bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification and nucleotide sequences of two similar tandem direct repeats in Epstein-Barr virus DNA
Authors: Dambaugh T R , Kieff E
Published: January 2020

Epstein-Barr virus DNA is known to have partially homologous segments, designated DL and DR, near the left and right ends of the long unique region (Raab-Traub et al., Cell 22:257-267, 1980). DL and DR are each partially composed of tandem direct repeat sequences. DL contains 11 to 14 repeats of a 124-base-pair sequence designated IR2. DR contains approximately 30 direct repeats of a 103-base-pair sequence designated IR4. The DL and DR sequences have colinear partial homology for approximately 2.4 and 1.5 kilobase pairs to the right of IR2 and IR4, respectively. IR2 and IR4 are similar sequences and evolved in part from a common ancestor. Both sequences are 84% guanine and cytosine and have limited homology to Epstein-Barr virus IR1 and to the herpes simplex virus type 1 inverted terminal repeat "a" sequence. IR2 encodes part of an abundant 2.5-kilobase persistent early EBV RNA expressed in productively infected cells, but does not encode part of the 3-kilobase Epstein-Barr virus RNA which is transcribed from the adjacent IR1-U2 region of the Epstein-Barr virus genome in latently infected cells.
Novel forms of woodchuck hepatitis virus DNA isolated from chronically infected woodchuck liver nuclei
Authors: Rogler C E , Summers J
Published: January 2020

We cloned several unique forms of woodchuck hepatitis virus, a DNA virus closely related to hepatitis B virus, from a chronically infected woodchuck liver. Each of the three clones contained more than two genome equivalents of viral sequences with extensive rearrangements and no detectable cellular sequences. From the frequency by which they were isolated from a library of recombinant clones, we estimate that they are present in approximately one copy per cell. Of a total of 11 sites at which rearrangements were mapped in the clones, 10 occurred between segments of opposite polarity, and 1 occurred between segments of the same polarity. The possible significance of these findings to the persistence of virus production in infected cells is discussed.
Development of a mouse mammary tumor virus-negative mouse strain: a new system for the study of mammary carcinogenesis
Authors: Cohen J C , Traina V L , Breznik T , Gardner M
Published: January 2020

All inbred strains of mice transmit one or more copies of mouse mammary tumor virus (MMTV) DNA integrated as proviral sequences. This complicates efforts to define viral-induced mammary carcinogenesis. Here we report the use of surgical nonlethal splenectomy in tissue typing mice and the development of an MMTV-negative mouse strain. The MMTV-negative strain allows study of the involvement of non-MMTV genes in mammary carcinogenesis. In addition, it can be used as a sterile background into which MMTV variants can be introduced. Through the techniques described here, mice containing single MMTV loci or specific combinations can be specially chosen and rapidly developed. In this manner, the oncogenecity of particular MMTV variants may be assessed.
Translational control of vesicular stomatitis virus protein synthesis: isolation of an mRNA-sequestering particle
Authors: Rosen C A , Ennis H L , Cohen P S
Published: January 2020

An mRNA-ribonucleoprotein particle (mRNP) was found in vesicular stomatitis virus (VSV)-infected Chinese hamster ovary cells. The particle was present 3 and 4.5 h after infection but was barely discernible at 2 h. The mRNP (buoyant density, 1.56 g/cm3), which cosedimented with viral nucleocapsid in a sucrose density gradient at approximately 120 to 160S, was separable from nucleocapsid (buoyant density, 1.31 g/cm3) by CsCl density gradient centrifugation. It contained all five VSV mRNAs and, almost exclusively, viral N protein. Some host mRNA and host protein was also present in the particle. The intact mRNP was incapable of stimulating protein synthesis in an in vitro protein-synthesizing system, although the VSV mRNA isolated from the particle by phenol extraction was functional in vitro. In contrast, intact polysomes stimulated cell-free protein synthesis to the same extent as purified polysomal mRNA. By 4.5 h after infection, 97% of the functional mRNA in vivo was associated with the mRNP, and only 3% was on polysomes. The amount of polysomal mRNA at 4.5 h after infection was only 31% of that found at 2 h after infection; this was reflected by the 76% decrease observed in the rate of in vivo protein synthesis at 4.5 h relative to that found at 2 h. Thus, it appears that the mRNP serves as an organelle which sequesters the large excess of VSV mRNA that is normally made during secondary transcription.
Alteration in the simian virus 40 maturation pathway after butyrate-induced hyperacetylation of histones
Authors: Roman A
Published: January 2020

The role of histone acetylation in the replication and maturation pathways of simian virus 40 was assessed. Histones were hyperacetylated by briefly exposing infected cells to sodium butyrate. Viral DNA in cells exposed to butyrate was found to reenter replication to a greater extent and mature to the previrion form to a lesser extent than viral DNA in control cells. Previrions formed in the presence of butyrate had altered sedimentation properties. These data suggest that increased acetylation of histones is not the signal for removal of DNA from the pool of molecules available for replication. It appears, in fact, that hyperacetylation retards entry into and progression along the maturation pathway.
Transcription of herpes simplex virus genes in vivo: overlap of a late promoter with the 3' end of the early thymidine kinase gene
Authors: Sharp J A , Wagner M J , Summers W C
Published: January 2020

