pinecone/core-2020-05-10-deduplication huggingface dataset:
HTLV-1 Cell Lines Differ in Constitutively Activated Signaling Pathways That Can Be Altered by Cytokine Exposure

AbstractExamination of signaling pathways used by HTLV-1-infected rabbit cell lines revealed differences between one, RH/K30, that mediates asymptomatic infection and another, RH/K34, that causes lethal experimental leukemia. Both lines are IL-2 independent; RH/K30 produces IL-4 while RH/K34 produces IL-10. Examination of the Jak/STAT (Janus kinase/signal transducer and activator of transcription) activation of the lines revealed constitutive phosphorylation of Jak1 in both. STAT6 phosphorylation, not previously reported for HTLV-1 cells, was observed in RH/K30; STAT1 and STAT3 were phosphorylated in RH/K34. Treatment with cytokines altered the activation of the STAT proteins: IL-2 induced STAT5 phosphorylation in both lines. Supernatant from RH/K34 or IL-10 induced STAT3 phosphorylation in RH/K30 cells. Supernatant from RH/K30 or IL-4 induced STAT6 phosphorylation in RH/K34 cells, which could be reversed with a Jak kinase inhibitor—AG-490
Functional Expression and Membrane Fusion Tropism of the Envelope Glycoproteins of Hendra Virus

AbstractHendra virus (HeV) is an emerging paramyxovirus first isolated from cases of severe respiratory disease that fatally affected both horses and humans. Understanding the mechanisms of host cell infection and cross-species transmission is an important step in addressing the risk posed by such emerging pathogens. We have initiated studies to characterize the biological properties of the HeV envelope glycoproteins. Recombinant vaccinia viruses encoding the HeV F and G open reading frames were generated and glycoprotein expression was verified by metabolic labeling and detection using specific antisera. Glycoprotein function and cellular tropism were examined with a quantitative assay for HeV-mediated membrane fusion. Fusion specificity was verified through specific inhibition by anti-HeV antiserum and a peptide corresponding to one of the α-helical heptad repeats of F. HeV requires both F and G to mediate fusion. Permissive target cells have been identified, including cell lines derived from cat, bat, horse, human, monkey, mouse, and rabbit. Fusion negative cell types have also been identified. Protease treatments of the target cells abolished fusion activity, suggesting that the virus is employing a cell-surface protein as its receptor
The Avian Reovirus Genome Segment S1 Is a Functionally Tricistronic Gene That Expresses One Structural and Two Nonstructural Proteins in Infected Cells

AbstractThe avian reovirus S1 gene contains three partially overlapping, out-of-phase open reading frames (ORFs) that are highly conserved in all avian reovirus strains examined to date. The three S1 ORFs of the avian reovirus strain S1133 were individually expressed in bacterial cells, and their purified translation products used as antigens to raise specific polyclonal antibodies. With these antibodies we were able to demonstrate that all three S1 ORFs from different avian reovirus strains are translatable in infected cells. Proteins p10 and p17, which are specified by ORF1 and ORF2, respectively, are nonstructural proteins which associate with cell membranes, whereas ORF3 directs the synthesis of protein ςC, a structural oligomeric protein responsible for cell attachment. While intracellular synthesis of protein ςC was demonstrated a long time ago and that of protein p10 was reported recently, this is the first time that expression of the S1 ORF2 has been demonstrated experimentally. Thus, the previously reported coding capacity of the avian reovirus genome is now expanded to 14 proteins, of which ten are structural (λA, λB, λC, μA, μB, μBC, μBN, ςA, ςB, and ςC) and four are nonstructural (μNS, ςNS, p17, and p10). Finally, protein p10, but not p17 or ςC, induces cell–cell fusion when transiently expressed in mammalian cells, supporting a previously published observation that the polypeptide encoded by the S1 ORF1 plays an important role in the syncytial phenotype displayed by avian reoviruses
Isolation and Characterization of Two Viruses with Large Genome Size Infecting Chrysochromulina ericina (Prymnesiophyceae) and Pyramimonas orientalis (Prasinophyceae)

