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Volume 67,
Issue 7,
1986
Volume 67, Issue 7, 1986
- Animal
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Characterization of Glycoprotein Complexes Present in Human Cytomegalovirus Envelopes
More LessSummaryThree disulphide cross-bridged glycoprotein complexes were immunoprecipitated from purified human cytomegalovirus envelopes using a monoclonal antibody with a specificity for a glycoprotein of mol. wt. 52 × 103. These complexes were isolated by electroelution after polyacrylamide gel electrophoresis (PAGE) under non-reducing conditions. Compositional analysis of each complex by PAGE under reducing conditions showed that at least two distinct complexes,one containing glycoproteins with mol. wt. of 52 × 103 and 95 × 103 and the other with glycoproteins of 52 × 103 and 130 × 103, were present. The results obtained indicated that one of these complexes could also exist as a dimer.
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Fc Receptor(s) Induced by Human Cytomegalovirus Bind Differentially with Human Immunoglobulin G Subclasses
More LessSummaryThe IgG subclass specificity of Fc receptor(s) induced on cells by infection with human cytomegalovirus (HCMV) was studied in a binding assay by using infected cells and purified iodinated IgG of various subclasses from HCMV seronegative healthy adult donors. All four human IgG subclasses bound to HCMV-infected cells, with the following relative magnitudes: IgG1 ⩾ IgG4 > IgG2 > IgG3. The IgG subclass specificity of the Fc receptor was further analysed in an inhibition assay by using fragments prepared from purified human IgG by papain digestion, and using unlabelled subclass proteins. Fc but not Fab fragments inhibited the binding of 125I-labelled human IgG to HCMV-infected cells. The biological role of the Fc receptor in HCMV infection is discussed.
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A Neutralizing Monoclonal Antibody to Respiratory Syncytial Virus which Binds to Both F1 and F2 Components of the Fusion Protein
More LessSummaryA virus-neutralizing monoclonal antibody (1E3) specifically immunoprecipitated the 70000 mol. wt. (70K) fusion (F) protein from respiratory syncytial (RS) virus-infected HeLa cells. Western blotting analysis of polypeptides from such cells separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions revealed that 1E3 was peculiar in that it bound to both F1 (50K) and F2 (20K) components of the F protein. Antibody subsequently eluted from either the F1 or the F2 regions of immunoblots re-bound to both F1 and F2 regions of the SDS-PAGE blot. These results show that monoclonal antibody 1E3 reacts with an epitope which is found on both F1 and F2 subunits of RS virus fusion protein.
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Persistence of BK Virus in Human Foetal Pancreas Cells
More LessSummaryHigh multiplicity BK virus (BKV) infection of primary cells derived from human foetal pancreas resulted in massive cytopathology and subsequent outgrowth of cells. Intranuclear BKV T-antigen was present in all cells and viral antigen was detected in 10 to 30% of these cells. The subcultured cells yielded BKV in the supernatant (approx. 105 TCID50/ml) and in the cells free viral DNA was present (approx. 10% of total cellular DNA content). Analysis of the viral DNA indicated the presence of deleted and rearranged BKV DNA molecules. Although all cells continuously expressed BKV T-antigen they did not exhibit the transformed phenotype. This persistent infection of human foetal pancreas cells represents a novel type of in vitro interaction between BKV and human cells which may correspond to the in vivo findings on BKV tropism for pancreatic cells.
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- Plant
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Comparison of the in vitro Translation Products of Wild-type and a Deletion Mutant of Soil-borne Wheat Mosaic Virus
More LessSummaryA wild-type isolate (WT) of soil-borne wheat mosaic virus (SBWMV, isolate JT) (RNA I 6.9 kb and RNA II 3.6 kb) was successively transferred to wheat plants by mechanical inoculation and a deletion mutant (DM) (RNA I 6.9 kb and RNA II 2.1 kb) was produced. In wheat germ extracts, RNA I of both WT and DM directed synthesis of polypeptides having mol. wt. of 220000 (220K) and 150K, and RNA II of both WT and DM directed synthesis of 25K and 19K polypeptides. Incubation in wheat germ extracts of WT or DM virions purified with an alkaline buffer also gave 220K, 25K and 19K polypeptides as major products and a 150K polypeptide as a minor product. In rabbit reticulocyte lysates, RNA I of both WT and DM directed synthesis of only the 220K polypeptide, whereas WT RNA II produced 100K, 46K, 25K and 19K polypeptides and DM RNA II, 31K, 25K and 19K polypeptides. SBWMV DM antiserum precipitated polypeptides of 100K, 31K, 25K and 19K but not 220K and 46K. The 19K polypeptide was identified as the capsid protein from its electrophoretic mobility and its reaction with antiserum to virus particles. Thus, the differences between the in vitro translation products of WT and DM SBWMV were confined only to those coded by RNA II.
