pinecone/core-2020-05-10-deduplication huggingface dataset:
Cell-to-Cell Movement of Turnip Crinkle Virus Is Controlled by Two Small Open Reading Frames That Functionin trans

AbstractPrevious studies on turnip crinkle virus (TCV) have suggested that the two small, centrally located ORFs, conserved in all Carmoviruses, are both required for cell-to-cell movement (Hackeret al.,1992). We now demonstrate that the cell-to-cell movement of TCV is mediated byin transcomplementation of the two proteins. First, both of the putative movement proteins (MPs p8 and p9) were shown to be translatedin vitrofrom transcripts representing the 1.7-kb subgenomic RNA. Western blot analysis, using antisera prepared against GST fusion proteins of both genes, was then used to show that the p8 but not the p9 protein accumulated to detectable levels in particulate fractions of infected cells. Cell-to-cell movement of various MP mutants inArabidopsiswas evaluated byin situhybridization of inoculated leaves. Changes in either of the two MP genes resulted in failure of the mutants to move cell-to-cell. Coat protein was found to be unnecessary for cell-to-cell movement. Complementation of cell-to-cell movement by co-inoculating p8-defective mutants with a p9-defective mutant resulted in delayed systemic infection. In contrast, efficient cell-to-cell movement was achieved when the MP mutants were inoculated into transgenic plants expressing the corresponding functional gene(s). These experiments provide further evidence that both MP genes encoded by Carmoviruses must functionin transin the same cell in order to mediate cell-to-cell movement
The Hepatitis B Virus Seroconversion to Anti-HBe Is Frequently Associated with HBV Genotype Changes and Selection of preS2-Defective Particles in Chronically Infected Children

AbstractIn order to investigate the generation and selection of hepatitis B virus mutants and the influence of interferon on their evolution, a longitudinal study including 22 patients was performed. The complete preS1/S2 open reading frame was analyzed by direct sequencing from serum samples obtained before and after seroconversion to anti-HBe in 11 children without α-interferon treatment. Furthermore, in 11 cases with therapy additional samples obtained during interferon therapy were investigated. The comparison of each patient's preS sequences analyzed before and during therapy did not show any nucleotide change, while in both groups numerous silent and missense mutations were found immediately after seroconversion. Surprisingly, in 7 cases the hepatitis B virus changed genotype from A to D (subtype adw to ayw) after seroconversion. Additional rearrangements were observed in 4 patients. In 3 cases the selection of preS2 start codon mutants was detected after seroconversion and in 1 individual a 183-nucleotide deletion was found during and after HBeAg positivity. In conclusion, the emergence of preS rearrangements and numerous base exchanges provide evidence for a strong selection process focused against the preS region. Moreover, the appearance of genotype changes after anti-HBe seroconversion reveales a thus far unrecognized event during the natural course of HBV infection in childhood
Mice Expressing the E7 Oncogene of HPV16 in Epithelium Show Central Tolerance, and Evidence of Peripheral Anergising Tolerance, to E7-Encoded Cytotoxic T-Lymphocyte Epitopes

AbstractIn order to derive mice which expressed both the E7 open reading frame transgene of human papillomavirus type 16 in skin and MHC class 1 restriction elements for several E7-encoded cytotoxic T-lymphocyte (CTL) epitopes, K14.HPV16E7 mice which express E7 in basal keratinocytes were crossed to the F1 generation with A2.1 Kbtransgenic mice which express the MHC binding cleft domains of human HLA A*0201, and murine H-2b. F1 mice (denoted K14E7×A2.1) expressed E7 in the thymus at least as early as 2–5 days before birth. Immunisation of FVB×A2.1 control mice (transgenic for HLA A*0201 and H-2bbut not for E7), with two HLA A*0201-restricted epitopes of E7 and one H-2b-restricted CTL epitope of E7, gave strong primary CTL responses recognising epitope-pulsed or constitutively E7-expressing syngeneic target cells. In contrast, in immunised K14E7×A2.1 mice, the CTL responses to the H-2bepitope and one of the HLA A*0201 CTL epitopes were strongly down-regulated, and to the other HLA A*0201 epitope, completely abolished, as demonstrated by percentage specific killing by bulk splenocyte cultures in cytotoxicity assays, and by CTL precursor frequency analysis. In thymus-transplanted bone marrow radiation chimeras in which the immune system of K14E7×A2.1 mice was replaced by a FVB×A2.1 immune system, specific immunisation did not result in reemergence of strong E7-directed CTL responses. In agreement with thesein vitrofindings, specific immunisation failed to significantly alter the course of E7-associated tumour development in K14E7×A2.1 mice. These data are consistent with a model of central deletional CTL tolerance to E7-encoded epitopes recognised in the context of two distinct MHC class 1 restriction elements, and with the possibility of peripheral T-cell anergy maintained by expression of E7 in the skin
The Exonuclease Activity of HSV-1 UL12 Is Required forin VivoFunction