AbstractTwo lytic viruses specific for Chrysochromulina ericina (Prymnesiophyceae) and for Pyramimonas orientalis (Prasinophyceae) were isolated from Norwegian coastal waters in June 1998. The lytic cycle was 14–19 h for both viruses; the burst size was estimated at 1800–4100 viruses per host cell for the Chrysochromulina virus and 800–1000 for the Pyramimonas virus. Thin sections of infected cells show that both viruses replicate in the cytoplasm and that they have a hexagonal cross section, indicating icosahedral symmetry. The Chrysochromulina virus had a particle size of 160 nm and a genome size of 510 kbp; the size of the major polypeptide was 73 kDa. The Pyramimonas virus had a particle size of 220 × 180 nm and a genome size of 560 kbp; the size of the major polypeptide was 44 kDa. The genome sizes of these viruses are among the largest ever reported for viruses and they are larger than the minimum required for cellular life. The Chrysochromulina virus clone CeV-01B and the Pyramimonas virus clone PoV-01B described in this study have several properties in common with other viruses infecting microalgae, suggesting that they belong to the Phycodnaviridae
Characterization of the Developmental Switch Region of Bacteriophage P2 Hy dis

AbstractIn this work, the DNA sequence of the transcriptional switch that affects the development of the P2 Hy dis bacteriophage was determined. The switch contains two face-to-face-located promoters and two repressors, Cox and C. The locations of the Pc and Pe promoters were determined by primer extension analysis. The P2 Hy dis homolog of the P2 multifunctional Cox protein was shown to be able to substitute for P2 Cox in repression of the P2 Pc promoter, excision of the P2 prophage, and activation of the satellite phage P4 PLL promoter. A directly repeated sequence, flanking the −35 region of the Pe promoter, was found to be important for C repressor binding as well as for repression. The P4 E protein was shown to derepress the developmental switch of P2 Hy dis in a plasmid-based derepression assay
Quantitative Evaluation of HIV-1 Coreceptor Use in the GHOST(3) Cell Assay

AbstractThe utility of the GHOST(3) cell assay has been evaluated for testing coreceptor use of primary human immunodeficiency virus type 1 (HIV-1) isolates. GHOST(3) cells were derived from the human osteosarcoma cell line, HOS, and have been engineered to stably express CD4 and one or another of the chemokine receptors CCR3, CCR5, CXCR4, Bonzo, or the orphan receptor BOB. The indicator cell line carries the HIV-2 long terminal repeat-driven green fluorescence protein (GFP) gene, which becomes activated upon infection with HIV or simian immunodeficiency virus. Viral entry is followed by Tat activation of transcription and GFP becomes expressed. Infected cells can be detected 2 or 3 days after infection by simple fluorescence microscopic observation. This simplicity is the main advantage of the GHOST(3) cell system and makes it particularly suitable for screening of a large number of isolates. In addition, the efficiency of coreceptor use can be accurately quantitated with flow cytometric analysis. Here, we evaluated the coreceptor use of 59 primary HIV-1 isolates of different subtypes
Coreceptor Usage of Sequential Isolates from Cynomolgus Monkeys Experimentally Infected with Simian Immunodeficiency Virus (SIVsm)

AbstractSequential isolates from eight cynomolgus monkeys experimentally infected with simian immunodeficiency virus (SIVsm, of sooty mangabey origin) were tested for coreceptor use in the human osteosarcoma indicator cell line, GHOST(3), expressing CD4 and one or another of the chemokine receptors CCR3, CCR5, CXCR4, BOB, or the orphan receptor Bonzo. The indicator cell line carries the human immunodeficiency virus type 2 long terminal repeat-driven green fluorescence protein gene that becomes activated upon infection with HIV or SIV and fluorescence can be quantitated by flow cytometric analysis. The methodological details are described in the accompanying paper (Vödrös et al., 2001, Virology 290, in press). All SIVsm inoculum viruses and reisolates used CCR5 with a high level of efficiency. CCR5 use was stable over time. BOB and Bonzo use was less efficient than CCR5 use and, in particular, late isolates obtained at the time of immunodeficiency varied greatly in their coreceptor use and often could not establish a productive infection in BOB- or Bonzo-expressing cells. Unexpectedly, early reisolates obtained 12 days postinfection could infect the entire GHOST(3) panel including the parental cells. In one case this was due to use of CXCR4, either transfected or endogenously expressed on the GHOST(3) cells. Our results demonstrate the complex coreceptor use of SIVsm isolates. Moreover, they focus attention on the initial phase of virus replication when the availability of target cells may govern the replication pattern of the virus
The Baculovirus Transcriptional Transactivator ie0 Produces Multiple Products by Internal Initiation of Translation

