bluuebunny/crossref_metadata_2025 huggingface dataset:
Junin virus structural proteins
Authors: De Martínez Segovia Z M , De Mitri M I
Published: January 2020

Polyacrylamide gel electrophoresis of purified Junin virus revealed six distinct structural polypeptides, two major and four minor ones. Four of these polypeptides appeared to be covalently linked with carbohydrate. The molecular weights of the six proteins, estimated by coelectrophoresis with marker proteins, ranged from 25,000 to 91,000. One of the two major components (number 3) was identified as a nucleoprotein and had a molecular weight of 64,000. It was the most prominent protein and was nonglycosylated. The other major protein (number 5), with a molecular weight of 38,000, was a glucoprotein and a component of the viral envelope. The location on the virion of three additional glycopeptides with molecular weights of 91,000, 72,000, and 52,000, together with a protein with a molecular weight of 25,000, was not well defined.
Free and membrane-bound polyribosomes in BHK cells infected with Sindbis virus
Authors: Martire G , Bonatti S , ALIPERTI G , De Giuli C , Cancedda R
Published: January 2020

The data presented in the paper demonstrate that in BHK cells infected with Sindbis virus virtually all the 42S mRNA not in nucleocapsid is associated with free polyribosomes, whereas the 26S mRNA is distributed between free and membrane-bound polyribosomes. We suggest that the 26S RNA polyribosomes are bound to the membranes through the nascent chains of the B1 protein and that a large percentage of 26S RNA polyribosomes free in the cytoplasm may be due to the small amount of rough endoplasmic reticulum in BHK cells. In addition, we found that intracellular nucleocapsid is in the nonmembrane fraction of the cytoplasm of infected cells.
Distinguishable transformation-defective phenotypes among temperature-sensitive mutants of Rous sarcoma virus
Authors: Becker D , Kurth R , Critchley D , Friis R , Bauer H
Published: January 2020

Eight transformation-defective, temperature-sensitive (ts) mutants of the Prague strain of Rous sarcoma virus, subgroup A, have been isolated after mutagenesis with 5-bromodeoxyuridine followed by selection on the basis of focus tests. Five of these mutants, ts GI201, GI202, GI203, GI204, and GI205, exhibit properties like most previously reported isolates in that they show a temperature-sensitive response to each of a variety of transformation-specific parameters tested. Interestingly, GI201, in addition to the temperature-sensitive defect, carries a lesion that was observed as a nonconditional loss of expression of plasminogen activator protease. Three mutants, ts GI251, GI252, and GI253 have been disignated partial transformation-defective (PTD) mutants since they behave as ts mutants according to some tests for transformation and as wild type according to others. These three mutants fail to form foci at the nonpermissive temperature (41 degrees C) and art nontumorigenic in 3-week-old chickens (body temperature, 42 degrees C). The agglutinability by concanavalin A of cells infected with these mutants shows a definite temperature sensitivity, as do the rate of 2-deoxyglucose uptake and the disappearance of the 250, 000-dalton normal cell glycoprotein (large, external, transformation sensitive [LETS]). Although the PTD mutant-infected cells, unlike cells infected with other transformation mutants, exhibit a cell-bound plasminogen activator protease at the nonpermissive temperature, this activator is not detectable as a free protease in the medium, as it is with wild-type, virus-infected cells. The PTD mutants behave like the wild-type parent in their ability to induce transformed growth properties in the infected cells, i.e., growth beyond normal cell saturation density with or without serum-supplemented medium and growth leading to colony formation in soft-agar- or methyl cellulose-containing suspension media.
RNase activity in human interferon preparations
Authors: Winchurch R A , Karpetsky T P , Levy C C , Cantell K
Published: January 2020

The level of RNase activity in human interferon preparations was examined. Although sequential purification of interferon resulted in nearly a 300-fold increase in specific activity, RNase-specific activity remained more or less constant. The implications of this finding for the analyses of the mode of action of interferon are discussed.
Pseudotypes of vesicular stomatitis virus with the mixed coat of reticuloendotheliosis virus and vesicular stomatitis virus
Authors: Kang C Y , Lambright P
Published: January 2020

Vesicular stomatitis virus (VSV) forms pseudotypes with envelope components of reticuloendotheliosis virus (REV). The VSV pseudotype possesses the limited host range and antigenic properties of REV. Approximately 70% of the VSV, Indiana serotype, and 45% of VSV, New Jersey serotype, produced from the REV strain T-transformed chicken bone marrow cells contain mixed envelope components of both VSV and REV. VSV pseudotypes with mixed envelope antigens can be neutralized with excess amounts of either anti-VSV antiserum or anti-REV antiserum.
Cell killing by simian virus 40: impairment of membrane formation and function
Authors: Norkin L C
Published: January 2020

