bluuebunny/crossref_metadata_2025_split huggingface dataset:
African Swine Fever Virus Is Enveloped by a Two-Membraned Collapsed Cisterna Derived from the Endoplasmic Reticulum
Authors: Andrés Germán , García-Escudero Ramón , Simón-Mateo Carmen , Viñuela Eladio 
Published: December 2019

ABSTRACTDuring the cytoplasmic maturation of African swine fever virus (ASFV) within the viral factories, the DNA-containing core becomes wrapped by two shells, an inner lipid envelope and an outer icosahedral capsid. We have previously shown that the inner envelope is derived from precursor membrane-like structures on which the capsid layer is progressively assembled. In the present work, we analyzed the origin of these viral membranes and the mechanism of envelopment of ASFV. Electron microscopy studies on permeabilized infected cells revealed the presence of two tightly apposed membranes within the precursor membranous structures as well as polyhedral assembling particles. Both membranes could be detached after digestion of intracellular virions with proteinase K. Importantly, membrane loop structures were observed at the ends of open intermediates, which suggests that the inner envelope is derived from a membrane cisterna. Ultraestructural and immunocytochemical analyses showed a close association and even direct continuities between the endoplasmic reticulum (ER) and assembling virus particles at the bordering areas of the viral factories. Such interactions become evident with an ASFV recombinant that inducibly expresses the major capsid protein p72. In the absence of the inducer, viral morphogenesis was arrested at a stage at which partially and fully collapsed ER cisternae enwrapped the core material. Together, these results indicate that ASFV, like the poxviruses, becomes engulfed by a two-membraned collapsed cisterna derived from the ER.
Functional Differences between BHRF1, the Epstein-Barr Virus-Encoded Bcl-2 Homologue, and Bcl-2 in Human Epithelial Cells
Authors: Dawson Christopher W. , Dawson Joanne , Jones Richard , Ward Kim , Young Lawrence S. 
Published: December 2019

ABSTRACT
BHRF1, a component of the restricted early antigen complex of the Epstein-Barr virus lytic cycle, encodes a 17-kDa protein with both sequence and functional homology to the antiapoptotic Bcl-2 oncogene. Recent work has suggested that BHRF1 behaves like Bcl-2 in protecting cells from apoptosis induced by a range of stimuli. In this study, the effect of BHRF1 and Bcl-2 on the growth and differentiation of the SCC12F human epithelial cell line was examined. The levels of stable transfected BHRF1 expression achievable in SCC12F cells was consistently lower than that obtained with Bcl-2. While both BHRF1 and Bcl-2 inhibited epithelial differentiation, the effect of Bcl-2 was more pronounced, resulting in an almost complete blockade of differentiation in organotypic raft cultures. However, BHRF1-expressing SCC12F cells proliferated at a much higher rate than SCC12F cells expressing Bcl-2, and this effect was supported by cell cycle analysis which demonstrated that BHRF1, but not Bcl-2, promotes rapid transit through the cell cycle. These data highlight important differences between BHRF1 and Bcl-2 and suggest that BHRF1 may function to promote the survival and proliferation of lytically infected cells. The proliferative properties of BHRF1 described in this study, together with the demonstration that other oncogenic gamma herpesviruses encode Bcl-2 homologues, suggests that these proteins may serve to increase the susceptibility of virus-infected cells to oncogenic transformation, thereby contributing to the development of virus-associated tumors.
Oral Immunization of Macaques with Attenuated Vaccine Virus Induces Protection against Vaginally Transmitted AIDS
Authors: Joag Sanjay V. , Liu Zhen Qian , Stephens Edward B. , Smith Marilyn S. , Kumar Anil , Li Zhuang , Wang Chunyang , Sheffer Darlene , Jia Fenglan , Foresman Larry , Adany Istvan , Lifson Jeff , McClure Harold M. , Narayan Opendra 
Published: December 2019