We identified in herpes simplex virus type 1-infected cells six cytoplasmic transcripts which were complementary to BamHI restriction endonuclease fragment Q. Two transcripts appeared in major amounts compared with the other four. One major transcript of about 1.4 kilobases was the mRNA for the viral thymidine kinase, was synthesized at intermediate times, and was classified as a beta transcript. The other major transcript was synthesized at late times and was classified as a gamma transcript. This late transcript was about 3 kilonucleotides long and was transcribed in the same direction as the gene for thymidine kinase. The 5' end of this late RNA was located by RNA sequence analysis and was 23 nucleotides downstream from the polyadenylation site for the thymidine kinase mRNA. This finding led to the conclusion that the control region for the 3-kilobase gamma transcript is contained within the 3' untranslated region of the thymidine kinase transcript.
Cell-free synthesis and assembly of vesicular stomatitis virus nucleocapsids
Authors: Patton J T , Davis N L , Wertz G W
Published: January 2020

The association of newly synthesized vesicular stomatitis virus proteins into nucleocapsid structures was examined in a cell-free system that supports concurrent viral protein synthesis, transcription, and RNA replication. The vesicular stomatitis virus proteins synthesized by this system associated with the newly replicated RNA to form structures that banded in CsCl gradients with marker nucleocapsids. In reactions lacking nucleocapsid templates to program RNA synthesis, the newly synthesized proteins did not associate into nucleocapsid structures. The newly synthesized proteins associated with nucleocapsids were analyzed by electrophoresis on polyacrylamide gels containing sodium dodecyl sulfate after separation from non-associated proteins by chromatography on Bio-Gel A15M agarose columns. The results of this analysis showed that newly synthesized L, NS, and N proteins associated into nucleocapsids in the in vitro system. In addition, a small amount of newly synthesized M protein was stably bound to the nucleocapsids. The molar ratio of the associated, newly synthesized proteins was 2:350:1,000:10 (L:NS:N:M). More than 90% of the newly synthesized NS protein that associated with nucleocapsids in vitro was of the NS2 subspecies, as assayed by DEAE-cellulose column chromatography. The stability of the association of the newly synthesized proteins with nucleocapsids in the system mimicked that of the association of viral proteins with nucleocapsids from infected cells as measured by salt sensitivity. These data indicate that nucleocapsids were assembled from newly synthesized proteins within our in vitro system and that the molar ratio of assembled proteins was similar to that observed for virion nucleocapsids.
Parvovirus genome: nucleotide sequence of H-1 and mapping of its genes by hybrid-arrested translation
Authors: Rhode S L , Paradiso P R
Published: January 2020

The nucleotide sequence of the parvovirus H-1 has been determined by the chain-terminating method of Sanger. The sequence is 5,176 nucleotides long. Two large open reading frames (1 and 2) and two smaller open reading frames (3 and 4) of potential importance were identified in the plus-strand sequence. Promoter sequences are located at map positions 4 and 38 when map positions are expressed as percent of genome length from the 3' end of the virion minus strand. The locations for the genes for the parvovirus capsid proteins and a 76,000-dalton noncapsid protein (NCVP1) were mapped by hybrid-arrested translation. The gene for the capsid proteins VP1 and VP2' is located in the 5' half of the virus genome. The gene for NCVP1 is located in the 3' half of the viral DNA.
Genetic analysis of temperature-sensitive mutants which define the genes for the major herpes simplex virus type 2 DNA-binding protein and a new late function
Authors: Dixon R A , Sabourin D J , Schaffer P A
Published: January 2020

Eleven temperature-sensitive mutants of herpes simplex virus type 2 (HSV-2) exhibit overlapping patterns of complementation that define four functional groups. Recombination tests confirmed the assignment of mutants to complementation groups 1 through 4 and permitted the four groups to be ordered in an unambiguous linear array. Combined recombination and marker rescue tests (A. E. Spang, P. J. Godowski, and D. M. Knipe, J. Virol. 45:332-342, 1983) indicate that the mutations lie in a tight cluster near the center of UL to the left of the gene for DNA polymerase in the order 4-3-2-1-polymerase. The seven mutants that make up groups 1 and 2 fail to complement each other and mutants in HSV-1 complementation group 1-1, the group thought to define the structural gene for the major HSV-1 DNA-binding protein with a molecular weight of 130,000. At 38 degrees C, mutants in groups 1 and 2 synthesize little or no viral DNA, and unlike cells infected with the wild-type virus, mutant-infected cells exhibit no detectable nuclear antigen reactive with monoclonal or polypeptide-specific antibody to the major HSV-2 DNA-binding protein. The four mutants that make up groups 3 and 4 do not complement each other, nor do they complement mutants in group 2. They do, however, complement mutants in group 1 as well as representative mutants of HSV-1 complementation group 1-1. At 38 degrees C, mutants in groups 3 and 4 are phenotypically DNA+, and nuclei of mutant-infected cells contain the HSV-2 DNA-binding protein. Thus, the four functional groups appear to define two closely linked genes, one encoding an early viral function affecting both viral DNA synthesis and expression of the DNA-binding protein with a molecular weight of 130,000 (groups 1 and 2), and the other encoding a previously unidentified late viral function (groups 3 and 4). The former gene presumably represents the structural gene for the major HSV-2 DNA-binding protein.
Genomic structure of human polyoma virus JC: nucleotide sequence of the region containing replication origin and small-T-antigen gene
Authors: Miyamura T , Jikuya H , Soeda E , Yoshiike K
Published: January 2020

