bluuebunny/crossref_metadata_2025 huggingface dataset:
Avian reticuloendotheliosis virus: characterization of the high-molecular-weight viral RNA in transforming and helper virus populations
Authors: Gonda M A , Rice N R , Gilden R V
Published: January 2020

Reticuloendotheliosis virus strain T (REV-T) is a type C retrovirus known to transform avian fibroblasts, spleen cells, and bone marrow cells and to produce virulent reticuloendotheliosis in young chicks. Analysis of REV-T high-molecular-weight RNA by electrophoresis in denaturing gels and by electron microscopy revealed the presence of at least two classes of molecules. One class appeared in CH3HgOH gels to have a monomer length of 9.3 kilobases (kb); in electron microscopic spreads under mildly denaturing conditions, it existed as a typical retrovirus dimer, having a monomer length of 8.8 +/- 0.7 kb. The second class also existed as a dimer, with a monomer length of 5.7 kb in CH3HgOH gels. Hybridization with REVA-A 32P-labeled complementary DNA revealed a third size class of molecules (4.7 kb), which were not resolvable from the 5.7-kb class in electron microscope spreads and which comigrated with chicken 28S rRNA in denaturing gels. Only the 9.3-kb class was found in the reportedly nontransforming virus produced after infection of canine thymus cell line with REV-T. Thus, REV-T appears to be similar to the murine and feline sarcoma viruses and the avian acute leukemia viruses in that it consists of a nontransforming helper virus genome and a defective genome responsible for oncogenicity. Our previous results demonstrated the presence in REV-T and in uninfected chicken cellular DNA of some nucleotide sequences not found in virus produced by the canine line (S. Simek and N. Rice, J. Virol. 33:320--329, 1980). In this report we show by hybridization with highly specific 32P-labeled complementary DNAs that REV-T-specific sequences exist within the 5.7-kb genome. Since 32P-labeled complementary DNA synthesized from the canine-derived virus genome hybridized with all three classes of RNAs, we conclude that the 5.7-kb genome is a recombinant between some sequences found in the putative helper and some sequences specific to REV-T. As with the other oncogenic viruses mentioned above, these specific sequences appear to be derived from host DNA.
Transformation of rat cells by fusion-infection with Rous sarcoma virus
Authors: Kawai S
Published: January 2020

The transformation of a rat cell line, 3Y1, by nonmammalian tropic strains of avian sarcoma virus was tested using cell-virus fusion mediated by Sendai virus or polyethylene glycol. Furthermore, the establishment of several transformed 3Y1 cell clones induced by the Schmidt-Ruppin strain of Rous sarcoma virus (RSV), its derivative mutants, and the Bryan high-titer strain of RSV is reported. The presence and expression of the viral genomes in these cells were examined, and all transformed cell clones tested were found to contain rescuable RSV genomes when they had been fused with normal chicken embryo fibroblast cells or those preinfected with Rous-associated virus type 1. However, the gag gene product, pr76, was barely detectable in wild-type RSV-transformed cells, whereas it was produced in considerable amounts in cells transformed by env-deleted mutants, the Bryan high-titer strain of RSV and NY8 derived from the Schmidt-Ruppin strain of RSV.
Proposal for a uniform nomenclature for defective interfering viruses of vesicular stomatitis virus
Authors: Reichmann M E , Bishop D H , Brown F , Crick J , Holland J J , Kang C Y , Lazzarini R , Moyer S , Perrault J , Prevec L , Pringle C R , Wagner R R , Youngner J S , Huang A S
Published: January 2020

Defective interfering particles of vesicular stomatitis virus have been named according to their parental derivation and to their genomic length and physical properties. This suggested uniform nomenclature can be adapted for other virus systems.
Shutoff of HeLa cell protein synthesis by encephalomyocarditis virus and poliovirus: a comparative study
Authors: Jen G , Detjen B M , Thach R E
Published: January 2020

Previous experimental results have suggested that poliovirus and encephalomyocarditis (EMC) virus employ very different mechanisms for shutting off host protein synthesis. However, this conclusion is suspect, inasmuch as different cell types were used for the two viruses; hence the apparent mechanistic differences might be specific for cell type and not virus type. To test this possibility we compared shutoff mechanisms in poliovirus- and EMC virus-infected HeLa cells. Striking differences were seen: poliovirus-induced shutoff was much more rapid and extensive than that induced by EMC virus; relative translation rates of certain host proteins were inhibited to different extents by the two viruses; initiation factors prepared from poliovirus-infected cells were specifically defective for translation of capped mRNA's in vitro, whereas those from EMC virus-infected cells were not. These results indicate that EMC virus and poliovirus employ different mechanisms for the shutoff of HeLa cell protein synthesis. This conclusion is consistent with much earlier work and indicates that many differences previously reported are specific to virus type.
Shutoff of lambda gene expression by bacteriophage T4: role of the T4 alc gene
Authors: Pearson R E , Snyder L
Published: January 2020

