bluuebunny/crossref_metadata_2025 huggingface dataset:
Incorporation of In Vitro Synthesized Reovirus Double-Stranded Ribonucleic Acid into Virus Corelike Particles
Authors: Sakuma S. , Watanabe Y.
Published: January 2020

When reovirus double-stranded ribonucleic acid (dsRNA) was synthesized in vitro by using a large-particulate fraction (LP-fraction) from reovirus-infected L cells, a significant amount of the
3
H-labeled dsRNA product was incorporated into reovirus corelike particles bound to the LP fraction. These corelike particles were found to be indistinguishable from virus core derived by chymotryptic digestion of virions when compared on the basis of their (i) resistance to chymotryptic digestion, (ii) buoyant density in CsCl, (iii) particle size as determined by agarose chromatography, (iv) elution characteristics from diethylaminoethyl-Sephadex, and (v) resistance of the incorporated
3
H-dsRNA to ribonuclease digestion in 0.01
m
NaCl. When the replicase reaction was partially inhibited by NaCl, there was an accumulation of particles that were less dense than the virus core. All of the results indicate that some virus core assembly takes place during the in vitro replicase reaction.
Transient Inhibition of Polyoma Virus Synthesis by Sendai Virus (Parainfluenza I) I. Demonstration and Nature of the Inhibition by Inactivated Virus
Authors: Smith Gary L. , Consigli Richard A.
Published: January 2020

Superinfection of polyoma virus-infected mouse embryo cells by beta-propiolactone-inactivated Sendai virus resulted in a 90% inhibition of the synthesis of infectious polyoma progeny. The interference is dependent upon the time of superinfection and the concentration of the inactivated virus. The inhibition of polyoma virus synthesis is transient in nature since normal synthesis of polyoma progeny virus is seen upon prolonged incubation. Interferon does not appear to be implicated in the interference. Various aspects of the biological and synthetic capabilities of beta-propiolactone-inactivated Sendai virus are also described.
Adenovirus-Associated Virus Multiplication IX. Extent of Transcription of the Viral Genome In Vivo
Authors: Carter Barrie J. , Khoury George , Rose James A.
Published: January 2020

Nucleic acid hybridization procedures were used to measure the extent of transcription of adenovirus-associated virus (AAV) deoxyribonucleic acid (DNA) in KB cells in the presence of either adenovirus or herpes simplex virus as the helper. Annealing of AAV ribonucleic acid to AAV DNA was monitored by a hybridization inhibition assay on nitrocellulose filters or by hydroxyapatite chromatography. These experiments confirmed the previous observation that, in the presence of either type of helper virus, only one strand of AAV DNA (the thymidine-rich or “minus” strand) is transcribed in vivo. However, it was found that only 70 to 80% of this strand appears to be transcribed in vivo. Furthermore, studies with minus strands employing hydroxyapatite chromatography and nuclease S
1
, which specifically degrades single-stranded DNA, indicated that up to 20% of the minus strand is self-complementary. It seems likely that these self-complementary sequences account for the bulk of that portion of the minus strand (20 to 30%) which is not transcribed in vivo.
Binding of Bacteriophage P22 Tail Parts to Cells
Authors: Israel Vance , Rosen Howard , Levine Myron
Published: January 2020

Purified base-plate parts of bacteriophage P22 can bind to the host cell,
Salmonella typhimurium
. Although the reaction is reversible, a stable equilibrium is not formed between bound and unbound base-plate parts. This is because the binding sites on the cell, presumably the O antigens, are destroyed. The destruction of binding sites does not kill the cells, and, in fact, the binding sites are soon regenerated. The site-destroying activity reacts with P22 heads to make active phage and with antiserum made against purified phage. Therefore site-destroying activity is a characteristic of the base-plate parts and not some contaminant of the preparation.
Transcription During the Development of Bacteriophage φ29: Production of Host-and φ29-Specific Ribonucleic Acid
Authors: Schachtele Charles F. , De Sain Carol V. , Hawley Louise A. , Anderson Dwight L.
Published: January 2020