The nucleotide sequence of the region of human polyoma virus JC DNA between 0.5 and 0.7 map units from a unique EcoRI cleavage site was determined and compared with those of the corresponding regions of another human polyoma virus, BK, and simian virus 40 DNAs. Within this region consisting of 945 base pairs, we located the origin of DNA replication near 0.7 map units, the entire coding region for small T antigen, and the splice junctions for large-T-antigen mRNA. The deduced amino acid sequences for small T antigen and the part of large T antigen markedly resembled those of polyoma virus BK and simian virus 40. The results strongly suggest that polyoma virus JC has the same organization of early genome as polyoma virus BK and simian virus 40 on the physical map, with the EcoRI site as a reference point.
Polyoma virus minichromosomes: poly ADP-ribosylation of associated chromatin proteins
Authors: Prieto-Soto A , Gourlie B , Miwa M , Pigiet V , Sugimura T , Malik N , Smulson M
Published: January 2020

The host nuclear enzyme poly(ADP-ribose) polymerase has been shown to be associated with the replicative intermediate and mature forms of polyoma virus minichromosomes. Minichromosome-associated histones H2A and H2B as well as several nonhistone proteins were poly ADP-ribosylated by endogenous poly(ADP-ribose) polymerase. In addition, minichromosome fractions catalyzed the formation in vitro of dimers of endogenous histone H1 linked by poly(ADP-ribose). Poly ADP-ribosylated polyoma virus minichromosome chromatin labeled in vivo with [3H]thymidine could be retained and eluted from anti-poly(ADP-ribose) immunoglobulin G-Sepharose. Pulse-labeled replicative intermediate minichromosomes were retained better on the antibody columns than were mature minichromosomes labeled for 2.5 h. The possible role of poly ADP-ribosylation of viral nucleosomes during polyoma replication or transcription is discussed.
Structural and nonstructural proteins of a rabbit parvovirus
Authors: Matsunaga Y , Matsuno S
Published: January 2020

The structural and nonstructural polypeptides of a rabbit parvovirus (RPV) (F-7-9 strain) were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The virion contained three polypeptide components, A (molecular weight, 96,000), B (85,000), and C (75,000). A part of the polypeptide C was cleaved into the smaller-molecular-weight polypeptide C' by proteolysis during purification steps. The major polypeptide C together with C' constituted about 87% of the total viral proteins, and the minor polypeptides, A and B, constituted 4 and 9%, respectively. The structural polypeptides of empty particles were similar in size and composition to those of the virion, but the content of the C' polypeptide was very low. When rabbit kidney cell cultures were infected with RPV, the C polypeptide was detected as early as 15 h postinfection, whereas A and B were first demonstrated at 18 h. The C' polypeptide was not detected for 44 h. In addition to the three structural polypeptides, at least three nonstructural polypeptides, E, F, and G, were demonstrated in the RPV-infected cells. Polypeptide E (molecular weight, 49,000), detected mostly in cytoplasm, seemed to be a cellular protein. The F (25,000) and G (22,000) polypeptides seemed to be virus-coded proteins since they were precipitated with the anti-RPV rabbit immunoglobulin. According to partial proteolysis and peptide mapping, the F and G polypeptides shared the same peptide components.
Genomic and copy-back 3' termini in Sendai virus defective interfering RNA species
Authors: Re G G , Gupta K C , Kingsbury D W
Published: January 2020

Direct sequencing of nine Sendai virus defective interfering RNA species revealed two kinds of 3'-terminal sequences. Six RNA species had 3' termini identical to the virus genome (negative strand), confirming that internal deletions are a frequent cause of Sendai virus defectiveness. The other three RNA species had 3'-terminal sequences identical to that described as the complement of the 5' terminus of the virus genome (R. A. Lazzarini, J. D. Keene, and M. Schubert, Cell 26:145-154, 1981), indicating that they are of the copy-back type. Extensive homology between these two types of 3' sequences evidently accounts for the ability of the copy-back sequence to function as an initiation signal for viral RNA replication. There may not be a selective advantage of one type of terminus over the other, since one defective interfering strain possessed two RNA species, one of which had the genomic 3' terminus and the other copy-back type.
Antigenic variants of herpes simplex virus selected with glycoprotein-specific monoclonal antibodies
Authors: Holland T C , Marlin S D , Levine M , Glorioso J
Published: January 2020

Monoclonal antibodies specific for herpes simplex virus type 1 (HSV-1) glycoproteins were used to demonstrate that HSV undergoes mutagen-induced and spontaneous antigenic variation. Hybridomas were produced by polyethylene glycol-mediated fusion of P3-X63-Ag8.653 myeloma cells with spleen cells from BALB/c mice infected with HSV-1 (strain KOS). Hybrid clones were screened for production of HSV-specific neutralizing antibody. The glycoprotein specificities of the antibodies were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of immunoprecipitates of radiolabeled infected-cell extracts. Seven hybridomas producing antibodies specific for gC, one for gB, and one for gD were characterized. All antibodies neutralized HSV-1 but not HSV-2. Two antibodies, one specific for gB and one specific for gC, were used to select viral variants resistant to neutralization by monoclonal antibody plus complement. Selections were made from untreated and bromodeoxyuridine- and nitrosoguanidine-mutagenized stocks of a plaque-purified isolate of strain KOS. After neutralization with monoclonal antibody plus complement, surviving virus was plaque purified by plating at limiting dilution and tested for resistance to neutralization with the selecting antibody. The frequency of neutralization-resistant antigenic variants selected with monoclonal antibody ranged from 4 X 10(-4) in nonmutagenized stocks to 1 X 10(-2) in mutagenized stocks. Four gC and four gB antigenic variants were isolated. Two variants resistant to neutralization by gC-specific antibodies failed to express gC, accounting for their resistant phenotype. The two other gC antigenic variants and the four gB variants expressed antigenically altered glycoproteins and were designated monoclonal-antibody-resistant, mar, mutants. The two mar C mutants were tested for resistance to neutralization with a panel of seven gC-specific monoclonal antibodies. The resulting patterns of resistance provided evidence for at least two antigenic sites on glycoprotein gC.
Genetic relatedness of the genomes of equine herpesvirus types 1, 2, and 3
Authors: Staczek J , Atherton S S , O'Callaghan D J
Published: January 2020

