bluuebunny/crossref_metadata_2025 huggingface dataset:
In vitro transcription of two human rotaviruses
Authors: Flores J , Myslinski J , Kalica A R , Greenberg H B , Wyatt R G , Kapikian A Z , Chanock R M
Published: January 2020

The RNA polymerase activities of a cultivatable (Wa) and a noncultivatable (DS-1) strain of human rotavirus were studied. Under optimal conditions, transcription of all of their RNA segments occurred, as evidenced by the hybridization of labeled transcripts to genomic RNA. Cross-hybridization between the two viruses showed that none of their 11 genes were completely homologous. The transcription products could be translated in vitro, yielding proteins with an electrophoretic pattern resembling that obtained with proteins labeled in vivo during infection with the Wa virus.
Acquisition of proviral DNA of mouse mammary tumor virus in thymic leukemia cells from GR mice
Authors: Michalides R , Wagenaar E , Hilkens J , Hilgers J , Groner B , Hynes N E
Published: January 2020

Male mice of strain GR develop T-cell leukemia at a low frequency late in life. These leukemia cells contain large amounts of mouse mammary tumor virus (MMTV) RNA and MMTV proteins in a precursor form (Nusse et al., J. Virol. 32:251-258, 1979). We used restriction enzyme analysis and molecular hybridization to identify MMTV proviruses in the DNA of these leukemia cells. GR leukemia cells contained additional integrated MMTV proviruses at various sites in the genome. This amplification of MMTV proviruses in GR leukemia cells is not restricted to one particular endogenous MMTV provirus of strain GR. The number and location of the extra MMTV proviruses present in transplants of GR leukemia cells did not change upon serial transplantation of the leukemia cells. Acquisition of MMTV proviruses was also found in a similar leukemia, L1210 of the DBA/2 mouse strain, but not in three other leukemias, SL2 of DBA/2, BW5147 of AKR, and a spontaneous thymoma of BALB/c. The two main classes of MMTV RNA, 35S and 24S, were present in the cytoplasmic RNA of GR leukemia cells, indicating that the aberrant processing of MMTV precursor proteins is not due to anomolously sized RNAs. We could not detect extra RNAs in GR leukemia cells which would represent read-through transcripts of cellular genes adjacent to the extra MMTV proviruses, initiated by a promoter signal in the right MMTV long terminal repeat sequence. These data suggest that acquisition of MMTV proviruses may coincide with the onset of leukemogenesis in GR male mice.
Characterization of c-lil, a chicken cellular sequence associated with a stock of B77 avian sarcoma virus
Authors: Boccara M , Pluquet N , Coll J , Rommens C , Stehelin D
Published: January 2020

Using biochemical methods, we have shown that a new specific sequence, v-lil, is associated with a given stock of B77 avian sarcoma virus (clone 9). We prepared a DNA complementary to v-lil sequences, using substractive hybridizations, and investigated the properties of this sequence. v-lil has a genetic complexity of ca. 2,000 nucleotides and is not present in various stocks of avian sarcoma virus, avian leukosis virus, or defective leukemia virus. v-lil is not associated with B77 avian sarcoma virus isolated from the original tumor and thus has been acquired by in vitro passage of the virus on chicken embryo fibroblasts. A search for the origin of the v-lil sequence among the DNAs of different avian species has shown that a similar sequence, c-lil, is present in normal chicken DNA (1 to 2 copies per haploid genome). c-lil is not highly conserved but is present in the DNA of all chickens from the genus Gallus. The c-lil sequence is transcribed at a low level (1 to 3 copies per cell) in normal chicken embryo fibroblasts. The biological function, if any, of v-lil or its cellular equivalent has yet to be determined.
Control of expression of the herpes simplex virus-induced deoxypyrimidine triphosphatase in cells infected with mutants of herpes simplex virus types 1 and 2 and intertypic recombinants
Authors: Wohlrab F , Garrett B K , Francke B
Published: January 2020

Infection of cells with herpes simplex virus type 1 (HSV-1) induces high levels of deoxypyrimidine triphosphatase. The majority of the enzyme activity is found in infected cell nuclei. A similar activity is induced by HSV type 2 (HSV-2) which, in contrast to the HSV-1 enzyme, fractionates to more than 99% in the soluble cytoplasmic extract. Of a series of temperature-sensitive mutants of HSV-1 studied, only the immediate-early mutants in complementation group 1-2 (strain 17 mutants tsD and tsK and strain KOS mutant tsB2) induced reduced levels of triphosphatase at nonpermissive temperature. Of a series of temperature-sensitive mutants of HSV-2 strain HG52, ts9 and ts13 failed to induce wild-type levels of the enzyme at nonpermissive temperature; ts9 was the most defective mutant with regard to triphosphatase expression of both herpes simplex virus serotypes. After shift-up from permissive to nonpermissive temperature, triphosphatase activity in cells infected with ts9 decreased rapidly, whereas all other mutants continued to exhibit enzyme levels comparable with controls kept at the permissive temperature. The type 1-specific nuclear expression of the triphosphatase was mapped physically by the use of HSV-1 x HSV-2 intertypic recombinants, based on enzyme levels different by more than two orders of magnitude found in nuclei of HSV-1- and HSV-2-infected cells. The locus for the type-specific expression maps between 0.67 and 0.68 fractional length on the HSV genome.
Antibodies against a synthetic peptide of the poliovirus replicase protein: reaction with native, virus-encoded proteins and inhibition of virus-specific polymerase activities in vitro
Authors: Baron M H , Baltimore D
Published: January 2020

