Heli; Rajala, Nina; Pentikainen, Olli; Johnson, Mark; Heino, Jyrki; Oker-Blom, Christian

    2005-08-01

    The use of baculovirus vectors shows promise as a tool for gene delivery into mammalian cells. These insect viruses have been shown to transduce a variety of mammalian cell lines, and gene transfer has also been demonstrated in vivo. In this study, we generated two recombinant baculovirus vectors displaying an integrin-specific motif, RKK, as a part of two different loops of the green fluorescent protein (GFP) fused with the major envelope protein gp64 of Autographa californica M nucleopolyhedrovirus. By enzyme linked immunosorbent assays, these viruses were shown to bind a peptide representing the receptor binding site of an alpha2 integrin, the alpha2I-domain. However, the interaction was not strong enough to overcome binding of wild type gp64 to the unknown cellular receptor(s) on the surface of alpha2 integrin-expressing cells (CHO-alpha2beta1) or enhance the viral uptake. After treatment of these cells with phospholipase C, internalization of all viruses was blocked or decreased significantly. However, one of the RKK displaying viruses, AcGFP(K)gp64, was still able to internalize into CHO-alpha2beta1 cells, although at a lower level as compared to non-treated cells. This may indicate the possible utilization of a PLC independent alternative route via, in this case, the alpha2beta1 integrin. PMID:16029062

  20. Novel nuclear localization and potential function of insulin-like growth factor-1 receptor/insulin receptor hybrid in corneal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Yu-Chieh Wu

    Full Text Available BACKGROUND: Type I insulin-like growth factor receptor (IGF-1R and insulin receptor (INSR are highly homologous molecules, which can heterodimerize to form an IGF-1R/INSR hybrid (Hybrid-R. The presence and biological significance of the Hybrid-R in human corneal epithelium has not yet been established. In addition, while nuclear localization of IGF-1R was recently reported in cancer cells and human corneal epithelial cells, the function and profile of nuclear IGF-1R is unknown. In this study, we characterized the nuclear localization and function of the Hybrid-R and the role of IGF-1/IGF-1R and Hybrid-R signaling in the human corneal epithelium. METHODOLOGY/PRINCIPLE FINDINGS: IGF-1-mediated signaling and cell growth were examined in a human telomerized corneal epithelial (hTCEpi cell line using co-immunoprecipitation, immunoblotting and cell proliferation assays. The presence of Hybrid-R in hTCEpi and primary cultured human corneal epithelial cells was confirmed by immunofluorescence and reciprocal immunoprecipitation of whole cell lysates. We found that IGF-1 stimulated Akt and promoted cell growth through IGF-1R activation, which was independent of the Hybrid-R. The presence of Hybrid-R, but not IGF-1R/IGF-1R, was detected in nuclear extracts. Knockdown of INSR by small interfering RNA resulted in depletion of the INSR/INSR and preferential formation of Hybrid-R. Chromatin-immunoprecipitation sequencing assay with anti-IGF-1R or anti-INSR was subsequently performed to identify potential genomic targets responsible for critical homeostatic regulatory pathways. CONCLUSION/SIGNIFICANCE: In contrast to previous reports on nuclear localized IGF-1R, this is the first report identifying the nuclear localization of Hybrid-R in an epithelial cell line. The identification of a nuclear Hybrid-R and novel genomic targets suggests that IGF-1R traffics to the nucleus as an IGF-1R/INSR heterotetrameric complex to regulate corneal epithelial homeostatic

  1. Early Detection of Baculovirus Expression and Infection in Lepidopteran Larvae Fed Occlusion Bodies of an AcMNPV Recombinant Carrying a Red Fluorescent Protein Gene

    Science.gov (United States)

    A method has been devised utilizing a baculovirus recombinant (AcMNPV hsp70Red) carrying a red fluorescent protein (RFP) gene under the early heat shock promoter (hsp70) to assess potential infectivity of larvae fed occlusion bodies. A time study was employed whereby first and third instars of Trich...

  2. Enhanced recombinant protein production and differential expression of molecular chaperones in sf-caspase-1-repressed stable cells after baculovirus infection

    Directory of Open Access Journals (Sweden)

    Lai Yiu-Kay

    2012-11-01

    Full Text Available Abstract Background There are few studies that have examined the potential of RNA inference (RNAi to increase protein production in the baculovirus expression vector system (BEVS. Spodoptera frugiperda (fall armyworm (Sf-caspase-1-repressed stable cells exhibit resistance to apoptosis and enhancement of recombinant protein production. However, the mechanism of recombinant protein augmentation in baculovirus-infected Caspase-repressed insect cells has not been elucidated. Results In the current study, we utilized RNAi-mediated Sf-caspase-1-repressed stable cells to clarify how the resistance to apoptosis can enhance both intracellular (firefly luciferase and extracellular (secreted alkaline phosphatase [SEAP] recombinant protein production in BEVS. Since the expression of molecular chaperones is strongly associated with the maximal production of exogenous proteins in BEVS, the differential expression of molecular chaperones in baculovirus-infected stable cells was also analyzed in this study. Conclusion The data indicated that the retention of expression of molecular chaperones in baculovirus-infected Sf-caspase-1-repressed stable cells give the higher recombinant protein accumulation.

  3. Survey the of building and dietary structure of local residents around Yangjiang and Hongyanhe nuclear power plants

    International Nuclear Information System (INIS)

    Objective: To provide a guide in the course of the daily operation and accident emergency response. Methods: The survey was conducted with questionnaires among heads of households collected by stratified radom sampling. The head of a household was asked about residential type and structure, the sources of drinking water, milk, type and frequency of main vegetables. Results: Two-storied and more than two-storied houses were dominant around Yangjiang nuclear power plant, single-storey houses and tile-roofed houses were dominant around Hongyanhe nuclear power plant. The top three of wall construction materials around the two nuclear power plants were orderly brick, stone and beton. Glass window shelters were in the majority. Drinking water of residents was mainly from wells and waterworks. Milk comes from nonlocal packaged products. The top five high frequency vegetables the residents around Yangjiang nuclear power plants eat were orderly cabbage, zucchini, string beans, tong dish, cucumber, and those around Yangjiang nuclear power plant were orderly chinese cabbage, leek, celery, cabbage, spinach. Conclusion: Building and dietary structure around Yangjiang and Hongyanhe nuclear power plants are incongruence. Government can make decisions to collect foods for nuclides monitoring according to building and dietary structure baseline data, and take emergency measures to direct nuclear safety radiation protection for residents when nuclear accident takes place. (authors)

  4. A case study of economic incentives and local citizens' attitudes toward hosting a nuclear power plant in Japan: Impacts of the Fukushima accident

    International Nuclear Information System (INIS)

    The attitude of local communities near a nuclear power plant (NPP) is a key factor in nuclear policy decision making in Japan. This case study compared local citizens' attitudes in 2010 and 2011 toward the benefits and drawbacks of hosting Kashiwazaki–Kariwa NPP. The Fukushima accident occurred in this period. After the accident, benefit recognition of utility bill refunds clearly declined, while that of public facilities did not, suggesting the influence of a bribery effect. The negative shift of attitudes about hosting the NPP after the accident was more modest in Kariwa Village, which saw a large expansion of social welfare programs, than in the other two areas, which lacked such a budget expansion. Policy implications of these results regarding the provision of economic incentives in NPP host areas after the Fukushima accident were discussed. - Highlights: • The Fukushima accident shocked Japan's nuclear policy. • Citizens' attitudes toward incentives of hosting a nuclear power plant surveyed. • More citizens thought negatively about incentives after the Fukushima accident. • The bribery effect, mode and amount of incentives affected citizens' attitudes

  5. Nuclear PKG localization is regulated by G₀ alpha and is necessary in the AWB neurons to mediate avoidance in Caenorhabditis elegans.

    Science.gov (United States)

    He, Chao; O'Halloran, Damien M

    2013-10-11

    Neural circuits interpret sensory information from their environment and use this information to influence behavioral outputs. In Caenorhabditis elegans two bilaterally symmetric cephalic amphid organs each contain the ciliated termini of twelve classes of sensory neurons. Two of these sensory neuron pairs are called the AWB and the AWC neurons. The AWC neurons confer an ability to detect attractive volatile odors such as benzaldehyde, whereas the AWB neurons endow the animal with an ability to detect repulsive volatile odors such as 2-nonanone. Previously, it has been shown that transient nuclear localization of a Protein Kinase G (PKG) called EGL-4 in the AWC neuron facilitates adaptation after sustained odor input, and here we show that constitutively nuclear EGL-4 is required in the AWB neurons for the detection of repellent odors. Furthermore, we show that the G₀ alpha subunit protein GOA-1 regulates the nuclear localization of EGL-4 in both AWB and AWC neurons. These data reveal novel insight into how the localization of an individual kinase can form a turnout switch to modify output in different neurons to drive opposite sensory behaviors. PMID:23954825

  6. 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.

  7. Metabolite Regulation of Nuclear Localization of Carbohydrate-response Element-binding Protein (ChREBP): ROLE OF AMP AS AN ALLOSTERIC INHIBITOR.

