pinecone/core-2020-05-10-deduplication huggingface dataset:
Immunity to Rotavirus in T Cell Deficient Mice

AbstractRotavirus infection was studied in adult nude mice (BALB/c background), αβ or γδ and αβ/γδ T cell receptor (TCR) knockout (−/−) mice (C57BL/6 and C57BL/6×129 backgrounds), and SCID mice (C57BL/6 background). The γδ TCR −/− mice cleared infection just like control mice. All of the nude mice, αβ, and αβ/γδ TCR −/− mice cleared primary rotavirus infection, with a short delay, compared to immunocompetent control mice and developed a rotavirus-specific intestinal IgA measured by ELISA. Elispot analysis with spleen and lamina propia cells showed that the virus-specific intestinal IgA response in immunocompetent C57BL/6 mice was similar to the γδ TCR −/− mice and 7- to 60-fold higher than in the αβ TCR −/− and αβ/γδ TCR −/− mice. Likewise, the response of nude +/− mice was 20 times greater than that of nude −/− littermates. While the intestinal IgA antibodies of C57BL/6 mice, γδ TCR −/− mice, and nude +/− mice recognized insect cells infected with recombinant baculovirus expressing rotavirus VP6 and VP4 proteins, those of the αβ TCR −/−, αβ/γδ TCR −/−, and nude −/− mice recognized only VP6. Immunocompetent C57BL/6 mice depleted of CD4+T cell developed similar levels of rotavirus-specific intestinal IgA as the αβ TCR −/− mice, suggesting that this T cell-independent IgA response is present in normal mice. In contrast to previously published results with BALB/c SCID and RAG 2 −/− (C57BL/6×129 background) mice, all of which become chronically infected with murine rotavirus, 40% of the C57BL/6 SCID mice cleared primary rotavirus infection. These results suggest that both a T cell-independent antibody response and innate mechanisms can contribute to immunity to murine rotavirus and show that γδ T cells are not necessary for efficient clearance of primary rotavirus infection in mice
Host Range and Cytopathogenicity of the Highly Attenuated MVA Strain of Vaccinia Virus: Propagation and Generation of Recombinant Viruses in a Nonhuman Mammalian Cell Line

AbstractModified vaccinia virus Ankara (MVA), attenuated by over 500 passages in primary chick embyro fibroblasts (CEF), is presently being used as a safe expression vector. We compared the host ranges of MVA and the parental Ankara strain in CEF and 15 permanent cell lines. The cells could be grouped into three categories: permissive, semipermissive, and nonpermissive. For MVA, the permissive category consisted of primary CEF, a quail cell line derived from QT6, and the Syrian hamster cell line BHK-21. Only in BHK-21 cells did the virus yield approach that occurring in primary CEF. The semipermissive category included two African green monkey cell lines: BS-C-1 and CV-1. The nonpermissive category for MVA consisted of three human cell lines HeLa, 293, and SW 839; one rhesus monkey cell line FRhK-4; two Chinese hamster cell lines CHO and CHL; one pig cell line PK(15); and three rabbit cell lines RK13, RAB-9, and SIRC. The grouping for MVA with a restored K1L host range gene was similar except for the inclusion of RK13 cells among permissive lines. The grouping for the Ankara strain, however, was quite different with more permissive and semipermissive cell lines. Nevertheless, in cells that were permissive for MVA, the virus replicated to higher levels than Ankara, consistent with both positive and negative growth elements associated with the adaptation of MVA. The cell lines were also characterized according to their susceptibilty to MVA-induced cytopathic effects, expression of a late promoter regulated reporter gene by an MVA recombinant, and stage at which virion morphogenesis was blocked. Finally, the permissive BHK-21 cell line was shown to be competent for constructing and propagating recombinant MVA, providing an alternative to primary CEF
Interactions between Tat of HIV-2 and Transcription Factor Sp1