ABSTRACTThe chimeric simian-human immunodeficiency virus SHIVKU-1, bearing the envelope of human immunodeficiency virus type 1 (HIV-1), causes fulminant infection with subtotal loss of CD4+T cells followed by development of AIDS in intravaginally inoculated macaques and thus provides a highly relevant model of sexually transmitted disease caused by HIV-1 in human beings. Previous studies using this SHIV model had shown that thevpuandnefgenes were important in pathogenesis of the infection, and so we deleted portions of these genes to create two vaccines, ΔvpuΔnefSHIV-4 (vaccine 1) and ΔvpuSHIVPPc(vaccine 2). Six adult macaques were immunized subcutaneously with vaccine 1, and six were immunized orally with vaccine 2. Both viruses caused infection in all inoculated animals, but whereas vaccine 1 virus caused only a nonproductive type of infection, vaccine 2 virus replicated productively but transiently for a 6- to 10-week period. Both groups were challenged 6 to 7 months later with pathogenic SHIVKU-1by the intravaginal route. All four unvaccinated controls developed low CD4+T-cell counts (<200/μl) and AIDS. The 12 vaccinated animals all became infected with SHIVKU-1, and two in group 1 developed a persistent productive infection followed by development of AIDS in one. The other 10 have maintained almost complete control over virus replication even though spliced viral RNA was detected in lymph nodes. This suppression of virus replication correlated with robust antiviral cell-mediated immune responses. This is the first demonstration of protection against virulent SHIV administered by the intravaginal route. This study supports the concept that sexually transmitted HIV disease can be prevented by parenteral or oral immunization.
Studies of the Neutralizing Activity and Avidity of Anti-Human Immunodeficiency Virus Type 1 Env Antibody Elicited by DNA Priming and Protein Boosting
Authors: Richmond J. F. L. , Lu S. , Santoro J. C. , Weng J. , Hu Shiu-Lok , Montefiori D. C. , Robinson H. L. 
Published: December 2019

ABSTRACTDNA vaccination is an effective means of eliciting strong antibody responses to a number of viral antigens. However, DNA immunization alone has not generated persistent, high-titer antibody and neutralizing antibody responses to human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env). We have previously reported that DNA-primed anti-Env antibody responses can be augmented by boosting with Env-expressing recombinant vaccinia viruses. We report here that recombinant Env protein provides a more effective boost of DNA-initiated antibody responses. In rabbits primed with Env-expressing plasmids, protein boosting increased titer, persistence, neutralizing activity, and avidity of anti-Env responses. While titers increased rapidly after boosting, avidity and neutralizing activity matured more slowly over a 6-month period following protein boosting. DNA priming and protein immunization with HIV-1 HXB-2 Env elicited neutralizing antibody for T cell line-adapted, but not primary isolate, viruses. The most effective neutralizing antibody responses were observed after priming with plasmids which expressed noninfectious virus-like particles. In contrast to immunizations with HIV-1 Env, DNA immunizations with the influenza virus hemagglutinin glycoprotein did not require a protein boost to achieve high-titer antibody with good avidity and persistence.
Development of a Heterologous Model in Germfree Suckling Rats for Studies of Rotavirus Diarrhea
Authors: Guerin-Danan C. , Meslin J. C. , Lambre F. , Charpilienne A. , Serezat M. , Bouley C. , Cohen J. , Andrieux C. 
Published: December 2019

ABSTRACTGermfree suckling rats were infected with an SA11 rotavirus strain. Infected pups developed diarrhea associated with histopathological changes. The virus was detected in feces and in the small intestine. Cellular vacuolation was observed in the villi of the jejunum. These results provide a new model for further investigations of group A rotavirus infection.
Use of a Prenylation Inhibitor as a Novel Antiviral Agent
Authors: Glenn Jeffrey S. , Marsters James C. , Greenberg Harry B. 
Published: December 2019

ABSTRACT
No specific therapy exists for hepatitis delta virus (HDV), which can cause severe liver disease. Molecular genetic studies have implicated the prenylation site of large delta antigen as a critical determinant of HDV particle assembly. We have established a cell culture model which produces HDV-like particles, and we show that delta antigen prenylation can be pharmacologically inhibited by the prenylation inhibitor BZA-5B. Furthermore, BZA-5B specifically abolishes particle production in a dose-dependent manner. These results demonstrate that the use of such a prenylation inhibitor-based antiviral therapy may be feasible and identify a novel class of potential antiviral agents.
A Map of Interactions between the Proteins of a Retrotransposon
Authors: Steele Scott J. S. , Levin Henry L. 
Published: December 2019