    Science.gov (United States)

    Sato, Shogo; Jung, Hunmin; Nakagawa, Tsutomu; Pawlosky, Robert; Takeshima, Tomomi; Lee, Wan-Ru; Sakiyama, Haruhiko; Laxman, Sunil; Wynn, R Max; Tu, Benjamin P; MacMillan, John B; De Brabander, Jef K; Veech, Richard L; Uyeda, Kosaku

    2016-05-13

    The carbohydrate-response element-binding protein (ChREBP) is a glucose-responsive transcription factor that plays an essential role in converting excess carbohydrate to fat storage in the liver. In response to glucose levels, ChREBP is regulated by nuclear/cytosol trafficking via interaction with 14-3-3 proteins, CRM-1 (exportin-1 or XPO-1), or importins. Nuclear localization of ChREBP was rapidly inhibited when incubated in branched-chain α-ketoacids, saturated and unsaturated fatty acids, or 5-aminoimidazole-4-carboxamide ribonucleotide. Here, we discovered that protein-free extracts of high fat-fed livers contained, in addition to ketone bodies, a new metabolite, identified as AMP, which specifically activates the interaction between ChREBP and 14-3-3. The crystal structure showed that AMP binds directly to the N terminus of ChREBP-α2 helix. Our results suggest that AMP inhibits the nuclear localization of ChREBP through an allosteric activation of ChREBP/14-3-3 interactions and not by activation of AMPK. AMP and ketone bodies together can therefore inhibit lipogenesis by restricting localization of ChREBP to the cytoplasm during periods of ketosis. PMID:26984404

  8. 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)

    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

  9. Overexpression and Constitutive Nuclear Localization of Cohesin Protease Separase Protein Correlates with High Incidence of Relapse and Reduced Overall Survival in Glioblastoma Multiforme

    Science.gov (United States)

    Mukherjee, Malini; Byrd, Tiara; Brawley, Vita S.; Bielamowicz, Kevin; Li, Xiao-Nan; Merchant, Fatima; Maitra, Saurabh; Sumazin, Pavel; Fuller, Greg; Kew, Yvonne; Sun, David; Powell, Suzanne Z.; Ahmed, Nabil M.; Zhang, Nenggang; Pati, Debananda

    2014-01-01

    Separase, an enzyme that cleaves the chromosomal cohesin during mitosis, is overexpressed in a wide range of human epithelial cancers of breast, bone and prostate [Meyer et al, 2009, Clin Cancer Res 15(8): 2703-2710] [1]. Overexpression of Separase in animal models results in aneuploidy and tumorigenesis. We have examined the expression and localization of Separase protein in adult and pediatric glioblastoma and normal brain specimens. Immunofluorescence microscopy and Western blot analysis showed significant overexpression of Separase in all adult and a subset of pediatric glioblastoma cells. Tumor status and patient survival strongly correlate with the mislocalization of Separase into the nucleus throughout all stages of the cell cycle. Unlike exclusively nuclear localization in mitotic control cells, glioblastoma samples have a significantly higher number of resting (interphase) cells with strong nuclear Separase staining. Additionally, patient survival analysis demonstrated a strong correlation between overexpression of Separase protein in adult glioblastoma and a high incidence of relapse and reduced overall survival. These results further strengthen our hypothesis that Separase is an oncogene whose overexpression induces tumorigenesis, and indicate that Separase overexpression and aberrant nuclear localization are common in many tumor types and may predict outcome in some human malignancies. PMID:24792645

  10. Coordinate regulation/localization of the carbohydrate responsive binding protein (ChREBP) by two nuclear export signal sites: Discovery of a new leucine-rich nuclear export signal site

    International Nuclear Information System (INIS)

    Carbohydrate response element binding protein (ChREBP) is responsible for conversion of dietary carbohydrate to storage fat in liver by coordinating expression of the enzymes that channel glycolytic pyruvate into lipogenesis. The activation of ChREBP in response to high glucose is nuclear localization and transcription, and the inactivation of ChREBP under low glucose involves export from the nucleus to the cytosol. Here we report a new nuclear export signal site ('NES1') of ChREBP. Together these signals provide ChREBP with two NES sequences, both the previously reported NES2 and now the new NES1 coordinate to interact together with CRM1 (exportin) for nuclear export of the carbohydrate response element binding protein.

  11. Intracellular distribution of an integral nuclear pore membrane protein fused to green fluorescent protein--localization of a targeting domain.

    Science.gov (United States)

    Söderqvist, H; Imreh, G; Kihlmark, M; Linnman, C; Ringertz, N; Hallberg, E

    1997-12-15

    The 121-kDa pore membrane protein (POM121) is a bitopic integral membrane protein specifically located in the pore membrane domain of the nuclear envelope with its short N-terminal tail exposed on the luminal side and its major C-terminal portion adjoining the nuclear pore complex. In order to locate a signal for targeting of POM121 to the nuclear pores, we overexpressed selected regions of POM121 alone or fused to the green fluorescent protein (GFP) in transiently transfected COS-1 cells or in a stably transfected neuroblastoma cell line. Microscopic analysis of the GFP fluorescence or immunostaining was used to determine the intracellular distribution of the overexpressed proteins. The endofluorescent GFP tag had no effect on the distribution of POM121, since the chimerical POM121-GFP fusion protein was correctly targeted to the nuclear pores of both COS-1 cells and neuroblastoma cells. Based on the differentiated intracellular sorting of the POM121 variants, we conclude that the first 128 amino acids of POM121 contains signals for targeting to the continuous endoplasmic reticulum/nuclear envelope membrane system but not specifically to the nuclear pores and that a specific nuclear pore targeting signal is located between amino acids 129 and 618 in the endoplasmically exposed portion of POM121. PMID:9461306

  12. The herpes simplex virus 1 U(L)34 protein interacts with a cytoplasmic dynein intermediate chain and targets nuclear membrane.

    Science.gov (United States)

    Ye, G J; Vaughan, K T; Vallee, R B; Roizman, B

    2000-02-01

    To express the function encoded in its genome, the herpes simplex virus 1 capsid-tegument structure released by deenvelopment during entry into cells must be transported retrograde to the nuclear pore where viral DNA is released into the nucleus. This path is essential in the case of virus entering axons of dorsal root ganglia. The objective of the study was to identify the viral proteins that may be involved in the transport. We report the following findings. (i) The neuronal isoform of the intermediate chain (IC-1a) of the dynein complex pulled down, from lysates of [(35)S]methionine-labeled infected cells, two viral proteins identified as the products of U(L)34 and U(L)31 open reading frames, respectively. U(L)34 protein is a virion protein associated with cellular membranes and phosphorylated by the viral kinase U(S)3. U(L)31 protein is a largely insoluble, evenly dispersed nuclear phosphoprotein required for optimal processing and packaging of viral DNA into preformed capsids. Reciprocal pulldown experiments verified the interaction of IC-1a and U(L)34 protein. In similar experiments, U(L)34 protein was found to interact with U(L)31 protein and the major capsid protein ICP5. (ii) To determine whether U(L)34 protein is transported to the nuclear membrane, a requirement if it is involved in transport, the U(L)34 protein was inserted into a baculovirus vector under the cytomegalovirus major early promoter. Cells infected with the recombinant baculovirus expressed U(L)34 protein in a dose-dependent manner, and the U(L)34 protein localized primarily in the nuclear membrane. An unexpected finding was that U(L)34-expressing cells showed a dissociation of the inner and outer nuclear membranes reminiscent of the morphologic changes seen in cells productively infected with herpes simplex virus 1. U(L)34, like many other viral proteins, may have multiple functions expressed both early and late in infection. PMID:10627546

  13. Burnout and Psychiatric Distress in Local Caregivers Two Years After the 2011 Great East Japan Earthquake and Fukushima Nuclear Radiation Disaster.

    Science.gov (United States)

    Fujitani, Kenji; Carroll, Matt; Yanagisawa, Robert; Katz, Craig

    2016-01-01

    The 2011 Great East Japan Earthquake precipitated a triple disaster of earthquake, tsunami and nuclear radiation disaster. To quantify the prevalence of burnout and psychiatric distress in local healthcare providers (caregivers) more than 2 years after the disaster, this study surveyed caregivers from affected areas through interviews about topics of concerns and two questionnaires: Maslach Burnout Inventory and General Health Questionnaire. Concerns listed by respondents were primarily radiation related: additional stress, concern for children, concern for local food, and sleep difficulties. We found significant number of caregivers to have signs of emotional exhaustion, low personal accomplishment, and psychological distress. Our findings suggest that local caregivers are experiencing substantial mental health burdens, which have unfortunately remained static from the year prior, even 2 years after the fact. Therefore, long term psychological support and improvement in caregiver work conditions are essential to maintain sustainable care in rebuilding disaster stricken areas. PMID:26303904

  14. Collaboration of local government and experts responding to increase in environmental radiation level due to the nuclear disaster: focusing on their activities and latest radiological discussion.

    Science.gov (United States)

    Iimoto, T; Nunokawa, J; Fujii, H; Takashima, R; Hashimoto, M; Fukuhara, T; Yajima, T; Matsuzawa, H; Kurosawa, K; Yanagawa, Y; Someya, S

    2015-11-01

    Activities were introduced in Kashiwa city in the Tokyo metropolitan area to correspond to the elevated environmental radiation level after the disaster of the Fukushima Daiichi nuclear power plant. These were based on a strong cooperation between local governments and experts. Ambient dose rate and radioactivity of foodstuff produced inside of the city have been monitored. Representative ambient dose rates around living environments have almost already become their original levels of the pre-accident because of the decontamination activity, natural washout and effective half-lives of radioactivity. The internal annual dose due to radioactive cesium under the policy of 'Local Production for Local Consumption' is estimated as extremely low comparing the variation range due to natural radioactivity. Systematic survey around a retention basin has been started. All of these latest monitoring data would be one of the core information for the policy making as well as a cost-benefit discussion and risk communication. PMID:25982790

  15. Collaboration of local government and experts responding to increase in environmental radiation level due to the nuclear disaster: focusing on their activities and latest radiological discussion

    International Nuclear Information System (INIS)

    Activities were introduced in Kashiwa city in the Tokyo metropolitan area to correspond to the elevated environmental radiation level after the disaster of the Fukushima Daiichi nuclear power plant. These were based on a strong cooperation between local governments and experts. Ambient dose rate and radioactivity of foodstuff produced inside of the city have been monitored. Representative ambient dose rates around living environments have almost already become their original levels of the pre-accident because of the decontamination activity, natural washout and effective half-lives of radioactivity. The internal annual dose due to radioactive cesium under the policy of 'Local Production for Local Consumption' is estimated as extremely low comparing the variation range due to natural radioactivity. Systematic survey around a retention basin has been started. All of these latest monitoring data would be one of the core information for the policy making as well as a cost-benefit discussion and risk communication. (authors)

  16. A study on the institutional design concerning role-sharing between national and local governments about nuclear safety regulation in Japan

    International Nuclear Information System (INIS)

    In this paper, we consider the concept and the concrete direction of the institutional design concerning role-sharing between national and local governments with respect to nuclear safety regulation, and propose the following four institutional options: 1) strengthen the regulatory body's independence and clarify its accountability, 2) establish a local information committee in reference to the French CLI (Commission Locale d'Information), 3) institutionalize the environmental monitoring of local governments, and 4) institutionalize consultation among the regulatory body and local governments. Options 2, 3, and 4 are designed on the assumption that option 1 is implemented, because it is thought to be essential for fundamental improvements to enhance the regulatory body's ability as well as to reinforce bridges between safety regulation and society. Each option has some advantages and disadvantages, so there is room for further research into achieving reforms. However, it is believed that these institutional options can lead to the development of a trusting relationship between organizations responsible for safety regulation and siting areas and to the reduction in consensus-building costs in local areas. (author)

  17. The hepatitis E virus ORF3 protein regulates the expression of liver-specific genes by modulating localization of hepatocyte nuclear factor 4.