Bacteriophage T4 normally contains 5-hydroxymethylcytosine instead of cytosine in its DNA. Multiple mutants of T4 which synthesize DNA with cytosine do not transcribe their late genes due to the action of the T4 alc gene (Snyder et al., Proc. Natl. Acad. Sci. U.S.A. 73:3098--3102, 1976), which is also responsible for unfolding the host nucleoid after T4 infection (Sirotkin et al., Nature [London] 265:28--32, 1977; Tigges et al., J. Virol. 24:775--785, 1977). It seems reasonable that T4 alc function plays a role in shutting off host transcription, and the observation that some of the RNA made after infection with a T4 alc mutant hybridizes to Escherichia coli DNA (Sirotkin et al., Nature [London] 265:28--32, 1977; Tigges et al., J. Virol. 24:775--785, 1977) supports this hypothesis. Although it is likely that the roles of the alc function in the blocking of some types of transcription and in the unfolding of the host nucleoid are related, it is not known how these effects are achieved or, in fact, whether all types of transcription are affected equally by the alc function. In an attempt to answer these questions, we studied the effect of T4 alc function on bacteriophage lambda transcription and on the structure of intracellular lambda DNA. We found that the alc function is responsible for the shutoff of lambda late transcription but probably not for the shutoff of lambda early transcription. We also found that alc does not block lambda transcription by directly removing the supercoils from circular lambda DNA via either a nicking or topoisomerase activity. Furthermore, we conclude that T4 infection also prevents the translation of non-T4 mRNA because late lambda mRNA's were made after superinfection by a T4 alcs mutant and were of normal length but were not translated into lambda late proteins.
Phenotype of polyoma-induced hamster tumor cells lines
Authors: Israel M A , Martin M A , Miyamura T , Takemoto K K , Rifkin D , Pollack R
Published: January 2020

Cell lines from polyoma-induced hamster tumors exhibit a fully transformed phenotype despite the absence of the 105K (105,000-dalton) form of polyoma T-antigen.
Transformation of Primary Hamster Brain Cells with JC Virus and Its DNA
Authors: Frisque R. J. , Rifkin D. B. , Walker D. L.
Published: January 2020

We transformed primary hamster brain cells with four isolates of JC virus and JC virus DNA. Several properties of these transformants were characterized and compared to those of simian virus 40 transformants isolated under identical conditions.
Enzyme Activities Associated with an Invertebrate Iridovirus: Protein Kinase Activity Associated with Iridescent Virus Type 6 (
Chilo
Iridescent Virus)
Authors: Monnier Chantal , Devauchelle Gerard
Published: January 2020

Iridescent virus type 6 was found to contain an endogenous protein kinase activity which can phosphorylate some viral proteins and exogenous basic proteins. The enzyme required a divalent metal ion but was not stimulated by cyclic nucleotides. Procedures which are known to solubilize the viral envelope indicated that the protein kinase was an internal component of the virion. Conditions for protein kinase activity are described.
Polyoma virus middle t antigen: a tumor progression factor
Authors: Seif R
Published: January 2020

Polyoma virus (PyV) deletion mutant dl23 (affecting both large T and middle t but not small t antigens) was used to study transformation of 3T3 rat cells. This mutant generated stable transformants in the agar assay at a frequency similar to that of wild-type virus (WT). However, WT-induced transformants were detected 3 weeks after infection, whereas those induced by the mutant could not be detected until 6 to 8 weeks after infection. In this respect, dl23 PyV behaved like WT simian virus 40 (SV40). Cells transformed by WT SV40 or by dl23 PyV were similar in all their transformed properties. Those transformed by WT PyV were different from the others on the basis of morphology, cell adhesion to the substrate, release of protease activity, efficiency of doubling in agar, growth rate, and time required for tumor formation. Saturation density, the ability to grow in agar, the serum requirement for cloning, and the ability to grow on a cell monolayer were similar for all transformants. Middle t antigen enhanced membrane alterations and growth rate of the transformed cells, shortening the time required for tumor formation in rats.
Mutations in Coliphage P1 Affecting Host Cell Lysis
Authors: Walker Jean Tweedy , Walker Donald H.
Published: January 2020

A total of 103 amber mutants of coliphage P1 were tested for lysis of nonpermissive cells. Of these, 83 caused cell lysis at the normal lysis time and have defects in particle morphogenesis. Five amber mutants, with mutations in the same gene (gene 2), caused premature lysis and may have a defect in a lysis regulator. Fifteen amber mutants were unable to cause cell lysis. Artificially lysed cells infected with five of these mutants produced viable phage particles, and phage particles were seen in thin sections of unlysed, infected cells. However, phage production by these mutants was not continued after the normal lysis time. We conclude that the defect of these five mutants is in a lysis function. The five mutations were found to be in the same gene (designated gene 17). The remaining 10 amber mutants, whose mutations were found to be in the same gene (gene 10), were also unable to cause cell lysis. They differed from those in gene 17 in that no viable phage particles were produced from artificially lysed cells, and no phage particles were seen in thin sections of unlysed, infected cells. We conclude that the gene 10 mutants cannot synthesize late proteins, and it is possible that gene 10 may code for a regulator of late gene expression for P1.
Shared antigenic determinants between two distinct classes of proteins in cells infected with herpes simplex virus
Authors: Zweig M , Heilman C J , Rabin H , Hampar B
Published: January 2020

Guinea pig antisera and mouse monoclonal antibodies against a 40,000-molecular-weight nucleocapsid protein (p40) of herpes simplex virus types 1 and 2 immunoprecipitated 40,000- and 80,000-molecular-weight classes of soluble proteins from infected cell extracts. The soluble 40,000-molecular-weight protein class (intracellular p40) appeared as a cluster of three to four closely spaced bands of proteins having molecular weights ranging between 39,000 and 45,000, whereas the soluble 80,000-molecular-weight protein class (intracellular p80) appeared as a doublet of bands. The peptide map of intracellular p40 closely resembled the maps of the p40 and p45 proteins of nucleocapsids, but it showed both differences and similarities when compared with the peptide map of intracellular p80. Pulse-chase experiments suggested that intracellular p80 was not a precursor of intracellular p40. We conclude that the intracellular p40 and p80 protein classes share common antigenic determinants, presumably reflecting similar amino acid sequences, although they have distinct differences in protein structure.
Transcriptional map for Newcastle disease virus
Authors: Collins P L , Hightower L E , Ball L A
Published: January 2020