AbstractIe0 is the only gene of the baculovirus Orgyia pseudotsugata multiple nucleopolyhedrovirus (OpMNPV) that is known to be spliced. In this study, cDNAs of ie0 were isolated, cloned, and sequenced. It was observed that IE0 contains 35 amino acids (aa) added to the N-terminus of IE1. In addition, it was found that the leader sequence of ie0 contains a 4-aa minicistron. To functionally characterize IE0, ie0 cDNAs were expressed under control of either the ie1 or the ie0 promoter. Unexpectedly, examination of ie0 translation products revealed that the predominant product from ie0 mRNAs was not IE0, but IE1. Mutation analysis showed that IE1 translation was preferentially initiated from either of two AUGs found in the first 15 nucleotides (nt) of the ie1 ORF that are internal to the ie0 ORF. It is unknown whether the internal translation initiation occurs via a leaky scanning mechanism or by an internal ribosomal entry site. Transactivation analysis with constructs that had point mutations in the ie1 AUGs and were translated only as IE0 revealed that OpMNPV IE0 is a 14- to 15-fold stronger transactivator than IE1. IE0 was also shown to be autoregulatory and to transactivate early genes in an enhancer-independent or -dependent manner. These results suggest that differential expression of baculovirus early genes can be obtained by coexpression of IE0 and IE1 in infected cells, which may permit subtle regulation of specific sets of viral genes
Association of Vpu-Binding Protein with Microtubules and Vpu-Dependent Redistribution of HIV-1 Gag Protein

AbstractThe efficient exit of HIV-1 particles from cells requires the action of the viral encoded protein Vpu. Vpu-binding protein (Ubp) is a cellular protein that interacts with both Vpu and the major structural component of the viral capsid (Gag) and appears to affect the efficiency of particle exit. Elucidation of the function of Ubp and characterization of the spatial distribution of Ubp may provide information pertinent to understanding the role of Ubp in virus replication. To investigate the subcellular location of Ubp, and to see whether Vpu affects the intracellular distribution of Gag, we carried out immunofluorescence localization in conjunction with confocal microscopy. Based on this analysis Ubp is present in both the nucleus and the cytoplasm. In the cytoplasm, Ubp appeared to be associated with microtubules as evidenced by cofluorescence with tubulin in the absence and in the presence of colchicine. However, cytoskeletal isolation and detergent extraction of cells resulted in association of Ubp with the soluble fractions, indicating that Ubp is not in tight association with microtubules. Moreover, flotation gradient analysis demonstrated that Ubp is cytoplasmic and not stably associated with the plasma membrane. Interestingly, expression of Vpu in cells resulted in redistribution of both Ubp and Gag to a location near the periphery of the cell. The effect of Vpu on both Ubp and Gag protein has implications for Vpu-mediated particle exit from cells
Inactivation of Expression of Gene 4 of Mouse Hepatitis Virus Strain JHM Does Not Affect Virulence in the Murine CNS

AbstractThe protein encoded by ORF 4 of mouse hepatitis virus (MHV) is not required for growth of some strains in tissue culture cells, but its role in pathogenesis in the murine host has not been defined previously in a controlled manner. MHV strain JHM causes acute and chronic neurological diseases in susceptible strains of rodents. To genetically manipulate the structural proteins of this and other strains of MHV, we have generalized an interspecies-targeted RNA recombination selection that was originally developed for the A59 strain of MHV. Using this approach, a recombinant MHV–JHM was constructed in which gene 4 was genetically inactivated. Virus lacking gene 4 expression replicated in tissue culture cells with similar kinetics to recombinant virus in which gene 4 expression was not disrupted. Both types of viruses exhibited similar virulence when analyzed in a murine model of encephalitis. These results establish a targeted recombination system for inserting mutations into MHV–JHM. Furthermore, the protein encoded by ORF 4 is not essential for growth in tissue culture cells or in the CNS of the infected host
Phenotypic and Molecular Analyses of Yellow Fever 17DD Vaccine Viruses Associated with Serious Adverse Events in Brazil