    Directory of Open Access Journals (Sweden)

    Vivek Chandra

    Full Text Available The hepatitis E virus (HEV is a small RNA virus and the cause of acute viral hepatitis E. The open reading frame 3 protein (pORF3 of HEV appears to be a pleiotropic regulatory protein that helps in the establishment, propagation and progression of viral infection. However, the global cellular effects of this protein remain to be explored. In the absence of traditional in vitro viral infection systems or efficient replicon systems, we made an adenovirus based ORF3 protein expression system to study its effects on host cell gene expression. We infected Huh7 hepatoma cells with recombinant adenoviruses expressing pORF3 and performed microarray-based gene expression analyses. Several genes down regulated in pORF3-expressing cells were found to be under regulation of the liver-enriched hepatocyte nuclear factor 4 (HNF4, which regulates hepatocyte-specific gene expression. While HNF4 localizes to the nucleus, its phosphorylation results in impaired nuclear localization of HNF4. Here we report that pORF3 increases HNF4 phosphorylation through the ERK and Akt kinases, which results in impaired nuclear translocation of HNF4 and subsequently the down modulation of HNF4-responsive genes in pORF3-expressing cells. We propose that modulation of several hepatocyte specific genes by pORF3 will create an environment favorable for viral replication and pathogenesis.

  18. Local fallout from nuclear test detonations. Volume 2. Compilation of fallout patterns and related test data. Supplement. Foreign nuclear tests. Sanitized

    Energy Technology Data Exchange (ETDEWEB)

    Morgenthau, M.; Showers, R.L.

    1964-10-01

    The available fallout patterns and related test data for nuclear weapon tests conducted by the United Kingdom, the Republic of France, and the Union of Soviet Socialist Republics, are included in this supplement to NDL-TR-34. The related test data for the British and French tests include: date and time of detonation, location of test site, total yield, fission yield, type of burst and placement, height of burst, cloud-top and -bottom heights, crater data, and wind information up to nuclear cloud-top height. No fallout patterns are available for any of the Soviet detonations. The list of Soviet detonations, which is as comprehensive as possible, contains the chronological order of the detonations, date, yield, type of burst and location of test site.

  19. Production and purification of VP2 protein of porcine parvovirus expressed in an insect-baculovirus cell system

    Directory of Open Access Journals (Sweden)

    Cui Shangjin

    2010-12-01

    Full Text Available Abstract The porcine parvovirus (PPV VP2 protein was expressed in an insect-baculovirus cell system and was purified using Ni-NTA affinity column chromatography. The recombinant 6-His-tagged VP2 protein with molecular mass (Mr of about 64 kDa was detected by anti-his antibody and anti-PPV serum. Electron microscopy showed that the purified VP2 protein assembled into spherical particles with diameters ranging from 20 to 22 nm. The expressed VP2 was antigenically similar to the native capsid protein according to HA and a Western blotting assay performed with polyclonal antibodies collected from an outbreak of PPV in one farm. This study provides a foundation for the application of VP2 protein in the clinical diagnosis of PPV or in the vaccination against PPV in the future.

  20. Baculovirus display of single chain antibody (scFv using a novel signal peptide

    Directory of Open Access Journals (Sweden)

    Gonzalez Gaëlle

    2010-11-01

    Full Text Available Abstract Background Cells permissive to virus can become refractory to viral replication upon intracellular expression of single chain fragment variable (scFv antibodies directed towards viral structural or regulatory proteins, or virus-coded enzymes. For example, an intrabody derived from MH-SVM33, a monoclonal antibody against a conserved C-terminal epitope of the HIV-1 matrix protein (MAp17, was found to exert an inhibitory effect on HIV-1 replication. Results Two versions of MH-SVM33-derived scFv were constructed in recombinant baculoviruses (BVs and expressed in BV-infected Sf9 cells, N-myristoylation-competent scFvG2/p17 and N-myristoylation-incompetent scFvE2/p17 protein, both carrying a C-terminal HA tag. ScFvG2/p17 expression resulted in an insoluble, membrane-associated protein, whereas scFvE2/p17 was recovered in both soluble and membrane-incorporated forms. When coexpressed with the HIV-1 Pr55Gag precursor, scFvG2/p17 and scFvE2/p17 did not show any detectable negative effect on virus-like particle (VLP assembly and egress, and both failed to be encapsidated in VLP. However, soluble scFvE2/p17 isolated from Sf9 cell lysates was capable of binding to its specific antigen, in the form of a synthetic p17 peptide or as Gag polyprotein-embedded epitope. Significant amounts of scFvE2/p17 were released in the extracellular medium of BV-infected cells in high-molecular weight, pelletable form. This particulate form corresponded to BV particles displaying scFvE2/p17 molecules, inserted into the BV envelope via the scFv N-terminal region. The BV-displayed scFvE2/p17 molecules were found to be immunologically functional, as they reacted with the C-terminal epitope of MAp17. Fusion of the N-terminal 18 amino acid residues from the scFvE2/p17 sequence (N18E2 to another scFv recognizing CD147 (scFv-M6-1B9 conferred the property of BV-display to the resulting chimeric scFv-N18E2/M6. Conclusion Expression of scFvE2/p17 in insect cells using a BV

  1. Development and Implementation of a High-Throughput High-Content Screening Assay to Identify Inhibitors of Androgen Receptor Nuclear Localization in Castration-Resistant Prostate Cancer Cells.

    Science.gov (United States)

    Johnston, Paul A; Nguyen, Minh M; Dar, Javid A; Ai, Junkui; Wang, Yujuan; Masoodi, Khalid Z; Shun, Tongying; Shinde, Sunita; Camarco, Daniel P; Hua, Yun; Huryn, Donna M; Wilson, Gabriela Mustata; Lazo, John S; Nelson, Joel B; Wipf, Peter; Wang, Zhou

    2016-05-01

    Patients with castration-resistant prostate cancer (CRPC) can be treated with abiraterone, a potent inhibitor of androgen synthesis, or enzalutamide, a second-generation androgen receptor (AR) antagonist, both targeting AR signaling. However, most patients relapse after several months of therapy and a majority of patients with relapsed CRPC tumors express the AR target gene prostate-specific antigen (PSA), suggesting that AR signaling is reactivated and can be targeted again to inhibit the relapsed tumors. Novel small molecules capable of inhibiting AR function may lead to urgently needed therapies for patients resistant to abiraterone, enzalutamide, and/or other previously approved antiandrogen therapies. Here, we describe a high-throughput high-content screening (HCS) campaign to identify small-molecule inhibitors of AR nuclear localization in the C4-2 CRPC cell line stably transfected with GFP-AR-GFP (2GFP-AR). The implementation of this HCS assay to screen a National Institutes of Health library of 219,055 compounds led to the discovery of 3 small molecules capable of inhibiting AR nuclear localization and function in C4-2 cells, demonstrating the feasibility of using this cell-based phenotypic assay to identify small molecules targeting the subcellular localization of AR. Furthermore, the three hit compounds provide opportunities to develop novel AR drugs with potential for therapeutic intervention in CRPC patients who have relapsed after treatment with antiandrogens, such as abiraterone and/or enzalutamide. PMID:27187604

  2. AHM1, a novel type of nuclear matrix-localized, MAR binding protein with a single AT hook and a J domain-homologous region.

    Science.gov (United States)

    Morisawa, G; Han-Yama, A; Moda, I; Tamai, A; Iwabuchi, M; Meshi, T

    2000-10-01

    Interactions between the nuclear matrix and special regions of chromosomal DNA called matrix attachment regions (MARs) have been implicated in various nuclear functions. We have identified a novel protein from wheat, AT hook-containing MAR binding protein1 (AHM1), that binds preferentially to MARs. A multidomain protein, AHM1 has the special combination of a J domain-homologous region and a Zn finger-like motif (a J-Z array) and an AT hook. For MAR binding, the AT hook at the C terminus was essential, and an internal portion containing the Zn finger-like motif was additionally required in vivo. AHM1 was found in the nuclear matrix fraction and was localized in the nucleoplasm. AHM1 fused to green fluorescent protein had a speckled distribution pattern inside the nucleus. AHM1 is most likely a nuclear matrix component that functions between intranuclear framework and MARs. J-Z arrays can be found in a group of (hypothetical) proteins in plants, which may share some functions, presumably to recruit specific Hsp70 partners as co-chaperones. PMID:11041885

  3. Measurement of 2-dimensional local instantaneous liquid film thickness around simulated nuclear fuel rod by ultrasonic transmission technique

    International Nuclear Information System (INIS)

    An accurate knowledge of hydro-dynamic behavior of a liquid film flow on nuclear fuel rods is indispensable for analysis of the CHF under postulated loss-of-coolant-accidents in boiling water reactors. This work is concerned with a new development of ultrasonic transmission technique for film flow measurements. The technique adopted a rotating reflector, capable of measuring time-dependent spatial distribution of liquid film thickness around a simulated nuclear fuel rod. The scanning time is currently 4 ms for reconstruction of one image of the circumferential film thickness distribution. (orig.)

