pinecone/core-2020-05-10-deduplication huggingface dataset:
Regulation of Human Papillomavirus Type 1 Replication by the Viral E2 Protein

AbstractThe E1 and E2 proteins encoded by papillomaviruses are required for viral replication. Earlier studies have shown that the viral E2 protein plays an important role in replication by targeting the E1 helicase to the origin of replication (ori). We have previously shown that the E1 protein of human papillomavirus (HPV) type 1 is sufficient for the in vivo replication of ori plasmids, although the E2 protein stimulates replication. In this study, we have further analyzed the role of the E2 protein in HPV-1 replication. The optimal ori of HPV-1 contains one putative E1 binding site (E1BS) and two putative E2 binding sites, E2BS-3 and E2BS-4. Plasmid pori171, containing the optimal ori, replicates to much higher levels than plasmid pori312, which includes an additional upstream E2 binding site, E2BS-2, located 75 nucleotides upstream of E2BS-3. To study the possible role of E2BS-2 and other upstream sequences in E2-dependent downregulation of replication, transient replication analysis was done in the presence of increasing levels of the E2 protein. Interestingly, inhibition of pori312 replication was more severe at higher levels of E2, suggesting that this protein may also negatively regulate HPV-1 replication. Deletion of sequences from pori312 containing an additional putative E2BS, E2BS-2A, relieved the repression of replication to a significant extent, while replacement of E2BS-2 with a different sequence of the same length had a modest effect. These results suggest that E2BS-2A plays a major and E2BS-2 a minor role in the negative regulation of HPV-1 replication at high E2 levels. Electrophoretic mobility-shift assays showed that the purified E2 protein bound with high affinity to E2BS-3 and weakly to the other putative E2BSs located within the viral long control region. EMSA using various ori fragments showed the formation of multiple E2–DNA complexes which likely represent binding of E2 to multiple E2BSs present within the HPV-1 ori. Our data are consistent with the assembly of ori–protein complexes at high E2 levels that are impaired for replication and further suggest that E2 may regulate HPV-1 replication by a mechanism involving interaction between the E2 protein bound to E2BSs at a distance
The Nucleotide Sequence and Genomic Organization of Citrus Leaf Blotch Virus: Candidate Type Species for a New Virus Genus

AbstractThe complete nucleotide sequence of Citrus leaf blotch virus (CLBV) was determined. CLBV genomic RNA (gRNA) has 8747 nt, excluding the 3′-terminal poly(A) tail, and contains three open reading frames (ORFs) and untranslated regions (UTR) of 73 and 541 nucleotides at the 5′ and 3′ termini, respectively. ORF1 potentially encodes a 227.4-kDa polypeptide, which has methyltransferase, papain-like protease, helicase, and RNA-dependent RNA polymerase motifs. ORF2 encodes a 40.2-kDa polypeptide containing a motif characteristic of cell-to-cell movement proteins. The 40.7-kDa polypeptide encoded by ORF3 was identified as the coat protein. The genome organization of CLBV resembles that of viruses in the genus Trichovirus, but they differ in various aspects: (i) in trichoviruses ORF2 overlaps ORFs 1 and 3, whereas in CLBV, ORFs 2 and 3 are separated and ORFs 1 and 2 overlap in one nucleotide; (ii) CLBV gRNA and CP are larger than those of trichoviruses; and (iii) the CLBV 3′ UTR is larger than that of trichoviruses. Phylogenetic comparisons based on CP amino acid signatures clearly separates CLBV from trichoviruses. Also contrasting with trichoviruses, CLBV could not be transmitted to Chenopodium quinoa Willd. Considering these singularities, we propose that CLBV should be included in a new virus genus
Transcription from the Gene Encoding the Herpesvirus Entry Receptor Nectin-1 (HveC) in Nervous Tissue of Adult Mouse

AbstractBoth human and murine forms of nectin-1 (HveC, Prr1) can serve as entry receptors for several neurotropic herpesviruses, including herpes simplex viruses 1 and 2 (HSV-1, HSV-2), porcine pseudorabies virus (PRV), and bovine herpesvirus 1. HSV-1, HSV-2, and PRV can cause lethal neurological disease in mice whether inoculation is directly into the central nervous system or by peripheral routes. Expression of nectin-1 transcripts in cells of the adult mouse nervous system was assessed by in situ hybridization. Specific hybridization signals were detected in neurons in sensory, sympathetic, and parasympathetic ganglia of the peripheral nervous system. In addition, specific signals were observed in neurons of the ventral and dorsal horns of the spinal cord and of the brain stem, cerebellum, cerebral cortex, hippocampus, dentate gyrus, and olfactory bulb. These results show that the nectin-1 gene is widely transcribed in neurons in adult mouse. Nectin-1 is the only known receptor capable of mediating the entry of all three viruses, HSV-1, HSV-2, and PRV. Its pattern of expression in the nervous system suggests a key role in neurological disease caused by these viruses
TNF-α-Induced Cell Death in Feline Immunodeficiency Virus-Infected Cells Is Mediated by the Caspase Cascade