Genomic DNAs of equine herpesvirus type 1 (EHV-1), EHV-2 (equine cytomegalovirus), and EHV-3 were examined by reassociation kinetic and thermal denaturation analyses to determine the extent and degree of homology among the three viral DNAs. Results of reassociation analyses indicated a limited homology among the three EHV genomes. Homologous DNA sequences equivalent to 1.8 to 3.7 megadaltons between EHV-1 and equine cytomegalovirus, 7.6 to 8.2 megadaltons between EHV-1 and EHV-3, and 1.3 to 1.9 megadaltons between equine cytomegalovirus and EHV-3 were detected. Examination by thermal denaturation of the DNA homoduplexes and heteroduplexes formed during reassociation revealed a high degree of base pairing within the duplexes, suggesting that closely related sequences may be conserved among the genomes of EHV.
Identification of B-lymphotropic papovavirus-coded proteins
Authors: Segawa K , Takemoto K K
Published: January 2020

The lymphotropic papovavirus (LPV)-specific mRNAs were translated in vitro in rabbit reticulocyte lysates. The specific products were 84,000-dalton (84K), 41K, 35K, and 26K proteins. Immunoprecipitation with anti-LPV hamster sera and analysis of partially purified LPV virions showed that the last three proteins were the LPV capsid proteins, and we designated the 41K, 35K, and 26K proteins VP1 (major capsid protein), VP2, and VP3, respectively. Several characteristics, such as the small amount of mRNA for the 84K protein at late stages of infection, its absence from partially purified virus preparations, no common tryptic peptides between the 84K and 41K proteins, and the pattern of in vivo phosphorylation, suggest that the 84K protein is not a simple dimer of the 41K protein. Normal human sera and sera from certain leukemic patients positive for antibody to LPV viral antigens immunoprecipitated the 41K protein.
Genomic RNA of mengovirus. VI. Translation of its two cistrons in lysates of interferon-treated cells
Authors: Degener A M , Pagnotti P , Facchini J , Pérez-Bercoff R
Published: January 2020

The addition of low levels (40 ng/ml) of the synthetic double-stranded polyribonucleotide poly I:C to lysates of interferon-treated L-cells resulted in a strong inhibition (70 to 75%) of the in vitro translation of mengovirus RNA. Under these conditions, the rates of incorporation of [35S]methionine or formyl-[35S]methionine were depressed to a comparable extent. The sequences of mengovirus RNA recognized by ribosomes of interferon-treated cells at initiation of translation were compared with those present in initiation complexes formed by ribosomes of untreated controls. Fingerprint analysis revealed that the same sequences of mengovirus RNA were protected against nuclease attack by the 80S and the 40S initiation complexes formed in vitro in lysates of control or interferon-treated L-cells. Mengovirus RNA-coded proteins were labeled at their N-terminal end with formyl-[35S]methionine and digested to completion with trypsin. The resulting fragments were separated by high-voltage paper electrophoresis. Two different formyl-[35S]methionine-labeled N termini were resolved. Further analyses supported the notion that the two radioactive peaks originated in the initiation of translation at two different sites. This pattern did not change when mengovirus RNA was translated in lysates of interferon-treated cells.
Differential antiviral effects of interferon in three murine cell lines
Authors: Sen G C , Herz R E
Published: January 2020

Infectious leukemia virus production by two chronically infected NIH/MOL lines was strongly inhibited by interferon treatment of the cells. The corresponding degree of inhibition in JLSV-11 cells was much lower. Multiplication of encephalomyocarditis virus in all three cell lines was barely affected by interferon treatment. Replication of vesicular stomatitis virus, on the other hand, was highly sensitive to interferon in the JLSV-11 line and in one NIH/MOL line but was practically insensitive in the other NIH/MOL line. Anticellular actions of interferon were more pronounced in the JLSV-11 line than in the others. In response to interferon treatment, 2',5'-oligoadenylate synthetase activity was induced to a high level in JLSV-11 cells and to lower levels in the NIH/MOL lines. We failed to detect any 2',5'-oligoadenylate-dependent endonuclease activity in extracts of these cells. Double-stranded RNA-dependent protein kinase activity was present in extracts of interferon-treated NIH/MOL cells, but it was barely detectable in extracts of interferon-treated JLSV-11 cells. The above studies demonstrated that interferon could differentially affect the replication of three different viruses in three different cell lines, including two seemingly identical NIH/MOL lines, and that certain tentative conclusions can be drawn regarding the roles of different interferon-inducible enzyme markers in the different antiviral actions of interferons.
Protein-RNA interaction in encephalomyocarditis virus as revealed by UV light-induced covalent linkages
Authors: Smirnov Y A , Rodrigues-Molto M P , Famadas M T
Published: January 2020