Simian virus 40 infection of the CV-1 line of green monkey kidney cells results in the release of mitochondrial malic dehydrogenase as early as 24 h. Released malic dehydrogenase is detected in the cytoplasm prior to its appearance in the overlay medium. Infected cells lose the ability to consume oxygen between 48 and 56 h, and damage to the elctron transport system is indicated. Nevertheless, cellular ATP levels remain high as late as 72 h. Infection leads to a stimulation of membrane phospholipid synthesis, which reaches a peak at about 32 h. This is followed by a severe decline in new membrane synthesis, which correlates in time with the release of cytoplasmic lactic dehydrogenase into the overlay media. Lactic dehydrogenase release precedes the accumulation of trypan blue-stainable cells by about 6 h. Infection had no effect on the turnover of prelabeled membrane phospholipids. An early simian virus 40 mutant, tsA58, and a late mutant, tsB11, are both less effective than wild-type virus at causing reduced levels of phospholipid synthesis, enzyme release, and the accumulation of trypan blue-stainable cells. Another late mutant, tsB8, is similar to wild-type virus in these respects. At 64 h, there is no detectable cell-associated lactic dehydrogenase and nearly all the cells are trypan blue stainable. Nevertheless, at concentrations of deoxyglucose in the medium below the transport Km, deoxyglucose uptake was similar in infected and control cultures. With higher concentrations of deoxyglucose in the medium, uptake by the infected cultures exceeded that by the control cultures.
Mechanism of restriction of ecotropic and xenotropic murine leukemia viruses and formation of pseudotypes between the two viruses
Authors: Besmer P , Baltimore D
Published: January 2020

Ecotropic and xenotropic murine leukemia viruses (MuLV's) constitute separate interference groups; within each group there is cross-interference, but between the groups there is no detectable interference. Interference is manifest against pseudotypes in which the vesicular stomatitis virus genome is contained within the coat of one of the murine leukemia viruses. The pseudotypes display the cell specificity of the leukemia viruses: pseudotypes with an ecotropic MuLV coat infect mouse cells but not rabbit or mink cells; pseudotypes with a xenotropic MuLV coat infect rabbit or mink cells well but mouse cells very poorly. Efficient pseudotype formation also occurs between the two MuLV classes, and both the interference patterns and the cell specificity of these pseudotypes are entirely determined by their envelope. Using these pseudotypes, ecotropic MuLV infection could be established in xenogeneic cells, and the resulting progeny could be scored by using a conventional XC cell assay. Also, xenotropic MuLV infection could be established in a mouse cell, showing that no absolute intracellular barrier against xenotropic virus growth exists in murine cells. The major barriers against both xenotropic and ecotropic MuLV therefore are cell surface barriers. Xenogeneic cells probably lack receptors for ecotropic MuLV, but murine cells may either lack receptors for xenotropic MuLV or have receptors that are blocked by endogenous expression of the glycoprotein of endogenous xenotropic MuLV.
Folded, concatenated genomes as replication intermediates of bacteriophage T7 DNA
Authors: Paetkau V , Langman L , Bradley R , Scraba D , Miller R C
Published: January 2020

A complex form of bacteriophage T7 DNA, containing up to several hundred phage equivalents of DNA, arises during replication of T7. The complex was stable to treatment with ionic detergent, Pronase, and phenol. The complex form normally exists for only a short time, corresponding to the phase of rapid T7 DNA synthesis. It is then converted to shorter molecules, both concatemers and unit-size DNA. The complex was stable up to the temperature of denaturation of the bihelix. It consisted of a series of loops amanating from a dense central core, as shownby electron microscopy. The complex form is similar to the relaxed Escherichia coli folded chromosome ('nucleoid'). The loops contained an average of 0.7 to 0.8 phage equivalent of DNA. During infection by phage with an amber mutation in gene 3 (endonuclease), formation of the complex occurred normally, but its maturation to unit-size DNA blocked. Before treatment with phenol, the complex contained short fragments of newly replicated DNA. These were released as single-stranded pieces during phenol treatment. A pathway for T7 DNA replication is indicated in which the flow of material is from unit-size DNA to linear concatemers to the complex form, and then back to unit-size DNA by way of linear concatemers.
Biophysical studies of infectious pancreatic necrosis virus
Authors: Dobos P , Hallett R , Kells D T , Sorensen O , Rowe D
Published: January 2020

The molecular weight of infectious pancreatic necrosis virus (IPNV) has been determined by analytical ultracentrifugation and dynamic light scattering. The sedimentation coefficient of the virus was found to be 435S. The average value for molecular weight is (55 +/- 7) x 106. The virus genome consists of two segments of double-stranded RNA (molecular weights, 2.5 x 106 and 2.3 x 106), which represents 8.7% of the virion mass. The capsid protein moiety of IPNV consists of four species of polypeptides, as determined by polyacrylamide gel electrophoresis. The number of molecules of each polypeptide in the virion has been determined. There are 22 molecules of the internal polypeptide alpha (molecular weight, 90,000), 544 molecules of the outer capsid polypeptide beta (molecular weight, 57,000), and 550 and 122 molecules, respectively, of the internal polypeptides gamma1 (molecular weight, 29,000) and gamma2 (molecular weight, 27,000). IPNV top component contains only the beta polypeptide species, and its molecular weight is estimated to be 31 x 106. The hydrodynamic diameter and electron microscopic diameter (calculated by catalase crystal-calibrated electron microscopy) of IPNV was compared with those of reovirus and encephalomyocarditis virus. Due to the swelling of the outer capsid, reovirus particles were found to be much larger when hydrated (96-nm diameter) than when dehydrated (76-nm diameter), having a large water content content and low average density. In contrast, IPNV particles are more rigid, having nearly the same average diameter under hydrous (64 nm) as under anhydrous conditions (59.3 nm). Encephalomyocarditis virus has a very low water content and does not shrink at all when prepared for electron microscopy.
RNase H and RNA-directed DNA polymerase: associated enzymatic activities of murine mammary tumor virus
Authors: Dion A S , Williams C J , Moore D H
Published: January 2020