AbstractTNF-α induced apoptosis in a feline fibroblastic cell line (CRFK) infected with FIV but not in its uninfected control. In this study, to understand the molecular basis of the different susceptibilities to TNF-α between FIV-infected and uninfected cells, we examined the expression of TNF receptors and the activation of the caspase and NF-κB pathways. Expression levels of TNFR I and TNFR II mRNAs were similar between uninfected and FIV-infected CRFK cells. To understand the role of caspases in TNF-α-induced apoptosis, we examined the effect of three different classes of caspase inhibitors, Z-VAD-FMK, Ac-YVAD-CMK, and Z-DEVD-FMK, on the TNF-α-induced apoptosis in FIV-infected cells. Pretreatment with each of these caspase inhibitors protected FIV-infected CRFK cells from TNF-α-induced cell death. Moreover, one of the caspase substrates, poly(ADP-ribose) polymerase, was shown to be cleaved after TNF-α treatment in FIV-infected CRFK cells but not in uninfected CRFK cells. Electrophoretic mobility shift assay using an NF-κB motif oligonucleotide and promoter assay using an NF-κB luciferase reporter construct indicated that TNF-α treatment had induced activation of NF-κB in both FIV-infected and uninfected CRFK cells. The present study indicates that TNF-α-induced apoptosis in FIV-infected CRFK cells is mediated by the activation of the caspase cascade, but not by either upregulation of TNF receptor or inhibition of NF-κB
Pathogenesis of SIV Pneumonia: Selective Replication of Viral Genotypes in the Lung

AbstractLymphocytic interstitial pneumonia of HIV-infected individuals and SIV pneumonia of macaques are both characterized by diffuse infiltration of the lungs with lymphocytes, plasma cells, and macrophages. This study was undertaken to determine whether there are specific, macrophage-tropic genotypes that selectively replicate in the lung of macaques with SIV pneumonia, as in SIV encephalitis. Using a rapid, reproducible SIV/macaque model of AIDS, 11 pig-tailed macaques were intravenously inoculated with an immunosuppressive viral strain, SIV/DeltaB670, and a macrophage-tropic molecule clone, SIV/17E-Fr, and euthanized at 3 months postinoculation. All 11 macaques had severe (6 macaques) or moderate (5 macaques) pneumonia. To identify the viral genotypes that were replicating in the lung parenchyma, bronchoalveolar lavage (BAL) cells, and peripheral blood mononuclear cells (PBMC) of each macaque, RNA was isolated and the SIV env V1 region was amplified, cloned, and sequenced. Lung homogenates and BAL cells contained a more limited repertoire of viral genotypes than PBMC. SIV/17E-Fr was the major genotype in the lungs of 5 macaques and in BAL cells of 6 macaques. The remainder of the macaques had SIV/17E-Fr and the macrophage-tropic strains of SIV/DeltaB670 clones 2 and 12. In contrast, SIV/17E-Fr was the predominant strain in the PBMC of only 3 of 11 macaques. The viral strain that predominated in PBMC was rarely the strain that predominated in the lungs (only 3 of 11 macaques). The severity of pulmonary lesions did not correlate with the levels of viral RNA in lung homogenates or in plasma. However, when only SIV/17E-Fr was expressed in the lung, the viral load in the lung was significantly higher (P = 0.016) than when SIV/DeltaB670 was present alone or in combination with SIV/17E-Fr. These data suggest that SIV pneumonia is associated with selective replication of specific macrophage-tropic genotypes in the lung and that SIV/17E-Fr has a selective advantage for replication in the lung
The Variability of Hop Latent Viroid as Induced upon Heat Treatment