UV irradiation of encephalomyocarditis virus led to an increase in the buoyant density of the virus in CsCl gradients from 1.34 to 1.46 g/cm3. Heat treatment of the irradiated virus (20 min at 54 degrees C) reduced the density to 1.40 g/cm3 and led to the loss of approximately 55% of the labeled RNA from the virions. The non-irradiated virions were converted by such heating into empty capsids. Irradiation also resulted in an increase in the accessibility of RNA inside the virions to the action of pancreatic RNase. An increase in the UV dose did not enlarge the fraction of RNA molecules covalently linked to protein; this was revealed by the lack of any secondary increase in the apparent RNase resistance of the labeled RNA in the irradiated virions. Destruction of the irradiated virus with sodium dodecyl sulfate and 2-mercaptoethanol allowed the isolation of a 40S structure containing viral RNA and RNA-linked proteins. The latter comprised no more than 2.5% of the whole protein content of the virion. Polyacrylamide gel electrophoretic analysis of the RNase-treated 40S structure revealed at least three viral structural proteins in the same ratio as was present in the intact virions.
Inhibition of vaccinia virus growth and virus-specific RNA synthesis by 3'-O-methyl adenosine and 3'-O-methyl guanosine
Authors: Goswami B B , Sharma O K
Published: January 2020

The 5' triphosphates of the methylated nucleoside analogs 3'-O-methyl adenosine and 3'-O-methyl guanosine are RNA chain terminators in vitro. However, anticellular or antiviral effects of 3'-O-methylated nucleosides or nucleotides have not been investigated. This is presumably because of the assumption that cellular kinases will be unable to phosphorylate the nucleosides. We report here that contrary to this assumption, 3'-O-methyl adenosine and to a lesser extent 3'-O-methyl guanosine are potent inhibitors of vaccinia virus growth in L-cells and Vero cells, without having a significant effect on cell growth at concentrations required to inhibit virus growth. Experiments revealed that early virus-specific RNA synthesis was preferentially inhibited by both 3'-O-methyl adenosine and 3'-O-methyl guanosine.
Further Evidence for the Protein Coding Potential of the Mouse Mammary Tumor Virus Long Terminal Repeat: Nucleotide Sequence of an Endogenous Proviral Long Terminal Repeat
Authors: Donehower Lawrence A. , Fleurdelys Barbara , Hager Gordon L.
Published: January 2020

The 3′ half of an endogenous mouse mammary tumor virus from a C3H mouse was cloned in the Charon 4A vector phage. A comparison of the proviral clone with previously published endogenous mouse mammary tumor virus restriction maps identified it as endogenous unit II (J. Cohen and H. Varmus, Nature [London]
278
:418-423, 1979), which is present in all inbred mouse strains derived from the original Bagg albino × DBA cross. The nucleotide sequence of the unit II long terminal redundancy (LTR) was determined and compared with the sequence previously determined for the exogenous C3H virus LTR (Donehower et al., J. Virol.
37
:226-238, 1981). Virtually all sequence differences between the two LTRs were base substitutions. The total amount of sequence divergence was 6.6%. The large open reading frame reported previously in the exogenous LTR was preserved in the endogenous LTR. In addition, the pattern of sequence divergence was highly nonrandom with respect to the putative amino acid codons of the two open reading frames. Most of the base substitutions in this region resulted in silent or conservative amino acid codon changes. The nonrandom divergence pattern indicates that selective forces are operating on this segment of DNA and argues that the putative protein is functional in the life cycle of mouse mammary tumor virus. Possible roles for the protein and its mode of expression are discussed.
Human Adenovirus Type 2 but Not Adenovirus Type 12 Is Mutagenic at the Hypoxanthine Phosphoribosyltransferase Locus of Cloned Rat Liver Epithelial Cells
Authors: Paraskeva Christos , Roberts Carl , Biggs Paul , Gallimore Phillip H.
Published: January 2020

Using resistance to the base analog 8-azaguanine as a genetic marker, we showed that adenovirus type 2, but not adenovirus type 12, is mutagenic at the hypoxanthine phosphoribosyltransferase locus of cloned diploid rat liver epithelial cells. Adenovirus type 2 increased the frequency of 8-azaguanine-resistant colonies by up to ninefold over the spontaneous frequency, depending on expression time and virus dose.
Structural and Intracellular Proteins of the Nonoccluded Baculovirus HZ-1
Authors: Burand John P. , Stiles Brad , Wood H. Alan
Published: January 2020

A plaque-purified isolate of the baculovirus HZ-1 was used to examine the kinetics of replication of this persistent, nonoccluded virus in TN-368 cells. Twenty-eight virus structural proteins ranging in molecular weight from 153,000 to 14,000 were identified. Fourteen of these proteins were found to be glycosylated. The sequence of appearance of the 37 virus-induced intracellular polypeptides was determined by pulse-labeling with [
35
S]methionine.
N
-[
3
H]acetylglucosamine, [
3
H]mannose, and the glycosylation inhibitor tunicamycin were used to detect virus structural glycoproteins. Post-transcriptional modification of two virus-induced proteins was detected.
Topography of simian virus 40 A protein-DNA complexes: arrangement of pentanucleotide interaction sites at the origin of replication
Authors: DeLucia A L , Lewton B A , Tjian R , Tegtmeyer P
Published: January 2020

Investigation of the DNA binding properties of the simian virus 40 (SV40) A protein (large T antigen) and the hybrid adenovirus-SV40 D2 protein revealed that both viral proteins protect similar regions of SV40 DNA from digestion by DNase I or methylation by dimethyl sulfate. However, the interaction of D2 protein with DNA was more sensitive to increases of NaCl concentration than was the interaction of wild-type SV40 A protein. Dimethylsulfate footprinting identified 13 DNA pentanucleotide contact sites at the viral origin of replication. The sequences of these sites corresponded to the consensus family 5'-(G greater than T) (A greater than G)GGC-3'. The pentanucleotides were distributed in three regions of origin DNA. Region I contained three pentanucleotide contact sites arranged as direct repetitions encompassing a span of 23 base pairs. In region II, four pentanucleotides were oriented as inverted repetitions that also spanned a total of 23 base pairs. Region III had six recognition pentanucleotides arranged as direct repetitions in a space of 59 base pairs. These fundamental variations in DNA arrangement are likely to determine different patterns of protein binding in each region.
Identification of a subset of herpesvirus saimiri polypeptides synthesized in the absence of virus DNA replication
Authors: O'Hare P , Honess R W
Published: January 2020