The RNA-directed DNA polymerase of murine mammary tumor virus, a type B RNA tumor virus, was purified sequentially through DEAE-cellulose, phosphocellulose (step gradient), and phosphocellulose (linear salt gradient) chromatography followed by glycerol sedimentation centrifugation. During all stages of purification, coincident peaks of RNA-directed DNA polymerase activity, templated by polyribocytidylate-oligodeoxyguanidylate, and RNase H digestion of [3H]polyriboadenylate-polydeoxythymidylate were observed, and both enzymatic activities displayed a cation preference for magnesium. Under conditions that removed adventitiously associated nucleases, RNase H activity was found to co-purify with polymerase. The specificity of this nuclease was assayed with various prepared substrates, which indicated that the polymerase-associated RNase H activity was directed only against the RNA strand of an RNA-DNA hybrid. It is highly probable that RNase H (RNA-DNA hybrid: ribonucleotide-hydrolase, EC 3.1.4..34) and RNA-directed DNA polymerase of type B viruses are associated enzymatic activities analogous to those observed for avian and mammalian type C RNA tumor viruses.
Electron microscopy of the segmented RNA genome ofLa Crosse virus: absence of circular molecules
Authors: Dahlberg J E , Obijeski J F , Korb J
Published: January 2020

The three species of single-stranded RNA present in La Crosse virus were examined in the electron microscope. Because large amounts of contaminating cellular DNA are copurified with the virus despite extensive attempts to purify the virus, it was necessary to use procedures that eliminated the bulk of this DNA before the viral RNA was analyzed. When this was done, the modal lengths of La Crosse virus RNA were 0.4, 2.0, and 3.1 mum. These lengths correspond well to their known molecular weights of 0.4 x 106, 1.8 x 106, and 2.9 x 106. Under the denaturing conditions used to permit complete spreading of these single-stranded RNA molecules, no single-stranded circular molecules are observed. Therefore, the circular nucleocapsids present in La Crosse virus and some other bunyaviruses do not appear to be due to convalent linkage of the ends of the RNA genome.
Multiple Copies of Human Adenovirus 12 Genomes Are Integrated in Virus-Induced Hamster Tumors
Authors: Green Michael R. , Mackey Jesse K. , Green Maurice
Published: January 2020

Tumors induced in hamsters by highly oncogenic human adenovirus 12 contain multiple copies of 90 to 100% of the viral genome in an integrated form.
Control of protein synthesis by a temperature-sensitive mutant of reovirus 3. I. Temperature-sensitive function of ts261-b mutant
Authors: Ikegami N
Published: January 2020

The ability of a temperature-sensitive (ts) mutant of reovirus, ts261-b, to synthesize virus-specific RNAs and proteins during infection at the nonpermissive temperature (37 degrees C) was investigated. The relative amounts of the mutant virus-specific single-stranded (ss) RNA's and double-stranded (ds) RNA's synthesized in cells at 37 degrees C were 20 to 25% as much as those synthesized in the wild-type virus-infected cells. The 10 segments of the mutant ds RNAs and the three size classes of the ss RNAs were synthesized in the usual proportions. The methylation of the mutant viral mRNA's (ss RNAs) was not blocked at 37 degrees C in infected cells. A striking temperature-sensitive restricted function of the ts261-b mutant was expressed in the synthesis of the viral proteins. This study, which uses an in vitro protein-synthesizing system reconstituted with an endogenous polysomal fraction and a postribosomal supernatant from reovirus-infected cells, has demonstrated that the endogenous polysomes obtained from ts261-b mutant-infected cells at 37 degrees C are not active in the synthesis of the viral polypeptides of known molecular weights, and the amounts of the mutant viral polypeptides synthesized in vitro by these polysomes are 5 to 9% of those synthesized by the corresponding fraction from wild-type-infected cells. The impaired protein-synthesizing capacity of the mutant virus-specific polysomes can be restored during maintenance of the infected cells at 30 degrees C after shift-down from 37 degrees C. The in vitro synthesis of viral polypeptides of known size by the active endogenous polysomes derived from cells infected at the permissive temperature is accelerated by the addition of the postribosomal supernatant obtained from cells infected at the permissive temperature. The postribosomal supernatant from mutant-infected cells at 37 degrees C did not have a stimulatory effect, but rather, it inhibited in vitro viral protein synthesis.
In vitro transcription of theavian oncornavirus genome by the RNA-directed DNA polymerase: analysis of DNA transcripts synthesized in reconstructed enzymatic reactions
Authors: Collett M S , Faras A J
Published: January 2020

We have analyzed the DNA products synthesized in vitro in reconstructed reactions containing purified avian oncornavirus genome RNA and RNA-directed DNA polymerase. The results of these studies indicate that: (i) the initial DNA product synthesized on either 70S RNA or reconstituted 35S RNA-tRNAtrp template - primer complexes in the presence of low concentrations of deoxynucleoside triphosphates consists of several discrete size classes, none of which exceed 200 nucleotides in length; (ii) large DNA transcripts (about 2,000 nucleotides) can be synthesized on both 70S RNA and the 35S RNA - tRNAtrp complex by increasing the deoxynucleoside triphosphate concentration; and (iii) DNA synthesized by detergent-disrupted virus is considerably longer than DNA synthesized in reconstructed reactions.
Expression of tumor-specific transplantation antigen in cell lines transformed by wild-type of tsA mutant simian virus 40
Authors: Chang C , Anderson J L , Martin R G , Mora P T
Published: January 2020

