pinecone/core-2020-05-10-deduplication huggingface dataset:
Adenovirus VAI RNA Antagonizes the RNA-Editing Activity of the ADAR Adenosine Deaminase

AbstractThe virus-associated VAI RNA of adenovirus is a small highly structured RNA that is required for the efficient translation of cellular and viral mRNAs at late times after infection. VAI RNA antagonizes the activation of the interferon-inducible RNA-dependent protein kinase, PKR, an important regulator of translation. The RNA-specific adenosine deaminase, ADAR, is an interferon-inducible RNA-editing enzyme that catalyzes the site-selective C-6 deamination of adenosine to inosine. ADAR possesses three copies of the highly conserved RNA-binding motif (dsRBM) that are similar to the two copies found in PKR, the enzyme in which the prototype dsRBM motif was discovered. We have examined the effect of VAI RNA on ADAR function. VAI RNA impairs the activity of ADAR deaminase. This inhibition can be observed in extracts prepared from interferon-treated human cells and from monkey COS cells in which wild-type recombinant ADAR was expressed. Analysis of wild-type and mutant forms of VA RNA suggests that the central domain is important in the antagonism of ADAR activity. These results suggest that VAI RNA may modulate viral and cellular gene expression by modulating RNA editing as well as mRNA translation
Characterization of a Linear Epitope in the Nonstructural Region 4 of Hepatitis C Virus with Reactivity to Seroconversion Antibodies

AbstractMapping and possible diagnostic meaning of a highly conserved, linear NS4 epitope (NS4/3), located outside the C100-3 antigen within the carboxyl terminal proportion of the NS4 region, with major immunoreactivity with specimens of patients with HCV infection from various geographic origins is described. Transient, acute-phase IgM anti-HCV NS4/3 was detected coincidentally or earlier than active IgG anti-HCV NS4/3 response with four well-characterized seroconversion panels. GenBank alignment studies identified patch homologies between the NS4/3 sequence and a number of non-HCV proteins, which may explain part of the cross-reactivity of the NS4/3 epitope. Some of the “false positive reactivities” of the NS4/3 epitope with asymptomatic blood donors, not being confirmed with FDA-approved anti-HCV assays without the NS4/3 epitope, may be explained by recognition of very early seroconversion antibodies
Characterization of the Two Overlapping Papain-like Proteinase Domains Encoded in Gene 1 of the Coronavirus Infectious Bronchitis Virus and Determination of the C-Terminal Cleavage Site of an 87-kDa Protein

AbstractIn a previous report, we showed that proteolytic processing of an 87-kDa mature viral protein from the coronavirus infectious bronchitis virus (IBV) 1a and 1a/1b polyproteins was mediated by two putative overlapping papain-like proteinase domains (PLPDs) encoded within the region from nucleotides 4243 to 5553 of ORF 1a (Liuet al.,1995). In this study, we demonstrate that only the first domain, PLPD-1, is responsible for this cleavage, as deletion of the second domain did not affect the formation of the 87-kDa protein. Site-directed mutagenesis studies further showed that a previously predicted nucleophilic cysteine residue (Cys1274) and a histidine residue (His1437) were essential for the proteinase activity, indicating that they may be important components of the catalytic center of the proteinase. Meanwhile, expression of a series of deletion mutants revealed that the 87-kDa protein was encoded by the 5′-most 2.6 kb of ORF 1a. Deletion and amino acid substitution mutation studies demonstrated that the Gly673–Gly674dipeptide bond was most likely the cleavage site responsible for releasing the C-terminus of the 87-kDa protein from the 1a and 1a/1b polyproteins
Nuclear Import and Export of Viruses and Virus Genomes

AbstractMany viruses replicate in the nucleus of their animal and plant host cells. Nuclear import, export, and nucleo-cytoplasmic shuttling play a central role in their replication cycle. Although the trafficking of individual virus proteins into and out of the nucleus has been well studied for some virus systems, the nuclear transport of larger entities such as viral genomes and capsids has only recently become a subject of molecular analysis. In this review, the general concepts emerging are discussed and a survey is provided of current information on both plant and animal viruses. Summarizing the main findings in this emerging field, it is evident that most viruses that enter or exit the nucleus take advantage of the cell's nuclear import and export machinery. With a few exceptions, viruses seem to cross the nuclear envelope through the nuclear pore complexes, making use of cellular nuclear import and export signals, receptors, and transport factors. In many cases, they capitalize on subtle control systems such as phosphorylation that regulate traffic of cellular components into and out of the nucleus. The large size of viral capsids and their composition (they contain large RNA and DNA molecules for which there are few precedents in normal nuclear transport) make the processes unique and complicated. Prior capsid disassembly (or deformation) is required before entry of viral genomes and accessory proteins can occur through nuclear pores. Capsids of different virus families display diverse uncoating programs which culminate in genome transfer through the nuclear pores
Characterization of the Interactions among Vaccinia Virus Transcription Factors G2R, A18R, and H5R