ABSTRACTThe yeast two-hybrid system and in vitro binding assays were used to characterize 54 potential interactions between the proteins of Tf1, an LTR-retrotransposon found inSchizosaccharomyces pombe. The Tf1 integrase (IN) protein was found to interact strongly with itself and not with other control proteins. In addition, the IN core domain interacted strongly with itself and full-length IN. Interestingly, the two-hybrid analysis detected an interaction between the RNase H domain of reverse transcriptase and IN. The biological implications of these interactions are discussed.
Metabolic Labeling of Woodchuck Hepatitis B Virus X Protein in Naturally Infected Hepatocytes Reveals a Bimodal Half-Life and Association with the Nuclear Framework
Authors: Dandri Maura , Petersen Joerg , Stockert Richard J. , Harris Thomas M. , Rogler Charles E. 
Published: December 2019

ABSTRACT
In order to identify potential sites of hepadnavirus X protein action, we have investigated the subcellular distribution and the stability of woodchuck hepatitis virus (WHV) X protein (WHx) in primary hepatocytes isolated from woodchucks with persistent WHV infection. In vivo cell labeling and cell fractionation studies showed that the majority of WHx is a soluble cytoplasmic protein while a minor part of newly synthesized WHx is associated with a nuclear framework fraction (20%) and with cytoskeletal components (5 to 10%). Pulse-chase experiments revealed that cytoplasmic WHx has a short half-life and decays with bimodal kinetics (approximately 20 min and 3 h). The rates of association and turnover of nucleus-associated WHx suggest that compartmentalization may be responsible for the bimodal turnover observed in the cytoplasm.
Viral Cell Entry Induced by Cross-Linked Decay-Accelerating Factor
Authors: Shafren Darren R. 
Published: December 2019

ABSTRACTDecay-accelerating factor (DAF) mediates cellular attachment for many human picornaviruses. In most cases, viral binding to DAF is itself insufficient to permit cell infectivity, with a second, functional internalization receptor being required to facilitate this process. Previously, we postulated that the role of DAF in enterovirus cell infection is as a sequestration receptor, maintaining a reservoir of bound virus in an infectious state, awaiting interaction with functional internalization receptors. Many of these functional receptors possess the capacity to induce relatively rapid changes in capsid conformations, resulting in the formation of altered particles (A-type particles). In this report, we show that antibody-cross-linked DAF, in contrast to endogenous surface-expressed forms, can act as a functional virus receptor to mediate coxsackie A21 virus (CAV21) lytic cell infection. In contrast to the situation with ICAM-1-mediated CAV21 infection, in which high levels of A-type particles are formed, cross-linked DAF-induced CAV21 replication occurs in the absence of detectable A-particle formation.
Nineteen Baculovirus Open Reading Frames, Including LEF-12, Support Late Gene Expression
Authors: Rapp Jeffrey C. , Wilson Joyce A. , Miller Lois K. 
Published: December 2019

ABSTRACT

A set of 18 plasmid subclones of the
Autographa californica
nuclear polyhedrosis virus genome, each containing an identified late expression factor gene (
lef
), supports expression from a late viral promoter in transient expression assays in the SF-21 cell line derived from
Spodoptera frugiperda
. We have constructed a further set of plasmids in which each
lef
open reading frame (ORF) is controlled by the
Drosophila melanogaster
heat shock protein 70 (hsp70) promoter and epitope tagged. Failure of this set of plasmids to support transient late gene expression, and the inability of the
p47
ORF to replace the
p47
-containing plasmid supplied in the
lef
plasmid library, led to the identification of a 19th late expression factor gene (
lef-12
) located adjacent to the
p47
gene. The sequence of
lef-12
is predicted to encode a protein of 21 kDa with no homology to any previously identified protein. The set of 19 hsp70-controlled
lef
ORFs (HSEpiHis
lef
library) supports transient expression from a late viral promoter.
lef-12
did not affect expression from an early baculovirus promoter. In TN-368 cells, which are also permissive for virus replication,
lef-12
provided a stimulatory effect but did not appear to be essential.
An Endogenous Inhibitor of Human Immunodeficiency Virus in Human Lymphocytes Is Overcome by the Viral Vif Protein
Authors: Madani Navid , Kabat David 
Published: December 2019