  4. Qualification of the indentation test for the local characterization of nuclear facility materials. Final report; Qualifizierung des Eindruckversuchs zur lokalen Charakterisierung kerntechnischer Werkstoffe. Abschlussbericht

    Energy Technology Data Exchange (ETDEWEB)

    Tandler, Martin; Seifert, Thomas; Schlesinger, Michael; Mohrmann, Ralf; Kilgus, Normen; Venugopal, Ravula

    2007-12-21

    With the aid of the registrating indentation test, the project intends to characterise the operational changes in the local material properties of nuclear materials by a quasi-nondestructive indentation test. The focus was on the materials 22NiMoCr3-7 and X6CrNiNb18-10, both of which are widely used in nuclear engineering. As the accuracy of the method depends on experimental influencing factors like surface treatment, intrinsic stresses, or material anisotropy, these influences are to be quantified and will be considered in the evaluation of the material characteristics. The influencing parameters will be investigated experimentally and numerically by FE simulations so that their influence can be distinguished from the actual material behaviour. (orig.)

  5. The 14 october 1999 nuclear accident simulation drill at Nogent-sur-Seine NPP role of the local information committee (C.L.I.)

    International Nuclear Information System (INIS)

    The EDF nuclear power plant at Nogent-sur-Seine consists in two 1 300 MWe pressurised water reactor units. It is located 120 km south-east of Paris. The Local Information Committee, which brigs 120 members together, was created when the plant started operating in 1988. The 14 October 1999 nuclear accident simulation drill, the sequence of which started from an incident in the virtual Unit 3, was intended to testing the reacting capacities of the local and national emergency PCs (command post). The main objective was to actually implement the evacuation of a 500 inhabitants village near the plant in order to manage their rallying in a reception centre about 15 kilometers from there. The aim was to assess the speed and accuracy of the alert to the population, the requisitioning of human and practical resources, the affected area cordoning off, the evacuation of two schools, the activation of the reception centre with firemen, doctors and finally direct work with the media. The CLI was involved from the initial stages in the 6 preparatory meetings and, on the day of the drill, put observers in the strategic places. After the drill it was invited to give its opinion during feedback meetings organised by authorities, both the Aube Prefecture in Troyes and the Nuclear Installation Safety Directorate in Paris. Our CLI had prepared in 1997 reactor cards 'fiches reflexes' for local representatives, which in the light of this drill, could be improved. CLI participation to the rewriting - at national level - PPI (offsite emergency plans) is all the more active since it is done in the light of this drill. Il will be the same with the reissue of our public information brochure dealing with security instructions. In a more general way, it appears that such a drill would have been completely excluded only ten years ago. (author)

  6. Nuclear Localization of the Autism Candidate Gene Neurobeachin and Functional Interaction with the NOTCH1 Intracellular Domain Indicate a Role in Regulating Transcription

    Science.gov (United States)

    Tuand, Krizia; Stijnen, Pieter; Volders, Karolien; Declercq, Jeroen; Nuytens, Kim; Meulemans, Sandra; Creemers, John

    2016-01-01

    Background Neurobeachin (NBEA) is an autism spectrum disorders (ASD) candidate gene. NBEA deficiency affects regulated secretion, receptor trafficking, synaptic architecture and protein kinase A (PKA)-mediated phosphorylation. NBEA is a large multidomain scaffolding protein. From N- to C-terminus, NBEA has a concanavalin A-like lectin domain flanked by armadillo repeats (ACA), an A-kinase anchoring protein domain that can bind to PKA, a domain of unknown function (DUF1088) and a BEACH domain, preceded by a pleckstrin homology-like domain and followed by WD40 repeats (PBW). Although most of these domains mediate protein-protein interactions, no interaction screen has yet been performed. Methods Yeast two-hybrid screens with the ACA and PBW domain modules of NBEA gave a list of interaction partners, which were analyzed for Gene Ontology (GO) enrichment. Neuro-2a cells were used for confocal microscopy and nuclear extraction analysis. NOTCH-mediated transcription was studied with luciferase reporter assays and qRT-PCR, combined with NBEA knockdown or overexpression. Results Both domain modules showed a GO enrichment for the nucleus. PBW almost exclusively interacted with transcription regulators, while ACA interacted with a number of PKA substrates. NBEA was partially localized in the nucleus of Neuro-2a cells, albeit much less than in the cytoplasm. A nuclear localization signal was found in the DUF1088 domain, which was shown to contribute to the nuclear localization of an EGFP-DPBW fusion protein. Yeast two-hybrid identified the Notch1 intracellular domain as a physical interactor of the PBW domain and a role for NBEA as a negative regulator in Notch-mediated transcription was demonstrated. Conclusion Defining novel interaction partners of conserved NBEA domain modules identified a role for NBEA as transcriptional regulator in the nucleus. The physical interaction of NBEA with NOTCH1 is most relevant for ASD pathogenesis because NOTCH signaling is essential for

  7. Local consequences and reactions to the closing down of nuclear reactors in Sweden, a KSO report, 1999

    International Nuclear Information System (INIS)

    The Swedish Government decision to close down the Barsebaeck-1 NPP resulted in a studi funded by the Organisation of Municipalities with Nuclear Operation (KSO). It was meant to evaluate the scenario if all nuclear reactors in Sweden were closed down by year 2005 or if the phase out took place in an orderly manner after 40 years i.e. between 2012 and 2025. Most important consequence would be the loss of approximately 8000 jobs mostly in five municipalities concerned. This should also be regarded from a qualitative viewpoint, namely as a significant loss of labour opportunities for highly educated personnel. The estimates show in addition, a dramatic increase in migration from nuclear power plant municipalities of particularly younger population. In conclusion it is mentioned that Sweden still has 11 nuclear reactors in operation supplying 50% of the electricity used in the country. If the government would demand immediate closure of all reactors, Sweden would be a dark and cold country at least during the six months of winter

  8. Some public finance aspects of nuclear energy centers. State and local public finance problems and prospects: Some implications of nuclear energy centers

    Energy Technology Data Exchange (ETDEWEB)

    Weinstein, P.A.

    1976-04-01

    Energy facilities characteristically have an impact on the areas where they are sited. A major generating station can alter the tax base in the county and the nature of public services to be provided. This particular study concentrates on the fiscal problems. It is intended as a first cut at the problem of how to apportion the revenues and the burdens which would exist if an energy park were to become a reality. Many of the problems which it explores apply equally to facilities in the energy field and elsewhere which are smaller than the nuclear park.

  9. SU-E-J-61: Electrodynamics and Nano-Scale Fluid Dynamics in Protein Localization of Nuclear Pore Complexes

    International Nuclear Information System (INIS)

    Purpose: To develop a simulation to catalyze a reevaluation of common assumptions about 3 dimensional diffusive processes and help cell biologists gain a more nuanced, intuitive understanding of the true physical hurdles of protein signaling cascades. Furthermore, to discuss the possibility of intracellular electrodynamics as a critical, unrecognized component of cellular biology and protein dynamics that is necessary for optimal information flow from the cell membrane to the nucleus. Methods: The Unity 3D gaming physics engine was used to build an accurate virtual scale model of the cytoplasm within a few hundred nanometers of the nuclear membrane. A cloud of simulated pERK proteins is controlled by the physics simulation, where diffusion is based on experimentally measured values and the electrodynamics are based on theoretical nano-fluid dynamics. The trajectories of pERK within the cytoplasm and through the 1250 nuclear pores on the nuclear surface is recorded and analyzed. Results: The simulation quickly demonstrates that pERKs moving solely by diffusion will rarely locate and come within capture distance of a nuclear pore. The addition of intracellular electrodynamics between charges on the nuclear pore complexes and on pERKs increases the number of successful translocations by allowing the electro-physical attractive effects to draw in pERKs from the cytoplasm. The effects of changes in intracellular shielding ion concentrations allowed for estimation of the capture radius” under varying conditions. Conclusion: The simulation allows a shift in perspective that is paramount in attempting to communicate the scale and dynamics of intracellular protein cascade mechanics. This work has allowed researchers to more fully understand the parameters involved in intracellular electrodynamics, such as shielding anion concentration and protein charge. As these effects are still far below the spatial resolution of currently available measurement technology this

  10. SU-E-J-61: Electrodynamics and Nano-Scale Fluid Dynamics in Protein Localization of Nuclear Pore Complexes

    Energy Technology Data Exchange (ETDEWEB)

    Cunningham, J; Gatenby, R [Moffitt Cancer Research Institute, Tampa, FL (United States)

    2014-06-01

    Purpose: To develop a simulation to catalyze a reevaluation of common assumptions about 3 dimensional diffusive processes and help cell biologists gain a more nuanced, intuitive understanding of the true physical hurdles of protein signaling cascades. Furthermore, to discuss the possibility of intracellular electrodynamics as a critical, unrecognized component of cellular biology and protein dynamics that is necessary for optimal information flow from the cell membrane to the nucleus. Methods: The Unity 3D gaming physics engine was used to build an accurate virtual scale model of the cytoplasm within a few hundred nanometers of the nuclear membrane. A cloud of simulated pERK proteins is controlled by the physics simulation, where diffusion is based on experimentally measured values and the electrodynamics are based on theoretical nano-fluid dynamics. The trajectories of pERK within the cytoplasm and through the 1250 nuclear pores on the nuclear surface is recorded and analyzed. Results: The simulation quickly demonstrates that pERKs moving solely by diffusion will rarely locate and come within capture distance of a nuclear pore. The addition of intracellular electrodynamics between charges on the nuclear pore complexes and on pERKs increases the number of successful translocations by allowing the electro-physical attractive effects to draw in pERKs from the cytoplasm. The effects of changes in intracellular shielding ion concentrations allowed for estimation of the “capture radius” under varying conditions. Conclusion: The simulation allows a shift in perspective that is paramount in attempting to communicate the scale and dynamics of intracellular protein cascade mechanics. This work has allowed researchers to more fully understand the parameters involved in intracellular electrodynamics, such as shielding anion concentration and protein charge. As these effects are still far below the spatial resolution of currently available measurement technology this

  11. Case-study session 2: Nuclear facilities and local information. The way we work with public relations at the Oskarshamn nuclear power plant

    International Nuclear Information System (INIS)