AbstractTat of HIV-2 (Tat-2) requires host cellular factors for optimal function. We show that transactivation by Tat-2 of the HIV promoter requirescis-acting binding sites for Sp1 or Sp1 brought to the promoter via a heterologous system. We demonstrate that an activation domain in Tat-2 consists of one of two potential α-helices in the amino-terminal region, the cysteine-rich region, and the core region and that this independent activation domain requirescis-acting Sp1-binding sites for function. Tat-2 interacts with Sp1 inin vitrobinding assays, and these interactions require basic residues outside of the Tat-2 activation domain. The regions in Sp1 sufficient for functional synergy with Tat are the Sp1 activation domains, while the DNA-binding region is dispensable. Substitution mutations of a glutamine-rich region in one Sp1 activation domain, which eliminate interactions with a TBP-associated factor, also significantly decrease synergy with Tat. Thus, the functional synergy between Tat-2 and Sp1 localizes to domains in each activator that interact with components of the transcription complex. We suggest that these interactions, rather than direct Tat/Sp1 binding, result in highly processive RNA polymerase II complexes and full-length viral transcripts
Formation of Cytomegalovirus DNA Replication Compartments Defined by Localization of Viral Proteins and DNA Synthesis

AbstractTo characterize the formation of replication compartments in human cytomegalovirus-infected cells, and to determine the fate of newly synthesized DNA, we localized viral replication proteins and DNA synthesis at early and late times during infection. As expected, ppUL57 (single-stranded DNA binding protein) and ppUL44 (DNA polymerase processivity factor) both localized to replication compartments beginning at 48 hpi. BrdU was incorporated into viral DNA in these compartments that was found to mature into progeny virus based on our ability to chase the label into the cytoplasm and out of the cell over the ensuing 72-h period. Although the pattern of BrdU incorporation at early times (20 or 24 hpi) was punctate, and distinct from the replication compartment that formed later during infection, viral DNA synthesized at this time also matured into progeny virus during a chase. Interestingly, sites of ppUL57 localization did not overlap completely with sites of BrdU incorporation at early times. Products from the UL112-113 gene localized to subnuclear regions by 6 hpi, earlier than ppUL57. Between 12 and 24 hpi, both ppUL57 and ppUL44 joined UL112-113 gene products at sites that subsequently developed into replication compartments. When infection was carried out in the presence of phosphonoformate or ganciclovir, replication compartment formation was blocked. A viral mutant deficient in uracil DNA glycosidase, previously shown to exhibit a delay in the initial phase of DNA replication, also exhibited delayed formation of replication compartments. These results raise the possibility that subnuclear sites defined by UL112-113 localization orchestrate the assembly of the CMV replication compartment and implicate punctate sites of BrdU incorporation as sites of early viral DNA replication that precedes the formation of the replication compartment
Molecular Studies on Bromovirus Capsid Protein

AbstractTwo members of the bromovirus group, brome mosaic virus (BMV) and cowpea chlorotic mottle virus (CCMV), selectively infect barley and cowpea, respectively, and also differ in their ability to systemically infect a common permissive host,Chenopodium quinoa.CCMV is confined to inoculated leaves ofC. quinoa,whereas BMV causes rapid systemic mottling. To examine whether host-specific determinants for systemic movement of BMV and CCMV in each of these hosts are localized in the coat protein (CP), sequences encoding this gene were exchanged between biologically active clones of BMV RNA3 (B3) and CCMV RNA3 (C3) to create chimera expressing heterologous CP genes (B3/CCP and C3/BCP). Inoculation of each chimera with its respective wild-type (wt) RNAs 1 and 2 to barley or cowpea orC. quinoaplants resulted in symptom phenotype and long distance movement characteristics similar to those of the parental virus donating RNAs 1 and 2. These observations suggest that neither BMV CP nor CCMV CP has host-specific determinants for long distance movement. Inoculation of additional recombinant viruses, constructed by reassorting wt genomic RNAs 1 and 2 of BMV and CCMV with either heterologous wt RNA3 (i.e., B1+B2+C3 and C1+C2+B3) or heterologous chimeric RNA3 (i.e., B1+B2+C3/BCP and C1+C2+B3/CCP), to susceptible hosts resulted only in localized infections. The significance of these observations in relation to bromovirus movement is discussed
Analysisin Vivoof Turnip Crinkle Virus Satellite RNA C Variants with Mutations in the 3′-Terminal Minus-Strand Promoter