A carboxy-terminal peptide of the poliovirus replicase protein (p63) was chemically synthesized, coupled to bovine serum albumin carrier, and injected into rabbits. The resulting antisera reacted with six virus-specific proteins from HeLa cells infected with poliovirus: NCVP 0b, NCVP 1b, NCVP 2, a protein of about 60,000 daltons, p63, and NCVP 6b. The identity of the 60,000-dalton protein is not known, but the other results were consistent with previous experimental approaches which demonstrated that p63 and the other four polypeptides have common coding sequences. An amino-terminal peptide of p63 failed to elicit an immune response in rabbits. Antibodies raised against the p63 carboxy-terminal peptide inhibited poliovirus replicase and polyuridylic acid polymerase activities in vitro, providing strong support for earlier suggestions that these activities are a property of a single virus-specific polypeptide.
Effect of simian virus 40 on the temporal and spatial organization of DNA replication in Muntjac cells
Authors: Jasny B R , Wright J M , Smith L A , Gershey E L , Tamm I
Published: January 2020

We examined the effects of simian virus 40 infection on the temporal and spatial organization of initiation sites for DNA replication in Muntjac cells by means of light microscopic DNA fiber autoradiography. Initiation at multiple sites along the DNA fiber in virus-infected confluent Muntjac cells was more nearly synchronous than in serum-deprived controls, although temporal control in the infected cells did not reach the level observed in cells incubated in serum-enriched medium. Initiation sites in virus-infected cells appeared to be spatially closer together than in either uninfected serum-deprived or uninfected serum-enriched cells. This change did not appear to be the result of the induction or repression by simian virus 40 of clusters of replication units with new and different organizations.
DNA sequence of the viral and cellular src gene of chickens. II. Comparison of the src genes of two strains of avian sarcoma virus and of the cellular homolog
Authors: Takeya T , Hanafusa H
Published: January 2020

The nucleotide sequence of the src gene and flanking regions of the Schmidt-Ruppin strain of Rous sarcoma virus (SR-A) was determined. The src region of SR-A was very homologous to that of recovered avian sarcoma virus (rASV1441), with only 17 differences among 1,578 nucleotides. The size of the predicted protein was 526 amino acids in both viruses, of which 6 amino acids were different. The differences in nucleotides and amino acids between the two viruses localized within the 5' two-thirds of the src coding region. There were also viruses localized within the 5' two-thirds of the src coding region. There were also some differences in the region flanking the 5' end of src. Since rASVs are considered to be recombinatns between deletion mutants of SR-A and cellular-src (c-src) sequences, several segments of c-src DNA were also sequenced to understand the molecular basis for the recombination. At 14 of 17 bases where SR-A and rASV1441 differed, rASV1441 had the same sequence as c-src. Three of these sequences corresponded to sequences of oligonucleotides which were previously identified in RNAs of nearly all isolates of rASV but which were absent in SR-A RNA. In the 5'-flanking sequences of the src gene, c-src was more similar to rASV1441 than to SR-A. These results confirm the cellular origin of the src sequences of rASVs and provide information about the possible sites of the recombination.
Novel phenotype of RNA synthesis expressed by vesicular stomatitis virus isolated from persistent infection
Authors: Frey T K , Youngner J S
Published: January 2020

Vesicular stomatitis virus (VSV) stocks isolated from two persistently infected mouse L-cell lines (designated VSV-PI stocks) express an altered phenotype of RNA synthesis. This phenotype is different from the RNA synthesis phenotype expressed by the viruses used to initiate the persistently infected lines, wild-type VSV and VSV ts-0-23 (a group III, ts-, RNA+ mutant). At 34 and 37 degrees C in L cells productively infected with VSV-PI stocks derived from the two cell lines, transcription of virus mRNA was significantly reduced, whereas replication of the 40S genomic RNA species was enhanced compared with wild-type VSV or ts-0-23. At 34 and 37 degrees C, both VSV-PI stocks replicated with equal or greater efficiency than wild-type VSV; 37 degrees C was the temperature at which the persistently infected cultures were maintained. At 40 degrees C, both VSV-PI stocks were temperature sensitive, and clonal VSV-PI isolates from both cell lines belong to complementation group I (RNA-). Standard ts- mutants (derived by mutagenesis of wild-type VSV) belonging to RNA- complementation groups I, II, and IV do not express the VSV-PI RNA synthesis phenotype at the permissive temperature, making this phenotype distinctive to persistent infection. Since the two VSV-PI populations from persistently infected cell lines initiated with different viruses both evolved this unique phenotype of RNA synthesis, the expression of this phenotype may play an important role in the maintenance of persistence.
Evidence that vesicular stomatitis virus produces double-stranded RNA that inhibits protein synthesis in a reticulocyte lysate
Authors: Thomas J R , Wagner R R
Published: January 2020

Cell-free protein synthesis by reticulocyte lysates was inhibited by a polyadenylated RNA fraction extracted from HeLa cells infected with vesicular stomatitis virus (VSV) but not by polyadenylated RNA from mock-infected HeLa cells. A similar inhibitor of cell-free protein synthesis was found in a polyadenylated fraction of RNA transcribed in vitro by VSV but not in untranscribed nucleocapsids. Fractionation of the VSV transcription product showed that the translation inhibitor segregated with nucleocapsids containing newly transcribed polyadenylated or non-polyadenylated RNA, as determined by oligodeoxythymidylate-cellulose chromatography. The inhibitors present in VSV-infected HeLa cells and in VSV in vitro transcripts both appeared to be double-stranded RNA, as judged by the characteristics for inhibition of reticulocyte cell-free protein synthesis described by Hunter et al. (J. Biol. Chem. 250:409-417, 1975). The double-stranded nature of the VSV RNA inhibitor was supported by the finding that the translational inhibitory effect was inactivated by melting the inhibitor in the absence of salt and by micrococcal nuclease.
Simple repeat sequence in Epstein-Barr virus DNA is transcribed in latent and productive infections
Authors: Heller M , van Santen V , Kieff E
Published: January 2020