    The Oskarshamn NPP is owned by OKG Aktiebolag, the only private company in Sweden operating NPPs. The site is situated 350 kilometres south of Stockholm, in the community of Oskarshamn with 30 000 inhabitants and in the county of Kalmar with about 250 000 inhabitants. OKG now owns and operates three BWRs of ABB Atom design, with a total net production of 15 TWh/year, i.e. 1 - 12% of the total electricity generation in Sweden. OKG also operates CLAB that is owned by SKB. On the site, SKB has now decided to build a hard-rock laboratory 500 metres down in the bedrock. OKG is the nuclear pioneer of Sweden, the first reactor has been in operation since 1972. Company policy has always been maximum open communication and encouraging the employees to engage themselves into debates about energy and nuclear and into social life as a whole. OKG also sponsors several cultural and sport events in Oskarshamn. The goal is to create confidence between society and the plant. Two different areas of influence are defined: organization of the information department taking care of both external and internal information which is considered an essential part of the work

  12. Nuclear Quadrupole Hyperfine Structure in HC14N/H14NC and DC15N/D15NC Isomerization: A Diagnostic Tool for Characterizing Vibrational Localization

    CERN Document Server

    Wong, Bryan M

    2010-01-01

    Large-amplitude molecular motions which occur during isomerization can cause significant changes in electronic structure. These variations in electronic properties can be used to identify vibrationally-excited eigenstates which are localized along the potential energy surface. This work demonstrates that nuclear quadrupole hyperfine interactions can be used as a diagnostic marker of progress along the isomerization path in both the HC14N/H14NC and DC15N/D15NC chemical systems. Ab initio calculations at the CCSD(T)/cc-pCVQZ level indicate that the hyperfine interaction is extremely sensitive to the chemical bonding of the quadrupolar 14N nucleus and can therefore be used to determine in which potential well the vibrational wavefunction is localized. A natural bonding orbital analysis along the isomerization path further demonstrates that hyperfine interactions arise from the asphericity of the electron density at the quadrupolar nucleus. Using the CCSD(T) potential surface, the quadrupole coupling constants of...

  13. A proposal on restart rule of nuclear power plants with piping having local wall thinning subjected to an earthquake. Former part. Aiming at further application

    International Nuclear Information System (INIS)

    Restart rule of nuclear power plants (NPPs) with piping having local wall thinning subjected to an earthquake was proposed taking account of local wall thinning, seismic effects and restart of NPPs with applicability of 'Guidelines for NPP Response to an Earthquake (EPRI NP-6695)' in Japan. Japan Earthquake Damage Intensity Scale (JEDIS) and Earthquake Ground Motion Level (EGML) were introduced. JEDIS was classified into four scales obtained from damage level of components and structures of NPPs subjected to an earthquake, while EGML was divided into four levels by safe shutdown earthquake ground motion (So), elastic design earthquake ground motion (Sd) and design earthquake ground motion (Ss). Combination of JEDIS and EGML formulated 4 x 4 matrix and determined detailed conditions of restart of NPPs. As a response to an earthquake, operator walk inspections and evaluation of earthquake ground motion were conducted to know the level of JEDIS. JEDIS level requested respective allowable conditions of restart of NPP, which were scale level dependent and consisted of weighted combination of damage inspection (operator walk inspections, focused inspections/tests and expanded inspections), integrity evaluation and repair/replacement. If JEDIS were assigned greater than 3 with expanded inspections, inspection of piping with local wall thinning, its integrity evaluation and repair/replacement if necessary were requested. Inspection and evaluation of piping with local wall thinning was performed based on JSME or ASME codes. Detailed work flow charts were presented. Carbon steel piping and elbow was chosen for evaluation. (T. Tanaka)

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

    International Nuclear Information System (INIS)

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

  15. Induction of robust immunity response in mice by dual-expression-system-based recombinant baculovirus expressing the capsid protein of porcine circovirus type 2

    OpenAIRE

    Ye, Yu; Cheng, Xiaoliang; Jie ZHANG; Tong, Tiezhu; Lin, Wenyao; Liao, Ming; Fan, Huiying

    2013-01-01

    Background Porcine circovirus type 2 (PCV2) is associated with post-weaning multisystemic wasting syndrome (PMWS), an emerging swine disease that causes progressive weight loss, dyspnea, tachypnea, anemia, jaundice, and diarrhea in piglets. Although baculovirus is an enveloped virus that infects insects in nature, it has emerged as a vaccine vector, and we used it to develop a novel candidate vaccine for a preventive or therapeutic strategy to control PCV2 infections. Methods Immunoblotting a...

  16. Expression of an Innate Immune Element (Mouse Hepcidin-1) in Baculovirus Expression System and the Comparison of Its Function with Synthetic Human Hepcidin-25

    OpenAIRE

    Yazdani, Yaghoub; Sadeghi, Hamid; Alimohammadian, Mohammad; Andalib, Alireza; Moazen, Fatemeh; Rezaei, Abbas

    2011-01-01

    Hepcidin is an innate immune element which decreases the iron absorption from diet and iron releasing from macrophage cell. In contrast to the chemical iron chelators, there has been limited effort applied to the specific use of hepcidin as a new drug for decreasing the iron overload. Hepcidin is produced in different biological systems. For instance, E-coli is used for human hepcidin expression, however, post-translational modification is impaired. We have used a simple baculovirus expressio...

  17. Differential contributions of porcine bocavirus NP1 protein N- and C-terminal regions to its nuclear localization and immune regulation.

    Science.gov (United States)

    Zhang, Ruoxi; Fang, Liurong; Cai, Kaimei; Zeng, Songlin; Wu, Wei; An, Kang; Chen, Huanchun; Xiao, Shaobo

    2016-05-01

    Porcine bocavirus (PBoV), a newly identified parvovirus in the family Parvoviridae, has been reported worldwide in swine with post-weaning multisystemic wasting syndrome, respiratory disease or diarrhoea and in asymptomatic swine. NP1 is a protein unique to the genus Bocavirus and its function is not fully understood. In this study, we show that the N-terminal region of PBoV NP1 contains two classical nuclear localization signals (cNLSs) and a non-classical NLS. The N-terminal region also inhibits the promoter activity of IFN-β and IFN-stimulated response element activity the same as full-length NP1 protein, but the PBoV NP1 C-terminal region does not. PBoV NP1 also induces NFκB activation by increasing the phosphorylation of p65, and we demonstrate that the C-terminal region (aa 168-218) is responsible for the induction of NFκB, although the cNLS region of NP1 enhances this activation. The data suggest that PBoV NP1 contains two functionally independent domains in its N- and C-terminal regions. Thus, the N-terminal region of PBoV NP1 is critical for its nuclear localization and IFN-related promoter inhibition, and the C-terminal region is critical for its induction of NFκB. PMID:26813332

  18. The Site Selected. The Local Decision-Making Regarding the Siting of the Spent Nuclear Fuel Repository in Olkiluoto

    International Nuclear Information System (INIS)

    In May 1999 Posiva, the company responsible for the final disposal of spent nuclear fuel in Finland, suggested that the Finnish Government considers only Olkiluoto in Eurajoki in its application of a decision in principle to be a final disposal site. In January 2000 the municipal council of Eurajoki made a positive statement on the decision in principle. The Government made the decision in principle in Dec 2000, and the Parliament ratified the decision in May 2001. The paper is focused on the decision making of Eurajoki municipality regarding the siting of the spent nuclear fuel repository. The paper shows how the interaction between the representatives of the candidate municipality and the nuclear energy industry was the crucial factor in the decision-making. Eurajoki serves as an example, in where the parties reached an agreement of the compensations for the final disposal repository. The negotiations between the Eurajoki municipality and the nuclear energy industry in reaching a positive decision are analysed from the beginning of the 1980s. The main emphasis is however on the years 1996-99, when the nuclear energy industry negotiated with the municipality on the compensation for the final disposal repository. The loss of income was an important reason why some of the councillors of Eurajoki were interested in having the final disposal repository in Olkiluoto. The industry's problem on the other hand was to safeguard the final disposal site. From the TVO's angle Olkiluoto was a potential final disposal site for example for its limited need for transport and for the existing infrastructure. The company used the financial benefits of the project as its trump card. The attitude of Eurajoki municipality to the final disposal of spent nuclear fuel turned positive with the Olkiluoto vision in December 1998, when still five years earlier the municipal council was prepared to act and prevent the final disposal. The future image presented by the municipality now matched

  19. Study of the influence of central nuclear power plants at Valdaliga (Civitavecchia) on the chemical composition of local rain

    International Nuclear Information System (INIS)

    Data from a wet deposition sampling over the area collocated nearby Civitavecchia power plants are here presented and discussed. In order to establish the possible influence of the power plants emission on the chemical composition of the rain collected, the experimental data have been correlated with synoptic and local meteorological conditions. (author)

  20. Expression and localization of nuclear proteins in autosomal-dominant Emery-Dreifuss muscular dystrophy with LMNA R377H mutation

    Directory of Open Access Journals (Sweden)

    Ewald Andrea

    2004-03-01

    Full Text Available Abstract Background The autosomal dominant form of Emery-Dreifuss muscular dystrophy (AD-EDMD is caused by mutations in the gene encoding for the lamins A and C (LMNA. Lamins are intermediate filament proteins which form the nuclear lamina underlying the inner nuclear membrane. We have studied the expression and the localization of nuclear envelope proteins in three different cell types and muscle tissue of an AD-EDMD patient carrying a point mutation R377H in the lamin A/C gene. Results Lymphoblastoid cells, skin fibroblasts, primary myoblasts and muscle thin sections were studied by immunocytochemistry and electron microscopy. Cellular levels of A-type lamins were reduced compared to control cells. In contrast, the amount of emerin and lamin B appeared unaltered. Cell synchronization experiments showed that the reduction of the cellular level of A-type lamin was due to instability of lamin A. By electron microscopy, we identified a proportion of nuclei with morphological alterations in lymphoblastoid cells, fibroblasts and mature muscle fibres. Immunofluorescence microscopy showed that a major population of the lamin B receptor (LBR, an inner nuclear membrane protein, was recovered in the cytoplasm in association with the ER. In addition, the intranuclear organization of the active form of RNA polymerase II was markedly different in cells of this AD-EDMD patient. This aberrant intranuclear distribution was specifically observed in muscle cells where the pathology of EDMD predominates. Conclusions From our results we conclude: Firstly, that structural alterations of the nuclei which are found only in a minor fraction of lymphoblastoid cells and mature muscle fibres are not sufficient to explain the clinical pathology of EDMD; Secondly, that wild type lamin A is required not only for the retention of LBR in the inner nuclear membrane but also for a correct localization of the transcriptionally active RNA pol II in muscle cells. We speculate that

