pinecone/core-2020-05-10-deduplication huggingface dataset:
Disruption of NF-κB Signaling and Chemokine Gene Activation by Retroviral Mediated Expression of IKKγ/NEMO Mutants

AbstractPhosphorylation of IκBs—the cytoplasmic inhibitors of the NF-κB transcription factors—is the key event which triggers activation of the NF-κB cascade. Signal-mediated phosphorylation of IκBα is mediated by a multiprotein complex, the IκB kinase (IKK) complex, which is composed of at least three identified subunits. Two of these polypeptides, IKKα and IKKβ, also known as IKK1 and IKK2, are the catalytic subunits of the kinase complex and phosphorylate IκBα and IκBβ. The third component, NEMO/IKKγ, does not exhibit kinase activity, but rather constitutes a regulatory subunit. In the present study, C-terminal truncated forms of IKKγ—ΔC-IKKγ 306 and ΔC-IKKγ 261—were stably expressed in the myeloid cell line U937 by retroviral-mediated gene transfer. Overexpression of ΔC-IKKγ resulted in a reduction in IKK kinase activity in vitro, a subsequent decrease in NF-κB DNA binding activity, and inhibition of chemokine gene induction in response to TNFα stimulation or paramyxovirus infection. This study demonstrates the efficacy of ΔC-IKKγ as a repressor of IKK signaling and NF-κB activation and suggests a potential gene therapy approach to limit chronic inflammation due to chemokine hyperactivation
Caprine Arthritis Encephalitis Virus: Evidence for a B/D-Type Assembly Pathway in a C-Type Lentivirus Replication

AbstractLentiviruses, among which is caprine arthritis encephalitis virus (CAEV), are known to concomitantly assemble and bud at the plasma membrane of infected cells, in a C-type defined pathway. Electron microscopy analysis of CAEV-infected cells demonstrated viral particles budding at the plasma membrane and into intracellular membrane-surrounded vesicles. Furthermore, nonenveloped immature virus-like particles, resembling intracytoplasmic type-A particles (ICAPs), accumulated within the cytoplasm of those cells. Fractionation on sucrose density gradients of cytoplasmic lysates from CAEV-infected cells revealed that enveloped immature or mature viral particles had a density of 1.16–1.17 g/ml, whereas ICAPs sedimented at a density of 1.2–1.27 g/ml. Endogenous reverse transcriptase activity was only associated with the 1.16–1.17 g/ml density particles despite the presence of viral RNA in both populations. The intracellular enveloped particles were found to be infectious. The CAEV Gag precursor by itself was shown to direct assembly, budding, and release of immature virus-like particles when expressed in goat primary synovial membrane cells using the same pathways of assembly and budding as observed in CAEV-infected cells. These data suggest that CAEV assembly, driven by the Gag precursor, could unusually proceed via two simultaneous pathways characteristic of type-C and type-B/D retroviruses
Evidence for the Presence of Two Novel Pestivirus Species

AbstractThe genus Pestivirus of the family Flaviviridae comprises four species, namely Bovine viral diarrhea virus-1 (BVDV-1), BVDV-2, Border disease virus (BDV), and Classical swine fever virus (CSFV). Comparative analyses of partial sequences have suggested that pestivirus isolates from giraffe (Giraffe-1) and reindeer (Reindeer-1) are distinct from the established species (Becher et al., Virology 262, 64–71, 1999). In this study, we report the complete genomic sequences of pestivirus strains Giraffe-1 and Reindeer-1. Comparative sequence analyses revealed considerable differences among Giraffe-1, Reindeer-1, and the currently recognized pestivirus species. Phylogenetic analysis of the complete coding sequences of these two strains, along with 13 other sequences representing the four established species, indicated that CSFV, BDV, and Reindeer-1 have bifurcated from one common branch and BVDV-1 and BVDV-2 from another. In the former branch BDV and the pestivirus from reindeer are more similar to each other than to CSFV. The giraffe pestivirus is equally distinct from both major branches. In addition, the antigenic relatedness of pestivirus isolates covering the observed major genetic groups was studied by cross-neutralization assays. A clustering procedure on the basis of antigenic differences indicated the presence of six major groups corresponding to the genetically defined groups. Taken together, the results of our analyses addressing both nucleotide sequence relatedness and serological relatedness argue for the inclusion of Giraffe-1 and Reindeer-1 as the first members of two separate novel species within the genus Pestivirus
The White Spot Syndrome Virus DNA Genome Sequence