A transcriptional map of Newcastle disease virus was determined by measuring the kinetics of UV inactivation of the transcription of individual genes and of viral infectivity. The inactivation of single genes was monitored by measuring the reduction in the accumulation of viral gene products in vivo and in vitro. In vivo, the accumulation of viral polypeptides in infected cells was measured after reversal of a cycloheximide treatment designed to inhibit secondary transcription. Actinomycin D and a hypertonic medium were used to decrease selectively the synthesis of host cell polypeptides in infected cells. In vitro, mRNA's synthesized by irradiated viruses were analyzed by translation in cell-free systems under conditions in which the amount of each polypeptide synthesized reflected the relative abundance of the corresponding mRNA. UV target sizes were obtained for the genes coding for the HN, F0, NP, M, L, and P polypeptides; the 47,000-dalton protein was not detected. A comparison of the UV target sizes with the corresponding gene sizes suggested that transcription of these genes initiated at a single promotor and proceeded in the order NP, P, (F0, M), HN, L. These experiments were performed with Newcastle disease virus strains Australia-Victoria and B1-Hitchner; for both strains, two forms of the P polypeptide which differed in electrophoretic mobility were detected. Proof that the P protein is virus specific was obtained. In addition, infection of chicken embryo cells with avirulent strain B1-Hitchner enhanced the accumulation of at least four polypeptides that appeared to be specified by the host cell rather than by the infecting virus.
Effect of defective interfering particles on plus- and minus- strand leader RNAs in vesicular stomatitis virus-infected cells
Authors: Leppert M , Kolakofsky D
Published: January 2020

Vesicular stomatitis virus-infected cells contain short RNA transcripts, called leader RNAs, which are coded by the exact 3' end of both the minus-strand and plus-strand nucleocapsid templates. The molar amounts of both the plus-strand leader RNA (which is templated from the minus-strand genome) and the minus-strand leader RNA (which is templated from the plus-strand antigenome) were determined both in standard-virus- and mixed-virus-infected cells by using end-labeled genome probes. The results demonstrate that the presence of defective interfering particles in the infecting virus stock decreases the amount of plus-strand leader RNA but increases the amount of minus-strand leader RNA found in the infected cells. In addition, considerably more minus-strand leader RNA per mole of nucleocapsid template is synthesized in mixed-virus-infected cells than plus-strand leader RNA per mole of nucleocapsid template in both standard-virus- and mixed-virus-infected cells.
Identification of poliovirus polypeptide P63 as a soluble RNA-dependent RNA polymerase
Authors: Van Dyke T A , Flanegan J B
Published: January 2020

A poliovirus-specific RNA-dependent RNA polymerase was isolated from a cytoplasmic extract of infected HeLa cells and was shown to copurify with a single virus-specific protein. The polymerase was isolated from cells labeled with [35S]-methionine and was fractionated from other soluble cytoplasmic proteins by ammonium sulfate precipitation, phosphocellulose chromatography, gel filtration on Sephacryl S-200, and chromatography on hydroxylapatite. The activity of the enzyme was measured by using either polyadenylic acid or poliovirion RNA as a template in the presence of an oligouridylic acid primer. A single virus-specific protein that had an apparent molecular weight of 63,000 (p63) was found to copurify with this activity. Host-coded proteins were present in reduced molar amounts relative to p63. Noncapsid viral protein 2 (NCVP2) and other viral proteins were clearly separated from p63 by gel filtration on Sephacryl S-200. Polymerase activity coeluted from the column precisely with p63. NCVP2 was totally inactive as an RNA polymerase and did not stimulate the polymerase activity of p63. The purified enzyme sedimented at about 4S on a glycerol gradient and thus appeared to be a monomer of p63. Two-dimensional gel electrophoresis of the polymerase protein indicated that it had an isoelectric point of about 7.5. Thus, the viral polypeptide, p63, as defined by the above physical parameters, is an RNA-dependent RNA polymerase that can copy poliovirion RNA when oligouridylic acid is used as a primer.
Activation of mouse lymphocytes by vesicular stomatitis virus
Authors: Goodman-Snitkoff G W , McSharry J J
Published: January 2020

Vesicular stomatitis virus (VSV) is a mitogen for mouse spleen cells, and infectious virus is not required for mitogenesis. At concentrations between 10 and 100 microgram per culture, VSV stimulated DNA synthesis and blast transformation. Maximal activation by VSV occurred 48 h after culture initiation. Spleen cells depleted of T-lymphocytes by treatment with anti-Thy 1.2 and complement and those obtained from congenitally athymic BALB/c nu/nu mice were activated by VSV, suggesting that VSV is a B-cell mitogen. Activation of spleen cells was independent of the host in which the virus was grown, since VSV grown in BHK-21, HKCC, or MDBK cells was mitogenic. The mitogenesis was specific for VSV, since MDBK cell-grown WSN influenza virus was not a mitogen in this in vitro activation system, VSV-specific antibody prevented VSV mitogenesis, and VSV was mitogenic for spleen cells from C3H/HeJ mice which were resistant to mitogenesis by endotoxin.
den V gene of bacteriophage T4 determines a DNA glycosylase specific for pyrimidine dimers in DNA
Authors: Seawell P C , Smith C A , Ganesan A K
Published: January 2020