AbstractPrior genetic analysis suggests that there may exist an interaction between the products of the vaccinia virus genes A18R, a putative negative transcription elongation factor, and G2R, a putative positive transcription elongation factor. In addition, affinity purification of polyhistidine-tagged G2R protein overexpressed in vaccinia virus-infected cells, reported here, results in copurification of the vaccinia H5R protein, previously characterized as a late viral transcription factor. We have therefore used several methods to screen further for interactions among the G2R, A18R, and H5R proteins. Methods include copurification or co-immunoprecipitation of proteins overexpressed during vaccinia virus infection, activation of the gal 4 promoter by gal 4 fusions in the yeast two-hybrid system, and co-immunoprecipitation of proteins synthesizedin vitroin a rabbit reticulocyte lysate. The results reveal interactions which include all possible pairwise combinations of the three proteins G2R, A18R, and H5R; however, not all possible permutations of the interactions are observed and the interactions are not observed in all environments tested. The results suggest that the vaccinia virus proteins G2R, A18R, and H5R interact as part of a higher order transcription complex
Identification of Two Replicons in Phage-Plasmid P4

AbstractDNA replication of phage-plasmid P4 proceeds bidirectionally from theori1site (previously namedori), but requires a secondcis-acting region,crr.Replication depends on the product of the P4 α gene, a protein with primase and helicase activity, that binds bothori1andcrr.A negative regulator of P4 DNA replication, the Cnr protein, is required for copy number control of plasmid P4. Using a plasmid complementation test for replication, we found that two replicons, both dependent on the α gene product, coexist in P4. The first replicon is made by thecnrand α genes and theori1andcrrsites. The second is limited to the α andcrrregion. Thus, in the absence of theori1region, replication can initiate at a different site. By deletion mapping, acis-acting region,ori2,essential for replication of the α-crrreplicon was mapped within a 270-bp fragment in the first half of the α gene. Theori2site was found to be dispensable in a replicon that containsori1.A construct that besidescrrand α carries also thecnrgene was unable to replicate, suggesting that Cnr not only controls replication fromori1,but also silencesori2
A Surface Immunodeterminant of Human Papillomavirus Type 16 Minor Capsid Protein L2

AbstractWe used human papillomavirus type 16 (HPV-16) particles composed of capsid proteins L1 and L2 (L1/L2 capsids) as an antigen to produce mouse monoclonal antibodies (MAbs). Of 18 MAbs recognizing surface epitopes of L1/L2 capsids, 1 was an anti-L1 MAb and 17 were anti-L2 MAbs. Seven of 11 anti-L2 MAbs recognizing linear epitopes were found to bind to a synthetic peptide with an HPV-16 L2 sequence of amino acids (aa) 69–81, which is within a highly conserved region among different HPVs. The synthetic peptide reacted with the human sera that had been shown to be positive for an antibody against HPV-16, -18, -58, or -6b capsids composed of L1 alone. The data suggest that the HPV-16 L2 region of aa 69–81 contains a type-common immunodeterminant exposed on the surface of HPV virions
Membrane-Anchored Incorporation of a Foreign Protein in Recombinant Influenza Virions

AbstractThe RNA polymerase I system forin vivosynthesis of recombinant influenza vRNA molecules was used for the expression of a chimeric protein, consisting of the 341-amino-acid ectodomain of the glycoprotein E2 of classical swine fever virus and the 37-amino-acid C-terminal membrane anchor of the influenza virus hemagglutinin (HA). During infection with an influenza A helper virus the amplified pseudo-viral RNA was packaged into progeny virions together with influenza vRNA segments. The foreign fusion protein E2-HA was shown to be physically incorporated into the viral envelope. Incorporation of a third major glycoprotein into the envelope did not affect biological functions of HA and neuraminidase that are required for the generation of infectious virus particles. Based on mutational analyses of the cytoplasmic tail of E2-HA fusion proteins three modes of interaction during virus budding have been observed: nonspecific low-level incorporation (truncated tails), specific full-level incorporation (wild-type amino acid sequence or minor variations of it), and exclusion from incorporation (elongated tails)
The Role of IL-10 in Mouse Hepatitis Virus-Induced Demyelinating Encephalomyelitis