  1. Mucosal Delivery of ACNPV Baculovirus Driving Expression of the Gal-Lectin LC3 Fragment Confers Protection against Amoebic Liver Abscess in Hamster

    Directory of Open Access Journals (Sweden)

    DM Meneses-Ruiz, JP Laclette, H Aguilar-Díaz, J Hernández-Ruiz, A Luz-Madrigal, A Sampieri, L Vaca, JC Carrero

    2011-01-01

    Full Text Available Mucosal vaccination against amoebiasis using the Gal-lectin of E. histolytica has been proposed as one of the leading strategies for controlling this human disease. However, most mucosal adjuvants used are toxic and the identification of safe delivery systems is necessary. Here, we evaluate the potential of a recombinant Autographa californica baculovirus driving the expression of the LC3 fragment of the Gal-lectin to confer protection against amoebic liver abscess (ALA in hamsters following oral or nasal immunization. Hamsters immunized by oral route showed complete absence (57.9% or partial development (21% of ALA, resulting in some protection in 78.9% of animals when compared with the wild type baculovirus and sham control groups. In contrast, nasal immunization conferred only 21% of protection efficacy. Levels of ALA protection showed lineal correlation with the development of an anti-amoebic cellular immune response evaluated in spleens, but not with the induction of seric IgG anti-amoeba antibodies. These results suggest that baculovirus driving the expression of E. histolytica vaccine candidate antigens is useful for inducing protective cellular and humoral immune responses following oral immunization, and therefore it could be used as a system for mucosal delivery of an anti-amoebic vaccine.

  2. ZNF198, a zinc finger protein rearranged in myeloproliferative disease, localizes to the PML nuclear bodies and interacts with SUMO-1 and PML

    International Nuclear Information System (INIS)

    The ZNF198/FGFR1 fusion gene in atypical myeloproliferative disease produces a constitutively active cytoplasmic tyrosine kinase, unlike ZNF198 which is normally a nuclear protein. We have now shown that the ZNF198/FGFR1 fusion kinase interacts with the endogenous ZNF198 protein suggesting that the function of ZNF198 may be compromised in cells expressing it. Little is currently known about the endogenous function of ZNF198 and to investigate this further we performed a yeast two-hybrid analysis and identified SUMO-1 as a binding partner of ZNF198. These observations were confirmed using co-immunoprecipitation which demonstrated that ZNF198 is covalently modified by SUMO-1. Since many of the SUMO-1-modified proteins are targeted to the PML nuclear bodies we used confocal microscopy to show that SUMO-1, PML and ZNF198 colocalize to punctate structures, shown by immunocytochemistry to be PML bodies. Using co-immunoprecipitation we now show that PML and sumoylated ZNF198 can be found in a protein complex in the cell. Mutation of the SUMO-1 binding site in wild-type ZNF198 resulted in loss of distinct PML bodies, reduced PML levels and a more dispersed nuclear localization of the PML protein. In cells expressing ZNF198/FGFR1, which also lack the SUMO-1 binding site, SUMO-1 is preferentially localized in the cytoplasm, which is associated with loss of distinct PML bodies. Recently, arsenic trioxide (ATO) was proposed as an alternative therapy for APL that was resistant to traditional therapy. Treatment of cells expressing ZNF198/FGFR1 with ATO demonstrated reduced autophosphorylation of the ZNF198/FGFR1 protein and induced apoptosis, which is not seen in cells expressing wild-type ZNF198. Overall our results suggest that the sumoylation of ZNF198 is important for PML body formation and that the abrogation of sumoylation of ZNF198 in ZNF198/FGFR1 expressing cells may be an important mechanism in cellular transformation

  3. Nuclear localization of lymphocyte-specific protein tyrosine kinase (Lck) and its role in regulating LIM domain only 2 (Lmo2) gene

    Energy Technology Data Exchange (ETDEWEB)

    Venkitachalam, Srividya; Chueh, Fu-Yu [Department of Microbiology and Immunology, H. M. Bligh Cancer Research Laboratories, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL 60064 (United States); Yu, Chao-Lan, E-mail: chaolan.yu@rosalindfranklin.edu [Department of Microbiology and Immunology, H. M. Bligh Cancer Research Laboratories, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL 60064 (United States)

    2012-01-20

    Highlights: Black-Right-Pointing-Pointer Lmo2 expression is elevated in Lck-transformed cells. Black-Right-Pointing-Pointer Both endogenous and exogenous Lck localize in the nucleus. Black-Right-Pointing-Pointer Nuclear Lck is active in Lck-transformed cells. Black-Right-Pointing-Pointer Lck binds to the promoter region of Lmo2 gene in vivo. Black-Right-Pointing-Pointer In contrast to JAK2, Lck does not increase histone H3 phosphorylation on Tyr 41. -- Abstract: LIM domain only protein 2 (Lmo2) is a transcription factor that plays a critical role in the development of T-acute lymphoblastic leukemia (T-ALL). A previous report established a link between Lmo2 expression and the nuclear presence of oncogenic Janus kinase 2 (JAK2), a non-receptor protein tyrosine kinase. The oncogenic JAK2 kinase phosphorylates histone H3 on Tyr 41 that leads to the relief of Lmo2 promoter repression and subsequent gene expression. Similar to JAK2, constitutive activation of lymphocyte-specific protein tyrosine kinase (Lck) has been implicated in lymphoid malignancies. However, it is not known whether oncogenic Lck regulates Lmo2 expression through a similar mechanism. We show here that Lmo2 expression is significantly elevated in T cell leukemia LSTRA overexpressing active Lck kinase and in HEK 293 cells expressing oncogenic Y505FLck kinase. Nuclear localization of active Lck kinase was confirmed in both Lck-transformed cells by subcellular fractionation and immunofluorescence microscopy. More importantly, in contrast to oncogenic JAK2, oncogenic Lck kinase does not result in significant increase in histone H3 phosphorylation on Tyr 41. Instead, chromatin immunoprecipitation experiment shows that oncogenic Y505FLck kinase binds to the Lmo2 promoter in vivo. This result raises the possibility that oncogenic Lck may activate Lmo2 promoter through direct interaction.

  4. Quantitative and subcellular localization analysis of the nuclear isoform dUTP pyrophosphatase in alkylating agent-induced cell responses

    International Nuclear Information System (INIS)

    Highlights: → MNNG-induced appearance of DUT-N in the extracellular fluid has cellular specificity. → MNNG alters the subcellular distribution of DUT-N in human cells in different ways. → DUT-N may be a potential biomarker to assess the risk of alkylating agents exposure. -- Abstract: Our previous proteome analysis showed that the nuclear isoform of dUTP pyrophosphatase (DUT-N) was identified in the culture medium of human amnion FL cells after exposure to the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). These results suggest that DUT-N may be a potential early biomarker to assess the risk of alkylating agents exposure. DUT-N is one of the two isoforms of deoxyuridine triphosphate nucleotidohydrolase (dUTPase). Our current knowledge of DUT-N expression in human cells is very limited. In the current study, we first investigated the appearance of DUT-N in the culture medium of different human cell lines in response to a low concentration of MNNG exposure. We verified that the MNNG-induced appearance of DUT-N in the extracellular environment is cell-specific. Western blot analysis confirmed that the intracellular DUT-N changes responded to MNNG in a concentration-dependent and cell-specific manner. Furthermore, subcellular fraction experiments showed that 0.25 μM MNNG treatment dramatically increased the DUT-N expression levels in the cytoplasmic extracts prepared from both FL and HepG2 cells, increased DUT-N levels in nuclear extracts prepared from HepG2 cells, and decreased DUT-N levels in nuclear extracts from FL cells. Morphological studies using immunofluorescence showed that a low concentration of MNNG could alter the distribution of DUT-N in FL and HepG2 cells in different ways. Taken together, these studies indicate a role of DUT-N in alkylating agent-induced cell responses.

  5. Specific nuclear localizing sequence directs two myosin isoforms to the cell nucleus in calmodulin-sensitive manner

    Czech Academy of Sciences Publication Activity Database

    Dzijak, Rastislav; Yildirim, Sukriye; Kahle, Michal; Novák, Petr; Hnilicová, Jarmila; Venit, Tomáš; Hozák, Pavel

    2012-01-01

    Roč. 7, č. 1 (2012), e30529. E-ISSN 1932-6203 R&D Projects: GA ČR(CZ) GA204/07/1592; GA ČR(CZ) GD204/09/H084; GA ČR GAP305/11/2232; GA MŠk LC545; GA MŠk(CZ) LC06063 Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 ; RVO:61388971 Keywords : NM1 * nuclear import * NLS * calmodulin Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.730, year: 2012

  6. Efficient Subcellular Targeting to the Cell Nucleus of Quantum Dots Densely Decorated with a Nuclear Localization Sequence Peptide.