AbstractTurnip crinkle virus and its associated RNA, sat-RNA C, share similar, but not identical hairpins near their 3′ ends and terminate with CCUGCCC-OH, which forms a single-stranded tail. With anin vitrotranscription system containing partially purified TCV RdRp, the 3′-terminal 29 bases making up the hairpin and single-stranded tail were previously demonstrated to be required for transcription, and alterations in the stem, but not the loop, could affect template activity (C. Song and A. E. Simon, 1995,J. Mol. Biol.254, 6–14). We have now analyzed sat-RNA C mutants in the 3′ hairpin for ability to accumulatein vivo.While active templatesin vitrowere able to accumulatein vivo,some very weak templatesin vitrowere also able to accumulatein vivowithout reversion or second-site alterations. Computer models of hairpin structure indicated that biologically active promoters could have hairpins less stable than wild type, with loops of variable length and sequence, and without a need for a 6-base single-stranded tail. In addition, transcripts containing compensatory exchanges in the upper stem region that had limited activityin vitrowere biologically activein vivo,indicating that positioning of specific bases in the stem is not required to produce an active minus-strand promoter
Destabilization of the RB Tumor Suppressor Protein and Stabilization of p53 Contribute to HPV Type 16 E7-Induced Apoptosis

AbstractCells that express the human papillomavirus (HPV) type 16 E7 oncoprotein are predisposed to undergo apoptosis. Transgenic mice that have E7 expression targeted to either the retinal photoreceptor cells or the lens cells exhibit signs of apoptosis in cells attempting to undergo differentiation. We established a cell culture system to study this process and have determined the domains of E7 that are required for predisposing cells to undergo apoptosis in response to growth arrest signals. Regions within the core pRB binding site of E7 were necessary but not sufficient for inducing apoptosis. Residues within the adenovirus conserved region 1 homology domain and the consensus casein kinase II phosphorylation site are also important for this effect on cell viability. Our data also demonstrate that the ability of E7 to induce destabilization of pRB and stabilization of p53 coincides with E7-mediated transformation and apoptosis
Epstein–Barr Virus Induces GM-CSF Synthesis by Monocytes: Effect on EBV-Induced IL-1 and IL-1 Receptor Antagonist Production in Neutrophils

AbstractNeutrophils play an important role in the control of viral infections by releasing a variety of potent agents. We previously demonstrated that Epstein–Barr virus (EBV) binds to human neutrophils and stimulates cytokine synthesis including interleukin-1 (IL-1) and IL-1 receptor antagonist (IL-1Ra). Since neutrophil functions are known to be modulated by the priming effect of granulocyte–macrophage colony-stimulating factor (GM-CSF), we therefore investigated the cellular source of GM-CSF synthesis following treatment of leukocytes with EBV and the effect of GM-CSF on the production of IL-1, IL-1Ra, and superoxide by EBV-treated neutrophils. In enriched-cell populations, only monocytes were found to produce GM-CSF in response to EBV, which was maximal after 12 h of incubation. The results obtained with UV-irradiated particles or EBV neutralized with monoclonal antibody 72A1 suggest that contact between the cell and the gp350 of the viral envelope is sufficient to induce the release of GM-CSF. On the other hand, GM-CSF differentially upregulated EBV-induced IL-1 and IL-1Ra production by neutrophils. Pretreatment of neutrophils with GM-CSF prior to EBV activation synergistically enhanced the production of IL-1α and IL-1β, but only marginally affected IL-1Ra synthesis. In addition, GM-CSF was also found to synergistically enhance the superoxide production by neutrophils in response to EBV. Molecular analysis showed that GM-CSF did not alter the IL-1β and IL-1Ra mRNA synthesis induced by EBV, suggesting that GM-CSF could act at a posttranslational level. Local production of GM-CSF by monocytes in tissues invaded by EBV could serve to potentiate the host defense mechanisms directed toward the destruction of the infectious virus
The Coat Protein of Turnip Crinkle Virus Is Involved in Subviral RNA-Mediated Symptom Modulation and Accumulation

AbstractSome satellite (sat-) and defective interfering (DI) RNAs associated with plant viruses intensify or ameliorate the symptoms of the virus. We recently demonstrated that the TCV coat protein (CP) is involved in symptom modulation by sat-RNA C. Two additional subviral RNAs have now been tested for effect of the CP on symptom modulation. DI RNA G, which normally intensifies the symptoms of TCV, is able to attenuate symptoms if the TCV CP is replaced with the CP of cardamine chlorotic fleck virus. DI RNA G had no effect on the symptoms of TCV with a single base alteration in the CP open reading frame, unlike sat-RNA C, which was able to ameliorate the symptoms of the mutant TCV. Using a hybrid sat-RNA constructed from sat-RNA C and TCV (which shares a similar 3′-end region with DI RNA G), the 3′-terminal 53 bases of sat-RNA C were found to be involved in symptom attenuation, which was directly correlated with the lack of detectable viral genomic RNA in whole plants. Sat-RNA D had no effect on the symptoms of mutant or wild-type TCV. The accumulation of TCV subviral RNAs in plants and protoplasts was also found to be strongly influenced by the presence or absence of the wild-type TCV CP
Transformation of Primary Chick Embryo Fibroblasts by Marek's Disease Virus