AbstractWe have previously shown that heat treatment of hop plants infected by hop latent viroid (HLVd) reduces viroid levels. Here we investigate whether such heat treatment leads to the accumulation of sequence variability in HLVd. We observed a negligible level of mutated variants in HLVd under standard cultivation conditions. In contrast, the heat treatment of hop led to HLVd degradation and, simultaneously, to a significant increase in sequence variations, as judged from temperature gradient–gel electrophoresis analysis and cDNA library screening by DNA heteroduplex analysis. Thirty-one cDNA clones (9.8%) were identified as deviating forms. Sequencing showed mostly the presence of quadruple and triple mutants, suggesting an accumulation of mutations in HLVd during successive replication cycles. Sixty-nine percent of base changes were localised in the left half and 31% in the right half of the secondary structure proposed for this viroid. No mutations were found in the central part of the upper conserved region. A “hot spot” region was identified in a domain known as a “pathogenicity domain” in the group representative, potato spindle tuber viroid. Most mutations are predicted to destabilise HLVd secondary structure. All mutated cDNAs, however, were infectious and evolved into complex progeny populations containing molecular variants maintained at low levels
Multiple Sclerosis Retrovirus Particles and Recombinant Envelope Trigger an Abnormal Immune Response in Vitro, by Inducing Polyclonal Vβ16 T-Lymphocyte Activation

AbstractA retroviral element (MSRV) defining a family of genetically inherited endogenous retroviruses (HERV-W) has recently been characterized in cell cultures from patients with multiple sclerosis (MS). To address the possible relationship with MS, direct detection of circulating virion RNA was proposed but revealed technically difficult to perform in standardized conditions, in the face of multiple endogenous HERV-W copies. A parallel approach has evaluated MSRV potential pathogenicity in relation to characteristic features of multiple sclerosis, in particular, T-lymphocyte-mediated immunopathology. We report here that MSRV particles induce T-lymphocyte response with a bias in the Vβ16 chain usage in surface receptor, whatever the HLA DR of the donor. A recombinant MSRV envelope—but not core—protein reproduced similar nonconventional activation. Molecular analysis of Vβ CDR3 showed that Vβ16 expansions are polyclonal. Our results thus provide evidence that MSRV envelope protein can trigger an abnormal immune response with similar characteristics to that of superantigens
Adaptation to Blockade of Human Immunodeficiency Virus Type 1 Entry Imposed by the Anti-CCR5 Monoclonal Antibody 2D7

AbstractThe second extracellular loop (ECL2) domain of CC-chemokine receptor 5 (CCR5) has been proposed as a specific target site for therapeutic agents aimed at blocking CCR5-dependent entry by human immunodeficiency virus type I (HIV-1). We have adapted two CCR5-using HIV-1 isolates, prototypic JR-CSF, and a primary isolate, 11-121, to replicate in vitro in the presence of high concentrations of a monoclonal antibody (MAb 2D7) specific for the CCR5 ECL2 domain. The 75% inhibitory concentrations (IC75) for the two 2D7-adapted isolates were approximately 100-fold higher than those for corresponding control isolates passaged without the MAb. Adapted isolates did not acquire the ability to use CXCR4, CCR3, or CCR1. Env clones derived from MAb 2D7-adapted JR-CSF showed several gp120 mutations that were not found in any of the control JR-CSF clones. The in vitro observations suggest that CCR5-using HIV-1 strains might also be able to adapt in vivo to evade an ECL2-blocking therapeutic agent
Enhanced Accumulation of Coronavirus Defective Interfering RNA from Expressed Negative-Strand Transcripts by Coexpressed Positive-Strand RNA Transcripts

AbstractExpression of negative-strand murine coronavirus mouse hepatitis virus (MHV) defective interfering (DI) RNA transcripts in MHV-infected cells results in the accumulation of positive-strand DI RNAs (M. Joo et al., 1996, J. Virol. 70, 5769–5776). However, the expressed negative-strand DI RNA transcripts are poor templates for positive-strand DI RNA synthesis. The present study demonstrated that DI RNA accumulation from the expressed negative-strand DI RNA transcripts in MHV-infected cells was enhanced by the coexpression of complementary RNA transcripts that correspond to the 5′ region of positive-strand DI RNA. The positive-strand RNA transcripts corresponding to the 5′ end-most 0.7–2.0 kb DI RNA had a similar enhancement effect. The coexpressed positive-strand RNA transcripts lacking the leader sequence or those containing only the leader sequence failed to demonstrate this enhancement effect, demonstrating that the presence of the leader sequence in the coexpressed positive-strand RNA transcripts was necessary, but not sufficient, for the enhancement of DI RNA accumulation from the coexpressed negative-strand DI RNA transcripts. Negative-strand DI RNA transcripts that were coexpressed with the partial-length positive-strand RNA transcripts were no more stable than those expressed alone, suggesting that a higher stability of the expressed negative-strand RNA transcripts was an unlikely reason for the higher DI RNA accumulation in cells coexpressing two complementary DI RNA transcripts. Sequence analyses unexpectedly demonstrated that the leader sequence of the majority of accumulated DI RNAs switched to helper virus derived leader sequence, suggesting that enhancement of DI RNA accumulation was mediated by the efficient utilization of helper virus derived leader sequence for DI RNA synthesis. Furthermore, our data suggested that this leader switching, a type of homologous RNA–RNA recombination, occurred during positive-strand DI RNA synthesis and that MHV positive-strand RNA synthesis mechanism may have a preference toward recognizing double-stranded RNA structures over single-stranded negative-strand RNA to produce positive-strand DI RNAs
Replication-Independent Assembly of an Insect Virus (Tetraviridae) in Plant Cells

