bluuebunny/crossref_metadata_2025 huggingface dataset:
Protease-sensitive transfection of Streptococcus pneumoniae with bacteriophage Cp-1 DNA
Authors: Ronda C , López R , Gómez A , García E
Published: January 2020

The transfecting activity of pneumococcal phage Cp-1 DNA was destroyed by treatment with proteolytic enzymes, although these enzymes did not affect transfection with bacteriophage Dp-4 DNA. This transfection was stimulated by calcium ions. Protease-treated Cp-1 DNA competes for binding and uptake with transforming pneumococcal DNA as well as with transfecting Dp-4 DNA to approximately the same extent as does untreated Cp-1 DNA. In addition, [3H]thymidine-labeled Cp-1 DNA, treated with proteases or untreated, was absorbed with the same efficiency. These data suggest that uptake of Cp-1 DNA is not affected by protease treatment. [3H]thymidine-labeled Cp-1 DNA showed remarkable resistance against surface nuclease activity of competent wild-type cells. The monomeric form of the Cp-1 DNA-protein complex showed a linear dose response in transfection.
Effects of ATP and inhibitory factors on the activity of vaccinia virus type I topoisomerase
Authors: Foglesong P D , Bauer W R
Published: January 2020

Vaccinia virus cores contain a type I topoisomerase which promotes the relaxation of superhelical DNA of either handedness (Bauer et al., Proc. Natl. Acad. Sci. U.S.A. 74:1841-1845, 1977). The activity of partially purified vaccinia virus topoisomerase (VV-Topo I) was determined in the presence of ATP, dATP, GTP, ADP, and ATP analogs in which hydrolysis of the alpha, beta or beta, gamma phosphate bond is restricted. Topoisomerase activity was stimulated 2.5-fold by the addition of 2 to 4 mM ATP or dATP to standard assay mixtures; 2 mM GTP produced no significant effect on enzyme activity. The addition of 2 mM beta, gamma-imido ATP or 2 mM gamma-thiophosphate ATP reduced VV-Topo I activity by 80 and 65%, respectively. In contrast, 4 mM alpha, beta-methylene ATP produced no significant change in topoisomerase activity compared to ATP itself. Assays performed in the presence of 4 mM ADP exhibited an 80% reduction in enzyme activity. The preparations of VV-Topo I used for these studies showed, however, no detectable DNA-dependent or -independent ATPase activity. The activity of VV-Topo I was similarly measured in the presence of the antibiotics novobiocin and coumermycin A1, which inhibited enzyme activity by 50% at concentrations of 180 and 40 microM, respectively. Comparable inhibition of VV-Topo I activity was observed in the presence of 1 mM beta, gamma-imido ATP. We determined that novobiocin inhibits vaccinia core transcription at the same concentrations which inhibit vaccinia core topoisomerase I activity. These results suggest that the vaccinia DNA topoisomerase may play a role in the ATP-dependent transcription of viral genes from intact core particles.
Two base changes restore infectivity to a noninfectious molecular clone of Moloney murine leukemia virus (pMLV-1)
Authors: Miller A D , Verma I M
Published: January 2020

The complete nucleotide sequence of a molecular clone of Moloney murine leukemia virus (pMLV-1) has previously been reported (Shinnick et al., Nature [London] 293:543-548, 1981). However, pMLV-1 does not generate infectious virus after transfection into cells (Berns et al., J. Virol. 36:254-263, 1980). The lesion in pMLV-1 has been localized by determining the biological activity of recombinants containing DNA from an infectious clone of Moloney murine leukemia virus (pMLV-48) and pMLV-1. Replacement of a 1.0-kilobase pair region which spans the gag-pol junction of pMLV-1 with the corresponding DNA fragment from the infectious clone restores its infectivity. Nucleotide sequence analysis of this fragment obtained from the infectious clone (pMLV-48) and pMLV-1 reveals two single base pair changes, one in the p30gag gene and the other in the 5' end of the pol gene. The mutation in the pol gene does not affect the production of infectious virus but renders them XC negative, whereas the mutation in the gag gene appears to be lethal. The complete nucleotide sequence of an infectious clone of Moloney murine leukemia virus can now be deduced.
Influenza virus hemagglutinin containing an altered hydrophobic carboxy terminus accumulates intracellularly
Authors: Sveda M M , Markoff L J , Lai C J
Published: January 2020

The influenza virus hemagglutinin (HA) glycoprotein synthesized from cloned DNA in a simian virus 40 vector is expressed on the surface of infected primate cells. Previously, it has been demonstrated that mutant HAs lacking the hydrophobic carboxy terminus fail to anchor on the cell surface and therefore are secreted extracellularly. During analysis of additional HA deletion mutants derived from an HA-simian virus 40 recombinant, we found a mutant with an altered hydrophobic carboxy terminus that exhibited another phenotype. This deletion mutant, dl-12, produced HA that was neither secreted nor expressed on the infected cell surface. The mutant HA was similar to the wild-type HA in apparent molecular weight and extent of glycosylation as assayed by endoglycosidase H sensitivity. The mutant HA localized near the perinuclear region of infected cells as indicated by an indirect immunofluorescence assay. Sequence analysis showed that a 5-base-pair deletion had occurred before the region encoding the hydrophobic carboxy terminus. Nevertheless, the physicochemical properties of the wild-type HA carboxy terminus were maintained in that the truncated HA carboxy terminus consisted of predominantly hydrophobic amino acids followed by several charged amino acids residues. This similarity in the carboxy terminus between the wild-type and mutant HAs may be responsible for the functional similarities observed. In spite of these similarities, the mutant HA failed to mature at the surface. These results suggest that the maturation of the mutant HA is blocked during a late stage in the transit to the cell surface.
Isolation and partial chemical characterization of a 64,000-dalton glycoprotein of human cytomegalovirus
Authors: Clark B R , Zaia J A , Balce-Directo L , Ting Y P
Published: January 2020