Endonuclease V of bacteriophage T4 has been described as an enzyme, coded for by the denV gene, that incises UV-irradiated DNA. It has recently been proposed that incision of irradiated DNA by this enzyme and the analogous "correndonucleases" I and II of Micrococcus luteus requires the sequential action of a pyrimidine dimer-specific DNA glycosylase and an apyrimidinic/apurinic endonuclease. In support of this two-step mechanism, we found that our preparations of T4 endonuclease V contained a DNA glycosylase activity that produced alkali-labile sites in irradiated DNA and an apyrimidinic/apurinic endonuclease activity that converted these sites to nicks. Both activities could be detected in the presence of 10 mM EDTA. In experiments designed to determine which of the activities is coded by the denV gene, we found that the glycosylase was more heat labile in extracts of Escherichia coli infected with either of two thermosensitive denV mutants than in extracts of cells infected with wild-type T4. In contrast, apyrimidinic/apurinic endonuclease activity was no more heat labile in extracts of the former than in extracts of the latter. Our results indicate that the denV gene codes for a DNA glycosylase specific for pyrimidine dimers.
Preparation and characterization of specific antisera to individual glycoprotein antigens comprising the major glycoprotein region of herpes simplex virus type 1
Authors: Eberle R , Courtney R J
Published: January 2020

The major glycoprotein complex (VP123) of herpes simplex virus type 1 resolved by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis was purified and further fractionated into two major and two minor components by chromatography of the isolated VP123 region on SDS-hydroxylapatite columns. The two major components (gC and gA/gB) were purified free of other polypeptides and used to prepare specific antisera to these glycoproteins. Radioimmune precipitation demonstrated that these antisera were specific for the antigens used in their production. These two antisera as well as an anti-VP123 serum were further characterized by immunoprecipitation, neutralization, and membrane immunofluorescence techniques. Results indicate that both of the major glycoprotein antigens are expressed on the surface of virions as well as on the surface of infected cells.
Expression of the gene for the polyoma small T antigen in Escherichia coli
Authors: Horwich A , Koop A H , Eckhart W
Published: January 2020

A cloned segment of the polyoma virus genome encoding the small T antigen has been fused, in the correct phase for translation, to the 5' end of the beta-galactosidase gene. The hybrid gene, cloned in Escherichia coli, produces a protein resembling the small T antigen.
Rabies mRNA translation in Xenopus laevis oocytes
Authors: Wunner W H , Curtis P J , Wiktor T J
Published: January 2020

Two rabies virus-specific mRNA species were identified by analysis of their encoded proteins after translation of the partially purified species in Xenopus laevis oocytes. One of these coded for the virion surface glycoprotein (G protein), and the other coded for the major structural protein of the virion nucleocapsid (N protein). The G-mRNA sedimented in a sucrose density gradient at about 18S, and the N-mRNA had a sedimentation coefficient of approximately 16S. Their respective translation products were identified in a radioimmunoassay with specific monoclonal antibody probes that recognized only G or N proteins. Immunoprecipitates formed between the radiolabeled viral antigens synthesized in programmed oocytes and their respective monoclonal antibodies were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The glycoprotein antigen translated from G-mRNA in oocytes migrated in the gel ahead of the virion G protein with a migration rate that was similar to that of nonglycosylated intracellular glycoproteins from virus-infected cells. The results suggested that the branched-chain carbohydrate of G protein was not required for recognition by the particular monoclonal antibody used. The nucleocapsid antigen translated from N-mRNA in oocytes migrated to the same position in the gel as marker virion N protein. Both the electrophoretic mobility of virus-specific antigens in sodium dodecyl sulfate-polyacrylamide gel and the antibody concentration dependence for immunoprecipitations were criteria for identifying the individual viral proteins encoded by the two rabies mRNA's.
Anogenital warts contain several distinct species of human papillomavirus
Authors: Krzyzek R A , Watts S L , Anderson D L , Faras A J , Pass F
Published: January 2020

Anogenital warts from 26 patients were examined for the presence of human papillomavirus (HPV). Although no whole, intact virus could be identified, varying amounts of nonintegrated HPV DNA were detected in 18 tissue specimens (70%) by employing both an agarose gel-ethidium bromide staining method and the Southern blot hybridization procedure. When hybridization analysis was performed under stringent conditions, six anogenital warts were observed to contain HPV genomic sequences related to either of the cutaneous viruses HPV type 1 (HPV-1) or HPV-2. In 12 tissue samples lacking sequence homology to either HPV-1 or HPV-2 under stringent conditions, HPV-related sequences were detected when the hybridization was performed under less stringent conditions, indicating that an HPV distinct from both HPV-1 and HPV-2 is also associated with these lesions. This anogenital HPV also appeared to be distinct from the other characterized types of HPV. These data indicate that at least three HPVs are associated with anogenital wart disease.
New endogenous herpesvirus of guinea pigs: biological and molecular characterization
Authors: Bia F J , Summers W C , Fong C K , Hsiung G D
Published: January 2020