AbstractWhite spot syndrome virus (WSSV) is at present a major scourge to worldwide shrimp cultivation. We have determined the entire sequence of the double-stranded, circular DNA genome of WSSV, which contains 292,967 nucleotides encompassing 184 major open reading frames (ORFs). Only 6% of the WSSV ORFs have putative homologues in databases, mainly representing genes encoding enzymes for nucleotide metabolism, DNA replication, and protein modification. The remaining ORFs are mostly unassigned, except for five, which encode structural virion proteins. Unique features of WSSV are the presence of a very long ORF of 18,234 nucleotides, with unknown function, a collagen-like ORF, and nine regions, dispersed along the genome, each containing a variable number of 250-bp tandem repeats. The collective information on WSSV and the phylogenetic analysis on the viral DNA polymerase suggest that WSSV differs profoundly from all presently known viruses and that it is a representative of a new virus family
Subcellular Targeting of the Coat Protein of African Cassava Mosaic Geminivirus

AbstractThe coat protein (CP) of geminiviruses is involved in a number of processes during the life cycle of the virus. The predominant function is encapsidation of single-stranded DNA and formation of the virus particle to protect viral DNA during transmission. The CP of monopartite geminiviruses is absolutely essential for virus movement, whereas CP mutants of bipartite geminiviruses are able to infect some host plants systemically, indicating an involvement of the CP in host specificity. During the life cycle of geminiviruses, the viral DNA enters the nucleus of the infected cell where virus replication, transcription, and encapsidation occur. For systemic infection, the virus moves cell-to-cell from the site of inoculation to vascular tissue and via phloem to other plant tissues. To move, viral DNA has to shuttle in and out of the nucleus and through plasmodesmata. Parts of the bipartite African cassava mosaic virus (ACMV) CP were fused with green fluorescent protein (GFP) or β-glucuronidase (GUS). CP domains were identified that mediate both nuclear import and export, as well as targeting of CP-fusion proteins to the cell periphery. These results indicate that domains of the CP facilitate several aspects of geminivirus movement, including nuclear import and export and transport of the viral genome to the cell periphery
An α-Helical Domain within the Carboxyl Terminus of Herpes Simplex Virus Type 1 (HSV-1) glycoprotein B (gB) is Associated with Cell Fusion and Resistance to Heparin Inhibition of Cell Fusion

AbstractPrevious studies from our laboratory indicated that a 28-amino-acid carboxyl-terminal truncation of gB caused extensive virus-induced cell fusion (Baghian et al., 1993, J Virol 67, 2396–2401). We tested the ability of additional truncations and mutations within gB to cause cell fusion in the recently established virus-free cell fusion assay (Turner et al., 1998, J. Virol. 72, 873–875). Deletion of the carboxyl-terminal 28 amino acids of gB (gBΔ28), which removed part of the predicted α-helical structure H17b, caused extensive cell fusion. A gB truncation specified by gBΔ36, which removed the entire H17b domain, caused as much cell fusion as the gBΔ28 truncation. Similarly, gB(A874P) containing a substitution of an Ala with Pro within H17b caused cell fusion. Heparin, a gB-specific inhibitor of virus-induced cell fusion, inhibited both wild-type gB and gB(syn3)-mediated cell fusion. In contrast, fusion of cells transfected with gB(Δ28), gB(Δ36), or gB(A874P) was resistant to heparin inhibition of cell fusion. We concluded the following: (1) The predicted α-helical structure of H17b within the carboxyl terminus of gB is involved in both virus-induced and virus-free cell fusion. (2) Heparin is a specific inhibitor of gB-mediated fusion in both systems. (3) Resistance to heparin inhibition of gB-mediated cell fusion is associated with the predicted α-helical structure H17b within the carboxyl terminus of gB
Human-Herpesvirus-8-Encoded K8 Protein Colocalizes with the Promyelocytic Leukemia Protein (PML) Bodies and Recruits p53 to the PML Bodies