A guanidinium chloride extract of [3H]glucosamine- and [35S]methionine-labeled virions plus dense bodies of human cytomegalovirus (Towne) was separated by reverse-phase high-pressure liquid chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the eluate revealed the major peak to be a glycoprotein with a relative mass of 64,000. This glycoprotein (HCMVgp64) was characterized by amino acid analysis and a high-pressure liquid chromatographic map of its tryptic peptides.
Autonomous parvovirus LuIII encapsidates equal amounts of plus and minus DNA strands
Authors: Bates R C , Snyder C E , Banerjee P T , Mitra S
Published: January 2020

Autonomous parvoviruses are thought to uniquely encapsidate single-stranded DNA of minus polarity. In contrast, the defective adeno-associated viruses separately encapsidate equal amounts of plus and minus DNA strands. We reexamined the uniqueness of minus strand encapsidation for the autonomous parvoviruses. Although we found that Kilham rat virus and H-1 virus encapsidate varying but small amounts of complementary-strand DNA, it was unexpected to find that LuIII virus encapsidated equal amounts of plus and minus DNA. The extracted LuIII DNA possessed properties of double-stranded replicative-form DNA, including insensitivity to S1 endonuclease, cleavage by restriction enzymes, and conversion to unit-length, single-stranded DNA when electrophoresed under denaturing conditions. However, the inability of this DNA to form single-stranded DNA circles when denatured and then renatured in the presence of formamide and the lack of double-stranded DNA circle formation after treatment with exonuclease III and reannealing shows a lack of sequence homology of the 3' and 5' termini of LuIII DNA, in contrast to adeno-associated virus DNA. Digestion of LuIII double-stranded DNA with EcoRI and HincII and separation of plus and minus DNA strands on composite agarose-acrylamide gels identified a heterogeneity present only in the plus DNA strand. These results suggest that strand specificity of viral DNA encapsidation is not a useful property for differentiation between the autonomous and defective parvoviruses. Furthermore, encapsidation by LuIII of equal amounts of complementary DNA strands in contrast to encapsidation of minus strands by H-1 virus, when propagated in the same host cell type, suggests that selection of strands for encapsidation is a virus-coded rather than host-controlled event.
Lack of neutralizing antibodies to caprine arthritis-encephalitis lentivirus in persistently infected goats can be overcome by immunization with inactivated Mycobacterium tuberculosis
Authors: Narayan O , Sheffer D , Griffin D E , Clements J , Hess J
Published: January 2020

The pathogenesis of the persistent progressive diseases of sheep (visna-maedi) and goats (arthritis-encephalitis) is dependent on continuous replication of the causative lentiviruses. One subgroup of these viruses, Icelandic visna virus, accomplishes this form of replication by undergoing antigenic mutation. Mutant viruses arising late in the infection escape neutralization by antibodies directed to the parental virus. In contrast, we show here that viruses obtained from persistently infected sheep and goats with natural disease in this country do not induce virus-neutralizing antibodies, although antibodies to virus core proteins were produced. The lack of neutralizing antibodies was not overcome by hyperimmunization of animals with concentrated preparations of live or inactivated virus. Thus, failure to produce these specific antibodies was not due to lack of sufficient antigen or interference with the immune response because of the ability of these viruses to infect macrophages. The hyporesponsive state, however, was overcome by immunization of animals with virus and large amounts of inactivated Mycobacterium tuberculosis. Induction of agglutinating and neutralizing antibodies by this method was probably due to a unique form of antigen processing by macrophages activated by M. tuberculosis. Neutralizing antibodies were produced for the first time against the caprine arthritis-encephalitis virus by this method. These antibodies have similar biological properties to those induced by Icelandic visna virus. They belong to the immunoglobulin G1 subclass, they are effective against a narrow range of caprine arthritis-encephalitis viruses, and they identify (for the first time) antigenic variants among these caprine agents.
Predominant immediate-early transcripts of human cytomegalovirus AD 169
Authors: Jahn G , Knust E , Schmolla H , Sarre T , Nelson J A , McDougall J K , Fleckenstein B
Published: January 2020

Transcription of the human cytomegalovirus genome (strain AD 169) was investigated at the immediate-early (IE) time after infection, by using cycloheximide to suppress virus-specific protein synthesis. In total cell RNA, four predominant IE transcripts were found which were encoded by one contiguous region of the long unique segment between map units 0.06 and 0.16 in prototype arrangement of human cytomegalovirus AD 169 DNA. Analysis by Northern blot hybridizations demonstrated that the transcripts possessed a size of 1.9, 2.2, 2.3, and 5.0 kilobases, respectively. Coding sequences and directions of transcriptions were mapped by Northern blots and hybridizations with oligodeoxythymidylic acid-primed and randomly primed cDNA. The 1.9-, 2.2-, and 2.3-kilobase RNAs were found in the polyadenylated fraction of IE RNA exclusively; in contrast, a part of the 5.0-kilobase RNA appeared polyadenylated, although the majority of the same transcript was found in the nonpolyadenylated pool. Also, different from the other IE genes, the DNA coding for the 5.0-kilobase IE RNA was transcribed in high quantities during the late phase of virus replication, suggesting an exemption from the temporal regulation of herpesvirus transcription.
Analysis of an influenza A virus mutant with a deletion in the NS segment
Authors: Buonagurio D A , Krystal M , Palese P , DeBorde D C , Maassab H F
Published: January 2020