The simian virus 40-induced tumor-specific surface antigen(s) (TSSA) and tumor-specific transplantation antigen(s) (TSTA)were detected in cells transformed by wild-type or temperature-sensitive mutant simian virus 40 by an antibody-mediated cytolytic assay for TSSA and an immunization test for TSTA. Cells transformed by tsA mutants, which lose their transformed phenotype when grown at nonpermissive temperatures, nonetheless do express TSSA and TSTA as well as T-antigen at both temperatures.
Epstein-barr virus-specific RNA. II. Analysis of polyadenylated viral RNA in restringent, abortive, and prooductive infections
Authors: Orellana T , Kieff E
Published: January 2020

The complexity and abundance of Epstein-Barr (EBV)-specific RNA in cell cultures restringently, abortively, and productively infected with EBV has been analyed by hybridization of the infected cell RNA with purified viral DNA. The data indicate the following. (i) Cultures containing productively infected cells contain viral RNA encoded by at least 45% of EBV DNA, and almost all of the species of viral RNA are present in the polyadenylated and polyribosomal RNA fractions. (ii) Restringently infected Namalwa and Raji cultures, which contain only intranuclear antigen, EBNA, and enhanced capacity for growth in vitro, contain EBV RNA encoded by at least 16 and 30% of the EBV DNA, respectively. The polyadenylated and polyribosomal RNA fractions of Raji and Namalwa cells are enriched for a class of EBV RNA encoded by approximately 5% of EBV DNA. The same EBV DNA sequences encode the polyadenylated and polyribosomal RNA of both Raji and Namalwa cells. (iii) After superinfection of Raji cultures with EBV (HR-1), the abortively infected cells contain RNA encoded by at least 41% of EBV DNA. The polyadenylated RNA of superinfected Raji cells is enriched for a class of EBV RNA encoded by approximately 20% of EBV HR-1 DNA. Summation hybridization experiments suggest that the polyadenylated RNA in superinfected Raji cells is encoded by the same DNA sequences as encode RNA present in Raji cells before superinfection, most of which is not polyadenylated. That the same EBV RNA sequences are present in the polyadenylated and polyribosomal fractions of two independently derived, restringently infected cell lines suggests that these RNAs may specify functions related to maintenance of the transformed state. The complexity of this class of RNA is adequate to specify a sequence of a least 5,000 amino acids. That only some RNA species are polyadenylated in restringent and abortive infection suggests that polyadenylation or whatever determines polyadenylation may play a role in the restricted expression of the EVB genome.
Structural polypeptides of adenovirus type 16 incomplete particles
Authors: Winberg G , Wadell G
Published: January 2020

The polypeptides of adenovirus type 16 incomplete particles, with average buoyant densities of CsCl of 1.33, 1.318, 1.305, and 1.299 g/cm3 and DNA content of less than 1 U genome size, were compared with the polypeptides of the complete virion (density, 1.344 g/cm3, and 22 x 10(6) daltons of DNA) and late polypeptides in infected cells by using sodium dodecyl sulfate-gel electrophoresis. In agreement with other serotypes studied (types 2 and 3), the light particles lack polypeptides V, VI, and VII. In adenovirus type 16, eight other major polypeptides are found, with apparent molecular weights of 59,000 (59K), 46K, 31K, 30K, 28K, 27K, 26K, and 19K. The 30K to 31K and 27K to 28K polypeptides are phosphorylated. The 27K and 19K polypeptides are precursors, whereas the 31K, 30K, and 26K polypeptides are chase products in the cell, as are polypeptides VI, VII, and VIII. The 26K polypeptide is proposed to be an intermediate in processing since it disappears from the young virions upon chasing. Although chase products, the 31K and 30K polypeptides are only associated with particles having buoyant density lower than 1.318 g/cm3. Polypeptides V and VII are only present in particles containing more than one quarter of the genome. No trace of cell histones could be detected in the purified incomplete particles. A major constituent of the incomplete particles, the 46K polypeptide, was rapidly labeled in the cell, and loss of radioactivity from this band was detectable only after 7 to 18 h of chase.
Precursors of the T4 internal peptides
Authors: Kurtz M B , Champe S P
Published: January 2020

The precursors of the two T4 internal peptides have been identified by in vitro cleavage of individual phage proteins eluted from sodium dodecyl sulfate-acrylamide gels. The precursor of internal peptide VII is p22, the product of T4 gene 22 and an essential component of the morphogenic core. The precursor of peptide II is a protein with a molecular weight of approximately 13,000, whose gene has yet to be defined by mutation. A newly detected protein of approximately 15,000 molecular weight is found to be cleaved and is, therefore, likely to be a component of precursor head structures.
Changes in synthesis of DNA-binding proteins during the onset of transformation in NRK cells transformed by a temperature-sensitive mutant of Rous sarcoma virus
Authors: Magun B E , Dorsett P H
Published: January 2020