The effects of phosphonoacetic acid on the synthesis of herpesvirus saimiri-specific polypeptides in productively infected cells were examined. At concentrations that inhibited virus DNA synthesis (greater than or equal to 150 micrograms/ml), phosphonoacetic acid prevented the synthesis of the majority of virus-specific polypeptides while allowing the synthesis of a subset of virus proteins (i.e., 110,000 [110K], 76K, 72K, 51K, 48K, 29K, 24K, and 20K or 21K) and the protracted synthesis of host-specified polypeptides. Other inhibitors of DNA synthesis (e.g., cytosine arabinoside) showed the same selective inhibition of late virus protein synthesis and identified the same resistant subset of early virus-specific polypeptides. This DNA synthesis-independent subset included the 51K phosphoprotein, which, together with the 110K, 48K, and 31K polypeptides, accumulated in the nuclear fraction of infected cells.
Molecular Cloning and Analysis of the Endogenous Retrovirus Chemically Induced from RFM/Un Mouse Cell Cultures
Authors: Liou R. S. , Boone L. R. , Kiggans J. O. , Yang D. M. , Wang T. W. , Tennant R. W. , Yang W. K.
Published: January 2020

We molecularly cloned and analyzed an N-tropic ecotropic retrovirus induced with iododeoxyuridine from RFM/Un mouse cell cultures. Based on the restriction map, the RFM/Un virus appears to be indistinguishable from other induced N-tropic retroviruses. A nucleotide sequence analysis of the long terminal repeat of an infectious clone revealed structural features characteristic of murine type C retrovirus long terminal repeats. The U3 region of the RFM/Un virus long terminal repeat, however, contained no short sequence duplication or insertion found in other murine leukemia virus isolates.
Abortive Infection of F-Plasmid-Containing
Escherichia coli
Cells by Bacterial Virus T7 Is Determined by the Right End of T7 Gene 1
Authors: Krüger Detlev H. , Hansen Sigrid , Chernin Leonid S.
Published: January 2020

Phage T7 infects male (F-plasmid-carrying)
Escherichia coli
cells abortively, whereas the closely related phage T3 grows normally. The inability or ability of phage to replicate in male host cells depends on whether the right end of gene 1 (coding for the phage-specific RNA polymerase) consists of T7 or T3 DNA base sequences.
Functional basis of poliovirus neutralization determined with monospecific neutralizing antibodies
Authors: Emini E A , Kao S Y , Lewis A J , Crainic R , Wimmer E
Published: January 2020

Antibody-mediated poliovirus neutralization was studied by using a series of 13 monospecific neutralizing antibodies. These antibodies were found to recognize seven individual viral epitopes, several of which functionally overlap one another. Each epitope was capable of undergoing variation so that the variant virus was no longer capable of being neutralized by antibody directed against that epitope. The measured degree of variation for each site varied from -3.1 to -4.2 log10 variant PFU per wild-type PFU. Under nonsaturating but neutralizing conditions, the antibodies, with the exception of those directed to one specific epitope, failed to completely inhibit the virion's binding to the cell. Similarly, none of the neutralizing antibodies completely inhibited viral penetration, but all prevented virus-specific transcription. A strong correlation was established between the binding of each of the neutralizing antibodies, with one exception, to the virion and a significant shift in the virion's pI from 7.0 to ca. 4.0.
A simian virus 40 dl884/tsA58 double mutant is temperature sensitive for abortive transformation
Authors: Sompayrac L , Danna K J
Published: January 2020

We have constructed a dl884/tsA58 double mutant and a t+T- early-region deletion mutant and have used these mutants to study the roles of the simian virus 40 tumor antigens (T and t) in transformation. Our major conclusions are that (i) although the mutant tsA58 is not temperature sensitive for the maintenance of transformation, the dl884/tsA58 double mutant is; (ii) small t antigen can provide at least one, but not all, of the functions required for the maintenance of transformation; and (iii) at least two different functions are required for the maintenance of simian virus 40 transformation.
Structural Relationships Between Minor and Major Proteins of Hepatitis B Surface Antigen
Authors: Stibbe Werner , Gerlich Wolfram H.
Published: January 2020

The minor glycoproteins from hepatitis B surface antigen, GP33 and GP36, contain at their carboxy-terminal part the sequence of the major protein P24. They have 55 additional amino acids at the amino-terminal part which are coded by the pre-S region of the viral DNA.
Expression of a transformation-related protein (p53) in the malignant stage of Friend virus-induced diseases
Authors: Ruscetti S K , Scolnick E M
Published: January 2020

Stage 1 (pre-malignant) and stage 2 (malignant) cells derived from mice infected with Friend murine leukemia virus or polycythemia-inducing Friend virus complex were examined and compared for the expression of a transformation-related cellular protein, p53. Stage 2 cells were found to express high levels of p53, whereas stage 1 cells did not express detectable levels of this protein. These results indicate that p53 may be a marker for transformed cells present in the second stage of diseases induced by Friend murine leukemia virus or polycythemia-inducing Friend virus complex.
Synthesis of multimeric polyoma virus DNA in mouse L-cells: role of the tsA1S9 gene product
Authors: Ganz P R , Sheinin R
Published: January 2020