AbstractThe yellow fever (YF) 17D virus is one of the most successful vaccines developed to date. Its use has been estimated to be over 400 million doses with an excellent record of safety. In the past 3 years, yellow fever vaccination was intensified in Brazil in response to higher risk of urban outbreaks of the disease. Two fatal adverse events temporally associated with YF vaccination were reported. Both cases had features similar to yellow fever disease, including hepatitis and multiorgan failure. Two different lots of YF 17DD virus vaccine were administered to the affected patients and also to hundreds of thousands of other individuals without any other reported serious adverse events. The lots were prepared from the secondary seed, which has been in continuous use since 1984. Nucleotide sequencing revealed minor variations at some nucleotide positions between the secondary seed lot virus and the virus isolates from patients; these differences were not consistent across the isolates, represented differences in the relative amount of each nucleotide in a heterogeneous position, and did not result in amino acid substitutions. Inoculation of rhesus monkeys with the viruses isolated from the two patients by the intracerebral (ic) or intrahepatic (ih) route caused minimal viremia and no clinical signs of infection or alterations in laboratory markers. Central nervous system histological scores of rhesus monkeys inoculated ic were within the expected range, and there were no histopathological lesions in animals inoculated ih. Altogether, these results demonstrated the genetic stability and attenuated phenotype of the viruses that caused fatal illness in the two patients. Therefore, the fatal adverse events experienced by the vaccinees are related to individual, genetically determined host factors that regulate cellular susceptibility to yellow fever virus. Such increased susceptibility, resulting in clinically overt disease expression, appears to be extremely rare
Transcription and Replication of a Natural Hepatitis B Virus Nucleocapsid Promoter Variant Is Regulated in Vivo by Peroxisome Proliferators

AbstractA hepatitis B virus (HBV) transgenic mouse containing a naturally occurring mutation in the nucleocapsid promoter (A1764T plus G1766A) that inhibits the retinoid X receptor α (RXRα) plus peroxisome proliferator-activated receptor α (PPARα) heterodimer from binding to the proximal nuclear hormone receptor recognition sequence has been generated. Viral transcription and replication occur in the liver and kidney. The nucleocapsid promoter mutation does not prevent peroxisome proliferators from increasing viral transcription and replication in the liver of these variant HBV transgenic mice. This suggests that peroxisome proliferators may enhance viral transcription directly in a PPARα-dependent manner through the nuclear hormone receptor recognition site in the enhancer 1 region of the HBV genome. Hepatocyte nuclear factor 4 (HNF4) binding to the proximal nuclear hormone receptor recognition sequence in the nucleocapsid promoter appears to limit RNA synthesis from the precore transcription initiation site. Consequently, the variant HBV transgenic mice transcribe very little precore RNA and secrete extremely low levels of hepatitis B e antigen (HBeAg) compared with the wild-type HBV transgenic mice. This is consistent with the suggestion that viruses expressing HBeAg are preferentially eliminated in infected individuals when they seroconvert from HBeAg positive to anti-HBe antibody-positive status, leaving escape HBV variants that have reduced HBeAg expression
A Family of Closely Related Bovine Viral Diarrhea Virus Recombinants Identified in an Animal Suffering from Mucosal Disease: New Insights into the Development of a Lethal Disease in Cattle