ABSTRACT

The
vif
gene of human immunodeficiency virus type 1 (HIV-1) encodes a basic
M
r
23,000 protein that is necessary for production of infectious virions by nonpermissive cells (human lymphocytes and macrophages) but not by permissive cells such as HeLa-CD4. It had been proposed that permissive cells may contain an unidentified factor that functions like the viral Vif protein. To test this hypothesis, we produced pseudotyped wild-type and
vif
-deleted HIV
gpt
virions (which contain the HIV-1 genome with the bacterial mycophenolic acid resistance gene
gpt
in place of the viral
env
gene) in permissive cells, and we used them to generate nonpermissive H9 leukemic T cells that express these proviruses. We then fused these H9 cells with permissive HeLa cells that express the HIV-1 envelope glycoprotein gp120-gp41, and we asked whether the heterokaryons would release infectious HIV
gpt
virions. The results clearly showed that the
vif
-deleted virions released by the heterokaryons were noninfectious whereas the wild-type virions were highly infectious. This strongly suggests that nonpermissive cells, the natural targets of HIV-1, contain a potent endogenous inhibitor of HIV-1 replication that is overcome by Vif.
Mta Has Properties of an RNA Export Protein and Increases Cytoplasmic Accumulation of Epstein-Barr Virus Replication Gene mRNA
Authors: Semmes O. John , Chen Lin , Sarisky Robert T. , Gao Zhigang , Zhong Ling , Hayward S. Diane 
Published: December 2019

ABSTRACT

The Epstein-Barr virus (EBV) Zta and Mta regulatory proteins were previously found to be required for efficient replication of oriLyt in cotransfection-replication assays, but the contribution of Mta to the replication process was unknown. We now demonstrate that Mta regulates replication gene expression. Using the polymerase processivity factor BMRF1 as an example, we found that in transfected cells, total BMRF1 mRNA levels were unaffected by Mta but that the amounts of cytoplasmic BMRF1 RNA and protein were greatly increased in the presence of Mta. Mta also increased cytoplasmic accumulation of the BALF2, BALF5, BSLF1, and BBLF4 replication gene mRNAs but did not affect cytoplasmic levels of BBLF2/3 mRNA. Thus, five of the six core replication genes require Mta for efficient accumulation of cytoplasmic RNA. The contribution of Mta to posttranscriptional RNA processing was examined. Examination of Mta localization in transfected cells by indirect immunofluorescence revealed that Mta colocalized with the splicing factor SC35. We also found that Mta has RNA binding activity. Glutathione
S
-transferase–Mta bound to BMRF1 and BMLF1 transcripts but not to a control cellular gene RNA. Mta contains a consensus leucine-rich nuclear export signal. Such signal sequences are characteristic of proteins that undergo nuclear export. Examination of Mta localization in a heterokaryon assay provided evidence that Mta shuttles between the nucleus and the cytoplasm. Our experiments indicate that Mta functions in RNA processing and transport and mediates cytoplasmic accumulation of a number of EBV early mRNAs.
Mutational Analysis of Residues in the Coiled-Coil Domain of Human Immunodeficiency Virus Type 1 Transmembrane Protein gp41
Authors: Weng Yongkai , Weiss Carol D. 
Published: December 2019

ABSTRACT
The envelope glycoprotein (Env) of human immunodeficiency virus mediates virus entry into cells by undergoing conformational changes that lead to fusion between viral and cellular membranes. A six-helix bundle in gp41, consisting of an interior trimeric coiled-coil core with three exterior helices packed in the grooves (core structure), has been proposed to be part of a fusion-active structure of Env (D. C. Chan, D. Fass, J. M. Berger, and P. S. Kim, Cell 89:263–273, 1997; W. Weissenhorn, A. Dessen, S. C. Harrison, J. J. Skehel, and D. C. Wiley, Nature 387:426–430, 1997; and K. Tan, J. Liu, J. Wang, S. Shen, and M. Lu, Proc. Natl. Acad. Sci. USA 94:12303, 1997). We analyzed the effects of amino acid substitutions of arginine or glutamic acid in residues in the coiled-coil (heptad repeat) domain that line the interface between the helices in the gp41 core structure. We found that mutations of leucine to arginine or glutamic acid in position 556 and of alanine to arginine in position 558 resulted in undetectable levels of Env expression. Seven other mutations in six positions completely abolished fusion activity despite incorporation of the mutant Env into virions and normal gp160 processing. Single-residue substitutions of glutamic acid at position 570 or 577 resulted in the only viable mutants among the 16 mutants studied, although both viable mutants exhibited impaired fusion activity compared to that of the wild type. The glutamic acid 577 mutant was more sensitive than the wild type to inhibition by a gp41 coiled-coil peptide (DP-107) but not to that by another peptide corresponding to the C helix in the gp41 core structure (DP-178). These results provide insight into the gp41 fusion mechanism and suggest that the DP-107 peptide may inhibit fusion by binding to the homologous region in gp41, probably by forming a peptide-gp41 coiled-coil structure.
An Adenovirus Vector with Genetically Modified Fibers Demonstrates Expanded Tropism via Utilization of a Coxsackievirus and Adenovirus Receptor-Independent Cell Entry Mechanism
Authors: Dmitriev Igor , Krasnykh Victor , Miller C. Ryan , Wang Minghui , Kashentseva Elena , Mikheeva Galina , Belousova Natalya , Curiel David T. 
Published: December 2019