Two known guinea pig herpesviruses, guinea pig cytomegalovirus (GPCMV) and guinea pig herpes-like virus (GPHLV), and well characterized. A third herpesvirus (GPXV) was originally isolated from leukocytes of healthy strain 2 guinea pigs. Growth of GPXV in guinea pig embryo fibroblastic cells produced a characteristic cytopathic effect. Electron microscopy of guinea pig cells infected with GPXV revealed the morphological development of a herpesvirus. Cross-neutralization tests and immunoferritin electron microscopy demonstrated that GPXV, GPCMV, and GPHLV were serologically distinct herpeviruses of guinea pigs. To confirm the distinction between these three herpesviruses, DNA genomes were compared by CsCl equilibrium buoyant density measurements and restriction endonuclease cleavage analysis. 32P-labeled viral DNA ws obtained from nucleocapsids isolated from virus-infected cells, and the buoyant density of GPXV DNA differed from that of GPCMV and GPHLV. Cleavage of viral DNAs with restriction endonucleases followed by gel electrophoresis revealed distinct patterns for each virus.
Simian virus 40-Chinese hamster kidney cell interaction. V. Cooperative effect of 5-iodo-2'-deoxyuridine and mitomycin C in the enhancement of virus replication in infected cells
Authors: Suárez H G , Lavialle C , Cassingena R
Published: January 2020

A cooperative effect of 5-iodo-2'-deoxyuridine and mitomycin C which amplified (10- to 50-fold) their stimulating effect on simian virus 40 replication was obtained in infected Chinese hamster kidney cells. The results indicate that these drugs act at different levels to modify the permissiveness of cells to simian virus 40.
Mechanism of inhibition of herpes simplex virus and vaccinia virus DNA polymerases by aphidicolin, a highly specific inhibitor of DNA replication in eucaryotes
Authors: Pedrali-Noy G , Spadari S
Published: January 2020

The inhibition in vitro of herpes simplex virus 1 and vaccinia virus DNA polymerases by aphidicolin is primarily noncompetitive with dGTP, dATP, dTTP, DNA, and Mg2+ and competitive with dCTP in analogy with the mode of inhibition of cellular alpha-polymerase. The degree of inhibition of viral or cellular growth in vivo can be quantitatively predicted by the degree of inhibition of the isolated replicative DNA polymerases at the same concentration of aphidicolin in suitable conditions (limiting dCTP concentration). Thus, the only in vivo target for aphidicolin is probably the replicative DNA polymerase, and aphidicolin is a highly specific inhibitor of replicative nuclear DNA synthesis in eucaryotes. This, coupled with the lack of mutagenic effect, represents a valuable property for an anticancer drug. The specificity of inhibition (contrary to the aspecific effect on almost all DNA polymerases by a true competitive inhibitor, such as 1-beta-D-arabinofuranosylcytidine 5'-triphosphate) and the structure of the drug, which does not resemble that of the triphosphates, suggest that aphidicolin must recognize a site common only to the replicative DNA polymerases of eucaryotes and different from the binding site for deoxyribonucleic triphosphates and DNA, which should be similar in reparative and procaryote-type DNA polymerase; the aphidicolin binding site is probably very near to, or even overlaping with, the binding site for dCTP so that the drug mimics a competitive effect with this nucleotide.
Abelson murine leukemia virus-induced tumors elicit antibodies against a host cell protein, P50
Authors: Rotter V , Witte O N , Coffman R , Baltimore D
Published: January 2020

When BALB/c mice were injected with a syngeneic cell line transformed by Abelson murine leukemia virus (A-MuLV), the tumor was usually lethal. In sera from tumor-bearing mice, and at highest levels in sera from mice that reject their tumors, was an antibody that immunoprecipitates a specific protein from [35S]-methionine-labeled A-MuLV-transformed BALB/c cells. This protein was not the previously characterized A-MuLV-specific protein (P120) but a 50,000-molecular-weight protein (P50). Such sera may also immunoprecipitate P120, but no other protein was reproducibly precipitated by them. A monoclonal antibody (RA3-2C2) that has been shown to stain normal B-lymphocytes also selectively immunoprecipitated P50. P50 was present in A-MuLV-transformed lymphoid and fibroblastic cells of a variety of mouse strains. One A-MuLV-transformed cell line had a very low P50 level, the L1-2 tumor of C57L origin. This tumor was previously shown to be rejected by C57L mice and is used to produce anti-P120 (anti-AbT) sera. P50 was not a Moloney MuLV protein and was found at low levels in normal cells of cells transformed by agents other than A-MuLV; thus, it was probably a host cell protein whose concentration was selectively accentuated by A-MuLV transformation. P50 was phosphorylated and, by using indirect immunofluorescence, anti-P50 serum stained live A-MuLV-transformed cells. The protein was not glycosylated and did not label by lactoperoxidase-catalyzed iodination. Thus, P50 was very like P120 in its cellular localization and properties, but it did not exhibit proptein kinase activity in vitro. The selective accentuation of this protein in A-MuLV transformants and its strong antigenicity in syngeneic animals suggest that it is a unique and functionally important protein.
Generation of nondefective Rous sarcoma virus by asymmetric recombination between deletion mutants
Authors: Martin G S , Lee W H , Duesberg P H
Published: January 2020

A replication-defective deletion mutant of Prague Rous sarcoma virus (RSV), which lacks functional gag, pol, and env genes, was crossed with a transformation-defective deletion mutant derived from Schmidt-Ruppin RSV. Transformation- and replication-competent viruses were generated in the cross. Characterization of one of these rescued viruses indicated that it was a nondefective recombinant containing the src gene of the replication-defective mutant plus the replicative genes of the transformation-defective virus. These results indicate that, contrary to previous reports, asymmetric recombination between RSV deletion mutants can result in the formation of nondefective RSV.
Leakage of nuclear transcripts late in simian virus 40-infected CV1 cells: quantitation of spliced and unspliced late 19S RNAs
Authors: Villarreal L P , Jovanovich S
Published: January 2020