The influenza virus host range mutant CR43-3, derived by recombination from the A/Alaska/6/77 and the cold-adapted and temperature-sensitive A/Ann Arbor/6/60 viruses, has previously been shown to possess a defect in the NS gene. To characterize this defect, nucleotide sequence data were obtained from cloned cDNAs. The CR43-3 NS gene was found to be 854 nucleotides long and to derive from the NS gene of the A/Alaska/6/77 parent virus by an internal deletion of 36 nucleotides. Direct sequencing of RNA 8 of CR43-3 virus confirmed that the deletion in the NS1-coding region was not an artifact that was generated during the cloning procedure. Protein analysis indicated that the NS1 protein of CR43-3 virus was synthesized in equal amounts in the restrictive (MDCK) cells as well as in the permissive (PCK) host cells. Also, indirect immunofluorescence studies of virus-infected cells showed that the NS1 protein of CR43-3 virus, like that of the parent viruses, accumulates in the nuclei of both cell systems. Although no differences in synthesis or localization of the NS1 protein could be detected, a consistent reduction in M1 protein was noted in CR43-3 virus-infected, nonpermissive cells as compared with that of the permissive host. Since analysis of the CR43-3 virus required us to obtain the NS nucleotide sequence of the 1977 isolate A/Alaska/6/77, we were able to compare this sequence with those of corresponding genes of earlier strains. The result of this analysis supports the idea of a common lineage of human influenza A viruses isolated over a 43-year period.
Sequence homology between bovine and human adenoviruses
Authors: Hu S L , Hays W W , Potts D E
Published: January 2020

Cross-hybridization has been detected between corresponding regions of the genomes of bovine adenovirus type 3 and human adenovirus type 2. The most conserved region on the viral genomes encodes the hexon polypeptide. The nucleotide sequence of this region in bovine adenovirus type 3 has been determined. Comparison of the predicted amino acid sequences of the bovine adenovirus type 3 and human adenovirus type 2 hexon polypeptides reveals three regions of nonhomology.
Epstein-Barr virus-specific DNase activity in nonproducer Raji cells after treatment with 12-o-tetradecanoylphorbol-13-acetate and sodium butyrate
Authors: Ooka T , De Turenne M , De The G , Daillie J
Published: January 2020

An Epstein-Barr virus (EBV)-specific DNase was induced in EBV nonproducer Raji cells after treatment with 12-O-tetradecanoylphorbol-13-acetate and sodium butyrate. The increase in EBV DNase activity was related to the appearance of early antigen-positive cells. The enzyme had a sedimentation coefficient of 4S and was resistant to 300 mM KCl, and its induction did not depend on viral DNA synthesis. The EBV-specific DNase activity was specifically inhibited by sera from patients who had nasopharyngeal carcinoma with high early antigen activities but not by sera from normal, healthy individuals. There was a correlation between the degree of anti-EBV DNase activity and the titers of early antigen antibody.
Intertypic cross-reactions of nonneutralizing, monoclonal poliovirus antibodies
Authors: Vrijsen R , Rombaut B , Boeye A
Published: January 2020

Quantitative data are presented on (i) the intertypic cross-reactions of polyclonal, guinea pig antibodies directed against the N or H antigen of type 1 poliovirus and (ii) a set of five nonneutralizing, mouse hybridoma antibodies raised against N antigen or a mixture of capsid polypeptides VP1, VP2, and VP3. Three of these antibodies recognize H antigen and VP1, the fourth H antigen only, and the fifth VP3 only. The antibodies recognize either only homotypic antigens or the antigens of the three serotypes.
Two genetically transmitted BALB/c mouse mammary tumor virus genomes located on chromosomes 12 and 16
Authors: Callahan R , Gallahan D , Kozak C
Published: January 2020

We have examined EcoRI-restricted cellular DNA from BALB/c mouse-hamster somatic cell hybrids by blot hybridization for the presence of mouse mammary tumor virus-related sequences. Results of this analysis show that mouse mammary tumor virus-related proviral copies are located on chromosomes 16 (16-kilobase-pair fragment) and 12 (10.5- and 7.7-kilobase-pair fragments).
Attachment of Sendai virus particles to cell membranes: dissociation of adsorbed virus particles with dithiothreitol
Authors: Chejanovsky N , Beigel M , Loyter A
Published: January 2020

Sendai virus particles bind to human erythrocytes at 4 degrees C and fuse with them at 37 degrees C. The present work describes a new method by which adsorbed virus particles can be removed from human erythrocytes, allowing quantitative determination of the number of virus particles which can bind and fuse with human erythrocyte membranes. Through the use of 125I-labeled Sendai virus particles, it is shown that incubation with 50 mM dithiothreitol removed about 90 to 95% of adsorbed virus particles. Fused virus particles were resistant to treatment with dithiothreitol. Negligible amounts of 125I-labeled Sendai virus particles were removed by treatment with dithiothreitol after incubation of virus-cell complexes at 37 degrees C. Trypsinized virus particles were able to attach to, but not fuse with, human erythrocytes even after prolonged incubation at 37 degrees C. Treatment with dithiothreitol removed as much as 80 to 85% of trypsinized virus particles incubated with human erythrocytes at 37 degrees C. A quantitative determination revealed that about 1,000 to 1,200 and 600 to 800 Sendai virus particles can bind to or fuse with human erythrocytes, respectively.
DNA methylation inhibits the transfecting activity of replicative- form phi X174 DNA
Authors: Wang R Y , Shenoy S , Ehrlich M
Published: January 2020