The synthesis of ribonucleic acid (RNA) during development of the virulent
Bacillus subtilis
bacteriophage φ29 has been analyzed. Transcription of host deoxyribonucleic acid (DNA) continues at the preinfection rate throughout the latent period of viral growth. RNA-DNA hybridization was used to show that host messenger RNA synthesis continues late into the phage lytic cycle. Amino acid-labeling experiments show that this RNA is continuously used to produce protein. Ribosomal RNA production is not inhibited by phage infection. Small quantities of phage-specific RNA first appear between min 6 and 9 after infection. This RNA is made exclusively from one of the φ29 DNA strands. At 12 min postinfection, when phage DNA replication commences, large quantities of viral RNA start to be synthesized. This RNA appears to be transcribed from both strands of φ29 DNA. Studies with rifamycin and rifamycin-resistant host strains showed that the production of all phage φ29-specific RNA requires those components of the host RNA polymerase which are sensitive to this antibiotic. Thus, phage φ29 does not stop transcription of host DNA and may produce only one element for regulation of transcription of its own DNA. These findings may reflect the limited amount of genetic information carried by this phage.
Polypeptide Synthesis by Extracts from
Escherichia coli
Treated with T2 Ghosts
Authors: Nugent Kenneth , Kennell David
Published: January 2020

Infection of
Escherichia coli
B in inorganic salts-glycerol with a multiplicity of deoxyribonucleic acid-less T2 “ghosts” just sufficient to block all protein synthesis results in both viable and killed bacteria. We enriched for the viable cells by a combination of lysozyme treatment and filtration and measured the in vitro capacity of their extracts to synthesize polypeptides. Without added template ribonucleic acid (RNA), such “ghost extracts” incorporate amino acids (endogenous synthesis) at approximately one-half the rate as do extracts from uninfected bacteria. However, they are unable to use added synthetic or natural template RNAs for peptide synthesis. Some activity can be observed but only at high concentrations of Mg
2+
. These results suggest that ghost infection may result in a blockage of ribosomes during translation. Mixing experiments show that the incapacity of ghost extracts to translate added template RNA is due to a defect in the ribosomes.
Density Heterogeneity of Simian Virus 40 Ribonucleic Acid Late After Infection of Permissive Cells
Authors: Fried Allan H.
Published: January 2020

Ribonucleic acid (RNA) was isolated from CV-1 cells 44 hr after simian virus 40 (SV40) infection. The molecules containing SV40 base sequences were characterized with respect to their buoyant density distribution. The density of these molecules was compared to that of single- and double-stranded RNA synthesized by using SV40 DNA and
Escherichia coli
RNA polymerase. The results suggest that about 20% of the SV40 RNA in the infected cells consisted of partially double-stranded molecules.
Rescue of DNA Replication and Bacteriophage Production After Infection with T4 DNA Ligase Mutants
Authors: Cascino A. , Riva S.
Published: January 2020

By preventing phage DNA synthesis during a critical period, conditions have been found under which DNA replication and phage production are rescued after infection with T4 DNA ligase mutants.
Nonproductive Infection of Guinea Pig Cells with Human Cytomegalovirus
Authors: Fioretti Armanda , Furukawa Toru , Santoli Daniela , Plotkin Stanley A.
Published: January 2020

Human cytomegalovirus was capable of adsorbing to and penetrating guinea pig cells, but was unable to replicate new virus. Cultures infected with virus inoculum of high titer showed a cytopathic effect (CPE) characterized by cell rounding. This CPE depended upon the presence of infectious virus, and its extent was directly related to the multiplicity of infection. Staining by indirect immunofluorescence by using human convalescent sera was positive as early as 4 h postinfection. Maximal fluorescence was observed 24 h postinfection when 50% of the cells contained fluorescent antigens both in nuclei and cytoplasm. No evidence for viral replication was found, and no defective particles were detected by electron microscopy. Treatment with actinomycin D or with cycloheximide strongly inhibited both the fluorescent antigens and the CPE, whereas 5-fluorodeoxyuridine and bromodeoxyuridine were ineffective.
Polypeptide Composition of Poliovirions, Naturally Occurring Empty Capsids, and 14
S
Precursor Particles
Authors: Phillips Bruce A. , Fennell Robert
Published: January 2020