The BamHI K region of Epstein-Barr virus DNA is transcribed in latently infected cells from Burkitt tumors and in growth-transformed B-lymphocytes latently infected with Epstein-Barr virus. We determined the nucleotide sequence of a 1,153-base pair HinfI fragment in BamHI fragment K from the B95-8 Epstein-Barr virus isolate. The fragment contains a remarkable 708-base pair simple sequence repeat array, designated IR3, which is composed of only three nucleotide triplet elements: GGG, GCA, and GGA. The triplets are organized into three repeat units: GCAGGA, GCAGGAGGA, and GGGGCAGGA. Immediately 3' of IR3 are tandem nearly perfect direct repeats of two different 24-base pair sequences. IR3 is conserved at a colinear position in the DNAs of other Epstein-Barr virus isolates, and a homologous sequence maps at the same location in the genome of a genetically related baboon herpesvirus, herpesvirus papio. IR3 is transcribed from left to right in latently infected, growth-transformed IB4 cells. It encodes part of a 2.0-kilobase exon of the 3.7-kilobase cytoplasmic polyadenylated RNA previously detected in IB4 cells (van Santen et al., Proc. Natl. Acad. Sci. U.S.A. 78:1930-1934, 1981). IR3 also encodes parts of 2.4- and 1.0-kilobase RNAs in productively infected B95-8 cells.
Disulfide bonding in influenza virus proteins as revealed by polyacrylamide gel electrophoresis
Authors: Selimova L M , Zaides V M , Zhdanov V M
Published: January 2020

Disulfide bonding in the major proteins of influenza virus A, WSN strain, was studied by electrophoresis in sodium dodecyl sulfate-polyacrylamide gels under reducing and nonreducing conditions. The electrophoretic behavior of the proteins correlated with their localization in the virions and their chemical composition. The internal proteins of the viral particles, i.e. matrix and nucleoproteins, were shown to contain a relatively small number of cysteine residues. Electrophoresis under nonreducing conditions yielded multiple forms of the proteins which could be discriminated by small but readily observable, reproducible differences in their migration rates in the gel. the multiplicity of the protein forms was caused by the formation of intramolecular disulfide bonds in matrix and nucleoproteins that arose during or after solubilization in sodium dodecyl sulfate. On the other hand, we failed to detect native inter- and intramolecular linkages in matrix and nucleoproteins. External glycoproteins of the virions (HA and NA) had, in contrast to the internal ones, a higher number of cysteine residues and native disulfide bonds. At least three disulfide linkages were revealed in HA and NA in our experiments. In uncleaved HA all of the linkages were intramolecular. In NA at least one disulfide bond linked two identical polypeptides into a dimer. It was established that the reduction of the different disulfide linkages in HA and NA required different concentrations of the reducing agent.
Methylation state and DNase I sensitivity of chromatin containing Moloney murine leukemia virus DNA in exogenously infected mouse cells
Authors: Montandon P E , Montandon F , Fan H
Published: January 2020

The nature of Moloney murine leukemia virus (M-MuLV)-specific proviral DNA in exogenously infected mouse cells was studied. M-MuLV clone A9 cells, NIH-3T3 fibroblasts productively infected with M-MuLV, were used. These cells contain 10 to 15 copies of M-MuLV proviral DNA. The state of methylation of M-MuLV proviral DNA was examined by cleaving A9 cell DNA with restriction endonucleases which have the dinucleotide CpG in their cleavage sequences. Analysis with such enzymes, which recognized nine different sites in M-MuLV DNA, indicated that most if not all of the M-MuLV proviruses in A9 cells were completely unmethylated. An individual proviral integration was examined, using as probe adjacent single-copy cellular sequences. These sequences were obtained from a lambda phage recombinant clone containing an M-MuLV provirus from the A9 cells. This individual integration also showed no detectable methylation. In contrast, endogenous MuLV-related sequences present in NIH-3T3 cells before infection were largely methylated. The configuration chromatin containing M-MuLV proviruses was also investigated by digesting A9 nuclei with DNase I, followed by restriction analysis of the remaining DNA. Endogenous MuLV-related DNA was in chromatin relatively resistant to DNase I digestion, whereas the majority of M-MuLV-specific proviruses were in domains of intermediate DNase I sensitivity. Two proviral copies hypersensitive to DNase I digestion were identified. Analogy to the DNase I sensitivity of expressed and nonexpressed globin genes suggested that the proviral copies containing DNase I-hypersensitive sites were transcribed.
Localization of viral structural proteins in the cytoplasm and nucleus of Rous-associated virus-2-infected chicken embryo fibroblasts
Authors: Enrietto P J , Erikson R L
Published: January 2020