AbstractInfectious virions of the insect RNA virus Helicoverpa armigera stunt virus (HaSV; Omegatetravirus, Tetraviridae) were assembled in cultured plant protoplasts of Nicotiana plumbaginifolia in the absence of detectable replication. Assembly of the virus, which has not been grown in cell culture, required cotransfection of a DNA plasmid expressing the HaSV capsid gene in combination with either genomic RNA or with DNA plasmids carrying the complete cDNAs to the two HaSV genomic RNAs. Each cDNA was placed under the control of the cauliflower mosaic virus 35S promoter and followed by a cis-acting ribozyme so that the resultant transcripts corresponded precisely to the two genomic RNAs. Protoplast assembly of infectious particles was confirmed by EM and bioassay of host insect larvae, which became diseased and produced virus particles confirmed as HaSV. Variant transcripts carrying nonviral sequences at either or both termini of the RNAs showed no infectivity, except for RNA2 carrying only a 3′ terminal extension. No replication of HaSV in protoplasts was detected in pulse-labeling and blotting experiments. Insects showed less severe disease symptoms when fed protoplasts transfected with only the RNA1 and coat protein plasmids. The symptomatic larvae contained only RNA1 and failed to yield infectious progeny virus, suggesting that RNA1 is capable of self-replication. This novel plasmid-based system confirms that the reported sequence of HaSV represents an infective genome and establishes a procedure for the reverse genetics of a tetravirus
Reduced Macrophage Infiltration and Demyelination in Mice Lacking the Chemokine Receptor CCR5 Following Infection with a Neurotropic Coronavirus

AbstractStudies were performed to investigate the contributions of the CC chemokine receptor CCR5 in host defense and disease development following intracranial infection with mouse hepatitis virus (MHV). T cell recruitment was impaired in MHV-infected CCR5−/− mice at day 7 postinfection (pi), which correlated with increased (P ≤ 0.03) titers within the brain. However, by day 12 pi, T cell infiltration into the CNS of infected CCR5−/− and CCR5+/+ mice was similar and both strains exhibited comparable viral titers, indicating that CCR5 expression is not essential for host defense. Following MHV infection of CCR5+/+ mice, greater than 50% of cells expressing CCR5 antigen were activated macrophage/microglia (determined by F4/80 antigen expression). In addition, infected CCR5−/− mice exhibited reduced (P ≤ 0.02) macrophage (CD45highF4/80+) infiltration, which correlated with a significant reduction (P ≤ 0.001) in the severity of demyelination compared to CCR5+/+ mice. These data indicate that CCR5 contributes to MHV-induced demyelination by allowing macrophages to traffic into the CNS
Avian-to-Mammal Transmission of an Avian Rotavirus: Analysis of Its Pathogenicity in a Heterologous Mouse Model

AbstractIt has been suggested that group A avian rotaviruses can be transmitted to mammals, but there is no direct evidence that such viruses induce disease in mammals. Suckling mice were orally inoculated with two avian rotaviruses. A pigeon rotavirus, PO-13, was found to induce diarrhea, but a turkey rotavirus, Ty-3, did not. The diarrhea induced by PO-13 was dependent on the age of the mouse. In histopathological examinations, antigens of PO-13 were sporadically detected in absorptive cells in the ileum, and lesions were observed as ballooning degenerations of absorptive cells in a region from the duodenum to the ileum. However, the rotavirus antigen was not detected in the majority of these degenerative cells. These results indicated that PO-13 could infect and induce diarrhea in suckling mice. This is the first evidence of an avian rotavirus being experimentally transmissible to a mammal
Mammalian Reovirus L2 Gene and λ2 Core Spike Protein Sequences and Whole-Genome Comparisons of Reoviruses Type 1 Lang, Type 2 Jones, and Type 3 Dearing