Replacement of virtually all the cytosine residues with 5-methylcytosine residues in the complementary strand of the replicative form (RF) of phi X174 DNA caused a 300- to 500-fold loss in its transfecting activity. Similar results were obtained with analogously methylated M13 RF. Transfection experiments with phi X RF hemimethylated in only part of the molecule, as assessed by analysis with restriction endonucleases, indicated that gene A of phi X, which needs to be nicked at a specific site by the gene A protein for RF replication, was not the main target for this inhibition by DNA methylation. We propose that the loss of transfecting activity was due to hemimethylation of the phi X RF interfering with the processively catalyzed movement of the replication fork.
Adenovirus early region 1B 58,000-dalton tumor antigen is physically associated with an early region 4 25,000-dalton protein in productively infected cells
Authors: Sarnow P , Hearing P , Anderson C W , Halbert D N , Shenk T , Levine A J
Published: January 2020

In soluble protein extracts obtained from adenovirus productively infected cells, monoclonal antibodies directed against the early region 1B 58,000-dalton (E1B-58K) protein immunoprecipitated, in addition to this protein, a polypeptide of 25,000 molecular weight. An analysis of tryptic peptides derived from this 25K protein demonstrated that it was unrelated to the E1B-58K protein. The tryptic peptide maps of the 25K protein produced in adenovirus 5 (Ad5)-infected HeLa cells and BHK cells were identical, whereas Ad3-infected HeLa cells produced a different 25K protein. The viral origin of this 25K protein was confirmed by an amino acid sequence determination of five methionine residues in two Ad2 tryptic peptides derived from the 25K protein. The positions of these methionine residues in the 25K protein were compared with the nucleotide sequence of Ad2 and uniquely mapped the gene for this protein to early region 4, subregion 6 of the viral genome. A mutant of Ad5 was obtained (Ad5 dl342) which failed to produce detectable levels of the E1B-58K protein. In HeLa cells infected with this mutant, monoclonal antibodies directed against the E1B-58K protein failed to detect the associated 25K protein. In 293 cells infected with Ad5 dl342, which contain an E1B-58K protein encoded by the integrated adenovirus genome, the mutant produced an E4-25K protein which associated with the E1B-58K protein derived from the integrated genome. Extracts of labeled Ad5 dl342-infected HeLa cells (E1B-58K-) were mixed in vitro with extracts of unlabeled Ad5 wild type-infected HeLa cells or 293 cells (E1B-58K+). When the mixed extracts were incubated with the E1B-58K monoclonal antibody, a labeled E4-25K protein was coimmunoprecipitated. When extracts of Ad5 dl342-infected HeLa cells and uninfected HeLa cells (both E1B-58K-) were mixed, the E1B-58K monoclonal antibody failed to immunoselect the E4-25K protein. These data provide evidence that the E1B-58K antigen is physically associated with an E4-25K protein in productively infected cells. This is the same E1B-58K protein that was previously shown to be associated with the cellular p53 antigen in adenovirus-transformed cells.
Deletion and insertion mutations in early region 1a of type 5 adenovirus that produce cold-sensitive or defective phenotypes for transformation
Authors: Babiss L E , Fisher P B , Ginsberg H S
Published: January 2020

On the basis of earlier findings showing that H5hr1 (hr1) is cold sensitive for transformation, a series of mutants were constructed so that they contained deletions or insertions in different sites of early region 1a (E1a) to ascertain: (i) whether the cold-sensitive phenotype of hr1 was the result of the identified single-base pair deletion of nucleotide 1,055 or due to a missense mutation at another site and (ii) what region and how much of the E1a 51-kilodalton protein is actually required to produce cell transformation. A mutant, H5dl101 (dl101), was constructed to contain a 5-base pair deletion of nucleotides 1,008 to 1,012, which produced a frameshift and a subsequent stop codon at nucleotide 1,241. This mutant, which should encode a truncated 33-kilodalton protein in place of the wild-type 51-kilodalton protein, had a cold-sensitive phenotype for transformation essentially identical to hr1. Consonant with this finding, a mutant (H5in106) engineered to contain a 16-base pair insertion initiated after nucleotide 1,009, with a stop codon beginning at the newly inserted nucleotide 1,013, also had a cold-sensitive phenotype like hr1 and dl101. It is striking, however, that a mutant (H5dl105) with a 69-base pair deletion beginning at nucleotide 1,003, and having a stop codon at nucleotide 1,544, was totally defective for transformation at any temperature. Transfection studies with plasmids containing the E1a or E1a and E1b genes of sub309, hr1, and dl101 further revealed that the cold-sensitive transformation phenotype observed could be exhibited in the absence of viral E1b gene expression.
Mapping of the structural gene for the herpes simplex virus type 2 counterpart of herpes simplex virus type 1 glycoprotein C and identification of a type 2 mutant which does not express this glycoprotein
Authors: Zezulak K M , Spear P G
Published: January 2020