The cellular location of viral structural proteins was carried out by immunohistochemistry and by cell fractionation. Antibody against the structural protein p27 was used in immunohistochemical reactions to demonstrate the presence of viral proteins in the cytoplasm and nucleus of Rous-associated virus 2-infected chicken cells. Localization in the nucleus was found over heterochromatic regions; in the cytoplasm it was found in discrete particulate structures. These observations were extended in cell fractionation studies in which cytoplasmic and nuclear fractions were immunoprecipitated with antibody against the viral structural proteins.
Adeno-associated virus helper activity of adenovirus DNA binding protein
Authors: McPherson R A , Ginsberg H S , Rose J A
Published: January 2020

The requirement for the adenovirus (Ad) single-stranded DNA binding protein (DBP) in the expression of adeno-associated virus (AAV) proteins was studied by specific immunofluorescent staining of infected cells and in vitro translation of RNA from infected cells. The Ad5 mutant ts125, which carries a mutation in the DBP gene, helped AAV as efficiently as the Ad5 wild type (WT) did at both the permissive (32 degrees C) and nonpermissive (40.5 degrees C) temperatures in HeLa and KB cells. Furthermore, at 40.5 degrees C ts125 was as efficient as Ad5WT was in inducing the expression of AAV proteins in a line of Detroit 6 cells which is latently infected with AAV. However, little if any AAV protein was synthesized when coinfections were carried out with Ad5WT in CV-C cells, a monkey cell line that is highly restrictive for human Ad replication unless the cells are also infected with simian virus 40. On the other hand, AAV protein was efficiently produced in CV-C cells in coinfections with the Ad5 mutant hr404, whose growth is unrestricted in CV-C cells and whose mutation also maps in the DBP gene. Finally, preparations of cytoplasmic RNA extracted from CV-C cells infected with AAV and Ad5WT or from CV-C cells infected with AAV, Ad5WT, and simian virus 40 were each capable of directing the in vitro synthesis of abundant amounts of AAV proteins in a rabbit reticulocyte lysate system. These results indicate that the abnormal DBP of ts125 still retains its helper function for AAV replication, but that the molecular feature of the DBP which relates to the monkey cell host range restriction of Ad's may also account for the observed block to AAV protein translation in CV-C cells.
Rauscher murine leukemia virus: molecular cloning of infectious integrated proviral DNA
Authors: Habara A , Reddy E P , Aaronson S A
Published: January 2020

The integrated proviral genome of Rauscher murine leukemia virus was molecularly cloned in a bacteriophage Charon 4A vector after the proviral sequences were enriched by sequential RPC-5 column chromatography and sucrose gradient centrifugation. A recombinant DNA clone, lambda-RV-1, possessing a 12-kilobase-pair EcoRI insert, was shown to contain the entire 8.8-kilobase-pair leukemia virus genome flanked by rat cellular sequences at the 5' and 3' ends. This DNA fragment was biologically active, inducing the release of virion-associated reverse transcriptase activity with as little as 10 ng of DNA insert. The virus induced XC plaque formation at high titers on NIH/3T3 and BALB/3T3 cells and demonstrated identity with the parental virus in radioimmunoassays for the highly type-specific gag gene-coded p12 protein. The molecularly cloned Rauscher murine leukemia virus should be useful in studying the molecular mechanisms involved in the transformation of specific lymphoid target cells by chronic mouse leukemia viruses.
Molecular biology of rotaviruses. IV. Molecular cloning of the bovine rotavirus genome
Authors: McCrae M A , McCorquodale J G
Published: January 2020

A new cloning strategy has been developed for cloning the genomes of double-stranded RNA viruses by using bovine rotavirus as a test system. The major modification adopted was the use of denatured polyadenylated double-stranded RNA as the template for reverse transcriptase. This allowed the two complementary strands of cDNA to be synthesized in a single reaction and removed the need for S1 nuclease digestion to remove the 5' hairpin structure normally generated in cDNA synthesis.
Accurate transcription of simian virus 40 chromatin in a HeLa cell extract
Authors: Brady J N , Radonovich M , Salzman N P
Published: January 2020

During simian virus 40 viral maturation, a series of modifications occur which alter the composition of viral nucleoprotein complexes. As a consequence, the chromatin that is extracted from extracellular simian virus 40 virions exhibits properties that are similar to those of transcriptionally active eucaryotic chromatin. The influence of this chromatin structure on specific RNA initiation by RNA polymerase II was examined by using the in vitro HeLa cell extract of Manley et al. (Proc. Natl. Acad. Sci. U.S.A. 77:3855-3859, 1980). The 5' ends of RNA transcripts were positioned by the "run-off" assay, in which transcripts extend from the initiation site to termination sites created by restriction cleavage and by S1 nuclease analysis, using DNA probes labeled at their 5' termini. Two major early RNA transcripts, which originated at map positions 5,240 +/- 10 and 5,145 +/- 10, and two major late RNA transcripts, which originated at map positions 325 +/- 10 and 185 +/- 10, were identified. Transcripts were initiated with comparable relative efficiencies at the same 5' site when either purified DNA or chromatin was used as the template. Our results suggest that extracellular simian virus 40 virion chromatin modifications do not regulate simian virus 40 promoter selection but function to increase the accessibility of RNA promoter sequences to RNA polymerase II and allow efficient elongation of the RNA chain after the initiation event.
Replication of mycoplasma virus L51. VII. Effect of chloramphenicol on the synthesis of DNA replicative intermediates
Authors: Das J , Maniloff J
Published: January 2020