Synthesis of cytoplasmic DNA-binding proteins was investigated after a shift from the nonpermissive to the permissive temperature in NRK cells transformed by a temperature-sensitive mutant of Rous sarcoma virus [ts339(RSV)]. Cells were labeled for several generations in [3H]leucine and were pulse-labeled with [35S]methionine for 1 h at the nonpermissive temperature (39 degrees C) and at the permissive temperature (33 degrees C, 5 h after shift from 39 degrees C). Proteins binding to sequential columns of double-stranded and single-stranded DNA-cellulose were examined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, and the 35S/3H ratios were obtained for each column fraction and for individual polypeptides. The protein fractions binding to single-stranded, but not double-stranded, DNA and eluting at high salt concentrations (greater than 0.60 M NaCl) showed elevated 35S/3H ratios. This indicated increased synthesis of these proteins within 5 h after the onset of transformation. The majority of the polypeptides in these fractions showed increased synthesis as a consequence of transformation. One prominent polypeptide among them constituted 0.1% of the cytosol protein and had a molecular weight of 93,000. We conclude that the synthesis of proteins binding tightly to single-stranded DNA is increased early after the onset of transformation.
RNA synthesized in calicivirus-infected cells is atypical of picornaviruses
Authors: Ehresmann D W , Schaffer F L
Published: January 2020

RNA labeled with [3H]uridine from Vero cells infected with San Miguel sea lion virus in the presence of actinomycin D was analyzed by glycerol density gradient sedimentation and polyacrylamide gel electrophoresis. The predominant single-stranded RNA (36S, 2.6 x 10(6) molecular weight) was genome size. There was also a prominent 22S, 1.1 x 10(6)-molecular weight, single-stranded component and one or more double-stranded or partially double-stranded classes. Replicative forms, sedimenting at 18S, contained single-stranded RNA corresponding to the larger-molecular-weight class. All classes of intracellular RNA and virion RNA were polyadenylated. These findings and results with pig kidney cells infected with vesicular exanthema of swine virus and feline cells infected with feline calicivirus indicate that caliciviruses exhibit a strategy of replication different from typical picornaviruses and supports removal of the caliciviruses from the family Picornaviridae.
Adenovirus type 2 nuclear RNA accumulating during productive infection
Authors: Bachenheimer S L
Published: January 2020

The viral-specific nuclear RNA which accumulates early and late during productive infection of HeLa cells by adenovirus-type 2 (Ad2) has been characterized with respect to its size and stability after denaturation by Me2SO. Early nuclear transcripts, under nondenaturing conditions, sediment in the range 28 to 45S, but treatment with Me2SO prior to sedimentation results in a shift to about 20S. Later nuclear RNA accumulates as a composite of two populations of molecules: one with a broad size distribution centering on 45S under nondenaturing conditions and less than 32S after denaturation and a second having a narrow size distribution around 35S which is quite stable to Me2SO. Analysis of late RNA by hybridization to Sma fragments of Ad2 DNA suggests that the 35S RNA species is derived from a limited portion of the left half of the viral genome.
Analysis of precursors to the envelope glycoproteins of avian RNA tumor viruses in chicken and quail cells
Authors: Moelling K , Hayami M
Published: January 2020

Immune precipitation with monospecific antiserum was employed to study the intracellular synthesis of viral glycoproteins gp85 and gp37. Labeled gp85 and gp37 were detected from lysates of cells transformed with Rous sacroma virus, strain B77, after long-term labeling with radioactive glucosamine or phenylalanine. Immune precipitates prepared from lysates of cells pulse-labeled for a short time resulted in a glycoprotein of 92,000 molecular weight (gp92). This precursor was stable in B77-transformed Japanese quail cells for several hours, whereas in chicken cells it could be chased within a few hours into virion glycoproteins gp85 and gp37. Similarly, the precursor for the structural viral proteins, pr76, persisted in quail cells much longer than in chicken cells. During very short pulses or in the presence of a glucosamine block (25 mM glucosamine), the antiserum against the viral envelope glycoproteins detected a precursor of higher electrophoretic mobility of approximately 70,000 molecular weight, "p70." Fucose label entered gp92 and gp85 as well as "p70." Proteolytic treatment of virion-bound gp85 in vitro generated two discrete glycoproteins of 62,000 and 45,000 molecular weight, but did not result in an increase in the amount of gp37.
Donation of N- or B-tropic phenotype to NB-tropic murine leukemia virus during mixed infections
Authors: Kashmiri S V , Rein A , Bassin R H , Gerwin B I , Gisselbrecht S
Published: January 2020

The IC isolate of Moloney murine leukemia virus (MuLV), which is NB-tropic, was grown in cells producing conditionally defective or defective virus particles derived from N- or B-tropic MuLV. The infectious MuLV that was then released was found to be sensitive to Fv-1 restriction but produced NB-tropic progeny upon passage. These results indicate that this NB-tropic MuLV can acquire sensitivity to Fv-1 restriction by phenotypic mixing with N- or B-tropic MuLV. It is thus suggested that NB-tropic MuLV is insensitive to Fv-1 restriction simply because it lacks the determinants of tropism.
Characterization of visna virus envelope neuraminic acid
Authors: August M J , Harter D H , Compans R W
Published: January 2020

Visna virus particles inhibit influenza virus hemagglutination in an assay for neuraminic acid-containing viruses. Pretreatment of visna virus with neuraminidase abolished hemagglutination inhibition activity but did not significantly affect attachment, infectivity, or virus-induced cell fusion in sheep choroid plexus cell monolayers.
Virion RNA polymerases of two salmonid rhabdoviruses
Authors: McAllister P E , Wagner R R
Published: January 2020