AbstractPromyelocytic leukemia protein (PML) bodies are nuclear sites for both input viral genome deposition and immediate-early (IE) gene transcription during infection with certain human DNA viruses, such as human cytomegalovirus (HCMV), herpes simplex virus type 1, and adenovirus. In this study, we showed that the K8 (K-bZIP) protein, an early protein encoded by the human herpesvirus 8 (HHV-8), colocalized with the PML bodies in HHV-8-infected primary effusion lymphoma cells. Cotransfection of two plasmids expressing the K8 protein and green-fluorescence protein (GFP)-PML fusion protein into 293T cells revealed that the K8 protein colocalized with PML in cells with high PML expression. Overexpression of the K8 protein in Chinese hamster ovary (CHO) cells with stable GFP-PML expression did not induce the dispersion of the PML bodies, unlike the IE1 protein of HCMV. Transfection of a truncated K8 gene revealed that the leucine zipper domain of the K8 protein was required for the colocalization with PML. We also demonstrated that the K8 protein bound to p53 in vivo and in vitro, and that high expression of the K8 protein caused the accumulation of p53 to the PML bodies in CHO cells, suggesting that the K8 protein functions in the recruitment of p53 to the PML bodies. These data suggest that the K8 protein may be associated with the functional modulation of p53 in the nucleus during the lytic phase of HHV-8
Effects of Alanine Cluster Mutations in the D12 Subunit of Vaccinia Virus mRNA (Guanine-N7) Methyltransferase

AbstractThe (guanine-N7)-methyltransferase domain of the vaccinia virus mRNA capping enzyme is a heterodimer composed of a catalytic subunit D1(498–844) bound to a stimulatory subunit D12. To identify structural elements of the 287-amino-acid D12 subunit that participate in binding and activation of the catalytic subunit, we introduced 12 double-alanine mutations at vicinal residues that are conserved in the D12 homologs of other vertebrate poxviruses. His-tagged D12 mutants were coexpressed in bacteria with the D1(498–544) subunit, and the recombinant D1(498–844)/His-D12 heterodimers were purified. Eight of the mutants (K111A-R112A, N120A-N121A, N126A-N127A, F141A-R142A, K223A-D224A, H260A-S261A, E275A-N276A, and R280A-R281A) had no significant effect on methyltransferase activity. Three of the mutants (L61A-K62A, F176A-K177A, and F245A-L246A) displayed an intermediate level of cap methylation (35–50% of wild-type activity). Only one mutation, N42A-Y43A, elicited a significant loss of the methyltransferase activation function (<20% of the wild-type activity). Nine of the D12-Ala/Ala proteins were produced individually in bacteria and tested for reconstitution of methyltransferase activity in vitro by mixing with the catalytic subunit. K111A-R112A, N120A-N121A, F176A-K177A, F245A-L246A, and L61A-K62A displayed diminished affinity for the D1 catalytic subunit. N42A-Y43A was uniquely defective in its ability to activate cap methylation by the catalytic subunit. Our results suggest that the methyltransferase activation function of D12, though clearly dependent on the physical interaction with D1, also requires constituents of D12 that are engaged specifically in catalysis
HIV-1 Vpr Induces Cell Cycle G2 Arrest in Fission Yeast (Schizosaccharomyces pombe) through a Pathway Involving Regulatory and Catalytic Subunits of PP2A and Acting on Both Wee1 and Cdc25

AbstractViral protein R (Vpr) of human immunodeficiency virus type 1 induces G2 arrest in cells from distantly related eukaryotes including human and fission yeast through inhibitory phosphorylation of tyrosine 15 (Tyr15) on Cdc2. Since the DNA damage and DNA replication checkpoints also induce G2 arrest through phosphorylation of Tyr15, it seemed possible that Vpr induces G2 arrest through the checkpoint pathways. However, Vpr does not use either the early or the late checkpoint genes that are required for G2 arrest in response to DNA damage or inhibition of DNA synthesis indicating that Vpr induces G2 arrest by an alternative pathway. It was found that protein phosphatase 2A (PP2A) plays an important role in the induction of G2 arrest by Vpr since mutations in genes coding for a regulatory or catalytic subunit of PP2A reduce Vpr-induced G2 arrest. Vpr was also found to upregulate PP2A, supporting a model in which Vpr activates the PP2A holoenzyme to induce G2 arrest. PP2A is known to interact genetically in fission yeast with the Wee1 kinase and Cdc25 phosphatase that act on Tyr15 of Cdc2. Both Wee1 and Cdc25 play a role in Vpr-induced G2 arrest since a wee1 deletion reduces Vpr-induced G2 arrest and a direct in vivo assay shows that Vpr inhibits Cdc25. Additional support for both Wee1 and Cdc25 playing a role in Vpr-induced G2 arrest comes from a genetic screen, which identified genes whose overexpression affects Vpr-induced G2 arrest. For this genetic screen, a strain was constructed in which cell killing by Vpr was nearly eliminated while the effect of Vpr on the cell cycle was clearly indicated by an increase in cell length. Overexpression of the wos2 gene, an inhibitor of Wee1, suppresses Vpr-induced G2 arrest while overexpression of rad25, an inhibitor of Cdc25, enhances Vpr-induced G2 arrest. These two genes may be part of the uncharacterized pathway for Vpr-induced G2 arrest in which Vpr upregulates PP2A to activate Wee1 and inhibit Cdc25
Reverse Genetics of Influenza Virus