During the late phase of simian virus 40 infections of CV1 cells, the relative ratios of the spliced to the unspliced RNA molecules of the 19S family were measured. In the cytoplasm, unspliced 19S RNA represented between 1 and 2% of spliced 19S RNA. This ratio could be altered by the use of different methods of RNA extraction such that unspliced RNA was observed at 10 to 20% of spliced RNA. The nuclear ratios of spliced to unspliced 19S RNA were also determined. In contrast to cytoplasmic RNA, nuclear unspliced RNA was several hundred percent that of nuclear spliced 19S RNA. Cytoplasmic unspliced 19S RNA appears to be of nuclear origin, and its presence in the cytoplasmic fraction is due to nuclear leakage during RNA fractionation.
Dinucleotide Sequences at the 5′ Ends of Vaccinia Virus mRNA's Synthesized In Vitro
Authors: Keith Jerry M. , Gershowitz Alan , Moss Bernard
Published: January 2020

The diversity of dinucleotide sequences at the 5′ ends of vaccinia virus mRNA's was determined by a two-dimensional electrophoresis procedure. RNA labeled with
S
-adenosyl[
methyl
-
3
H]methionine was synthesized in vitro by enzymes present in vaccinia virus cores. The RNA, ending in m
7
G(5′)pppN
m
pN−, was β-eliminated and treated with alkaline phosphatase. After digestion with RNases T
2
, T
1
, and A, all eight possible dinucleotides containing G
m
and A
m
were identified. They are, in decreasing order of abundance: G
m
pUp (22%), A
m
pCp (18%), G
m
pAp (16%), G
m
pCp (15%), A
m
pAp (11%), A
m
pUp (10%), A
m
pGp (7%), and G
m
pGp (2%).
Mapping of adenovirus 12 mRNA's transcribed from the transforming region
Authors: Sawada Y , Fujinaga K
Published: January 2020

Adenovirus 12 mRNA's transcribed from the transforming region were analyzed and mapped on viral DNA by the nuclease S1 gel and diazobenzyloxymethyl paper blot techniques in cells transformed of adenovirus 12 DNA and in cells early and late after lytic infection with adenovirus 12. Two initiation sites of mRNA transcription and two kinds of splicing were found in each of early regions 1A and 1B. For early region 1A mRNA's, four species were found in lytically infected cells. Three of them were commonly found in cells transformed by either HindIII-G or EcoRI-C. Cells transformed by HindIII-G contained two additional 1A transcripts, which could be the 3' portions of chimeric mRNA's of cellular-viral or viral-viral sequences. Transcription in the 1B region diverged among the above cell lines. Early after lytic infection, no appreciable amount of 1B mRNA was detected, whereas two species of mRNA's, one corresponding to protein IX mRNA and another having splicing, were found at the late stage. In cells transformed by EcoRI-C, a distinct mRNA species with splicing was observed. Cells transformed by HindIII-G contained a transcript from the leftmost part of the 1B sequence at the 5' portion of chimeric mRNA species, suggesting the presence of tandem integration of viral DNA in the cells. Other mRNA species in the 1A and 1B regions were also detected in both transformed cell lines. The results are discussed in relation to the nucleotide sequence of the HindIII-G fragment of adenovirus 12 DNA.
Translational control of protein synthesis after infection by vesicular stomatitis virus
Authors: Lodish H F , Porter M
Published: January 2020

Four hours after infection of BHK cells by vesicular stomatitis virus (VSV), the rate of total protein synthesis was about 65% that of uninfected cells and synthesis of the 12 to 15 predominant cellular polypeptides was reduced to a level about 25% that of control cells. As determined by in vitro translation of isolated RNA and both one- and two-dimensional gel analyses of the products, all predominant cellular mRNA's remained intact and translatable after infection. The total amount of translatable mRNA per cell increased about threefold after infection; this additional mRNA directed synthesis of the five VSV structural proteins. To determine the subcellular localization of cellular and viral mRNA before and after infection, RNA from various sizes of polysomes and nonpolysomal ribonucleoproteins (RNPs) was isolated from infected and noninfected cells and translated in vitro. Over 80% of most predominant species of cellular mRNA was bound to polysomes in control cells, and over 60% was bound in infected cells. Only 2 of the 12 predominant species of translatable cellular mRNA's were localized to the RNP fraction, both in infected and in uninfected cells. The average size of polysomes translating individual cellular mRNA's was reduced about two- to threefold after infection. For example, in uninfected cells, actin (molecular weight 42,000) mRNA was found predominantly on polysomes with 12 ribosomes; after infection it was found on polysomes with five ribosomes, the same size of polysomes that were translating VSV N (molecular weight 52,000) and M (molecular weight 35,000) mRNA. We conclude that the inhibition of cellular protein synthesis after VSV infection is due, in large measure, to competition for ribosomes by a large excess of viral mRNA. The efficiency of initiation of translation on cellular and viral mRNA's is about the same in infected cells; cellular ribosomes are simply distributed among more mRNA's than are present in growing cells. About 20 to 30% of each of the predominant cellular and viral mRNA's were present in RNP particles in infected cells and were presumably inactive in protein synthesis. There was no preferential sequestration of cellular or viral mRNA's in RNPs after infection.
Structural relationships between the surface antigens of ground squirrel hepatitis virus and human hepatitis B virus
Authors: Gerlich W H , Feitelson M A , Marion P L , Robinson W S
Published: January 2020