Several different forms of progeny viral DNA can be identified in polyoma virus (Py)-infected mouse L-cells. The majority comprise mature form I superhelical DNA and the circular, double-stranded "theta" replicating intermediates in which the progeny DNA strands never exceed the unit genome length of the template. There is formed, in addition, a minority fraction of multimeric, linear, double-stranded Py DNA molecules that sediment heterogeneously at 28 to 35S and greater than 35S. Restriction enzyme analysis of these large Py DNA molecules reveals them to be tandem arrays of multiple unit genome lengths, covalently linked head to tail. It is estimated that the 28 to 35S multimeric DNA has an average size of about 20 megadaltons, made up of 6 to 20 Py genome units. The greater than 35S Py DNA is, of course, larger. Kinetic analysis indicates that formation of the monomeric progeny viral DNA and the 28 to 35S multimeric Py DNA reaches a peak at about 35 to 36 h postinfection. Synthesis of the very large linear molecules of greater than 35S is first detected after this interval and continues thereafter. The de novo synthesis of all of these progeny Py DNA molecules proceeds apparently normally in Py-infected tsA1S9 mouse L-cells incubated at 38.5 degrees C under conditions which restrict normal cellular DNA replication. These findings suggest that the cellular DNA topoisomerase II activity, encoded in the tsA1S9 locus (R. W. Colwill and R. Sheinin, submitted for publication), is not required for de novo formation of any form of Py DNA. However, the total amount made and the rate of synthesis of the large molecular weight Py DNA are affected very late in temperature-inactivated tsA1S9 cells.
Structure of glycosylated and unglycosylated gag and gag-pol precursor proteins of Moloney murine leukemia virus
Authors: Saris C J , van Eenbergen J , Liskamp R M , Bloemers H P
Published: January 2020

Precursor polyproteins containing translational products of the gag gene of Moloney murine leukemia virus were purified by gel electrophoresis and cleaved into large fragments by hydroxylamine, mild acid hydrolysis, or cyanogen bromide. The hydroxylamine cleavage method (specific for asparagine-glycine bonds) was adapted especially for this study. The electrophoretic mobility and antigenicity of the fragments and, in some cases, the presence or absence of [35S]methionine revealed detailed information on the structure of Pr65gag, gPr80gag, and Pr75gag (the unglycosylated variant of gPr80gag formed in vivo in the presence of tunicamycin or in vitro in a reticulocyte cell-free system). When compared with Pr65gag, gPr80gag contains 7,000 daltons of additional amino acids, presumably as, or as part of, a leader sequence at or very close to its N terminus. We present evidence that this leader may have replaced part of the p15 sequence. Furthermore, gPr80gag contains three separate carbohydrate groups. One is attached to the presumed leader sequence or to the p15 domain, and two are attached to the p30 domain. Each of the Moloney murine leukemia virus gag precursor proteins Pr65gag, gPr80gag, and Pr75gag corresponds with a read-through product into the pol gene. We designated these products Pr180gag-pol, gPr200gag-pol, and Pr190gag-pol (the unglycosylated variant of gPr200gag-pol), respectively. gPr200gag-pol contains all of the extra amino acids and carbohydrate groups present in gPr80gag and at least one carbohydrate group in its pol sequences.
Recombinants Between Bacteriophages T7 and T3 Which Productively Infect F-Plasmid-Containing Strains of
Escherichia coli
Authors: Molineux Ian J. , Mooney Patrick Q. , Spence Jean L.
Published: January 2020

Recombinant phages between T7 and T3 have been isolated that grow well on strains of
Escherichia coli
that contain the F factor. One phage that has been characterized physically and genetically is predominately of the T7 genotype. Within this hybrid phage, two separate regions of T3 DNA have been located which are necessary for the phenotype of productive growth on F-containing strains. One of these, designated M1, contains the right part of gene 1 and continues through gene 1.3; the second, M2, appears to lie between gene 3 and gene 4.
Effects of UV irradiation on the fate of 5-bromodeoxyuridine-substituted bacteriophage T4 DNA
Authors: Restifo L L , Vogelbacker H H , Madara T , Ling S K , Kozinski A W
Published: January 2020

We have carried out a series of experiments designed to characterize the impact of UV irradiation (260 nm) on 5-bromodeoxyuridine-labeled (heavy) T4 bacteriophage, both before and after infection of Escherichia coli. In many respects, these effects differ greatly from those previously described for non-density-labeled (light) phage. Moreover, our results have led us to propose a model for a novel mechanism of host-mediated repair synthesis, in which excision of UV-damaged areas is followed by initiation of replication, strand displacement, and a considerable amount of DNA replication. UV irradiation of 5-bromodeoxyuridine-labeled phage results in single-stranded breaks in a linear, dose-dependent manner (1.3 to 1.5 breaks per genomic strand per lethal hit). This damage does not interfere with injection of the phage genome, but some of the UV-irradiated heavy phage DNA undergoes additional intracellular breakdown (also dose dependent). However, a minority (25%) of the injected parental DNA is protected, maintaining its preinjection size. This protected moiety is associated with a replicative complex of DNA and proteins, and is more efficiently replicated than is the parental DNA not so associated. Most of the progeny DNA is also found with the replicative complex. The 5-bromodeoxyuridine of heavy phage DNA is debrominated by UV irradiation, resulting in uracil which is removed by host uracil glycosylase. Unlike the simple gap-filling repair synthesis after infection with UV-irradiated light phage, the repair replication of UV-irradiated heavy phage is extensive as determined by density shift of the parental label in CsC1 gradients. The newly synthesized segments are covalently attached to the parental fragments. The repair replication takes place even in the presence of chloramphenicol, a protein synthesis inhibitor, suggesting it is host mediated. Furthermore, the extent of the repair replication is greater at higher doses of UV irradiation applied to the heavy phage. This abundant synthesis results ultimately in dispersion of the parental sequences as short stretches in the midst of long segments of newly synthesized progeny DNA. Together, the extensive replication and the resulting distribution pattern of parental sequences, without significant solubilization of parental label, are most consistent with a model of repair synthesis in which the leading strand displaces, rather than ligates to, the encountered 5' end.
Cycloheximide induces expression of the human interferon beta 1 gene in mouse cells transformed by bovine papillomavirus-interferon beta 1 recombinants
Authors: Maroteaux L , Chen L , Mitrani-Rosenbaum S , Howley P M , Revel M
Published: January 2020