Chloramphenicol affects several steps in the DNA replication of mycoplasma virus L51, a noncytocidal, naked, bullet-shaped virion containing circular single-stranded (SS) DNA of 1.5 X 10(6) daltons (4.5 kilobases). In the presence of chloramphenicol, adsorption was normal and parental SS DNA was converted to double-stranded replicative forms (RF), but subsequent RF leads to RF replication was inhibited. Chloramphenicol added late in infection, when most viral nascent DNA is in progeny SS molecules, inhibited SS synthesis, but nascent RF molecules were formed. However, a chase experiment showed that these RF molecules could not be converted to SS DNA. Therefore, viral RF molecules made in the presence of chloramphenicol are not functional as SS DNA precursors.
In vitro synthesis and assembly of picornaviral capsid intermediate structures
Authors: Palmenberg A C
Published: January 2020

Cell-free translation of encephalomyocarditis RNA in extracts of rabbit reticulocytes results in the synthesis of viral proteins indistinguishable from those produced during virus infection of cells. The viral capsid proteins are produced in an active form capable of assembly into viral capsid intermediate structures. Protomers (5S), pentamers (14S), and shell-like structures (75 to 85S) can be detected after prolonged incubation in the extracts. Proteolytic cleavage of capsid precursor proteins appears to be a prerequisite for assembly, in apparent contrast to cell-associated assembly. Assembly of pentamers is also preceded by conversion of protein epsilon 1 to epsilon in a step which may reflect an amino-terminal blocking reaction.
Effect of intracellular vesicular stomatitis virus mRNA concentration on the inhibition of host cell protein synthesis
Authors: Schnitzlein W M , O'Banion M K , Poirot M K , Reichmann M E
Published: January 2020

Inhibition of host cellular protein synthesis by vesicular stomatitis virus (VSV) has been suggested to be primarily the result of competition for ribosomes between cellular and viral mRNAs (H. F. Lodish and M. Porter, J. Virol., 36:719-733, 1980; Lodish and Porter, J. Virol. 38:504-517, 1981). This hypothesis was investigated by regulating the extent of VSV mRNA synthesis through the use of defective interfering particles. Although intracellular VSV mRNA concentrations decreased by as much as a factor of 14 at high multiplicities of infection of defective interfering particles, the inhibition of host cell protein synthesis by VSV decreased by a maximum of only 10%. The data also indicated that under these conditions the protein-synthesizing capacity of the cells was not exhausted. We concluded that competition for cellular ribosomes could not have been the major factor in the inhibition of host cell protein synthesis by VSV. This conclusion was further supported by inhibition data obtained with VSV mutants. The ts G22 mutant, defective in replication but not in primary transcription, inhibited host protein synthesis at the nonpermissive temperature (39 degrees C) to the same extent as did wild-type virus, even though it generated only 30 to 50% of the amount of viral mRNA as did wild-type virus. Conversely, in infections with the R1 mutant, which did not inhibit host cell protein synthesis, the amount of total and polysome-bound viral mRNA was indistinguishable from that obtained in infections by wild-type virus.
Comparison of infectious JC virus DNAs cloned from human brain
Authors: Grinnell B W , Padgett B L , Walker D L
Published: January 2020

We cloned JC virus DNA obtained directly from brain tissue of 10 cases of progressive multifocal leukoencephalopathy and compared DNAs by restriction endonuclease mapping. Before cloning, each DNA preparation was homogeneous with respect to restriction patterns, but with the cloned DNAs we found variability in three regions of the genome among DNAs from different cases. There was a region of hypervariability between 0.67 and 0.725 map units; no two DNAs were exactly alike in this region. We determined that the origin of DNA replication also was in this region at 0.69 +/- 0.02 map units. In 4 of the 10 DNAs examined there was a deletion of approximately 75 base pairs between 0.14 and 0.235 map units, the region presumed to contain the codons for the C-terminal ends of the structural protein Vpl and for T antigen. JC virus DNA from these same four cases had an additional HincII-HpaI site at 0.895 map units in the presumptive Vp3 and Vp2 coding regions. Overall, no two JC virus genomes were identical although all were from fatal central nervous system infections and were infectious in vitro. Our restriction patterns suggest that there are two subtypes of JC virus circulating in the population.
Isolation and Characterization of Mink Cell Focus-Inducing Murine Leukemia Viruses with Xenotropic Host Range from Mouse Strain SL
Authors: Adachi Akio , Sakai Koji , Ishimoto Akinori
Published: January 2020

A new type of mink cell focus-inducing virus was persistently isolated from the leukemic tissues of SL mice. In contrast to the dual tropic mink cell focus-inducing viruses reported to date, the new virus has the host range of the xenotropic murine leukemia virus. Analysis of RNase T
1
fingerprints of genomic RNAs suggested that the mink cell focus-inducing virus with the xenotropic host range isolated from SL mice is a recombinant virus deriving from xenotropic murine leukemia virus.
Solubilization of a protein synthesis inhibitor from vaccinia virions
Authors: Ben-Hamida F , Person A , Beaud G
Published: January 2020

The protein synthesis inhibitor previously demonstrated to be associated with vaccinia cores was quantitatively solubilized from vaccinia virions or cores after an endogenous protein kinase reaction at pH 10. The presence of the inhibitor in the soluble extract correlated with the presence of soluble virion proteins phosphorylated in vitro. Partially purified inhibitor blocked methionyl-tRNAfMet-40S initiation complex formation, as was the case in cell-free extracts derived from vaccinia virus-infected cells.
Development of Anti-pp60

src

Serum with Antigen Produced in
Escherichia coli
Authors: Gilmer Tona M. , Erikson R. L.
Published: January 2020