AbstractInterleukin 10 (IL-10) is an important anti-inflammatory cytokine. To examine its role in virus-induced encephalomyelitis, IL-10-deficient (IL-10 −/−) mice were infected with a neurotropic strain of mouse hepatitis virus (JHMV). JHMV-infected IL-10 −/− mice, compared to IL-4 −/− and syngeneic C57BL/6 mice, exhibited increased morbidity and mortality. Virus was cleared from the CNS of all groups of mice with equal kinetics by day 9 postinfection and the lack of either IL-4 or IL-10 did not alter the distribution of viral antigen, suggesting a lack of correlation between viral replication and the increased clinical disease in IL-10 −/− mice. In moribund IL-10 −/− mice, a moderate increase in mononuclear cell infiltration was correlated with increased expression of tumor necrosis factor-α, interferon-γ, and inducible nitric oxide synthase mRNAs. In the small percentage of IL-10 −/− mice that survived, no differences in either demyelination or inflammation were observed. Together, these results suggest that IL-10 is not required for viral clearance, and although it appears to be one of the mechanisms responsible for inhibiting the extent of inflammation in the CNS during acute JHMV infection, it has little role in the eventual resolution of CNS inflammatory responses
The Predominant eIF4G-Specific Cleavage Activity in Poliovirus-Infected HeLa Cells Is Distinct from 2A Protease

AbstractHuman enteroviruses and rhinoviruses rapidly and selectively abolish translation from cellular mRNA upon infection of susceptible cells. Expression of the poliovirus 2A protease (PV 2Apro) is sufficient to cause host translation shutoff through cleavage of eIF4G (formerly p220, eIF4γ) either directly or indirectly through activation of a cellular factor. Evidence exists for both direct and indirect cleavage mechanisms; however, factors presumed to participate in an indirect mechanism have not yet been purified or defined. Here we show that the dominant eIF4G cleavage activity in lysates from infected HeLa cells was separable from PV 2Aproby size exclusion chromatography. 2Aproseparated into two peak fractions which contained activity which cleaved a peptide substrate derived from the poliovirus polyprotein. These peak 2Aprofractions did not cleave eIF4G or an eIF4G-derived peptide, as expected, due to the poor efficiency of direct cleavage reactions. Conversely, fractions which contained peak eIF4G cleavage activity and only trace amounts of 2Aproefficiently cleaved a peptide substrate derived from the previously mapped eIF4G cleavage site and also cleaved a peptide derived from the poliovirus 1D2A region. The dominant eIF4G cleavage activity was highly purified through four chromatography steps and found to be devoid of all traces of 2Aproor its precursors. Quantitation of 2Aprofrom lysates of infected cells showed that during infections in HeLa cells, 2Aprodoes not reach molar excess over eIF4G, as previously shown to be required for direct eIF4G cleavagein vitro.Further, infection of HeLa cells in the presence of 2 mM guanidine–HCl, a potent inhibitor of viral RNA replication, suppressed accumulation of 2Aproand its precursor 2ABC below detectable levels but was unable to delay the onset of eIF4G proteolysisin vivo.The eIF4G cleavage activity was still easily detectable inin vitroassays using fractions from guanidine-treated cells. Thus, the data suggest that poliovirus utilizes two catalytic activities to ensure rapid cleavage of eIF4Gin vivo.Although it was not directly measurable here, 2Aprolikely does cleave a portion of eIF4G in cells. However, the data suggest that a cellular factor which can be activated by small quantities of 2Aproconstitutes the bulk of the eIF4G-specific cleavage activity in infected cells and is responsible for the rapid and efficient eIF4G cleavage activity observedin vivo
Direct Cleavage of eIF4G by Poliovirus 2A Protease Is Inefficientin Vitro