Several physical, chemical, and serological properties of surface antigen particles from ground squirrel hepatitis virus (GSHsAg) and human hepatitis B virus (HBsAg) were compared. GSHsAg and HBsAg particles were purified from positive sera by gel chromatography and isopycnic centrifugation. Both antigens consisted mainly of spherical particles with an average diameter of approximately 20 nm and a buoyant density in CsCl of approximately 1.19 g/ml. Their UV absorption spectra indicated the presence of more tryptophane than tyrosine and the absence of detectable nucleic acid. GSHsAg was found to contain two major polypeptides of approximately 23,000 and 27,000 daltons, with electrophoretic migration rates distinctly faster than those of the two major polypeptides of HBsAg particles. After radiolabeling of purified antigen preparations with Bolton-Hunter reagent, the two major polypeptides of GSHsAg showed almost identical tryptic peptide maps. The tryptic peptide map of the major polypeptide from GSHsAg contained 13 of 37 spots also present in the map of the major HBsAg polypeptide, and 13 of 27 spots in the map of the major HBsAg polypeptide were also present in the map of the major GSHsAg polypeptide. This suggests considerable sequence homology between the major surface antigen polypeptides of the two viruses. However, there was only a weak serological cross-reactivity between antigens of the two viruses. Using an anti-HBs-containing serum with a relatively strong cross-reactivity, GSHsAg was found to consist of at least two antigenically different subspecies. The more strongly cross-reacting from had a slightly higher buoyant density than the other antigenic form.
Molecular genetics of herpes simplex virus. VII. Characterization of a temperature-sensitive mutant produced by in vitro mutagenesis and defective in DNA synthesis and accumulation of gamma polypeptides
Authors: Conley A J , Knipe D M , Jones P C , Roizman B
Published: January 2020

We report on the properties of a temperature-sensitive mutant produced by transfection of cells with intact DNA and a specific DNA fragment mutagenized with low levels of hydroxylamine. The plating efficiency of the mutant at 39 degrees C relative to that at 33.5 degrees C was 5 X 10(-6). The pattern of polypeptides produced at the nonpermissive temperature was similar to that seen with wild-type virus in infected cells treated with inhibitory concentrations of phosphonoacetic acid in that alpha and beta polypeptides were produced, whereas most gamma polypeptides were either reduced or absent. Consistently, the mutant did not make viral DNA, although temperature sensitivity of the viral DNA polymerase could not be demonstrated. Marker rescue studies with herpes simplex virus type 2 (HSV-2) DNA mapped the mutant in the L component within map positions 0.385 and 0.402 in the prototype (P) arrangement of the HSV-1 genome. Analysis of the recombinants permitted the mapping of the genes specifying infected cell polypeptides 36, 35, 37, 19.5, 11, 8, 2, 43, and 44, but only the infected cell polypeptide 8 of HSV-2 was consistently made by all recombinants containing demonstrable HSV-2 sequences. Marker rescue studies with cloned HSV-1 DNA fragments mapped the temperature-sensitive lesion within less than 10(3) base pairs between 0.383 and 0.388 map units. Translation of the RNA hybridizing to cloned HSV-1 DNA, encompassing the smallest region containing the mutation, revealed polypeptide 8 (128,000 molecular weight), which was previously identified as a beta polypeptide with high affinity for viral DNA, and a polypeptide (25,000 molecular weight) not previously identified in lysates of labeled cells.
Hybridization selection and cell-free translation of mRNA's encoded within the inverted terminal repetition of the vaccinia virus genome
Authors: Cooper J A , Wittek R , Moss B
Published: January 2020

Early polypeptides encoded within the 10,000-base pair terminally repeated region of the vaccinia virus genome were mapped by cell-free translation of mRNA that was selected by hybridization to restriction fragments and to separated strands of a recombinant lambda phage. The results, which were confirmed by hybrid arrest of translation, indicated that polypeptides of 7,500 (7.5K), 19,000 (19K), and 42,000 (42K) daltons mapped at approximately 3.2 to 4.3, 6.5 to 7.2, and 7.2 to 8.3 kilobase pairs from the end of the genome, respectively. mRNA's for the 42K and 7.5K polypeptides were transcribed towards the end of the genome, whereas mRNA for the 19K polypeptide was transcribed in the opposite direction. Including polyadenylic acid tails, the lengths of the mRNA's for the 7.5K, 19K, and 42K polypeptides, determined by gel electrophoresis of denatured RNA, hybridization selection, and cell-free translation, were approximately 1,200, 680, and 1,280 nucleotides, respectively. mRNA's for the 42K and 19K polypeptides were only about 100 nucleotides longer than the minimums required to code for their respective polypeptides, whereas mRNA for the 7.5K polypeptide contained 900 nucleotides of untranslated sequence. This long untranslated portion of the latter mRNA was probably located near the 3' end, because this gene was only inactivated by high doses of UV irradiation. This small target size also excluded certain models for RNA processing involving formation of the mRNA's for the 42K and 7.5K polypeptides from a common promoter. Rabbitpox virus, which has an inverted terminal repetition approximately half that of vaccinia virus, was also shown to encode mRNA's that hybridized to the cloned terminal segment of vaccinia virus DNA.
Noncytopathic mutants of Newcastle disease virus are defective in virus-specific RNA synthesis
Authors: Madansky C H , Bratt M A
Published: January 2020