The gene encoding glycoprotein F (gF) of herpes simplex virus type 2 (HSV-2) was mapped to the region of the viral genome from 0.62 to 0.64 map units. This region is colinear with, and partially homologous to, the region of the HSV-1 genome previously shown to encode gC. Mapping of the gF gene was done by insertion of HSV-2 DNA fragments into the thymidine kinase gene of an HSV-1 virus and screening of the resultant recombinant viruses for the expression of gF. In this way, DNA sequences necessary for the expression of gF in infected cells were also delimited. Because several plaque morphology mutants (syncytial mutants) of HSV-1 have previously been shown to be gC-, a syncytial mutant of HSV-2 (GP) was tested for the expression of gF. It was found to be gF-, indicating that gF is not essential for replication of HSV-2 in cell culture, just as gC is not essential for replication of HSV-1. This result also suggests that the gF- and gC- phenotypes are related in the same, as yet undefined, way to the expression of a syncytial marker. A proposal to change the name of HSV-2 gF to gC (gC-2) is discussed.
Interferon-induced 2-5A synthetase activity in human peripheral blood mononuclear cells after immunization with influenza virus and rubella virus vaccines
Authors: Penn L J , Williams B R
Published: January 2020

The interferon-induced enzyme 2-5A synthetase can be a sensitive indicator of activation of the human interferon system during viral infection or interferon therapy. To determine the response of the human interferon system to viral antigens, the level of 2-5A synthetase activity was monitored in peripheral blood mononuclear cells of healthy adults before and after immunization with influenza or rubella virus vaccine. The influenza virus-vaccinated individuals demonstrated increases in enzyme activity on days 1 and 11 in vivo, whereas those vaccinated with rubella virus vaccine showed an increase only on day 11. The difference in the day 1 in vivo 2-5A synthetase response in the two vaccinated groups could be demonstrated by in vitro incubations of peripheral blood mononuclear cells isolated approximately 90 days postvaccination with the two vaccines. The day 11 increase of enzyme activity in the rubella virus group showed a positive correlation with an increase in serum antibody titer, suggesting activation of the interferon system during antibody production in vivo after human exposure to virus antigens. The demonstration of increased 2-5A synthetase activity at specific times postimmunization in this investigation indicates that the interferon system is involved in the human in vivo response to virus vaccination.
In vitro morphogenesis of foot-and-mouth disease virus
Authors: Grubman M J
Published: January 2020

Foot-and-mouth disease virion RNA is translated efficiently and completely in a rabbit reticulocyte lysate cell-free system. Treatment of cell-free lysates with monospecific serum prepared against the individual viral structural proteins or with monoclonal antibodies prepared against the inactivated virus or against a viral structural protein precipitated all of the structural proteins, suggesting that structural protein complexes were formed in vitro. Sucrose gradient analysis of the cell-free lysate indicated that complexes sedimenting at 5, 14, 60 to 70, and ca. 110S were assembled in vitro. Structural proteins VP0, VP1, and VP3 were the major polypeptides found in these complexes. The material sedimenting at 110S, i.e., containing VP0, VP1, and VP3, was precipitated by a 140S-specific monoclonal antibody but not by a 12S subunit-specific monoclonal antibody, suggesting that this capsid structure contained at least one epitope present on the intact virus.
Generation of mink cell focus-forming viruses by Friend murine leukemia virus: recombination with specific endogenous proviral sequences
Authors: Evans L H , Cloyd M W
Published: January 2020

A family of recombinant mink cell focus-forming viruses (MCF) was derived by inoculation of NFS mice with a Friend murine leukemia virus, and their genomes were analyzed by RNase T1-resistant oligonucleotide fingerprinting. The viruses were obtained from the thymuses and spleens of preleukemic and leukemic animals and were evaluated for dualtropism and oncogenicity. All these isolates induced cytopathic foci on mink cells but could be classified into two groups based on their relative infectivities for SC-1 (mouse) or mink (ATCC CCL64) cells. One group of Friend MCFs (F-MCFs) (group I) exhibited approximately equal infectivities for SC-1 and mink cells, whereas a second group (group II) infected mink cells 1,000- to 10,000-fold more efficiently than SC-1 cells. Structural analyses of the F-MCFs revealed that group I and group II viruses correlated with recombination of Friend murine leukemia virus with two distinct, but closely related, endogenous NFS proviral sequences. No correlation was found between the type of F-MCF and the tissue of origin or the disease state of the animal. Furthermore, none of the F-MCF isolates were found to be oncogenic in NFS/N or AKR/J mice. F-MCFs of both groups underwent extensive substitution of ecotropic sequences, involving much of the gag and env genes of group I F-MCFs and most of the gag, pol, and env genes of group II F-MCFs. All F-MCF isolates retained the 3' terminal U3 region of Friend murine leukemia virus. Comparison of the RNAs of the F-MCFs with RNAs of MCFs derived from NFS.Akv-1 or NFS.Akv-2 mice indicated that the F-MCFs were derived from NFS proviral sequences which are distinct from the sequences contained in NFS.Akv MCF isolates. This result suggested that recombination with particular endogenous proviral sequences to generate MCFs may be highly specific for a given murine leukemia virus.
Large-T-antigen-p53 complex formation is not cold sensitive in a cold-sensitive transformant induced by simian virus 40 mutant tsA1499
Authors: Bouck N , Fikes J , Rundell M K
Published: January 2020

F111 rat cells transformed by simian virus 40 mutant tsA1499 are cold sensitive for the expression of transformation. Yet, unlike F111 cells transformed by tsA58, they do not lose the ability to stabilize the transformation-associated host cell protein p53 at the temperature at which transformation is extinguished.
Lack of Nucleotide Sequence Relationship Among the Temperate Bacteriophage SPO2,
Bacillus subtilis
, and Virulent Bacteriophages β3 and β22
Authors: Yehle Clifford O. , Brenner Don J.
Published: January 2020