We have purified p60

src

from bacterial recombinants which direct the synthesis of the Rous sarcoma virus transforming gene (
src
) product. This protein was injected into rabbits, and they produced a highly cross-reactive serum which can recognize the
src
protein from many different strains of Rous sarcoma virus.
Control of RFM strain endogenous retrovirus in RFM mouse cells
Authors: Tennant R W , Otten J A , Wang T W , Liou R S , Brown A , Yang W K
Published: January 2020

RFM/Un mice express an endogenous type C retrovirus throughout their life span in many tissues; primary or established embryo fibroblast cell cultures do not express a virus but can be induced by exposure to 5-iodo-2'-deoxyuridine. All of our sources yielded a single ecotropic virus (RFV) which appeared to be related more closely to the endogenous N-tropic virus (WN1802N) of BALB/c mice than to Gross leukemia virus on the basis of two-dimensional gel electropherograms of virion proteins. No xenotropic or recombinant viruses were isolated by cocultivation techniques. RFV is N-tropic, and RFM/Un cells possess the Fv-1n allele, as indicated by restriction of B-tropic virus and susceptibility to Gross strain N-tropic virus. However, RFM cells are highly resistant to RFV and other endogenous N-tropic viruses. This resistance is expressed by two-hit titration kinetics and by inhibition of viral linear duplex DNA formation. This is similar to the effects of the Fv-1 locus, but preliminary work has shown no apparent genetic linkage between the two restrictions. The relative strength of the restriction, the presence of a single class of ecotropic virus, and the absence of recombinant viruses suggest that in RFM mice virus is expressed only in cells in which it is induced and not by cell-to-cell transmission.
In vitro transcription of defective interfering particles of influenza virus produces polyadenylic acid-containing complementary RNAs
Authors: Chanda P K , Chambers T M , Nayak D P
Published: January 2020

Influenza virus defective interfering (DI) RNAs, which originate from polymerase genes by simple internal deletion, can be transcribed in vitro. These DI RNA transcripts contain covalently linked polyadenylic acid, and their synthesis is dependent on ApG or capped RNAs as primers. Furthermore, like the standard viral RNA transcripts, they are complementary in nature and are slightly smaller in size compared with the corresponding DI RNAs. Hybridization of the specific DI RNA transcripts with the corresponding DI RNA segments and analysis of the duplex RNA by gel electrophoresis indicate that they are not incomplete polymerase gene transcripts, but rather the transcripts of the DI RNAs. Since influenza virus DI RNAs contain both the 5' and the 3' termini and transcribe polyadenylic acid-containing complementary RNAs in vitro the mechanism of interference may differ from that of the 5' DI RNAs of Sendai and vesicular stomatitis viruses.
Two Transcription Products of the Vesicular Stomatitis Virus Genome May Control L-Cell Protein Synthesis
Authors: Dunigan David D. , Lucas-Lenard Jean M.
Published: January 2020

When mouse L-cells are infected with vesicular stomatitis virus, there is a decrease in the rate of protein synthesis ranging from 20 to 85% of that in mock-infected cells. Vesicular stomatitis virus, irradiated with increasing doses of UV light, eventually loses this capacity to inhibit protein synthesis. The UV inactivation curve was biphasic, suggesting that transcription of two regions of the viral genome is necessary for the virus to become inactivated in this capacity. The first transcription product corresponded to about 373 nucleotides, and the second corresponded to about 42 nucleotides. Inhibition of transcription of the larger product by irradiating the virus with low doses of UV light left a residual inhibition of protein synthesis consisting of approximately 60 to 65% of the total inhibition. This residual inhibition could be obviated by irradiating the virus with a UV dose of greater than 20,000 ergs/mm
2
and was thus considered to represent the effect of the smaller transcription product. In the R1 mutant of C. P. Stanners et al. (Cell
11
:273-281, 1977), inhibition of transcription of the larger product sufficed to restore protein synthesis to the mock-infected level, suggesting that the smaller transcription product is nonfunctional with respect to protein synthesis inhibition. It thus appears that the inhibition of protein synthesis by wild-type vesicular stomatitis virus involved at least two separate viral transcription products, and the inhibition by the R1 mutant involved only one. Extracts from cells infected with virus irradiated with low doses of UV light showed a protein synthesis capacity quite similar to that of their in vivo counterparts, indicating that these extracts closely reflect the in vivo effects of virus infection.
Structural proteins of Western equine encephalitis virus: amino acid compositions and N-terminal sequences
Authors: Bell J R , Bond M W , Hunkapiller M W , Strauss E G , Strauss J H , Yamamoto K , Simizu B
Published: January 2020

The structural proteins of Western equine encephalitis virus, a member of the alphavirus group, have been characterized by the determination of their amino acid compositions and by N-terminal sequence analysis. More than 60 residues of the N-terminal sequences of each of the envelope glycoproteins have been determined. A comparison of these sequences with the previously determined sequences of two related alphaviruses. Sindbis virus and Semliki Forest virus, strongly supports the view that all three viruses have evolved from a common ancestor and provides information on the pattern of this evolution. The analysis of the capsid proteins of Western equine encephalitis virus shows that the nucleocapsid of this virus can accommodate a considerable degree of variability in its protein component and that at least some regions of alphavirus capsid proteins show more extensive differences between different viruses than do the envelope glycoproteins.
Analysis of the recombination event generating a vesicular stomatitis virus deletion defective interfering particle
Authors: Yang F , Lazzarini R A
Published: January 2020