We have studied virus-specific RNA synthesis in cells infected by six noncytopathic (nc) mutants of the Australia-Victoria wild-type strain (AV-WT) of Newcastle disease virus (NDV) (19). The rates of NDV-specific RNA synthesis in mutant infection were three to sevenfold lower than those observed in wild-type infection. Velocity sedimentation of this NDV-specific RNA revealed that the lower rates of synthesis in mutant infection correlated with reduced accumulation of 18S and 35S mRNA. Electrophoresis in polyacrylamide-urea gels showed that accumulation of all of the 18S mRNA species was reduced and no new species could be detected. Primary transcription appeared unaltered in mutant infection. Cells infected with two naturally occurring avirulent strains of NDV also showed less accumulation of 18S mRNA. Electrophoresis of this RNA resulted in patterns which differed from those obtained with RNA from either AV-WT or nc mutant infection. Complementation for RNA accumulation between the nc mutants and RNA- temperature-sensitive mutants of AV-WT (32) suggested a common defect in the nc mutants. Analysis of plaque-forming revertants of five of the nc mutants revealed that viral RNA synthetic capacity, cell killing, and plaque-forming ability correlated absolutely. These results suggest that viral RNA synthesis and cytopathogenicity may be causally related. In addition, several of the plaque-forming (and cell-killing) revertants were found to be unable to induce fusion from within in infected cell cultures. This result, coupled with the finding that several of the nc mutants are capable of wild-type levels of fusion from within, suggests that the ability to cause such fusion does not correlate with the ability to kill cells.
Genetic analysis of gene 1.2 of bacteriophage T7: isolation of a mutant of Escherichia coli unable to support the growth of T7 gene 1.2 mutants
Authors: Saito H , Richardson C C
Published: January 2020

The product of gene 1.2 of bacteriophage T7 is not required for the growth of T7 in wild-type Escherichia coli since deletion mutants lacking the entire gene 1.2 grow normally (Studier et al., J. Mol. Biol. 135:917-937, 1979). By using a T7 strain lacking gene 1.2, we have isolated a mutant of E. coli that was unable to support the growth of both point and deletion mutants defective in gene 1.2. The mutation, optA1, was located at approximately 3.6 min on the E. coli linkage map in the interval between dapD and tonA; optA1 was 92% cotransducible with dapD. By using the optA1 mutant, we have isolated six gene 1.2 point mutants of T7, all of which mapped between positions 15 and 16 on the T7 genetic map. These mutations have also been characterized by DNA sequence analysis, E. coli optA1 cells infected with T7 gene 1.2 mutants were defective in T7 DNA replication; early RNA and protein synthesis proceeded normally. The defect in T7 DNA replication is manifested by a premature cessation of DNA synthesis and degradation of the newly synthesized DNA. The defect was not observed in E. coli opt+ cells infected with T7 gene 1.2 mutants or in E. coli optA1 cells infected with wild-type T7 phage.
Genetic interactions among viruses of the Bunyamwera complex
Authors: Iroegbu C U , Pringle C R
Published: January 2020

Seventy-seven temperature-sensitive (ts) mutants belonging to three antigenically distinct and geographically isolated members of the Bunyamwera complex--Batai virus, Bunyamwera virus, and Maguari virus--have been isolated after 5-fluorouracil treatment. High-frequency recombination was observed, and the mutants of each virus were classified into two groups, which were shown to be equivalent by heterologous recombination experiments. In most combinations heterologous recombination was less efficient than homologous recombination, but all crosses of group I and II mutants yielded viable recombinants. Recombination was an early event. Analysis by polyacrylamide gel electrophoresis of the proteins of the wild-type viruses and recombinant clones obtained from the six possible heterologous combinations of group I and II mutants indicated that recombination occurred by reassortment of genome subunits. Group I appeared to correspond to the genome subunit coding for the N protein, and group II corresponded to the G1/G2 determinant. The G1 (or G2 or both) protein was associated with neutralization specificity and plaque diameter, and the N protein was associated with plaque opacity. Complementation was observed between two nonrecombining mutants of Maguari virus belonging to group I, which may indicate that the N genome subunit codes for an additional protein. There appeared to be no genetic barrier to exchange of genetic material between Batai, Bunyamwera, and Maguari viruses in vitro, and it is concluded that the Bunyamwera complex is potentially a single gene pool if geographical and ecological constraints are discounted.
Evolutionary variants of simian virus 40 which are impaired in early lytic functions but transform nonpermissive cells
Authors: Sol C J , Hassing I , Maris W , Walig C , van der Noordaa J
Published: January 2020

From an undiluted passaged virus stock, two size classes of defective simian virus 40 (SV40) DNA were isolated from which two evolutionary variants were cloned. By means of restriction enzyme and heteroduplex analysis, physical maps of the mutants have been constructed. Both mutants contained the region of SV40 DNA coding for the early proteins plus some adjacent sequences (the region from 0.120 to 0.685 map unit, clockwise, on the standard SV40 DNA map). Furthermore, each mutant contained, in the form of two inverted repeats, four times the sequences from the region 0.625 to 0.685 map unit, clockwise. Some biological properties of the mutant DNA were examined, and we found that the mutant DNA (i) has, as compared with SV40 DNA, an impaired ability to induce T antigen in permissive and nonpermissive cells; (ii) does not complement a thermosensitive A mutant of SV40; (iii) replicates very inefficiently without a helper; and (iv), as an apparent contradiction, transforms nonpermissive baby rat kidney cells as well as SV40 DNA. A hypothetical mechanism for the expression of the mutant DNA that might explain the observed biological properties is presented.