AbstractPreviously, the purified recombinant 2A proteases (2Apro) of coxsackievirus B4 (CVB4) and human rhinovirus type 2 (HRV2) were shown to cleave synthetic peptides derived from human or rabbit eIF4G as well as eIF4G protein purified from rabbit reticulocytes. These results were in contrast to previous evidence which supported the view that eIF4G cleavage activity in poliovirus-infected HeLa cells required a cellular factor(s) activated by poliovirus (PV) 2Apro. In the present study, recombinant PV 2Aprowas shown to cleave either rabbit or human eIF4G or their derived peptides in direct cleavage reactions, but cleaved the 4G-derived peptides with 100-fold lower efficiency than with a peptide derived from the poliovirus polyprotein. In these experiments, up to 25-fold molar excess of 2Aproover eIF4G protein was required to cause greater than 50% cleavage. CVB4 2Aprowas also tested in peptide cleavage assays under the same conditions as PV 2Aproand was found to cleave all eIF4G substrates with efficiencies similar to PV 2Apro. Finally, cleavage reactions utilizing recombinant eIF4G containing a G486E substitution at the cleavage site for CVB4 and HRV2 proteases resulted in drastically reduced cleavage by PV 2Apro, similar to the reduction previously seen with HRV2 and CVB4 2Apro, confirming that all three viral 2A proteases recognize the same cleavage site on eIF4G. These data show that PV 2Aprocan directly cleave eIF4Gin vitrowith efficiencies similar to those of CVB 2Apro, but cleavage efficiency of eIF4G is approximately 1000-fold lower than cleavage of a peptide derived from the authentic 2A cleavage site on the poliovirus polyprotein
Three Active Forms of Aspartic Proteinase from Mason–Pfizer Monkey Virus

AbstractMason–Pfizer monkey virus (M-PMV) proteinase, released by the autocatalytic cleavage of Gag-Pro and Gag-Pro-Pol polypeptide precursors, catalyzes the processing of viral precursors to yield the structural proteins and enzymes of the virion. In retroviruses, usually only one proteolytically active form of proteinase exists. Here, we describe an unusual feature of M-PMV, the existence of three active forms of a retroviral proteinase with molecular masses of 17, 13, and 12 kDa as determined by mass spectroscopy. These forms arisein vitroby self-processing of a 26-kDa proteinase precursor. We have developed a process for isolation of each truncated product and demonstrate that all three forms display proteolytic activity. Amino acid analyses, as well as the determination of N- and C-terminal sequences, revealed that the N-termini of all three forms are identical, confirming thatin vitroautoprocessing of the 17-kDa form occurs at the C-terminus to yield the truncated forms. The 17-kDa form and the newly described 13-kDa form of proteinase were identified in virions collected from the rhesus monkey CMMT cell line chronically infected with M-PMV, confirming that multiple forms existin vivo
Primary SIVsm Isolates Use the CCR5 Coreceptor from Sooty Mangabeys Naturally Infected in West Africa: A Comparison of Coreceptor Usage of Primary SIVsm, HIV-2, and SIVmac

AbstractGenetically divergent strains of simian immunodeficiency virus (SIV) from macaques (mac), chimpanzees, and sooty mangabeys (SM) efficiently used rhesus and human CCR5 (R5), but not CXCR4 (xR4), for cell entry. Thus far, however, no studies have characterized primary SIVsm strains for their use of coreceptors derived from their own natural host. Coreceptor usage of two primary, blood-derived SIVsm isolates, SIVsmSL92b and SIVsmFNS from naturally infected sooty mangabeys, was determined. Primary SIVsm efficiently used SM-CCR5 expressed on HOS.CD4 and U87.CD4 cells. Sequence polymorphisms in CCR5 found in four sooty mangabeys did not alter viral entry. Unlike primary rhesus blood-derived R5-tropic SIVmac251, primary SM blood-derived R5-tropic SIVsm was strongly CD4 dependent. The SM-CXCR4 gene was fully functional for xR4-tropic primate lentiviruses, but was not used by primary SIVsm. Therefore, the lack of xR4 tropism among naturally occurring SIVsm strains was not due to CxCR4 gene defects in the natural host. SIVmac derived from four macaques with AIDS also did not use macaque- or SM-derived CXCR4, showing that xR4 tropism did not develop during progression to disease as for humans infected with HIV-1. Three of four primary HIV-2 strains used CCR5 from human, sooty mangabey, and macaque. The fourth, HIV-27924A, obtained from a patient with AIDS, was xR4-tropic. Because SIVmac is most closely related to HIV-2, SIVmac might be expected to mimic tropisms of HIV-2 infections. However, the correlation between xR4 tropism and AIDS may be a species-specific phenomenon limited to humans. The R5-tropic primary SIVsm and HIV-2 strains grew in CCR5-negative human PBMC, consistent with their use of non-CCR5 coreceptors. However, primary SIVsmSL92b did not use non-CCR5 coreceptors efficiently. The two primary SIVsm isolates replicated poorly in CEMx174 cells, which do not express CCR5, compared to CCR5-positive PM1 cells. SIVmac grew equally well in both cell lines. The findings show that SM-chemokine receptors are fully functional for virus entry and that multicoreceptor tropism is a common property of primary lentiviruses within the SIVsm/HIV-2 subfamily
The Biological Relevance of Virus Neutralisation Sites for Virulence and Vaccine Protection in the Guinea Pig Model of Foot-and-Mouth Disease