AbstractReverse genetics of negative-sense RNA viruses, which enables one to generate virus entirely from cloned cDNA, has progressed rapidly over the past decade. However, despite the relative ease with which nonsegmented negative-sense RNA viruses can now be produced from plasmids, the ability to generate viruses with segmented genomes has lagged considerably, largely because of the inherent technical difficulties in providing all viral RNAs and proteins from cloned cDNA. A breakthrough in reverse genetics technology in the influenza virus field came in 1999, when we (Neumann et al., 1999, Proc. Natl. Acad. Sci. USA 96, 9345–9350) and others (Fodor et al., 1999, J. Virol. 73, 9679–9682) exploited a new approach to viral RNA production. In this review, we discuss the background for this advance, the systems that are now available for the generation of influenza viruses, and the implications of these developments for the future of virus research
Correlation of Genetic Variability with Safety of Mumps Vaccine Urabe AM9 Strain

AbstractThe Urabe AM9 strain of mumps vaccine live is known for its genetic instability and some vaccines derived from this strain were withdrawn from the market due to an excessive number of vaccine-associated parotitis and meningitis cases. To identify the molecular basis of this instability, we determined complete nucleotide sequences of several stocks of the Urabe strain used for vaccine production by different manufacturers and of two clinical isolates from cases of vaccine-associated meningitis. In contrast to previously published studies relating the Lys335 → Glu mutation in the viral HN gene with neurovirulence of mumps virus, we could not confirm any association of this mutation with the safety of mumps vaccine. Each of the three vaccine stocks studied had its own characteristic profile of mutations that was identified by cDNA sequencing and quantitated by mutant analysis by PCR and restriction enzyme cleavage. Determination of the mutational profile of mumps vaccine lots could allow vaccine manufacturers to characterize seed viruses and monitor the consistency of vaccine production to prevent emergence of virulent revertants
Vaccine Potential of Ebola Virus VP24, VP30, VP35, and VP40 Proteins

AbstractPrevious vaccine efforts with Ebola virus Zaire (EBOV-Z) emphasized the potential protective efficacies of immune responses to the surface glycoprotein and the nucleoprotein. To determine whether the VP24, VP30, VP35, and VP40 proteins are also capable of eliciting protective immune responses, these genes were expressed from alphavirus replicons and used to vaccinate BALB/c and C57BL/6 mice. Although all of the VP proteins were capable of inducing protective immune responses, no single VP protein protected both strains of mice tested. VP24, VP30, and VP40 induced protective immune responses in BALB/c mice, whereas C57BL/6 mice survived challenge only after vaccination with VP35. Passive transfer of immune sera to the VP proteins did not protect unvaccinated mice from lethal disease. The demonstration that the VP proteins are capable of eliciting protective immune responses to EBOV-Z indicates that they may be important components of a vaccine designed to protect humans from Ebola hemorrhagic fever
Nonneutralizing Human Antibody Fragments against Hepatitis C Virus E2 Glycoprotein Modulate Neutralization of Binding Activity of Human Recombinant Fabs