Radiolabeled phage SPO2 fragments were tested for the abiity to form interspecies deoxyribonucleic acid (DNA) duplexes with DNA from
Bacillus subtilis
and from phages β3 and β22. No reassociation, above control values, occurred between the DNA of phage SPO2 and that of its host or either of the virulent
B. subtilis
phages. It appears that extensive nucleotide sequence similarities between portions of host and phage DNA species are not necessary for the formation of the lysogenic state. The thermal elution profile of reassociated SPO2 DNA exhibited a normal distribution, indicating no heterogeneity in base composition. This differs from temperate enterophages whose DNA molecules contain regions of differing base composition.
Proteins Specified by Herpes Simplex Virus
Authors: Spear Patricia G. , Keller John M. , Roizman Bernard
Published: January 2020

Membranes prepared from HEp-2 cells infected with herpes simplex virus and free from soluble proteins, virus, ribosomes, and other cellular constituents were solubilized and subjected to electrophoresis on acrylamide gels. The electropherograms showed the following. (i) The synthesis of host proteins and glycoproteins ceases after infection. However, the spectrum of host proteins in membranes remains unaltered. (ii) Between 4 and 22 hr postinfection, at least four glycoproteins are synthesized and bound to the smooth cytoplasmic membranes. On electrophoresis, these glycoproteins form two major and two minor bands in the gel and migrate with proteins ranging from 50,000 to 100,000 daltons in molecular weight. (iii) The same glycoproteins are present in all membranes fractionated by density and in partially purified virus. The implications of the data are discussed.
Purification, Characterization, and Attempts at Isotopic Labeling of Mouse Interferon
Authors: Paucker K. , Berman B. J. , Golgher R. R. , Stanček D.
Published: January 2020

Under optimal conditions which minimized the accumulation of extraneous proteins, interferon preparations were obtained in L cells containing from 2 × 10
4
to 5 × 10
4
units/mg of protein. The radiolabeled proteins were liberated simultaneously with interferon from cultures exposed to tritiated amino acids after viral stimulation and from corresponding controls, and were subsequently purified with the following results. Chromatography of interferon on carboxymethyl-Sephadex C-25 eliminated selectively unlabeled or poorly labeled proteins, resulting in a greater than sixfold increase in counts per minute per milligram of protein. Similarly purified control material harbored at least 12 times less tritium per milligram of protein than interferon, and the label was more diversely distributed among proteins of different molecular weights. On electrophoresis of interferon in polyacrylamide gels, labeled proteins were reduced further by a factor of at least 10 without loss in titer. Final purification was estimated at greater than 280-fold, representing a calculated specific activity of at least 1.4 × 10
7
units of interferon per milligram of protein.
Chemical Studies on Polyoma and Shope Papilloma Viruses
Authors: Kass Sidney J.
Published: January 2020

Polyoma and Shope papilloma viruses were purified and analyzed by chemical and physical methods. Disc electrophoresis of degraded virions indicated the presence, in both cases, of only one major species of polypeptide subunit. The weight of the peptide chain of polyoma virus was estimated in 8
m
urea to be about 45,000 avograms, based on the sedimentation rate in a sucrose-urea gradient and the diffusion coefficient estimated from the differential migration in electrophoresis in gels of different pore size. The presence of a minor peptide of smaller size was suggested by carboxyl-terminal and sedimentation analyses. The amino acid composition of polyoma capsid protein was reported. Chemical analyses showed that polyoma virus and Shope papilloma virus contained 16 and 17.5% deoxyribonucleic acid, respectively. Light scattering by the polyoma virion showed it to have a molecular weight of 22 × 10
6
and a diameter of 54 nm.
Trypsinized Placental Cell Cultures for the Propagation of Viruses and as “Feeder Layers”
Authors: Grogan Elizabeth A. , Enders John F. , Miller George
Published: January 2020

The preparation and properties of cell cultures derived from human placental tissue are described and their usefulness for the propagation of animal viruses and as “feeder layers” is indicated.
Deoxyribonucleic Acid Replication in Simian Virus 40-Infected Cells
Authors: Levine Arnold J. , Teresky Angelika K.
Published: January 2020

Purified simian virus 40 (SV40) virions, grown in primary African green monkey kidney cells labeled prior to infection with
3
H-thymidine, contain a variable quantity of
3
H-labeled deoxyribonucleic acid (DNA). This DNA is resistant to deoxyribonuclease, sediments at 250
S
, and is enclosed in a particle that can be precipitated with SV40-specific antiserum. DNA-DNA hybridization experiments demonstrate that this
3
H-labeled component in purified SV40 virions is cellular DNA. When this
3
H-labeled DNA is released from purified virus with sodium dodecyl sulfate, it has an average sedimentation constant of 14
S
. Sedimentation through neutral and alkaline sucrose gradients shows that this 14
S
DNA is composed of a collection of different sizes of DNA molecules that sediment between 11 and 15
S
. As a result of this size heterogeneity, SV40 virions containing cellular DNA (pseudovirions) have a variable DNA to capsid protein ratio and exhibit a spectrum of buoyant densities in a CsCl equilibrium gradient. Pseudovirions are enriched, relative to true virions, on the lighter density side of infectious SV40 virus banded to equilibrium in a CsCl gradient. Little or no cellular DNA was found in purified SV40 virus preparations grown in BSC-1 or CV-1 cells.
Phenotypic Mixing of Envelope Proteins of the Parainfluenza Virus SV5 and Vesicular Stomatitis Virus
Authors: Choppin Purnell W. , Compans Richard W.
Published: January 2020