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An RNA-dependent RNA Polymerase Associated with the Filamentous Nucleoproteins of Rice Stripe Virus
More LessSummaryFilamentous particles of rice stripe virus (RSV) were found to be associated with an RNA-dependent RNA polymerase activity. The enzyme catalysed the synthesis in vitro of single- and double-stranded RNAs corresponding in size with RNA found in RSV particles, as well as two other dsRNAs with mol. wt. of 1.0 × 106 and 0.56 × 106. The RSV polymerase activity required Mg2+, Mn2+ or Fe2+ but not Na+, K+ or NH+ 4 which were inhibitory at concentrations of more than 50 mm. The optimum pH of the polymerase was around 8.0 at the optimum temperature, 40°C. Detergent treatment of RSV particles did not enhance the polymerase activity. Purified RSV particles contained a minor polypeptide (mol. wt. 230000) as well as coat protein; the former may be the RSV polymerase. The taxonomy of RSV is discussed in relation to enveloped animal viruses with negative-stranded RNA, which also contain an RNA-dependent RNA polymerase in virus particles.
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The Nucleotide Sequence of Tomato Black Ring Virus RNA-2
More LessSummaryThe sequence of the 4662 nucleotides [excluding poly(A)] of RNA-2 of tomato black ring virus (TBRV) has been determined. Most of the sequence (4074 nucleotides) encodes a polypeptide of mol. wt. 150000 (150K). The 5′ and 3′ non-coding sequences are 287 and 301 nucleotides in length, differ from the coding sequence in base composition, and contain repeated sequences, some of which resemble oligonucleotides in the 3′ non-coding sequence of M-RNA of cowpea mosaic virus (CPMV). From its amino acid composition, the coat protein of TBRV was tentatively located in the C-terminal third of the 150K polypeptide. The amino acid sequence of the 150K polypeptide immediately N-terminal to the putative coat protein sequence was found to resemble parts of the 30K polypeptides of tobamoviruses and, to a lesser extent, part of the 105K polypeptide translation product of CPMV M-RNA.
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Serotype-specific and General Luteovirus Probes from Cloned cDNA Sequences of Barley Yellow Dwarf Virus
More LessSummaryGenome fragments from two serotypes of barley yellow dwarf virus (BYDV) were cloned and used as hybridization probes. Dot blot assays using 32P-labelled plasmid probes readily detected BYDV in crude sap extracts from infected plants and were at least as sensitive as ELISA. Some clones were specific for either the RPV or the PAV serotypes. Others hybridized not only with the RNA of both serotypes but also with those of three other luteoviruses (beet western yellows, potato leaf roll and soybean dwarf), but they did not hybridize with the RNA of a tobamovirus or Escherichia coli tRNA. These nucleic acid probes therefore have potential for the general diagnosis of infection by members of the luteovirus group as well as detection of specific BYDV serotype infections. The taxonomic implications of the observation that the genomes of different luteoviruses have some conserved regions are discused.
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Infectious and Non-infectious Mutants of Cauliflower Mosaic Virus DNA
SummaryMutants of cauliflower mosaic virus (CaMV), generated in vitro by modification of recombinant DNA plasmids containing the viral genome, either retained the ability to induce disease symptoms on turnip plants, produced less severe symptoms or failed to induce symptoms. Wild-type symptoms were produced by a variant CaMV DNA of the Cabbage S isolate that had 4 bp in open reading frame (ORF) III replaced with a 16 bp sequence. Less severe symptoms, due to a delay in symptom appearance relative to inoculation with wild-type DNA, were induced by a mutant with a frameshift mutation in ORF II (pSA103). CaMV DNA, recovered from plants infected with pSA103, contained a second mutation which restored the original translation reading frame. Nucleic acid hybridization to ‘squishes’ of leaf tissue from plants that had been inoculated with mutant DNAs that included DNAs modified in each of the six major ORFs of CaMV DNA revealed that only those plants that appeared diseased had detectable CaMV nucleic acid in uninoculated leaves. Replicated CaMV DNA was also not detected in non-encapsidated and virion DNA fractions from inoculated leaves of non-diseased plants.
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