AbstractThe reovirus L2 genome segment encodes the core spike protein λ2, which mediates enzymatic reactions in 5′ capping of the viral plus-strand transcripts. Complete nucleotide-sequence determinations were made for the L2 genome segments of eight mammalian reoviruses, including the prototype isolates of serotypes 1 and 2: Lang (T1L) and Jones (T2J), respectively. Each L2 segment was found to be 3912 or 3915 bases in length. Partial nucleotide-sequence determinations were also made for the 3916-base L2 segment of reovirus type 3 Dearing (T3D), the prototype isolate of serotype 3. The whole-genome sequence of reovirus T3D was reported previously. The T1L L2 analysis represents completion of the whole-genome sequence of that isolate as well. The T2J L2 analysis leaves only the sequence of the M1 segment yet to be reported from the genome of that isolate. The T2J M1 sequence made available from analysis in another lab was used for initiating whole-genome comparisons of reoviruses T1L, T2J, and T3D in this report. The nine L2 gene sequences and deduced λ2 protein sequences were used to gain further insights into the biological variability, structure, and functions of λ2 through comparisons of the sequences and reference to the crystal structure of core-bound λ2. Phylogenetic comparisons suggest the presence of three evolutionary lines of divergent L2 alleles among the nine isolates. Localized regions of conserved amino acids in the λ2 crystal structure include active-site clefts of the RNA capping enzyme domains, sites of interactions between λ2 domains within the pentameric spike structure, and sites of interaction between λ2 subunits and other proteins in viral particles
Antibodies That Neutralize SIVmac251 in T Lymphocytes Cause Interruption of the Viral Life Cycle in Macrophages by Preventing Nuclear Import of Viral DNA

AbstractPrevious reports from our lab had shown that sera obtained from SIVmac-infected animals neutralized SIVmac infectivity in CD4+ T cells but failed to protect monkey primary macrophages from infection with the virus. However, the antibodies could inhibit completion of the viral life cycle in the macrophages at the postentry stage(s). In this report we examined the mechanisms of the late effect of the antibodies. Using monoclonal antibodies (MAbs), we demonstrated that only antibodies to the SIV envelope protein (KK17 and KK42) but not antibody to the viral core protein (FA2) had the same inhibitory effect as that of the anti-SIV sera. To identify the stage of the viral replication cycle that was inhibited by anti-SIV antibodies in macrophages, we used various PCR techniques to study viral entry/reverse transcription (by amplifying the viral gag gene), viral genome nuclear transport (by amplifying 2-LTR circular forms), viral integration (by Alu-PCR assay), and viral protein expression (by RIPA). We found that in macrophage cultures inoculated with SIVmac251 that were preincubated with antienvelope MAbs, viral DNA was detected at 8 h postinoculation but the 2-LTR circular forms and integrated viral DNAs were undetectable, and viral proteins were not expressed in these infected macrophages. These results strongly suggested that anti-SIV antibodies inhibited SIVmac replication in macrophages by blocking nuclear transport of viral genomes since viral DNA could not be detected in the nuclei of treated cultures. Furthermore, we showed that although viral replication in macrophages was interrupted by the antibodies, when cocultured with permissive T cells, the viral genomes presented in the cytoplasm of the macrophages could readily transfer to T cells during cell–cell contact. Importantly, this transfer could not be prevented by the antibodies. These results might explain the failure of passive antibody immunization against SIVmac251—a critical obstacle in AIDS vaccine development
Identification of Sequences in Rotavirus mRNAs Important for Minus Strand Synthesis Using Antisense Oligonucleotides

AbstractThe core of the rotavirion consists of three proteins, including the viral RNA polymerase, and 11 segments of double-stranded (ds)RNA. The RNA polymerase of disrupted (open) cores is able to catalyze the synthesis of dsRNA from exogenous viral mRNAs in vitro. In this study, we have identified sequences in exogenous viral mRNAs important for RNA replication using antisense oligonucleotides. The results showed that oligonucleotides complementary to the highly conserved 3′-terminal sequence of rotavirus mRNAs prevented all but basal levels of dsRNA synthesis. Notably, we observed that the addition of oligonucleotides which were complementary to nonconserved sequences present either at the 5′- or 3′-end of a viral mRNA effectively inhibited its replication without interfering with the replication of other viral mRNAs present in the same replication assay. Thus, the nonconserved sequences in rotavirus mRNAs contain gene-specific information that promotes RNA replication. The fact that antisense oligonucleotides inhibited dsRNA synthesis indicates that the strandedness (single- versus double-stranded) and secondary structure of the viral mRNA template are factors that affect the efficiency of minus strand synthesis