RNA-dependent RNA polymerases were found to be associated with two salmonid rhabdoviruses: infectious hematopoietic necrosis (IHN) virus and the virus of hemorrhagic septicemia (VHS). The protein composition of these rhabdoviruses closely resembles that of rabies virus rather than that of vesicular stomatitis virus (McAllister and Wagner, 1975). The optimal temperature for in vitro transcription was found to be approximately 18 degrees C for IHN virus and approximately 15 degrees for VHS,, closely approximating optimal temperatures for growth of these viruses in salmonid cells. Unlike vesicular stomatitis virus, manganese ion (1 mM) could be used as a divalent cation substitute for magnesium ion (5 mM). The in vitro transcription products of IHN and VHS viruses hybridized completely to the homologous genome but not at all to the heterologous genome.
Phosphorylation and nucleic acid binding properties of m1 Moloney murine sarcoma virus-specific pP60gag
Authors: Oskarsson M K , Long C W , Robey W G , Scherer M A , Vande Woude G F
Published: January 2020

The pP60gag polyprotein of the feline leukemia virus pseudotype of m1 Moloney murine sarcoma virus [m1MSV(FeLV)] was previously shown to be MSV specific and to contain murine p30 and smaller structural polypeptides. This protein was detected in m1MSV-transformed cells, and in pulse-chase studies it was found to be stable. In this study virion P60 was shown to contain murine pp12, to be phosphorylated, and to bind to nucleic acids. 32P-labeled m1MSV[FeLV) was fractionated by guanidine agarose chromatography and analyzed by gel electrophoresis. Both P60 and pp12 were found to be the major phosphoproteins, phosphorylated in both serine and threonine residues. Virion P60 bound preferentially to single-stranded DNA and RNA in a competition filter binding assay, using 125I-labeled single-stranded calf thymus DNA and various unlabeled nucleic acids. Similar phosphorylation and DNA binding properties were demonstrated for cellular P60. Thus, immunoprecipitation of cellular extracts showed that P60 was phosphorylated in both producer and nonproducer transformed cells, indicating that phosphorylation occurs independently of virus assembly. Moreover, P60 from cytoplasmic extracts was retained on single-stranded DNA-Sepharose columns, demonstrating that cellular P60 binds to DNA.
Membrane damage in abortive infections of colicin Ib-containing Escherichia coli by bacteriophage T5
Authors: Cheung A K , Duckworth D H
Published: January 2020

Strains of Escherichia coli K-12 containing the colicin Ib (Col Ib) factor did not produce progeny phage when infected by T5 bacteriophage. The cells were killed but did not lyse. If sodium dodecyl sulfate (SDS) was added to T5-infected E. coli (Col Ib), lysis occurred prematurely, but no phage were produced. SDS had no effect on infected cells that did not contain the Col Ib factor or on uninfected cells with or without the Col Ib factor. Cells that contained a mutant Col Ib factor that allowed phage production were not prematurely lysed after infection in the presence of SDS. When the Col Ib-containing cells were infected, protein and RNA synthesis stopped at about 10 min postinfection, and the cells released abnormal amounts of 32P-containing material, ATP, and beta-galactosidase into the medium. They also became inhibited in their ability to accumulate thiomethyl-beta-D-galactopyranoside and to utilize glycerol. Two alternative hypotheses are presented to explain these results.
Structure of Hepatitis B Dane Particle DNA and Nature of the Endogenous DNA Polymerase Reaction
Authors: Landers Terry A. , Greenberg Harry B. , Robinson William S.
Published: January 2020

The circular DNA of hepatitis B Dane particles, which serves as the primer/template for an endogenous DNA polymerase, was analyzed by electrophoresis before and after a polymerase reaction and after digestion by restriction endonuclease or single-strand-specific endonuclease S1. The unreacted molecules extracted from the particles were electrophoretically heterogeneous, and treatment with S1 nuclease produced double-stranded linear DNA ranging in length from 1,700 to 2,800 base pairs (bp). After an endogenous DNA polymerase reaction, two discrete species of DNA molecules were found: a circular form and a linear form 3,200 bp long. The reaction resulted in a population of molecules with an elongated and more homogeneous double-stranded region. These results suggest that the circular molecules in Dane particles have single-stranded regions of varying lengths that are made double stranded during the DNA polymerase reaction. The endogenous DNA polymerase was found to initiate apparently at random in a region spanning more than a third of the molecule. Analysis of restriction endonuclease cleavage fragments of the fully elongated DNA revealed that although the molecules were of a uniform length, they were somewhat heterogeneous in sequence. The sum of the sizes of the 10 major endonuclease
Hae
III-generated fragments, detected by ethidium bromide, was 3,880 bp. Two additional fragments (B and G) detected by autoradiography after an endogenous DNA polymerase reaction with
32
P-labeled deoxynucleoside triphosphates made the total 4,910 bp.
In Vitro Packaging of UV Radiation-Damaged DNA from Bacteriophage T7
Authors: Kuemmerle Nancy B. , Masker Warren E.
Published: January 2020