Mouse cells transformed by a bovine papillomavirus recombinant vector containing the human interferon (IFN) beta 1 (IFN-beta 1) gene could be induced to produce human as well as mouse IFNs. The optimal conditions for induction of human IFN and of its mRNA in these transformants resembled those needed for mouse IFN: high concentrations of DEAE-dextran and low concentrations of polyriboinosinic acid-polyribocytidylic acid. Superinduction by inhibitors of protein synthesis which strongly stimulate IFN-beta 1 induction in human cells had only a small effect on human IFN induction in bovine papillomavirus IFN-beta 1-transformed mouse cells. In contrast, cycloheximide without double-stranded RNA could induce significant levels of human IFN in the bovine papillomavirus IFN-beta 1 mouse transformants. After cycloheximide treatment, these cells contained IFN-beta 1 mRNA whose 5' ends originated in the authentic start site of the human IFN-beta 1 gene, as shown by S1 nuclease mapping. The transferred human gene, propagated extrachromosomally in the mouse cells, was, therefore, inducible under conditions different from those in human cells. The results also confirmed that the inhibitor of protein synthesis, cycloheximide, can induce expression of a human IFN gene.
Acquisition of Host Cell DNA Sequences by Baculoviruses: Relationship Between Host DNA Insertions and FP Mutants of
Autographa californica
and
Galleria mellonella
Nuclear Polyhedrosis Viruses
Authors: Fraser M. J. , Smith Gale E. , Summers Max D.
Published: January 2020

Mutants of
Autographa californica
and
Galleria mellonella
nuclear polyhedrosis viruses, which produce an altered plaque phenotype as a result of reduced numbers of viral occlusions in infected cells, were isolated after passage in
Trichoplusia ni
(TN-368) cells. These mutants, termed FP (few-polyhedra) mutants, had acquired cell DNA sequences ranging from 0.8 to 2.8 kilobase pairs in size. The insertions of cell DNA occurred in a specific region between 35.0 and 37.7 map units of the
A. californica
viral genome. A cloned viral fragment containing one of the host DNA inserts was homologous to host DNA inserts in two other mutant viruses and to dispersed, repetitious sequences in
T. ni
cell DNA. Most of the homology between the cloned insert and cell DNA was contained within a 1,280-base-pair
Alu
I fragment. Marker rescue studies and analysis of infected-cell-specific proteins suggested that the insertion of cell DNA into the viral genomes resulted in the FP plaque phenotype, possibly through the inactivation of a 25,000-molecular-weight protein.
Positive control of the herpes simplex virus thymidine kinase gene requires upstream DNA sequences
Authors: Smiley J R , Swan H , Pater M M , Pater A , Halpern M E
Published: January 2020

We examined the ability of deleted versions of the herpes simplex virus type 1 thymidine kinase gene, present in biochemically transformed mouse cells, to be induced in trans to a higher level of expression by superinfecting herpes simplex virus immediate early gene products. The results demonstrate that sequences mapping between -200 and -80 and between -70 and -12 are required for induction. As these regions are largely coincident with the previously identified thymidine kinase gene promoter, the results suggest that herpes simplex virus immediate early gene products or their metabolic product activate thymidine kinase expression by acting at the promoter region to increase the rate of transcription.
Localization of the leukemogenic determinants of SL3-3, an ecotropic, XC-positive murine leukemia virus of AKR mouse origin
Authors: Lenz J , Haseltine W A
Published: January 2020

SL3-3 is a potent leukemogenic retrovirus that closely resembles the non-leukemogenic virus Akv. Both viruses were isolated from AKR mice, have ecotropic host ranges, and form plaques in the XC assay. They differ at only 1 to 2% of the nucleotides in the viral genomes but differ markedly in virulence properties. SL3-3 induces leukemia in a high percentage of inoculated AKR, C3H, CBA, and NFS mice, whereas Akv does not induce disease in any of these strains. To determine which region of the genome accounts for the leukemogenic potential of SL3-3, we constructed recombinant genomes between molecular clones of SL3-3 and Akv. Recombinant, viral DNA genomes were cloned and then were transfected onto NIH 3T3 fibroblasts to generate infectious virus. The recombinant viruses were tested for leukemogenicity in AKR/J, CBA/J, and C3Hf/Bi mice. We localized the primary leukemogenic determinant to a 3.8-kilobase fragment of the SL3-3 genome containing the viral long terminal repeat, 5' untranslated sequences, gag gene, and 5', 30% of the pol gene. Reciprocal recombinants containing the equivalent region from Akv, linked to the env gene and the remainder of the pol gene from SL3-3, did not induce leukemia. We conclude that the primary virulence determinant of SL3-3 lies outside the region of the genome that encodes the envelope proteins gp70 and p15E.