AbstractThe herpes simplex virus type 1 (HSV-1) UL12 gene encodes an alkaline pH-dependent deoxyribonuclease termed alkaline nuclease. A recombinant UL12 knockout mutant, AN-1, is severely compromised for growth, and analysis of this mutant suggests that UL12 plays a role in processing complex DNA replication intermediates (R. Martinez, R. T. Sarisky, P. C. Weber, and S. K. Weller, (1996)J. Virol.70, 2075–2085). This processing step may be required for the generation of capsids that are competent for egress from the nucleus to the cytoplasm. In this report, we address the question of whether the AN-1 growth phenotype is due to the loss of UL12 catalytic activity. We constructed two point mutations in a highly conserved region (motif II) of UL12 and purified wild-type and mutant enzymes from a baculovirus expression system. Both mutant proteins are stable, soluble, and competent for correct nuclear localization, suggesting that they have retained an intact global conformation. Neither mutant protein, however, exhibits exonuclease activity. In order to examine thein vivoeffects of these mutations, we determined whether expression of mutant proteins from amplicon plasmids could complement AN-1. While the wild-type plasmid complements the growth of the null mutant, neither UL12 mutant can do so. Loss of exonuclease activity therefore correlates with loss ofin vivofunction
Evidence That both HIV and HIV-Induced Immunodeficiency Enhance HCV Replication among HCV Seroconverters

AbstractThe objective of this retrospective cohort study is to assess the mechanism by which human immunodeficiency virus type 1 (HIV) influences hepatitis C virus (HCV) replication in injecting drug users. Virological (HCV and HIV RNA levels) and immunological (CD4+, CD8+cell counts, and anti-CD3 reactivity) parameters were determined in 19 HCV seroconverters in sequential samples over a period of 1 to 9 years. Among these subjects, 10 were HIV-seronegative (HIVneg), 4 were HIV-seropositive (HIVpos), and 5 seroconverted for HIV (HIVsc) during the observation period. HCV RNA levels were higher in HIVpos subjects than in HIVneg subjects. In subjects seroconverting for HIV, HCV RNA levels increased significantly immediately after HIV seroconversion (P< 0.0001), while they remained stable over time in HIVpos and HIVneg subjects. HCV RNA correlated inversely with CD4+cell counts in both the HIVpos population (R= −0.22,P< 0.05) and the HIVneg population (R= −0.45,P< 0.0001). In addition, when subjects were stratified according to CD4+cell counts a significant difference was found in HCV RNA levels between HIVpos and HIVneg subjects with CD4+cell counts >500 cells/μl (P= 0.001), but not in the population with CD4+cell counts <500 cells/μl. In no population was a correlation found between HCV RNA levels and CD8+cell counts or anti-CD3 reactivity. Both HIV infection and CD4+cell counts are apparently associated with HCV RNA levels. The direct association, independent of CD4+cell counts, between HIV infection and HCV replication appears to be stronger than the association between HIV-induced CD4+cell decline and HCV replication. We conclude that (i) HCV replication is in some way directly influenced by the presence of HIV; (ii) HCV-specific host immunity controls, in part, HCV replication; and (iii) HCV replication increases when the immune system is impaired by HIV
Tomato Spotted WiltTospovirusGenome Reassortment and Genome Segment-Specific Adaptation