AbstractFive neutralisation epitopes have been defined for the O1Kaufbeuren strain of foot-and-mouth disease virus (FMDV) by neutralising murine monoclonal antibodies (Mabs). A mutant virus which is resistant to all these Mabs also resists neutralisation by bovine polyclonal sera, and this characteristic was exploited in the current study to investigate the biological relevance of neutralisation sites in FMDV virulence and vaccine protection. The five site neutralisation-resistant mutant was shown to be as pathogenic as wild-type virus in the guinea pig model of FMD. Guinea pigs were protected in cross-challenge studies from virulent wild-type and mutant viruses using either wild-type or mutant 146S antigen as inactivated whole virus vaccine. Furthermore, hyperimmune sera raised to either wild-type or mutant antigen offered passive protection against wild-type challenge, in spite of the serum raised against the mutant antigen having minimal neutralising activityin vitro.These results imply that virus neutralisation, at least as defined by thein vitroassay, may not play an essential role in the mechanism of immunity induced by whole inactivated FMDV vaccines
DNA Vaccines Expressing either the GP or NP Genes of Ebola Virus Protect Mice from Lethal Challenge

AbstractDNA vaccines expressing the envelope glycoprotein (GP) or nucleocapsid protein (NP) genes of Ebola virus were evaluated in adult, immunocompetent mice. The vaccines were delivered into the skin by particle bombardment of DNA-coated gold beads with thePowderJect-XRgene gun. Both vaccines elicited antibody responses as measured by ELISA and elicited cytotoxic T cell responses as measured by chromium release assays. From one to four vaccinations with 0.5 μg of the GP DNA vaccine resulted in a dose-dependent protection from Ebola virus challenge. Maximal protection (78% survival) was achieved after four vaccinations. Mice were completely protected with a priming dose of 0.5 μg of GP DNA followed by three or four subsequent vaccinations with 1.5 μg of DNA. Partial protection could be observed for at least 9 months after three immunizations with 0.5 μg of the GP DNA vaccine. Comparing the GP and NP vaccines indicated that approximately the same level of protection could be achieved with either vaccine
Vaccinia Virus Nucleoside Triphosphate Phosphohydrolase I Is an Essential Viral Early Gene Transcription Termination Factor

AbstractDeng and Shuman (J. Biol Chem.271, 29386 (1996)) reported that an ATPase different from the known viral termination factor, VTF, is required for vaccinia virus early gene transcription termination. Properties of this ATPase were similar to those of a known vaccinia virus enzyme, nucleoside triphosphate phosphohydrolase I (NPH I) the product of gene D11L. Transcription-competent cell-free extracts were prepared from A549 cells infected with wild-type or mutant vaccinia virus harboring ts mutations in gene D11L. These extracts were employed to investigate the role of NPH I in early gene transcription termination. Extracts prepared under nonpermissive conditions from both wild-type virus and ts mutant virus-infected cells exhibited high levels of early and intermediate gene transcription activity but were incapable of supporting late gene transcription. ts mutant extracts lacked signal-dependent early gene transcription termination activity, which was restored by the addition of either free NPH I or a GST-NPH I fusion protein. A comparison of the NPH I amino acid sequence to the protein databases revealed the presence of a set of sequences characteristic of nucleic acid helicase superfamily II members. A series of site-specific mutations in the helicase motifs and N-terminal and C-terminal deletion mutations were expressed as GST fusion proteins and their activities assessed. Of the mutations in helicase motifs I to VI, alteration of all but motif III reduced the ATPase activity. Removal of as few as 24 amino acids from the N-terminal end eliminated ATPase activity, while deletion of 68 C-terminal amino acids exhibited only a modest decrease in ATP hydrolysis. Larger C-terminal deletions eliminated ATPase activity. Each deletion mutation, and site-specific mutations other than the motif III mutation, failed to support transcription terminationin vitro.Mutations in motifs I, II, V, and VI inhibit wild-type NPH I transcription termination activity. However, deletion of up to 68 amino acids from the C-terminal end eliminates this inhibitory property. This observation is particularly interesting since these C-terminal deletions retain both ATPase activity and single-stranded DNA binding activity. Their failure to inhibit transcription termination suggests that these C-terminal deletion mutations eliminate a site required for a function other than from DNA binding or ATP hydrolysis