AbstractMarek's disease virus (MDV) is an alphaherpesvirus, which can mediate the malignant transformation of lymphocytes to form lymphomas in chickens. In this study, we demonstrate that MDV can transform primary chick embryo fibroblasts (CEF). The cell line derived from primary CEF infected with the GA strain of MDV was called CEM(MDV). The fibroblast nature of CEM(MDV)was verified by absence of cytokeratin type II. The CEM(MDV)phenotype differed from either primary CEF or MDV-infected CEF. CEM(MDV)were extensively vacuolated, with unusual multilamellar structures in the cytoplasm. The nuclei were considerably larger than those in primary CEF and were uniformly positive for proliferating cell nuclear antigen. The cell line was subcultured for more than 10 generations; however, CEM(MDV)did not support a fully productive MDV infection, because complete nucleocapsids were not detected and infectivity assays showed that the cell line produced no infectious virus. PCR analyses demonstrated that this cell line carried both polypeptide 38 (pp38) and Meq DNA, MDV-specific genes associated with transformation. In addition, examination by laser scanning confocal microscopy revealed that CEM(MDV)constitutively produced MDV MEQ protein in nuclei and pp38 as well as glycoprotein B in the cytoplasm and on the plasma membrane. Growth in soft agar assay demonstrated that CEM(MDV)formed colonies, similar to HeLa and human melanoma cells. Retroviral insertion was not detected in DNA from the CEM(MDV)line
Negative Regulation of a Heterologous Promoter by Human Cytomegalovirus Immediate-Early Protein IE2

AbstractThe HCMV IE2 protein promiscuously activates transcription of many viral and cellular genes. IE2 also negatively autoregulates its own expression by binding to a strategically positioned IE2 binding site, called CRS, located immediately downstream of the TATA box of the HCMV major IE promoter. Here we show that IE2 is able to repress transcription driven by a heterologous promoter, RSV LTR. Repression of RSV LTR by IE2 is completely dependent of DNA sequences downstream of the TATA box of RSV LTR. A DNA sequence, 5′-CGATACAATAAACG-3′, evidently matching the proposed CRS consensus sequence, is located between nucleotides −13 and +1 (relative to the transcription start site) of RSV LTR. Three lines of evidence support the notion that this RSV CRS element is involved in the IE2-mediated repression of RSV LTR. First, introduction of mutation to the RSV CRS element renders to the mutant RSV LTR resistance to IE2-mediated repression. Second, a mutant IE2 defective in DNA binding cannot downregulate transcription from RSV LTR. Third, IE2 specifically binds to the wild-type, but not the mutant, RSV CRS elementin vitro.These data, in conjunction with previous works, demonstrate that IE2 can passively repress transcription of homologous and heterologous promoters that contain a CRS element
Novel Defective Interfering DNAs Associated with Ageratum Yellow Vein Geminivirus Infection ofAgeratum conyzoides

AbstractDefective DNA forms of the geminivirus ageratum yellow vein virus (AYVV) have been identified in naturally infectedAgeratum conyzoidesplants. Several examples of the defective DNA have been cloned from purified virus-specific supercoiled DNA and characterized by sequence analysis. All are approximately half the size of AYVV genomic DNA, and all contain intergenic region sequences and the 5′ terminus of geneC1as well as additional sequences that are unrelated to the viral genomic DNA. The chimeric nature of the defective DNA distinguishes it from previously characterized geminivirus defective and satellite DNAs. The defective DNA ameliorates disease symptoms and causes a significant delay in the accumulation of viral DNA during the early stage of infection when coinoculated with the AYVV genomic DNA intoNicotiana benthamiana,suggesting a biological role as a defective interfering DNA
Equine Herpesvirus 1 Mutants Devoid of Glycoprotein B or M Are Apathogenic for Mice but Induce Protection against Challenge Infection