AbstractA system to associate specific genome segments with viral phenotypes and to study factors influencing genome reassortment was developed for tomato spotted wiltTospovirus(TSWV). Reassortant isolates were generated by co-inoculating a TSWV isolate, TSWV-D, with TSWV-10 or TSWV-MD. The parental origin of each genome segment in putative reassortant isolates was determined by segment-specific restriction fragment length polymorphisms. The TSWV isolates readily exchanged genome segments in a nonrandom fashion. The S RNA from TSWV-D was dominant over the S RNA from TSWV-10. The intergenic region (IGR) of the S RNA was correlated with competitiveness of this genome segment in reassortant isolates. The less competitive S RNA contained a net increase of 62 nt, including a 33-nt duplication in the IGR. This duplicate sequence was highly conserved among isolates from the southeastern United States and an isolate from Bulgaria. The evidence supports the hypothesis that the IGR of the S RNA with an ambisense coding strategy serves a regulatory function which influences the occurrence of this segment in the viral population. In addition, it was demonstrated that stable parental phenotypes can be mapped to specific genome segments as well as generating novel phenotypes not associated with either parent
Ultrastructural and Temporal Observations of the Potyvirus Cylindrical Inclusions (CIs) Show That the CI Protein Acts Transiently in Aiding Virus Movement

AbstractA systematic ultrastructural study across the edge of an advancing infection in pea seed-borne mosaic potyvirus-infected pea cotyledons showed the cylindrical inclusion (CI) protein to exist in transient functional states. Initially, the characteristic CI pinwheel inclusion bodies were positioned centrally over the plasmodesmal apertures (including those of plasmodesmata connected to the previously infected cell), in agreement with a proposed role in virus movement (Carringtonet al.,1998,Plant J.,13, in press). The viral coat protein was associated with these structures and was seen within the modified plasmodesma, most notably in a continuous channel that passed along the axis of the pinwheel and through the plasmodesma. The CI protein was not detected within the plasmodesmal cavities. Later in the infection (i.e., behind the zone of active virus replication) the CI was no longer associated with cell walls, or with coat protein, and showed signs of structural degeneration. In contrast, the coat protein remained within plasmodesmal cavities. The role of the CI in assisting virus movement is not known but the presence of the CI was linked with an apparent transient reduction in callose in the vicinity of the plasmodesmata
The Tyrosine-17 Residue of Nef in SIVsmmPBj14 Is Required for Acute Pathogenesis and Contributes to Replication in Macrophages

AbstractThe variant simian immunodeficiency virus termed SIVsmmPBj14 induces a rapidly fatal disease in pig-tailed macaques. The acute pathogenic effects of this virus appear to be associated with at least twoin vitrocharacteristics: the ability to induce lymphocyte proliferation; and the ability to replicate in unstimulated PBMC. Two of the amino acids in Nef of PBj14 (the No. 17 residue, tyrosine, and the No. 18 residue, glutamic acid) appear to be linked to the virus' ability to induce lymphocyte activation. To further study the effects of these amino acids on PBj14-induced pathogenesis, we generated two mutant viruses from our molecular clone, PBj6.6, containing either changes in both the No. 17 and No. 18 residues (termed PBj6.6YE-RQ), or a single change in the No. 17 residue (termed PBj6.6Y-R).In vitroanalyses of these viruses showed that while their replicative abilities in stimulated peripheral blood mononuclear cells (PBMC) were altered, they still maintained the ability to replicate in unstimulated PBMC. Replication of these viruses in macrophage populations was impaired relative to the wild-type virus. Both mutant viruses were unable to induce proliferation of macaque PBMCin vitro.Virus derived from PBj6.6Y-R was unable to induce acute disease in macaques, but did maintain the ability to induce lymphopenia and intestinal lymphoid hyperplasia. These results show that the tyrosine-17 residue of Nef is linked to lymphocyte proliferation and disease development, but also suggest that the pathogenic characteristics of SIVsmmPBj14 are dependent upon multiple genetic determinants
Adaptation of Theiler's Virus to L929 Cells: Mutations in the Putative Receptor Binding Site on the Capsid Map to Neutralization Sites and Modulate Viral Persistence

AbstractPersistent strains of Theiler's virus, a murine picornavirus, produce a life-long infection of the central nervous system of the mouse and induce a chronic demyelinating disease. Strain DA1, a molecular clone of such a persistent strain, produces a prominent cytopathic effect in BHK-21 cells but is less efficient at infecting L929 cells. We cloned the cDNA of a derivative of virus DA1, adapted to promote a rapid cytopathic effect in L929 cells. Adaptation of the new variant (named KJ6) to L929 cells correlated with an enhanced viral entry rather than with an increased replication rate of the genome. Mutations responsible for L929 cells adaptation occurred in amino acids exposed at the surface of the capsid, in the CD loop of VP1 (100–102) and in the EF loop of VP2 (162–171–173), suggesting that these residues could be involved in receptor recognition. These two clusters of amino acids are precisely known to be part of neutralization epitopes. They also differentiate persistent from neurovirulent strains of Theiler's virus. Adaptation of the virus to L929 cells was accompanied by attenuation of its virulence for the mouse. Taken together, these data suggest a close relationship between receptor binding, virus neutralization, and virus phenotype
Adenovirus Death Protein, a Transmembrane Protein Encoded in the E3 Region, Is Palmitoylated at the Cytoplasmic Tail