cDNA clones of different portions of the L cistron and 5'-terminal region of the vesicular stomatitis virus genome have been prepared and used to identify the exact site of the deletion in the defective interfering particle, DI-LT. The deletion extends from nucleotide 251 from the beginning of the L gene to a position 342 nucleotides from the end of the genome. The nucleotide sequences flanking the deletion site, as well as those at the ends of the deleted segment, did not contain any obvious vesicular stomatitis virus initiation or termination signals as had been found near the recombination sites in other defective interfering particle RNAs. The results best fit a model for the origin of this type of defective interfering particle in which the polymerase interrupts its synthesis and moves with its nascent daughter strand to a new position on the template and resumes synthesis there, further extending the nascent strand. Neither the interruption nor the resumption of synthesis appears to be in response to the template nucleotide sequence. The sequences of two partial L cistron clones also reveal open reading frames that code for amino acid sequences likely to be the amino and carboxy termini of the L protein.
Early intermediates in bacteriophage T4 DNA replication and recombination
Authors: Dannenberg R , Mosig G
Published: January 2020

We investigated, by density gradients and subsequent electron microscopy, vegetative T4 DNA after single or multiple infection of Escherichia coli with wild-type T4. Our results can be summarized as follows. (i) After single infection (i.e., when early intermolecular recombination could not occur), most, if not all, T4 DNA molecules initiated the first round of replication with a single loop. (ii) After multiple infection, recombinational intermediates containing label from both parents first appeared as early as 1 min after the onset of replication, long before all parental DNA molecules had finished their first round and before secondary replication was detectable. (iii) At the same time, in multiple infections only, complex, highly branched concatemeric T4 DNA first appeared. (iv) Molecules in which two loops or several branches were arranged in tandem were only found after multiple infections. (v) Secondary loops within primary loops were seen after both single and multiple infections, but they were rare and many appeared off center. Thus, recombination in wild-type T4-infected cells occurred very early, and the generation of multiple tandem loops or branches in vegetative T4 DNA depended on recombination. These results are consistent with the previous finding (A. Luder and G. Mosig, Proc. Natl. Acad. Sci. U.S.A. 79:1101-1105, 1982) that most secondary growing points of T4 are not initiated from origin sequences but from recombinational intermediates. By these and previous results, the various DNA molecules that we observed are most readily explained as intermediates in DNA replication and recombination according to a model proposed earlier to explain various other aspects of T4 DNA metabolism (Mosig et al., p. 277-295, in D. Ray, ed., The Initiation of DNA Replication, Academic Press, Inc., New York, 1981).
Identification and Characterization of a Herpes Simplex Virus Gene Product Required for Encapsidation of Virus DNA
Authors: Preston Valerie G. , Coates Jonathan A. V. , Rixon Frazer J.
Published: January 2020

A mutant of herpes simplex virus type 1, 17
ts
VP1201, has a temperature-sensitive processing defect in a late virus polypeptide. Immunoprecipitation studies with monoclonal antibodies showed that the aberrant polypeptide in mutant virus-infected cells was the nucleocapsid polypeptide known as p40. Since a revertant, TS
+
for growth, processed the polypeptide normally under conditions restrictive for the mutant, the processing event must be essential for virus replication. Electron microscopic analysis of mutant virus-infected cells grown at the nonpermissive temperature revealed that the nuclei contained large aggregations of empty nucleocapsids possessing some internal structure. Therefore, although the mutant synthesized virus DNA at the nonpermissive temperature, the DNA was not packaged into nucleocapsids. When mutant virus-infected cells were shifted from 39 to 31°C in the presence of cycloheximide, the polypeptide p40 was processed to lower-molecular-weight forms, and full nucleocapsids were detected in the cell nuclei. The aberrant polypeptide of the mutant, however, was not processed in cells mixedly infected with 17
ts
VP1201 and a revertant at the nonpermissive temperature, suggesting that the defect of the mutant was in the gene encoding p40 rather than in a gene of a processing enzyme.
Biochemical properties of the 145,000-dalton super-T antigen from simian virus 40-transformed BALB/c 3T3 clone 20 cells
Authors: Chaudry F , Belsham G J , Smith A E
Published: January 2020

SV3T3 C120 cells contain a 145,000-dalton form of simian virus 40 (SV40) super-T antigen but little if any normal-sized large-T. The subcellular location of super-T, its DNA binding properties, and its interaction with nonviral tumor antigen (NVT) were examined. Immunofluorescence microscopy and subcellular fractionation indicated that super-T is almost exclusively nuclear. Chromatography on double-stranded DNA-cellulose showed that super-T binds to double-stranded DNA and has an elution profile indistinguishable from normal-sized large-T. Super-T also binds specifically to a fragment of SV40 DNA which contains the origin of DNA replication. However, immunoprecipitation of super-T or large-T either with anti-tumor cell serum or with anti-NVT serum from fractions obtained by sucrose density centrifugation of 32P-labeled or [35S]methionine-labeled extracts revealed clear differences in the sedimentation characteristics of these proteins. The bulk of labeled 145,000-dalton super-T sedimented between 4S and 10S, whereas the bulk of 32P-labeled large-T from normal SV40-transformed cells sedimented as two peaks at 23S to 25S and 16S to 18S. By contrast, the sedimentation properties of NVT from the SV3T3 C120 cells were similar to those normally observed with other SV3T3 cell lines. The reason for this apparent difference in complex formation between super-T and NVT and that normally observed with large-T is unclear, but it probably has no deleterious effect on the ability of super-T to maintain transformation.
Reexamination of the Morphogenetic Block of a Putative Late-Stage, Temperature-Sensitive Mutant of Moloney Murine Leukemia Virus
Authors: Demsey Anthony , Friedman Robert M.
Published: January 2020