    Science.gov (United States)

    Maity, Amit Ranjan; Stepensky, David

    2016-01-27

    Organelle-targeted drug delivery can enhance the efficiency of the intracellularly acting drugs and reduce their toxicity. We generated core-shell type CdSe-ZnS quantum dots (QDs) densely decorated with NLS peptidic targeting residues using a 3-stage decoration approach and investigated their endocytosis and nuclear targeting efficiencies. The diameter of the generated QDs increased following the individual decoration stages (16.3, 18.9, and 21.9 nm), the ζ-potential became less negative (-33.2, -17.5, and -11.9 mV), and characteristic changes appeared in the FTIR spectra following decoration with the linker and NLS peptides. Quantitative analysis of the last decoration stage revealed that 37.9% and 33.2% of the alkyne-modified NLS groups that were added to the reaction mix became covalently attached or adsorbed to the QDs surface, respectively. These numbers correspond to 63.6 and 55.7 peptides conjugated or adsorbed to a single QD (the surface density of 42 and 37 conjugated and adsorbed peptides per 1000 nm(2) of the QDs surface), which is higher than in the majority of previous studies that reported decoration efficiencies of formulations intended for nuclear-targeted drug delivery. QDs decorated with NLS peptides undergo more efficient endocytosis, as compared to other investigated QDs formulations, and accumulated to a higher extent in the cell nucleus or in close vicinity to it (11.9%, 14.6%, and 56.1% of the QDs endocytosed by an average cell for the QD-COOH, QD-azide, and QD-NLS formulations, respectively). We conclude that dense decoration of QDs with NLS residues increased their endocytosis and led to their nuclear targeting (preferential accumulation in the cells nuclei or in close vicinity to them). The experimental system and research tools that were used in this study allow quantitative investigation of the mechanisms that govern the QDs nuclear targeting and their dependence on the formulation properties. These findings will contribute to the

  7. Initial Evaluation of the Heat-Affected Zone, Local Embrittlement Phenomenon as it Applies to Nuclear Reactor Vessels

    International Nuclear Information System (INIS)

    The objective of this project was to determine if the local brittle zone (LBZ) problem, encountered in the testing of the heat-affected zone (HAZ) part of welds in offshore platform construction, can also be found in reactor pressure vessel (RPV) welds. Both structures have multipass welds and grain coarsening along the fusion line. Literature was obtained that described the metallurgical evidence and the type of research work performed on offshore structure welds

  8. Detection of localized and general corrosion of mild steel in simulated defense nuclear waste solutions using electrochemical noise analysis

    International Nuclear Information System (INIS)

    Underground waste tanks fabricated from mild steel store more than 60 million gallons of radioactive waste from 50 years of weapons production. Leaks are suspected in a significant number of tanks. The probable modes of corrosion failures are reported to be localized corrosion (e.g. nitrate stress corrosion cracking and pitting). The use of electrochemical noise (EN) for the monitoring and detection of localized corrosion processes has received considerable attention and application over the last several years. Proof of principle laboratory tests were conducted to verify the capability of EN evaluation to detect localized corrosion and to compare the predictions of general corrosion obtained from EN with those derived from other sources. Simple, pre-fabricated flat and U-bend specimens of steel alloys A516-Grade 60 (UNS K02100) and A537-CL 1 (UNS K02400) were immersed in temperature controlled simulated waste solutions. The simulated waste solution was either 5M NaNO3 with 0.3M NaOH at 90 C or 11M NaNO3 with 0.15M NaOH at 95 C. The electrochemical noise activity from the specimens was monitored and recorded for periods ranging between 140 and 240 hours. At the end of each test period, the specimens were metallographically examined to correlated EN data with corrosion damage

  9. Concentrations of Radiocesium in Local Foods Collected in Kawauchi Village after the Accident at the Fukushima Dai-ichi Nuclear Power Station

    Science.gov (United States)

    Orita, Makiko; Nakashima, Kanami; Hayashida, Naomi; Endo, Yuuko; Yamashita, Shunichi; Takamura, Noboru

    2016-06-01

    We evaluated the current concentrations of radiocesium in local foods collected in Kawauchi Village, which is located less than 30 km from Fukushima Daiichi Nuclear Power Station, to minimize public anxiety regarding internal radiation exposure through the consumption of locally produced foods after the 2011 Fukushima accident. The number of samples exceeding the regulatory radiocesium limit (100 Bq/kg for general foods) was five out of 4,080 vegetables (0.1%), 652 of 1,986 (32.8%) among edible wild plants and fungi, and eight of 647 (1.2%) in fruits. Our study confirmed that the internal radiation doses of ingesting these foods are acceptably low compared to the public dose limit, ranging from 24.4 to 42.7 μSv for males and from 21.7 to 43.4 μSv for females, although the potential for radiation exposure still exists. Long-term comprehensive follow-up should take place to clarify trends in radiocesium concentrations in local foods and the committed effective doses found in Fukushima-area residents. By constructing a system that allows residents to access information on radiocesium concentration in foods, a risk communication model between specialists and residents could be developed in the recovery phase after the Fukushima accident.

  10. Concentrations of Radiocesium in Local Foods Collected in Kawauchi Village after the Accident at the Fukushima Dai-ichi Nuclear Power Station.

    Science.gov (United States)

    Orita, Makiko; Nakashima, Kanami; Hayashida, Naomi; Endo, Yuuko; Yamashita, Shunichi; Takamura, Noboru

    2016-01-01

    We evaluated the current concentrations of radiocesium in local foods collected in Kawauchi Village, which is located less than 30 km from Fukushima Daiichi Nuclear Power Station, to minimize public anxiety regarding internal radiation exposure through the consumption of locally produced foods after the 2011 Fukushima accident. The number of samples exceeding the regulatory radiocesium limit (100 Bq/kg for general foods) was five out of 4,080 vegetables (0.1%), 652 of 1,986 (32.8%) among edible wild plants and fungi, and eight of 647 (1.2%) in fruits. Our study confirmed that the internal radiation doses of ingesting these foods are acceptably low compared to the public dose limit, ranging from 24.4 to 42.7 μSv for males and from 21.7 to 43.4 μSv for females, although the potential for radiation exposure still exists. Long-term comprehensive follow-up should take place to clarify trends in radiocesium concentrations in local foods and the committed effective doses found in Fukushima-area residents. By constructing a system that allows residents to access information on radiocesium concentration in foods, a risk communication model between specialists and residents could be developed in the recovery phase after the Fukushima accident. PMID:27334847

  11. A Deoxynivalenol-Activated Methionyl-tRNA Synthetase Gene from Wheat Encodes a Nuclear Localized Protein and Protects Plants Against Fusarium Pathogens and Mycotoxins.

    Science.gov (United States)

    Zuo, Dong-Yun; Yi, Shu-Yuan; Liu, Rong-Jing; Qu, Bo; Huang, Tao; He, Wei-Jie; Li, Cheng; Li, He-Ping; Liao, Yu-Cai

    2016-06-01

    Fusarium graminearum is the fungal pathogen that causes globally important diseases of cereals and produces mycotoxins such as deoxynivalenol (DON). Owing to the dearth of available sources of resistance to Fusarium pathogens, characterization of novel genes that confer resistance to mycotoxins and mycotoxin-producing fungi is vitally important for breeding resistant crop varieties. In this study, a wheat methionyl-tRNA synthetase (TaMetRS) gene was identified from suspension cell cultures treated with DON. It shares conserved aminoacylation catalytic and tRNA anticodon binding domains with human MetRS and with the only previously characterized plant MetRS, suggesting that it functions in aminoacylation in the cytoplasm. However, the TaMetRS comprises a typical nuclear localization signal and cellular localization studies with a TaMetRS::GFP fusion protein showed that TaMetRS is localized in the nucleus. Expression of TaMetRS was activated by DON treatment and by infection with a DON-producing F. graminearum strain in wheat spikes. No such activation was observed following infection with a non-DON-producing F. graminearum strain. Expression of TaMetRS in Arabidopsis plants conferred significant resistance to DON and F. graminearum. These results indicated that this DON-activated TaMetRS gene may encode a novel type of MetRS in plants that has a role in defense and detoxification. PMID:26882849

  12. The C-terminal region of Rad52 is essential for Rad52 nuclear and nucleolar localization, and accumulation at DNA damage sites immediately after irradiation

    International Nuclear Information System (INIS)

    Highlights: •Rad52 might play a key role in the repair of DSB immediately after irradiation. •EYFP-Rad52 accumulates rapidly at DSB sites and colocalizes with Ku80. •Accumulation of Rad52 at DSB sites is independent of the core NHEJ factors. •Localization and recruitment of Rad52 to DSB sites are dependent on the Rad52 CTR. •Basic amino acids in Rad52 CTR are highly conserved among vertebrate species. -- Abstract: Rad52 plays essential roles in homologous recombination (HR) and repair of DNA double-strand breaks (DSBs) in Saccharomyces cerevisiae. However, in vertebrates, knockouts of the Rad52 gene show no hypersensitivity to agents that induce DSBs. Rad52 localizes in the nucleus and forms foci at a late stage following irradiation. Ku70 and Ku80, which play an essential role in nonhomologous DNA-end-joining (NHEJ), are essential for the accumulation of other core NHEJ factors, e.g., XRCC4, and a HR-related factor, e.g., BRCA1. Here, we show that the subcellular localization of EYFP-Rad52(1–418) changes dynamically during the cell cycle. In addition, EYFP-Rad52(1–418) accumulates rapidly at microirradiated sites and colocalizes with the DSB sensor protein Ku80. Moreover, the accumulation of EYFP-Rad52(1–418) at DSB sites is independent of the core NHEJ factors, i.e., Ku80 and XRCC4. Furthermore, we observed that EYFP-Rad52(1–418) localizes in nucleoli in CHO-K1 cells and XRCC4-deficient cells, but not in Ku80-deficient cells. We also found that Rad52 nuclear localization, nucleolar localization, and accumulation at DSB sites are dependent on eight amino acids (411–418) at the end of the C-terminal region of Rad52 (Rad52 CTR). Furthermore, basic amino acids on Rad52 CTR are highly conserved among mammalian, avian, and fish homologues, suggesting that Rad52 CTR is important for the regulation and function of Rad52 in vertebrates. These findings also suggest that the mechanism underlying the regulation of subcellular localization of Rad52 is

  13. The C-terminal region of Rad52 is essential for Rad52 nuclear and nucleolar localization, and accumulation at DNA damage sites immediately after irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Koike, Manabu, E-mail: m_koike@nirs.go.jp [DNA Repair Gene Res., National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan); Yutoku, Yasutomo [DNA Repair Gene Res., National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan); Graduate School of Science, Chiba University, Yayoicho, Inage-ku, Chiba 263-8522 (Japan); Koike, Aki [DNA Repair Gene Res., National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan)