AbstractThe 11.6-K protein of human adenovirus 2 (Ad2), which was recently renamed as adenovirus death protein (ADP), is a type III membrane glycoprotein that ultimately localizes to the nuclear membrane. ADP is encoded in the E3 transcription unit of Ad2 and migrates as a set of multiple bands in SDS–PAGE with three major forms. The corresponding gene product of adenovirus 5 (Ad5) has a slightly lower molecular weight and shows the same pattern in SDS–PAGE. We report here the covalent attachment of fatty acids to cysteine residues of ADP. In the case of Ad5-ADP all three major forms of this protein can be labeled by [3H]palmitic acid, but not by [3H]myristic acid, whereas only two [3H]palmitic acid-labeled Ad2-ADP species could be detected. The label is sensitive to treatment with 1 M hydroxylamine at pH 7 and with 20% β-mercaptoethanol indicating that the fatty acids are linked via a thioester bond. By thin layer chromatography, the vast majority of the incorporated label was identified as palmitic acid. Two cysteine residues at the boundary between transmembrane domain and cytoplasmic tail which could serve as acceptor sites were mutated to alanine residues by site-directed mutagenesis of the cloned Ad5-ADP gene. Expression of wild-type Ad5-ADP and the resulting mutants was performed in HeLa cells using the vaccinia virus T7 expression system. As demonstrated by labeling with [3H]palmitic acid, only the mutants with one remaining cysteine residue in the cytoplasmic tail were able to incorporate [3H]palmitic acid, indicating that either could serve as acceptor site. In contrast the double cysteine mutant could not be labeled by [3H]palmitic acid, clearly demonstrating that cysteines 53 and 54 are required for palmitoylation and probably represent the palmitoylation sites in Ad5-ADP
Identification of Transcripts and Promoter Regions of Ovine Adenovirus OAV287

AbstractThe ovine adenovirus isolate OAV287 represents a new group of adenoviruses that are distinct from the Mast- and Aviadenoviruses by several criteria, including genome arrangement. The OAV major late promoter and some late transcripts were previously mapped. To better define the probable coding sequences and to identify the approximate location of early promoters a partial transcription map of the genome was elucidated using a PCR-based approach. This was possible because the complete nucleotide sequence of the genome was known. The strategy permitted the identification of transcription start sites and RNA splice junctions and allowed the approximate location of promoters in the lefthand end, IVa2, E2, P32K, and E4 regions to be deduced. The data showed that lefthand end and E4 regions are controlled by three and two temporally distinct promoters, respectively. The E2 region is controlled by a single promoter, in contrast to Mastadenoviruses, where E2 expression is controlled by the E2A and E2B promoters. The p32kDa structural protein at the lefthand end and the IVa2protein are also expressed from their own promoters. These data contribute to the first overview of transcription from a non-Mastadenovirusgenome
Conversion of VP1 to VPg in Cells Infected by Infectious Pancreatic Necrosis Virus

AbstractThe RNA polymerase VP1 of IPNV (a bisegmented dsRNA containing virus) is present in the virion both as a free polypeptide and as a genome-linked protein (VPg). Virion VP1 primes viral RNA synthesisin vitro(P. Dobos, 1995,Virology208, 19–25), and here we present data which suggest that protein-primed RNA synthesis may also take place in infectious pancreatic necrosis virus (IPNV)-infected cells. Anti-VP1 serum immunoprecipitated several polypeptides larger than the 94-kDa VP1 of IPNV from [35S]methionine-labeled infected cell lysates. During denaturing, SDS–polyacrylamide gel electrophoresis these polypeptides formed a characteristic “ladder” which was resistant to alkaline phosphatase but sensitive to RNases, indicating that it consisted of VP1 polypeptides with oligoribonucleotides of various lengths attached to them. Probing the ladder with 5′ and 3′ end-specific, as well as plus-, or minus-strand-specific oligonucleotides revealed that they represent VP1 linked to 5′ terminal sequences of genome segment A- and B-specific plus strands. Pulse–chase experiments in combination with two-dimensional polyacrylamide gel electrophoresis revealed that labeled VP1 could be chased to replicative intermediate, to ssRNA, to dsRNA, and eventually to virion VPg-dsRNA and that VP1 could be released from all these structures by RNase treatment. We suggest that these results are most compatible with the model where a VP1-pN structure acts as a primer for viral RNA synthesisin vivo,a mechanism that has been shown to occurin vitro
HHR23A, the Human Homologue of the Yeast Repair Protein RAD23, Interacts Specifically with Vpr Protein and Prevents Cell Cycle Arrest but Not the Transcriptional Effects of Vpr