When DNA from bacteriophage T7 is irradiated with UV light, the efficiency with which this DNA can be packaged in vitro to form viable phage particles is reduced. A comparison between irradiated DNA packaged in vitro and irradiated intact phage particles shows almost identical survival as a function of UV dose when
Escherichia coli
wild type or
polA
or
uvrA
mutants are used as the host. Although
uvrA
mutants perform less host cell reactivation, the
polA
strains are identical with wild type in their ability to support the growth of irradiated T7 phage or irradiated T7 DNA packaged in vitro into complete phage. An examination of in vitro repair performed by extracts of T7-infected
E.coli
suggests that T7 DNA polymerase may substitute for
E. coli
DNA polymerase I in the resynthesis step of excision repair. Also tested was the ability of a similar in vitro repair system that used extracts from uninfected cells to restore biological activity of irradiated DNA. When T7 DNA damaged by UV irradiation was treated with an endonuclease from
Micrococcus luteus
that is specific for pyrimidine dimers and then was incubated with an extract of uninfected
E. coli
capable of removing pyrimidine dimers and restoring the DNA of its original (whole genome size) molecular weight, this DNA showed a higher packaging efficiency than untreated DNA, thus demonstrating that the in vitro repair system partially restored the biological activity of UV-damaged DNA.
Changes in macromolecular synthesis in Xanthomonas oryzae infected with bacteriophage XP-12
Authors: Ehrlich M , Lin F H , Ehrlich K , Brown S L , Mayo J A
Published: January 2020

Phage XP-12, which has complete substitution of the cytosine residues in its DNA with 5-methylcytosine residues, was shown to inhibit incorporation of uracil into host DNA and RNA during the latent period. This apparent inhibition of host macromolecular synthesis was not accompanied by extensive degradation of the host chromosome. Phage DNA synthesis in infected cells occurred at a faster rate than host DNA synthesis in analogous uninfected cells. However, phage DNA synthesis could not be accurately monitored by incorporation of [methyl-3H]thymidine into DNA because, soon after infection, there was a marked inhibition of utilization of exogenous thymidine for DNA synthesis. Phage infection conferred upon a thymine auxotrophic host the ability to synthesize thymine nucleotides for phage DNA synthesis. It is suggested that a phage-induced thymidylate synthetase activity is partially responsible for the inhibition of thymidine incorporation.
Integration of Proviral DNA in Chicken Cells Infected with Schmidt-Ruppin Rous Sarcoma Virus Is Not Enhanced by DNA Repair
Authors: Tsuruo T. , Baluda M. A.
Published: January 2020

The effect DNA repair might have on the integration of exogenous proviral DNA into host cell DNA was investigated by comparing the efficiency of proviral DNA integration in normal chicken embryonic fibroblasts and in chicken embryonic fibroblasts treated with UV or 4-nitroquinoline-1-oxide. The cells were treated with UV or 4-nitroquinoline-1-oxide at various time intervals ranging from 6 h before to 24 h after infection with Schmidt-Ruppin strain A of Rous sarcoma virus. The chicken embryonic fibroblasts were subsequently cultured for 18 to 21 days to ensure maximal integration and elimination of nonintegrated exogenous proviral DNA before DNA was extracted. Integration of proviral DNA into the cellular genome was quantitated by hybridization of denatured cellular DNA on filters with an excess of
3
H-labeled 35S viral RNA. The copy number of the integrated proviruses in normal cells and in infected cells was also determined from the kinetics of liquid RNA-DNA hybridization in DNA excess. Both RNA excess and DNA excess methods of hybridization indicate that two to three copies of the endogenous provirus appear to be present per haploid normal chicken cell genome and that two to three copies of the provirus of Schmidt-Ruppin strain A of Rous sarcoma virus become integrated per haploid cell genome after infection. The copy number of viral genome equivalents integrated per cell treated with UV or 4-nitroquinoline-1-oxide at different time intervals before or after infection did not differ from the copy number in untreated but infected cells. This finding supports our previous report that the integration of oncornavirus proviral DNA is restricted to specific sites in the host cell DNA and suggests a specific mechanism for integration.
Endogenous RD-114 virus genome expression in malignant tissues of domestic cats
Authors: Niman H L , Gardner M B , Stephenson J R , Roy-Burman P
Published: January 2020

Endogenous xenotropic cat type C virus (RD-114)- and infectious feline leukemia virus (FeLV)-specific gene expressions were measured in spontaneous sarcomas carcinomas, and nonmalignant cat tissues by molecular hybridization for virus-specific RNA and competition radio-immunoassays for the major internal protein (p30) of these two viruses. The results indicate that RD-114 gene expression in sarcomas and carcinomas at both RNA and p30 levels is significantly higher than histologically normal tissues from cats free of cancer. In contrast, the levels of FeLV viral RNA and p30 are fount to be low or undetectable in the majority of these tumored and normal tissues examined. Whereas variability in the amounts of RD-114 OR FeLV RNA and p30 expressed is found in tissues from different cats, their expression is fairly uniform in multiple malignant tissues of the same cat. The finding of widespread occurrence of elevated RD-114 gene expression in sarcomas and carcinomas is consistent with our similar observation with natural lymphomas of domestic cats and suggests that expression of certain functions of this endogenous virus may be etiologically involved in the development of many different spontaneous neoplasms of cats.
Localization of single-chain interruptions in bacteriophage T5 DNA I. Electron microscopic studies
Authors: Scheible P P , Rhoades E A , Rhoades M
Published: January 2020

Bacteriophage T5 DNA was examined in an electron microscope after limited digestion with exonuclease III from Escherichia coli. The effect of the exonuclease treatment was to convert each naturally occurring single-chain interruption in T5 DNA into a short segment of single-stranded DNA. The locations of these segments were determined for T5st(+) DNA, T5st(0) DNA, and fragments of T5st(0) DNA generated by EcoRI restriction endonuclease. The results indicate that single-chain interruptions occurr in a variable, but nonrandom, manner in T5 DNA. T5st(+) DNA has four principal interruptions located at sites approximately 7.9, 18.5, 32.6, and 64.8% from one end of the molecule. Interruptions occur at these sites in 80 to 90% of the population. A large number of additional sites, located primarily at the ends of the DNA, contain interruptions at lower frequencies. The average number of interruptions per genome, as determined by this method, is 8. A similar distribution of breaks occurs in T5st(0) DNA, except that the 32.6% site is missing. At least one of the principal interruptions is reproducibly located within an interval of 0.2% of the entire DNA.
Synthesis of Proteins and Glycoproteins in Cells Infected with Human Cytomegalovirus
Authors: Stinski Mark F.
Published: January 2020