Cells mixedly infected with parainfluenza virus SV5 and vesicular stomatitis virus (VSV) yield phenotypically mixed virions, in addition to both parental types. Two types of phenotypically mixed virions have been identified: 0.6 to 1.2% of the VSV plaque formers were neutralized by SV5 antiserum, but
not
by VSV antiserum, suggesting the presence of a VSV genome in an SV5 envelope; 9 to 45% of the VSV plaque formers were neutralized by
both
antisera, indicating the presence of both SV5 and VSV antigens in their envelopes. The presence of SV5 antigen in virions with the typical bullet-shaped appearance of VSV was confirmed with ferritin-labeled anti-SV5 antibody. In contrast to standard VSV, phenotypically mixed virions adsorbed to and eluted from chicken erythrocytes, indicating that these virions contained in their envelopes SV5 hemagglutinin, and possibly neuraminidase. Thus, the VSV nucleocapsid can interact with membranes which contain SV5 proteins in the manner which leads to virus maturation, and the production of a high yield of phenotypically mixed virions with the morphology of VSV indicates that this process can function efficiently. No evidence of genetic recombination between the two viruses was found. These results raise the possibility of an evolutionary relatedness between the paramyxoviruses and the rhabdoviruses.
Inhibition of Synchronized Cellular Deoxyribonucleic Acid Synthesis During Newcastle Disease Virus, Mengovirus, or Reovirus Infection
Authors: Ensminger William D. , Tamm Igor
Published: January 2020

Cultures of L cells were synchronized with respect to deoxyribonucleic acid (DNA) synthesis with thymidine and 5-fluoro-2′-deoxyuridine (FUdR) and infected with Newcastle disease virus (NDV), mengovirus, or reovirus 3. Inhibition of incorporation of
3
H-cytidine into the DNA of synchronized cells is partially inhibited 2 hr after infection with NDV or mengovirus and nearly completely suppressed 4 hr after infection. With NDV and mengovirus, no evidence was obtained of differences in sensitivity of cells during early S phase as compared to later stages in DNA synthesis. When cells were infected with reovirus at the time of release from FUdR block, inhibition of cellular DNA synthesis was evident at 2 to 3 hr, and it was complete at 4 to 5 hr after infection. However, when cells were infected several hours prerelease, synthesis of DNA occurred in early S phase in spite of the fact that the cells had been infected for up to 6 hr. The results indicate that DNA synthesis in early S phase is relatively insensitive to the inhibitory function of reovirus. Colorimetric determinations (diphenylamine reaction) of the amounts of DNA produced in synchronized cells have substantiated the inhibition of DNA synthesis observed by isotope incorporation techniques.
Morphogenesis of Bittner Virus
Authors: Gay Frederick W. , Clarke John K. , Dermott Evelyn
Published: January 2020

The morphogenesis of Bittner virus (mouse mammary tumor virus) was studied in sectioned mammary tumor cells. Internal components of the virus (type A particles) were seen being assembled in virus factories close to the nucleus and were also seen forming at the plasma membrane. The particles in virus factories became enveloped by budding through the membrane of cytoplasmic vacuoles which were derived from dilated endoplasmic reticulum. Complete virus particles were liberated from these vacuoles by cell lysis. Particles budding at the plasma membrane were released into intercellular spaces. Maturation of enveloped virus occurred after release, but mature internal components were rarely seen in the cytoplasm before envelopment. Direct cell-to-cell transfer of virus by pinocytosis of budding particles by an adjacent cell was observed, and unusual forms of budding virus which participated in this process are illustrated and described. There was evidence that some virus particles contained cytoplasmic constituents, including ribosomes. Certain features of the structure of internal components are discussed in relation to a recently proposed model for the internal component of the mouse leukemia virus. Intracisternal virus-like particles were occasionally seen in tumor cells, but there was no evidence that these structures were developmentally related to Bittner virus.
Characterization of a host range mutant of human adenovirus 12 defective in early region 1B
Authors: Fukui Y , Shiroki K , Saito I , Shimojo H
Published: January 2020

We isolated an adenovirus 12 early region 1B mutant (in206B) by ligation of the cleaved DNA-protein complex and transfection of human embryo kidney cells with the ligation products. By deduction from the DNA sequencing analysis, the two polypeptides (with molecular weights of 19,000 and 54,000) coded in the early region 1B were fused in this mutant to produce a large polypeptide. This mutant could replicate efficiently in 293 cells but not efficiently in KB or human embryo kidney cells. In KB cells, viral DNA replication could not be detected after infection with in206B. In human embryo kidney cells, viral DNA replication did occur, but the transition of viral mRNA patterns from early to late did not occur even after DNA replication, resulting in failure to produce the late polypeptides. These results indicate that the early region 1B products may be involved in both viral DNA replication and regulation of transcription.
Transcriptional control of the bovine leukemia virus genome: role and characterization of a non-immunoglobulin plasma protein from bovine leukemia virus-infected cattle
Authors: Gupta P , Kashmiri S V , Ferrer J F
Published: January 2020

Using cloned bovine leukemia virus (BLV) DNA as a probe in the dot blot hybridization technique, we demonstrated that the expression of the BLV genome in infected lymphocytes is blocked in vivo at the transcriptional level. This blocking effect is due to a non-immunoglobulin protein present in the plasma but not in the serum of BLV-infected cattle. The plasma BLV-blocking protein also blocks the expression of the BLV genome in fibroblast cells of bovine and nonbovine origin infected with BLV in vitro. The plasma BLV-blocking factor has no inhibitory effect on the expression of Rauscher murine leukemia virus and feline leukemia virus in monolayer culture. The plasma BLV-blocking factor is not an interferon molecule. As determined by gel filtration chromatography, the plasma BLV-blocking factor has an apparent molecular weight of ca. 150,000.
Fv-1 host cell restriction of friend leukemia virus: microinjection of unintegrated viral DNA
Authors: Chinsky J , Soeiro R , Kopchick J
Published: January 2020