AbstractYeast two-hybrid selection of proteins interacting with human immunodeficiency virus type 1 Vpr identified HHR23A, a human homologue of the yeast DNA repair protein RAD23, as a specific interactor. A small 57-amino-acid C-terminal portion of HHR23A was sufficient for Vpr interaction. When introduced into human cells by transfection, full-length HHR23A or its C-terminal fragments were able to alleviate Vpr-induced cell cycle arrest, suggesting that HHR23A may participate in the pathway leading to G2 arrest by Vpr. We have also examined the effects of HHR23 on the recently identified transcription coactivator function of Vpr. The two Vpr functions are independent, since we have identified mutants lacking either the cell cycle arrest or the coactivator function. Our analysis showed that excess of HHR23A does not affect the coactivator function of Vpr, while it affects the cell cycle arresting function. Therefore, a simple sequestering model for Vpr in the presence of excess HHR23A is not supported. We propose that the interaction of HHR23A with Vpr may affect specifically pathways leading to cell cycle regulation
Different HTLV-I Neutralization Patterns among Sera of Patients Infected with Cosmopolitan HTLV-I

AbstractTo determine if sequence variations observed in cosmopolitan HTLV-I interfered with viral recognition by neutralizing antibodies, we evaluated the neutralization potential of sera from persons infected by HTLV-I of this clade selected for amino acid changes in their env glycoproteins. Each serum was used to neutralize three previously described HTLV-I isolates, 2060, 2072, and 1010, that possess amino acid env sequences differing at several positions, one of them being located in the immunodominant and neutralizable domain (aa 187–199). The results obtained in syncytia and/or reporter gene inhibition assays showed that the neutralization pattern of the sera clearly differed and could be classified in three categories. Five sera completely neutralized the three viruses with an equivalent titer, two sera gave a maximum inhibition, with higher ID50on the 2072 virus than on the 2060 or 1010 viruses, and three sera had a stronger neutralization potential toward the 1010 virus than toward the 2060 virus. One of these sera partially neutralized the virus produced by 2072 cells, whereas neutralizing antibodies in the other two recognized the neutralizable epitopes on the 1010 or 2072 viruses equally well. Identification of amino acid sequences involved in induction of neutralizing antibodies with different recognition capacities could help identify new neutralizable epitopes of HTLV-I envelope glycoproteins and to better define the component(s) of an effective vaccine
Transmission and Propagation in Cell Culture of Virus Produced by Cells Transfected with an Infectious Molecular Clone of Bovine Leukemia Virus

AbstractA full-length molecular clone of bovine leukemia virus (BLV) pBLV-IF with two copies of a long terminal repeat (LTR) was constructed from a previously isolated, covalently closed, circular DNA clone, pB6490, that has one copy of the LTR and the pX region split at anEcoRI site. This molecular clone directed the synthesis of viral proteins and the induction of syncytia in transiently transfected cells. In addition, virus particles were released into the culture medium. Serial passages of transient transfectants also resulted in propagation of BLV. After transfection of five cell lines with linearized pBLV-IF and a neomycin-resistance gene, BLV-producing transfectants were established in cell lines COS-1 and 23CLN that did not form syncytia upon expression of BLV. In HeLa and FLK cells, BLV produced by a stable COS-1 transfectant was transmitted by both cell-free and cell-to-cell infection. Thus, pBLV-IF encoded an infectious provirus that successfully induced primary and secondary infections. This study indicates that the infectious molecular clone and the virus-producing transfectants could be useful for further examination of the biological properties of BLV