AbstractEquine herpesvirus 1 (EHV-1) mutants devoid of the open reading frames (ORFs) of either glycoprotein (g) B or M were constructed and tested for their immunogenic potential in a murine model of EHV-1 infection. The mutant viruses were engineered using the virulent EHV-1 strain RacL11 or the modified live vaccine strain RacH by inserting theEscherichia coli LacZgene into the viral ORFs. RacL11-infected mice showed signs typical of an EHV-1 infection, whereas mice infected with the EHV-1 gB- or gM-negative mutants or with RacH did not develop disease. No difference in the pathogenic potential of RacL11 gB- and gM-negative viruses was observed after application of either phenotypically complemented or negative viruses. However, revertant RacL11 viruses in which the gB or gM gene had been restored caused EHV-1-related symptoms that were indistinguishable from those induced by RacL11. Mice that had been immunized with phenotypically negative gB- and gM-deficient EHV-1 were challenged with the RacL11 virus 25 days after immunization. Mock-immunized mice developed EHV-1 disease and high virus loads in their lungs were observed. In contrast, mice immunized with the mutant viruses did not exhibit EHV-1-caused disease. It was concluded (i) that deletion of either gB or gM abolished the virulence of strain RacL11 and (ii) that immunization with gB- or gM-negative EHV-1 elicited a protective immunity that was reflected by both virus-neutralizing antibodies and EHV-1-specific T-cells in spleens of immunized mice
A Short Basic Domain Supports a Nucleic Acid-Binding Activity in the Rice Tungro Bacilliform Virus Open Reading Frame 2 Product

AbstractLittle is known about the features of badnavirus open reading frame 2 products (P2). So far, no consensus functional domain has been found in these proteins. However, they all have in common at their C-terminus amino acids which may have the capacity to bind nucleic acids. Such capacity has already been established for cacao swollen shoot virus protein P2. We have looked for such a binding capacity of rice tungro bacilliform virus (RTBV) ORF 2 product. For this purpose, the protein was expressed as full-length or truncated versions inEscherichia coli.When used in nucleic acid-binding assays, complete RTBV P2 was shown to bind both DNA and RNA. This property may be related to a basic sequence, PPKKGIKRKYPA, localized at its C-terminus. Mutations were introduced into this sequence and revealed that four of the five basic residues, including a crucial lysine, are required for the binding to nucleic acids. Moreover, this sequence can confer binding capacity when it is fused to the N-terminus of nonbinding proteins
A Comparative Analysis of the Interactions of the E6 Proteins from Cutaneous and Genital Papillomaviruses with p53 and E6AP in Correlation to Their Transforming Potential

AbstractA common necessity for all papillomaviruses is to induce DNA synthesis in quiescent cells. This is commonly achieved by the E7 gene product, which interferes with the function of members of the retinoblastoma family controlling transition from the G1-phase to the S-phase of the cell cycle. Uncontrolled entry into S-phase activates, however, negative growth control signals which have to be bypassed to achieve production of progeny viruses. In addition to inherent activities of the E7 protein, high risk genital types encode an E6 protein that overcomes p53-mediated G1-arrest and apoptosis in concert with the cellular factor E6AP by targeting p53 for the enhanced ubiquitin-dependent degradation. The key question, which of these functions of genital E6 and E7 proteins is responsible for the carcinogenic phenotype, is still not completely answered. In contrast to high risk genital types no immortalizing or transforming activities have been found for the E7 proteins of the high risk cutaneous HPV8 and 47. On the other hand the ability of the E6 protein to transform established rodent fibroblasts seems to be a property shared by high risk genital and cutaneous types. To examine the existence of a common E6-mediated transforming pathway for both virus groups we compared the properties of the cutaneous E6 proteins with already known functions of E6 proteins of genital viruses. For this we analyzed the E6 proteins of low risk and high risk cutaneous and genital papillomaviruses with respect to cell transformation, to their abilities to bind, degradate, and influence the activity of human p53, and to bind E6AP. The results of our study demonstrate a clear lack of interaction between the transforming E6 proteins of HPV1 and HPV8 and both cellular proteins p53 and E6AP. In contrast, we found E6AP-independent binding of HPV16 E6 and HPV6 E6 to p53, although both proteins were different in their transforming potential. Of all four proteins investigated, only HPV16 E6 was able to bind to p53 and E6AP and to induce degradation of the p53 protein in the reticulocyte system. When we investigated in frame deletion mutants of the E6 protein of HPV16 for their abilities to bind to p53 or E6AP, degradate, and inhibit the transactivation function of p53 and to transform rodent fibroblasts, no correlation between the different activities could be found. Mutants still able to bind p53 and E6AP lacked transforming ability and other mutants that were transformation-competent were deficient in p53 and E6AP binding