The murine gene Fv-1 has been shown to exert a major influence over the replication of ecotropic murine leukemia viruses. Studies of the replication of Friend murine leukemia virus have shown that the restriction of viral replication occurs intracellularly after the initiation of viral DNA synthesis. The precise mechanism of the block imposed by the Fv-1 gene product is not completely understood. Our studies of Fv-1 restrictive infection have shown a variable decrease in the accumulation of intracellular unintegrated form I viral DNA. Analysis by microinjection of the viral DNA formed in nonpermissively infected BALB/c cells indicates that this DNA is infectious. These studies indicate that the form I DNA accumulated in nonpermissively infected BALB/c cells contains the complete viral sequences necessary for the production of viral progeny, and therefore, they suggest that the Fv-1 host restrictive mechanism recognizes viral factors other than form I DNA alone. These results support the possibility that Fv-1 host restriction occurs after formation of infectious viral DNA, perhaps at the integration step itself.
Different bovine leukemia virus-induced tumors harbor the provirus in different chromosomes
Authors: Grégoire D , Couez D , Deschamps J , Heuertz S , Hors-Cayla M C , Szpirer J , Szpirer C , Burny A , Huez G , Kettmann R
Published: January 2020

DNAs from 25 independent hamster X bovine somatic cell hybrids were submitted to the Southern blot analysis with probes made of unique cellular fragments adjacent to single-copy proviruses from three different bovine tumors. It appeared that these cellular sequences, and thus the respective proviruses, belonged to three different chromosomes in the three different tumors examined.
Role of T-lymphocyte subsets in recovery from herpes simplex virus infection
Authors: Larsen H S , Feng M F , Horohov D W , Moore R N , Rouse B T
Published: January 2020

Our investigations probed the nature of different T-lymphocyte subsets effecting clearance of herpes simplex virus after infection of the pinna. Cell populations from animals recently infected subcutaneously or intraperitoneally (acute population) or from animals infected 6 weeks previously (primed population) or the latter cells reimmunized in vitro with virus (memory population) were studied. Viral clearance was a function of the Lyt 1+2- subset in the acute population, but with the memory population both Lyt 1+ and Lyt 2+ cells affected clearance. In primed populations, viral clearance was effected only by the Lyt 2+ subset. The ability of the various cell populations to adoptively transfer delayed-type hypersensitivity was also studied. Only acute population cells from animals infected subcutaneously and memory population cells transferred delayed-type hypersensitivity. In both cases, the cell subtype was Lyt 1+2-. Our results demonstrated that the delayed-type hypersensitivity response does not always correlate with immunity to herpes simplex virus. Multiple subsets of T cells participate in viral clearance, and their respective importances vary according to the stage of the virus-host interaction.
Isolation and characterization of monopinocytotic vesicles containing polyomavirus from the cytoplasm of infected mouse kidney cells
Authors: Griffith G R , Consigli R A
Published: January 2020

Monopinocytotic vesicles containing polyomavirus were isolated from the cytoplasm of mouse kidney cells infected with polyomavirus using sucrose density gradients. Nonenclosed, membrane-associated virions released by the action of neuraminidase separated from vesicle-enclosed virions in the sucrose gradient. Marker enzyme assays indicated the derivation of the vesicle membrane from the plasma membrane of the cell. The 125I-labeled virus enclosed in the vesicle sedimented more slowly in the gradient and was not observed unless infection and endocytosis had occurred. Detergent treatment of virion-containing vesicles caused the release of polyomavirus with sedimentation properties similar to those of purified polyoma virions. In addition, sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of virion proteins from vesicles containing virions demonstrated patterns of proteins similar to those of purified intact virions. Electron microscopy confirmed the presence of single intact virions inside vesicles. The study of these monopinocytotic virion-containing vesicles represents a further step in elucidating the early events of polyomavirus infection.
Detection of vesicular stomatitis virus RNA and its defective-interfering particles in individual mouse brains
Authors: Cave D R , Hagen F S , Palma E L , Huang A S
Published: January 2020

To develop a highly sensitive and direct assay for defective interfering (DI) particles of vesicular stomatitis virus (VSV), we reverse transcribed RNA from DI particles and cloned the DNA in pBR322 and used it as hybridization probes. At the lower limit, cDNA of about 850 nucleotides detected 150 pg of VSV RNA. For differentiation of hybridizable sequences found in the RNA of DI particles from complementary or identical sequences in the L mRNA or standard genomic RNA of VSV, RNA obtained from mouse brains was first separated by size, blotted onto nitrocellulose, and then hybridized to in vitro-labeled cDNA probe. Genomic VSV, DI, or L mRNA sequences from one-half of the brain of an infected mouse were detectable, whereas uninfected mice failed to react with this specific probe. When mice were infected intranasally with 10(8) PFU of standard VSV, most of them died between days 6 and 7, and the detection of standard genomic RNA correlated with paralysis and death. DI RNA was not detected in these mice. When mice were infected with 10(8) PFU of standard VSV together with an equivalent amount of DI particles, similar results were obtained. When fewer DI particles were inoculated together with standard virus, significant protection of mice occurred together with the detection of DI RNA. These results indicate that DI particles are protective in vivo and that the details of the virus-host interaction may resemble the cyclic growth patterns in cell cultures for standard VSV and its DI particles.