    2013-05-31

    Highlights: •Rad52 might play a key role in the repair of DSB immediately after irradiation. •EYFP-Rad52 accumulates rapidly at DSB sites and colocalizes with Ku80. •Accumulation of Rad52 at DSB sites is independent of the core NHEJ factors. •Localization and recruitment of Rad52 to DSB sites are dependent on the Rad52 CTR. •Basic amino acids in Rad52 CTR are highly conserved among vertebrate species. -- Abstract: Rad52 plays essential roles in homologous recombination (HR) and repair of DNA double-strand breaks (DSBs) in Saccharomyces cerevisiae. However, in vertebrates, knockouts of the Rad52 gene show no hypersensitivity to agents that induce DSBs. Rad52 localizes in the nucleus and forms foci at a late stage following irradiation. Ku70 and Ku80, which play an essential role in nonhomologous DNA-end-joining (NHEJ), are essential for the accumulation of other core NHEJ factors, e.g., XRCC4, and a HR-related factor, e.g., BRCA1. Here, we show that the subcellular localization of EYFP-Rad52(1–418) changes dynamically during the cell cycle. In addition, EYFP-Rad52(1–418) accumulates rapidly at microirradiated sites and colocalizes with the DSB sensor protein Ku80. Moreover, the accumulation of EYFP-Rad52(1–418) at DSB sites is independent of the core NHEJ factors, i.e., Ku80 and XRCC4. Furthermore, we observed that EYFP-Rad52(1–418) localizes in nucleoli in CHO-K1 cells and XRCC4-deficient cells, but not in Ku80-deficient cells. We also found that Rad52 nuclear localization, nucleolar localization, and accumulation at DSB sites are dependent on eight amino acids (411–418) at the end of the C-terminal region of Rad52 (Rad52 CTR). Furthermore, basic amino acids on Rad52 CTR are highly conserved among mammalian, avian, and fish homologues, suggesting that Rad52 CTR is important for the regulation and function of Rad52 in vertebrates. These findings also suggest that the mechanism underlying the regulation of subcellular localization of Rad52 is

  14. Case-study, session 2. Nuclear facilities and local information. Speech by Lennart Franzon (together with L.G. Wahlberg, OKG)

    International Nuclear Information System (INIS)

    Forsmark NPP is situated 100 miles north of Stockholm on the east coast. The plant has 3 units of about 1 000 MWe each. The annual net production is about 16% of the Swedish electric power consumption. Forsmark was the only Swedish nuclear power plant not yet operating at all at the time of te nuclear power referendum in 1980. The neighbourhood of Forsmark means a radius of about 50 miles from the station, which means about 100 000 inhabitants, including a lot of summer house owners. The first building in use in Forsmark 1973 - seven years before operation start - was a visitors center. Since then 400 000 people have visited Forsmark. The visitors programmes are the most important activities towards the Swedes. To let visitors into a reactor hall balcony, a few meters above the high level waste close to the reactor, is a good way to explain the nuclear power. Many visitors also feel privileged when they together with their friends are able to investigate the radiation protection monitors, where the Chernobyl disaster was detected for the first time outside the Soviet Union in April 1986. Since 1983 both positive and negative aspect of reactor operation are advertised. Data related to electricity supply availability, employment, radiation doses of the employees, radioactive release into air and sea, operational problems, events etc. are all published by the operator in the local newspapers quarterly. The experience is that negative publicity, based on authority reports has decreased. It was found that the most important for the owner/operator is to have the initiative all the time. Similar to Oskarshamn Power Group Forsmark edits a magazine - Forsmarks Tidning - twice a year since 1980, the year of operation start. The magazine is distributed to about 65.000 homes, to authorities, politicians, employees, private companies, etc. About 50% of the neighbours read the magazine. This is considered as success. Through the years of operation of Forsmark it was founds that

  15. Present state and some problem of health services for residents of local government neighboring Fukushima Daiichi Nuclear Power Station

    International Nuclear Information System (INIS)

    After nuclear disaster, residents of 8 towns and villages of Futaba county within Alert area moved to the west and further removed far. Public offices were also moved. On March 12 health services were started to accept refugee residents at the first refuge. From the end of March to the first part of April they moved to second refuge at specified hotel or inn prepared for a long-term refugee life and then they removed from second refuge to temporary dwelling or loan house. Some of them moved to temporary dwelling at Iwaki city and totaled to more than 20000. New organization of the Fukushima prefecture was established in Iwaki city to start health services for refugee including mental cares. (T. Tanaka)

  16. Ca2+ Channel Re-localization to Plasma-Membrane Microdomains Strengthens Activation of Ca2+-Dependent Nuclear Gene Expression

    Directory of Open Access Journals (Sweden)

    Krishna Samanta

    2015-07-01

    Full Text Available In polarized cells or cells with complex geometry, clustering of plasma-membrane (PM ion channels is an effective mechanism for eliciting spatially restricted signals. However, channel clustering is also seen in cells with relatively simple topology, suggesting it fulfills a more fundamental role in cell biology than simply orchestrating compartmentalized responses. Here, we have compared the ability of store-operated Ca2+ release-activated Ca2+ (CRAC channels confined to PM microdomains with a similar number of dispersed CRAC channels to activate transcription factors, which subsequently increase nuclear gene expression. For similar levels of channel activity, we find that channel confinement is considerably more effective in stimulating gene expression. Our results identify a long-range signaling advantage to the tight evolutionary conservation of channel clustering and reveal that CRAC channel aggregation increases the strength, fidelity, and reliability of the general process of excitation-transcription coupling.

  17. 16. national conference of the presidents of the local commissions of information (C.L.I.) around nuclear facilities

    International Nuclear Information System (INIS)

    This conference contained two plenary sessions. The first one concerned the epidemiology with the following subjects: the epidemiology, its contributions and its limits; the epidemiological surveillance of cancers in France: role of registers within the device; the aggregates of leukaemia near the nuclear installations; perception and management of the risk; the point of view of the scientific committee of the A.N.C.L.I.; Followed a work in a workshop on the subjects relative to the functioning of the C.L.I. (Organization, relations, information) then the second plenary session with as subjects: organization and functioning of C.L.I., relations and links with A.N.C.L.I., information received and diffused by the C.L.I. (N.C.)

  18. Dasatinib, a multi-kinase inhibitor increased radiation sensitivity by interfering with nuclear localization of epidermal growth factor receptor and by blocking DNA repair pathways

    International Nuclear Information System (INIS)

    Background and purpose: Although inhibition of epidermal growth factor receptor (EGFR) signaling during radiation led to improvement of tumor control and survival, novel strategies are needed to further improve the outcome of patients with locally advanced head and neck carcinoma. Because EGFR is known to interact with c-Src kinases, the present study investigated dasatinib (BMS-354825), an inhibitor of c-Src kinases, for its efficacy in enhancing radiosensitivity of human head and neck squamous cell carcinomas (HNSCC) in vitro and examined the underlying mechanisms for this effect. Materials and methods: Six HNSCC lines were exposed to dasatinib, radiation, or both, and assessed for c-Src and EGFR expression, cell survival and colony forming ability. Among these cell lines, HN-5 and FaDu lines were analyzed for induction of apoptosis, cell cycle re-distribution and for nuclear localization of EGFR, γ-H2AX and 53BP1 proteins. Immuno-precipitation and Western blots were performed to analyze the levels and binding of proteins involved in cell survival, apoptosis and DNA repair pathways. Suppression of c-Src by siRNA and subsequent clonogenic assay was performed in HN-5 cells. Results: All six HNSCC lines that were examined expressed high levels of c-Src. Two (HN-5 and MDA-183) expressed higher levels of EGFR than other lines. Dasatinib suppressed cell survival of all cell lines tested independent of c-Src or EGFR levels but enhanced the radiosensitivity of HN-5 and MDA-183. HN-5 and FaDu were analyzed further. Dasatinib suppressed phosphorylation of c-Src in both cell lines, but decreased repair of radiation-induced DNA damage in HN-5 cells only as evidenced by suppression of c-Abl and Nbs-1 activity, inhibition of the association between c-Src and EGFR or Her-2, prolongation of nuclear γ-H2AX and 53BP1 foci and inhibition of EGFR nuclear localization and its association with DNA-PKcs. Finally, partial suppression of c-Src resulted in a small increase in HN-5 cell

  19. Hydrophobic Interactions Involved in Attachment of a Baculovirus to Hydrophobic Surfaces

    OpenAIRE

    Small, Deirdre A.; Moore, Norman F.; Entwistle, Philip F.

    1986-01-01

    The hydrophobic interactions of Trichoplusia ni nuclear polyhedrosis virus were characterized by hydrophobic interaction chromatography. The determination of the hydrophobic force and some of the factors that influence its size is discussed in relation to the attachment to leaf surfaces of polyhedra during their use as biological control agents against insect pests.

  20. Developmental and adult expression patterns of the G-protein-coupled receptor GPR88 in the rat: Establishment of a dual nuclear-cytoplasmic localization.

    Science.gov (United States)

    Massart, Renaud; Mignon, Virginie; Stanic, Jennifer; Munoz-Tello, Paola; Becker, Jerôme A J; Kieffer, Brigitte L; Darmon, Michèle; Sokoloff, Pierre; Diaz, Jorge

    2016-10-01

    GPR88 is a neuronal cerebral orphan G-protein-coupled receptor (GPCR) that has been linked to various psychiatric disorders. However, no extensive description of its localization has been provided so far. Here, we investigate the spatiotemporal expression of the GPR88 in prenatal and postnatal rat tissues by using in situ hybridization and immunohistochemistry. GPR88 protein was initially detected at embryonic day 16 (E16) in the striatal primordium. From E16-E20 to adulthood, the highest expression levels of both protein and mRNA were observed in striatum, olfactory tubercle, nucleus accumbens, amygdala, and neocortex, whereas in spinal cord, pons, and medulla GPR88 expression remains discrete. We observed an intracellular redistribution of GPR88 during cortical lamination. In the cortical plate of the developing cortex, GPR88 presents a classical GPCR plasma membrane/cytoplasmic localization that shifts, on the day of birth, to nuclei of neurons progressively settling in layers V to II. This intranuclear localization remains throughout adulthood and was also detected in monkey and human cortex as well as in the amygdala and hypothalamus of rats. Apart from the central nervous system, GPR88 was transiently expressed at high levels in peripheral tissues, including adrenal cortex (E16-E21) and cochlear ganglia (E19-P3), and also at moderate levels in retina (E18-E19) and spleen (E21-P7). The description of the GPR88 anatomical expression pattern may provide precious functional insights into this novel receptor. Furthermore, the GRP88 nuclear localization suggests nonclassical GPCR modes of action of the protein that could be relevant for cortical development and psychiatric disorders. J. Comp. Neurol. 524:2776-2802, 2016. © 2016 Wiley Periodicals, Inc. PMID:26918661