AbstractEvidence from clinical and experimental studies indicates that hepatitis C virus E2 (HCV/E2) glycoprotein is the major target of a putatively protective immune response. However, even in the presence of a vigorous production of anti-HCV/E2 antibodies, reinfection can occur. Dissection of the human immune response against HCV/E2 indicated that blocking of binding of HCV/E2 to target cells [neutralization of binding (NOB) activity] varies widely among antibody clones. Moreover, in vivo, simultaneous binding of antibodies to distinct epitopes can induce conformational changes and synergies that may be relevant to understanding the anti-HCV immune response. In this study, human recombinant Fabs were generated by affinity-selecting a phage display repertoire library with antibody-coated HCV/E2. These Fabs, which share the same complementarity-determining region DNA sequences, had higher affinity than other anti-HCV/E2 Fabs but showed no NOB activity even at the highest concentrations. Binding of Fabs to HCV/E2 caused conformational changes modifying Fab-binding patterns and reducing, with a negative synergistic effect, Fab-mediated NOB activity. These data suggest that some antibody clones have the potential to modify HCV/E2 conformation and that, in this state, binding of this glycoprotein to its cellular target is less prone to inhibition by some antibody clones. This can explain why high anti-HCV/E2 antibody titers do not directly correlate with protection from infection. Information on the interactions among different antibody clones can contribute to understanding virus–host interplay and developing more effective vaccines
The 3′ Stem Loop of the West Nile Virus Genomic RNA Can Suppress Translation of Chimeric mRNAs

AbstractCis-acting elements that regulate translation have been identified in the 3′ noncoding regions (NCRs) of cellular and viral mRNAs. As one means of analyzing the effect on translation of the conserved 3′ terminal RNA structure of the West Nile virus (WNV) genome, the translation efficiencies of chimeric mRNAs composed of a CAT reporter gene flanked by viral or nonviral 5′ and 3′ terminal sequences were compared. In vitro, the WNV 3′(+) stem loop (SL) RNA reduced the translation efficiencies of chimeric mRNAs with either viral or nonviral 5′ NCRs, suggesting that a specific 3′–5′ RNA–RNA interaction was not involved. In contrast, the 3′ terminal sequence of a togavirus, rubella virus, enhanced translation efficiency. The WNV 3′(+)SL reduced translation efficiency both in cis and in trans and of both capped and uncapped chimeric mRNAs. We have previously reported that three cellular proteins bind specifically to the WNV 3′(+)SL RNA. Competition between the WNV 3′(+)SL and the 5′ terminus of the chimeric mRNAs for proteins involved in translation initiation could explain the translation inhibition observed
Expression of Kaposi's Sarcoma-Associated Herpesvirus G Protein-Coupled Receptor Monocistronic and Bicistronic Transcripts in Primary Effusion Lymphomas

AbstractKaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) encodes a G protein-coupled receptor (vGPCR) in open reading frame (ORF) 74, which is homologous to human chemokine receptors. KSHV vGPCR is constitutively active and induces VEGF-mediated angiogenesis. Previous studies have shown that ORF 74 is transcribed as part of a bicistronic message containing ORF K14 upstream of ORF 74, with an early lytic pattern of expression. We have now extended these studies by analyzing three different KSHV-positive primary effusion lymphoma (PEL) cell lines and three PEL clinical samples. In addition, we have identified another less abundant monocistronic transcript containing only ORF 74. Both transcripts were identified at low but similar levels in two PEL clinical samples. We evaluated the degree of sequence and functional conservation of ORF74 in three additional PELs and two KS clinical specimens, demonstrating complete identity at the amino acid level among all isolates. While it is expressed as an early lytic transcript in PEL cell lines, in primary clinical PEL samples transcription of KSHV vGPCR can be readily detected
Full-Genome Nucleotide Sequence of a Hepatitis E Virus Strain That May Be Indigenous to Japan

AbstractWe identified hepatitis E virus (HEV) RNA in serum from a Japanese patient with acute hepatitis, who had never been abroad. The full-genome nucleotide sequence of the HEV isolate (JRA1) from this patient was composed of 7227 nucleotides excepting the poly(A) tail and had ORF1 coding for 1703 amino acids (aa), ORF2 coding for 660 aa, and ORF3 coding for 122 aa. This Japanese strain showed approximately 87% nucleotide similarity to human and swine strains reported from the United States, while it had only 73–76% similarity to Asian and Mexican strains. Here we report the characteristics of the HEV-JRA1 isolate, which might be the first example of an indigenous strain(s) of HEV in Japan