In cytomegalovirus-infected cells, the rate of protein synthesis was detected as two peaks. One occurred during the early phase of infection, 0 to 36 h postinfection, and the other occurred during the late phase, after the initiation of viral DNA synthesis. Double-isotopic-label difference analysis demonstrated that host and viral proteins were synthesized simultaneously during both phases. In the early phase, approximately 70 to 90% of the total proteins synthesized were host proteins, whereas approximately 10 to 30% were viral, even at a multiplicity of infection of 20 PFU/cell. Virus-related proteins or glycoproteins were referred to as infected-cell specific (ICS). Two ICS glycoproteins (gp145 and 100) were clearly detectable and were synthesized preferentially in the early phase of infection. Their synthesis was concomitant with stimulation of the protein synthesis rate. In the late phase of infection, approximately 50 to 60% of the total protein synthesis was viral and approximately 40 to 50% was host. The ICS proteins and glycoproteins detected during the late phase of infection were viral structural proteins. Infectious virus was not detectable until 48 to 72 h postinfection. An inhibitor of viral DNA synthesis, phosphonoacetic acid, prevented the appearance of the late-phase ICS proteins and glycoproteins, but there was little or no effect on early ICS glycoprotein synthesis. Radiolabeled ICS proteins and glycoproteins were identified by their relative rates of synthesis, by their different electrophoretic mobilities compared with those of host proteins and host glycoproteins, and by their similar electrophoretic mobilities compared to those of proteins and glycoproteins associated with virions and dense bodies of cytomegalovirus. Structural viral antigens in the infected-cell extracts were removed by immunoprecipitation, using F(ab′)
2
fragments of cytomegalovirus-specific antibodies, and identified as described above. The last two criteria were used to identify viral structural ICS proteins and glycoproteins. Although approximately 35 structural proteins were found to be associated with purified virions and dense bodies, the continued synthesis of host cell proteins complicated their identification in infected cells. Nevertheless, seven of the nine structural glycoproteins were identified as ICS glycoproteins.
Common Precursor for Rauscher Leukemia Virus gp69/71, p15(E), and p12(E)
Authors: Karshin W. L. , Arcement L. J. , Naso R. B. , Arlinghaus R. B.
Published: January 2020

Rauscher murine leukemia virus glycoprotein gp69/71 and non-glycosylated p15(E) are synthesized by way of a 90,000-dalton precursor glycoprotein, termed Pr2
a
+
b
. Peptide mapping experiments showed that Pr2
a
+
b
contains all the tyrosine-containing tryptic peptides of gp69/71. Two additional tyrosine-containing tryptic peptides in Pr2
a
+
b
that are not detected in gp69/71 are found in p15(E). Thus, gp69/71 and p15(E) peptide sequences account for all the tyrosine tryptic peptides of Pr2
a
+
b
. The gene order of the two proteins was determined by pulse-labeling infected cells in the presence and absence of pactamycin at concentrations of the inhibitor that prevent initiation of translation, but not elongation. The gene order was found to be:
2
HN-gp69/71-p15(E)-COOH. A newly identified major viral protein, termed p12(E), migrates in sodium dodecyl sulfate-polyacrylamide gels in the “p12” region. It is related to p15(E) as determined by tryptic mapping experiments. p15(E) and p12(E) are not phosphorylated, and both can be separated from phosphoprotein p12 by guanidine hydrochloride-agarose chromatography. p12(E) and p15(E) elute in the void volume fraction, whereas phosphoprotein p12 elutes between p15 and p10. The two p12 proteins can also be separated from each other by two-dimensional gel electrophoresis involving isoelectric focusing in the first dimension and sodium dodecyl sulfate-gel electrophoresis in the second dimension.
Secondary structure of RNA from bacteriophages f2 Qbeta, and PP7
Authors: Edlind T D , Bassel A R
Published: January 2020

Electron microscopy of RNA-protein monolayers prepared under partial denaturing conditions has been used to compare the secondary structure of coliphage f2 and Qbeta and Pseudomonas aeruginosa phage PP7 RNAs. The secondary structure map of f2 RNA contains a central open loop and four symmetrically placed hairpins, which is similar to the pattern reported by Jacobson (A. B. Jacobson, Proc. Natl. Acad. Sci. U.S.A. 73:307-311, 1976) for the closely related phage MS2. With the same denaturing conditions, Qbeta RNA, which is 20% larger than f2 or PP7 RNA, has a central open loop and a smaller terminal loop. PP7 RNA has two large, closed secondary structures, one of which is nearly central. The base composition of PP7 RNA was determined and is similar to that of the group I coliphage RNAs. Thus, the greater amount of large base-paired structure is not related to an increased guanine-plus-cytosine content of PP7 RNA. With increased denaturing conditions, the central, closed structure of PP7 RNA is converted into an open loop. The central structures of all three phages include about 700 nucleotides. The relevance of these findings to the genetic maps of the coliphage RNAs is discussed.