AbstractInduction of lethal mucosal disease (MD) in cattle is linked to the generation of cytopathogenic (cp) bovine viral diarrhea virus (BVDV) in animals persistently infected with a noncytopathogenic BVDV. In most cases the cp variants are generated by recombination with cellular or viral sequences. BVDV was obtained from the serum of an MD animal and propagated in tissue culture without plaque purification. Analysis of cDNA clones established from RNA of these cells showed that apparently a variety of different viral RNAs were present. Seven of the cDNA clones contained a cellular sequence coding for light chain 3 (LC3) of microtubule-associated proteins 1A and 1B. This insertion had already been found in the cp virus JaCP obtained from the same animal and isolated by plaque purification. Analysis of further plaque-purified cp viruses showed that the diseased animal contained a family of closely related cp BVDV recombinants. A set of viruses with different duplications of viral sequences in their genomes and a variety of defective viral RNAs with deletions were found that all contained the LC3* insertion. For all the recombinants the 3′ recombination sites and, in all but one case, also the 5′ recombination sites between cellular and viral sequence were identical. Variation between the individual deduced genome structures resulted from different duplications or deletions of viral sequences located upstream of the cellular insertion. These results suggest that within the animal a primary recombinant with a genome containing the LC3* insertion was generated. In a trimming process a set of secondary virus recombinants was generated from this hypothetical primary recombined RNA. These secondary recombinants display genome structures that represent variations of the basic scheme already present in the primary recombinant. Apparently this trimming process that finally led to an outbreak of MD lasted a long time since recombined RNA with the basic genome structure of the cp viruses could be demonstrated in samples already taken a long time before outbreak of the disease
The Hantaan Virus Glycoprotein Precursor Is Cleaved at the Conserved Pentapeptide WAASA

AbstractThe medium segment of the tripartite negative-stranded RNA genome of hantaviruses encodes for the predicted glycoprotein precursor GPC. We have demonstrated here the expression of the glycoprotein precursor of Hantaan virus following transfection of mammalian cells. The cleavage of the precursor into the glycoproteins G1 and G2 followed the rules for signal peptides and seemed to occur directly at the pentapeptide motif “WAASA.” Our data indicate that the signal peptidase complex is responsible for the proteolytic processing of the precursor GPC of Hantaan virus. The comparison of this region of the glycoprotein precursor, including the absolutely conserved WAASA motif, suggests a similar cleavage event for all hantavirus glycoproteins
Cloning, Expression, and Crystallization of the Fusion Protein of Newcastle Disease Virus

AbstractWe have recently reported the X-ray crystal structure of a fragment of the fusion protein (F) of Newcastle disease virus (NDV). This work describes the methodology involved in the production and crystallization of that protein in recombinant form. The full-length cDNA of NDV-F was cloned and the ectodomain expressed in both CHO-K1 and Lec-3.2.8.1 cells. The recombinant protein, secreted as a single-chain polypeptide F0′, was purified using a c-myc antibody affinity column followed by gel filtration chromatography. Electron microscopic imaging showed the F0′ product to consist of unaggregated club-shaped particles. Trypsin treatment of F0′ could be used to produce disulfide-linked F2 and F1′ chains. However, imaging revealed extensive rosette-like aggregation of the trypsin-treated material, indicative of a conformational change. Only the non-trypsin-treated product was thus suitable for crystallization and two crystal forms were obtained, diffracting to ca. 3.5 and 4.0 Å, respectively. Both crystal forms were used in the structure determination
Varicella-Zoster Virus Gene Expression in Latently Infected Rat Dorsal Root Ganglia

AbstractLatent infection of human ganglia with Varicella-Zoster virus (VZV) is characterized by a highly restricted pattern of viral gene expression. To enhance understanding of this process we used in situ hybridization (ISH) in a rat model of VZV latency to examine expression of RNA corresponding to eight different VZV genes in rat dorsal root ganglia (DRG) at various times after footpad inoculation with wild-type VZV. PCR in situ amplification was also used to determine the cell specificity of latent VZV DNA. It was found that the pattern of viral gene expression at 1 week after infection was different from that observed at the later times of 1 and 18 months after infection. Whereas multiple genes were expressed at 1 week after infection, gene expression was restricted at the later time points when latency had been established. At the later time points after infection the RNA transcripts expressed most frequently were those for VZV genes 21, 62, and 63. Gene 63 was expressed more than any other gene studied. While VZV DNA was detected almost exclusively in 5–10% of neurons, VZV RNA was detected in both neurons and nonneuronal cells at an approximate ratio of 3:1. A newly described monoclonal antibody to VZV gene 63-encoded protein was used to detect this protein in neuronal nuclei and cytoplasm in almost half of the DRG studied. These results demonstrate that (1) this rat model of latency has close similarities in terms of viral gene expression to human VZV latency which makes it a useful tool for studying this process and its experimental modulation and (2) expression of VZV gene 63 appears to be the single most consistent feature of VZV latency