The
ts
3 temperature-sensitive mutant of Moloney murine leukemia virus has been reported to have a morphogenetic block in a late stage of the budding process. As evidence, previously published electron micrographs of cells maintained at the nonpermissive temperature (39°C) revealed numerous budding virions on the cell surface. However, it appears now that these micrographs reflected budding that occurred not at 39°C, but after cells were removed from the incubator before fixation. The morphogenesis of
ts
3 is actually blocked at an earlier stage of development.
Site-directed point mutation in the src gene oF rous sarcoma virus results in an inactive src gene product
Authors: Bryant D , Parsons J T
Published: January 2020

Site-directed mutagenesis techniques were used to construct defined point mutations within the src gene of the Prague A strain of Rous sarcoma virus. Bisulfite mutagenesis at a Bg/I restriction site in the src gene yielded three mutations which contained the same single base change, a guanine-to-adenine transition. The resulting genomes encoded an src protein containing a substitution of threonine for alanine at amino acid position 433. Transfection of chicken cells with mutagenized DNA did not result in cellular transformation even though the cells produced a pp60src. Immune complexes containing mutant pp60src did not phosphorylate immunoglobulin G heavy chain or casein.
Sequence Determination of the (+) Leader RNA Regions of the Vesicular Stomatitis Virus Chandipura, Cocal, and Piry Serotype Genomes
Authors: Giorgi Colomba , Blumberg Benjamin , Kolakofsky Daniel
Published: January 2020

Using 3′-end-labeled genome probes, cells infected with vesicular stomatitis virus Chandipura, Cocal, and Piry serotypes were shown to contain (+) leader RNAs of approximately 50 nucleotides in length. The nucleotide sequence of the leader RNA regions of these genomes was determined and compared with the previously reported sequences of both the (+) and (-) leader RNA regions of other vesicular stomatitis virus serotypes. Regions of strong conservation of nucleotide sequence among the various vesicular stomatitis virus serotypes suggest those nucleotides thought to be involved in control functions during vesicular stomatitis virus replication.
Nucleotide sequence of the env-specific segment of NFS-Th-1 xenotropic murine leukemia virus
Authors: Repaske R , O'Neill R R , Khan A S , Martin M A
Published: January 2020

The sequence of 863 contiguous nucleotides encompassing portions of the pol and env genes of NFS-Th-1 xenotropic proviral DNA was determined. This region of the xenotropic murine leukemia virus genome contains and env-specific segment that hybridizes exclusively to xenotropic and mink cell focus-forming but not to ecotropic proviral DNAs (C. E. Buckler et al., J. Virol. 41:228-236, 1982). The unique xenotropic env segment contained several characteristic deletions and insertions relative to the analogous region in AKR and Moloney ecotropic murine leukemia viruses. Portions of an endogenous env segment cloned from a BALB/c mouse embryo gene library that had a restriction map and hybridization properties typical of xenotropic viruses (A. S. Khan et al., J. Virol. 44:625-636, 1982) were also sequenced. The sequence of the endogenous env gene was very similar to the comparable region of the NFS-Th-1 xenotropic virus containing the characteristic deletions and insertions previously observed and could represent a segment of an endogenous xenotropic provirus.
Effects of Tunicamycin on Rotavirus Morphogenesis and Infectivity
Authors: Petrie Betty L. , Estes Mary K. , Graham David Y.
Published: January 2020

The functions of the two rotavirus glycoproteins were investigated by using tunicamycin and a variant of SA11 rotavirus having nonglycosylated VP7. Results showed that glycosylation of VP7 is not required for normal viral morphogenesis and infectivity and suggested that the nonstructural glycoprotein is involved in assembly of the outer capsid.
Genetic variability of herpes simplex virus: development of a pathogenic variant during passaging of a nonpathogenic herpes simplex virus type 1 virus strain in mouse brain
Authors: Kaerner H C , Schröder C H , Ott-Hartmann A , Kümel G , Kirchner H
Published: January 2020

Herpes simplex virus type 1 ANG (HSV-1 ANG) is originally nonpathogenic for inbred mice upon intraperitoneal intravenous, or intravaginal inoculation. In contrast, mice died of encephalitis within 4 to 5 days after intracerebral inoculation with this strain. HSV-1 ANG was serially passaged in mouse brains. In two independent series, peripherally pathogenic virus variants had developed and accumulated in the virus progeny after 12 to 15 intracerebral passages. In mixed infections both nonpathogenic and pathogenic viruses replicated at the primary site of infection and spread to various organs. However, only the pathogenic phenotype could be recovered from the spinal cord and the brain. Comparison of the restriction enzyme cleavage patterns of pathogenic ANG and nonpathogenic ANG virus DNAs revealed distinct alterations in the S-segment (US) sequences bounded by coordinates 0.953 and 0.958 in the prototype orientation and by coordinates 0.862 to 0.867 in the IS orientation of the viral genome. However, it is not known whether these alterations are physiologically relevant to the observed changes in pathogenicity. When coinjected intraperitoneally at 50 to 100-fold excess, the nonpathogenic HSV-1 ANG protected mice against its own pathogenic variant as well as against other pathogenic HSV-1 strains. Pathogenic HSV-1 ANG proved to be genetically and phenotypically stable for at least 25 serial passages in tissue culture at either high or low multiplicity of infection.