ABSTRACT
Recombinant adenoviruses (Ad) have become the vector system of choice for a variety of gene therapy applications. However, the utility of Ad vectors is limited due to the low efficiency of Ad-mediated gene transfer to cells expressing marginal levels of the coxsackievirus and adenovirus receptor (CAR). In order to achieve CAR-independent gene transfer by Ad vectors in clinically important contexts, we proposed modification of viral tropism via genetic alterations to the viral fiber protein. We have shown that incorporation of an Arg-Gly-Asp (RGD)-containing peptide in the HI loop of the fiber knob domain results in the ability of the virus to utilize an alternative receptor during the cell entry process. We have also demonstrated that due to its expanded tissue tropism, this novel vector is capable of efficient transduction of primary tumor cells. An increase in gene transfer to ovarian cancer cells of 2 to 3 orders of magnitude was demonstrated by the vector, suggesting that recombinant Ad containing fibers with an incorporated RGD peptide may be of great utility for treatment of neoplasms characterized by deficiency of the primary Ad type 5 receptor.
Two Proline-Rich Nuclear Localization Signals in the Amino- and Carboxyl-Terminal Regions of the Borna Disease Virus Phosphoprotein
Authors: Shoya Yuko , Kobayashi Takeshi , Koda Toshiaki , Ikuta Kazuyoshi , Kakinuma Mitsuaki , Kishi Masahiko 
Published: December 2019

ABSTRACT

Borna disease virus (BDV) uses a unique strategy of replication and transcription which takes place in the nucleus, unlike other known, nonsegmented, negative-stranded RNA viruses of animal origin. In this process, viral constituents necessary for replication must be transported to the nucleus from the cytoplasm. We report here the evidence that BDV P protein, which may play an important role in viral replication and transcription, is transported into the nucleus in the absence of other viral constituents. This transportation is accomplished by its own nuclear localization signals (NLSs), which are present in both N-terminal (
29
PRPRKIPR
36
) and C-terminal (
181
PPRIYPQLPSAPT
193
) regions of the protein. These two NLSs can function independently and both have several Pro residues as key amino acids.
Successful Readministration of Adeno-Associated Virus Vectors to the Mouse Lung Requires Transient Immunosuppression during the Initial Exposure
Authors: Halbert Christine L. , Standaert Thomas A. , Wilson Christopher B. , Miller A. Dusty 
Published: December 2019

ABSTRACTThe airway is an important target for gene transfer to treat cystic fibrosis and other diseases that affect the lung. We previously found that marker gene expression did not persist in the bronchial epithelium following adeno-associated virus (AAV) vector administration to the rabbit lung. In an attempt to promote continued expression, we tested repeat vector administration, but no additional transduction was observed, and the block to transduction correlated with the appearance of neutralizing antibodies to the viral capsid. Here we show that mice exhibit a similar response but that treatment with anti-CD40 ligand antibody (MR1) and a soluble CTLA4-immunoglobulin fusion protein (CTLA4Ig) at the time of primary AAV vector exposure allowed successful repeat transduction and prevented production of neutralizing antibodies. We also tested the possibility that an immune response caused the loss of marker-positive cells in the epithelial population in rabbits by evaluating AAV vector expression in immunocompetent and immunodeficient mice. In contrast to results in rabbits, marker protein expression persisted in the lung in both groups of mice. AAV vector transduction occurred in alveolar cells, airway epithelial cells, and smooth muscle cells, and vector expression persisted for at least 8 months. Although data on persistence of AAV vector expression in the human lung are not available, it is likely that repeat transduction will be necessary either due to loss of expression or to the need for repeat administration to deliver effective amounts of AAV vectors. Results presented here indicate that transient immunosuppression will allow such repeat vector treatment of the lung.