The three serotypes of poliovirus were compared with respect to their polypeptide composition. Type 1, 2, and 3 strains were clearly different from each other in the electrophoretic mobilities of their larger structural polypeptides. Some of the viral polypeptides formerly identified as single peaks (e.g., VP 2) were shown to contain multiple components, indicating that purified virions contain at least six polypeptides. Three type 1 strains were indistinguishable in their viral polypeptides. A quantitative estimate was made of the polypeptide composition of the type 1 Mahoney poliovirion, as well as of naturally occurring empty capsids and 14
S
precursor particles. The data are discussed in light of the antigenic differences among polioviruses and the possible modes of virion morphogenesis.
Effect of Viral Double-Stranded RNA on Mammalian Cells in Culture: Cytotoxicity Under Conditions Preventing Viral Replication and Protein Synthesis
Authors: Cordell-Stewart B. , Taylor Milton W.
Published: January 2020

Noninfectious bovine enterovirus double-stranded RNA induces cytopathic effects when added to mammalian cells in culture. This is demonstrated by
51
Cr release from prelabeled murine lymphoma cells and trypan blue uptake. Also, the induction of cell death by this viral RNA occurs in the presence of inhibitors of protein synthesis (cycloheximide and puromycin). The possible role and mechanism of viral, double-stranded RNA as a cytopathic agent in virally infected cells are discussed.
Variable Transforming Activity of Nonglucosylated and Fragmented T4 DNA
Authors: McNicol Lore' Anne
Published: January 2020

Nonglucosylated and fragmented T4 DNA shows a gene-specific variation in transformation efficiency.
Acquisition of Sequences Homologous to Host DNA by Closed Circular Simian Virus 40 DNA II. Further Studies on the Serial Passage of Virus Clones
Authors: Lavi Sara , Rozenblatt Shmuel , Singer Maxine F. , Winocour Ernest
Published: January 2020

Three plaque isolates of SV40 strain 777 and 1 plaque isolate of strain 776 were grown to high-titer stocks and serially passaged, undiluted, in monkey BS-C-1 cells. In each case, the serial passaging procedure resulted in the accumulation of closed-circular SV40 DNA molecules containing covalently linked sequences homologous to reiterated host cell DNA (called substituted virus DNA). The relative yields, at a given passage level, of SV40 DNA with measurable homology to host DNA varied in different sets of serial passages, including passages of the same virus clone. More reproducible yields of substituted viral DNA progeny were obtained when the serial passaging procedure was initiated from earlier passages rather than from the original plaque-purified stock. Fractionation of closed-circular SV40 DNA molecules on alkaline sucrose gadients indicated that the majority of substituted virus DNA molecules are not plaque producers and are slightly smaller in size than plaque-forming DNA molecules which display no detectable homology to host DNA. Evidence that substituted SV40 DNA molecules replicate during serial undiluted passage was obtained from experiments which demonstrated (i) the presence of host sequences in replicative forms of the viral DNA and (ii) the incorporation of
3
H-thymidine into host sequences isolated from the mature substituted virus DNA molecule.
Protein Kinase Activity from Vaccinia Virions: Solubilization and Separation into Heat-Labile and Heat-Stable Components
Authors: Kleiman Jay , Moss Bernard
Published: January 2020

A protein kinase was solubilized from whole vaccinia virions by using a solution containing deoxycholate, dithiothreitol, and sodium or potassium chloride. The released enzyme was completely dependent on Mg
2+
and was greatly stimulated by added basic proteins such as protamine or histones. Dithiothreitol was also stimulatory, whereas GTP, CTP, UTP, and P
i
at concentrations equimolar with ATP had little or no effect. Attempts to purify the protein kinase were initially unsuccessful, leading us to consider that either the enzyme was extremely labile or that two readily separable components were required for activity. The observation that the material extracted with NP-40 detergent during the preparation of viral cores stimulated the protein kinase activity of the intact cores supported the second possibility. As the protein kinase, now solubilized from viral cores, was passed through successive DEAE-cellulose columns, it became increasingly dependent for activity on addition of the NP-40 extract. A 30- to 40-fold stimulation of protein kinase activity, which afforded recovery of essentially all starting activity, could be effected by addition of the NP-40 extract to the partially purified enzyme. The NP-40 extract was shown to contain a heat stable, trypsin-sensitive protein, whose action could not be duplicated by cyclic nucleotides.
Basic Characterization of a
Pseudomonas aeruginosa
Pilus-Dependent Bacteriophage with a Long Noncontractile Tail
Authors: Bradley David E.
Published: January 2020

Bacteriophage PO4 has been found to depend on the presence of pili for the infection of its host organism,
Pseudomonas aeruginosa.
Unlike other pilus phages, which either contain RNA and are “spherical” or contain single-stranded DNA and are filamentous, PO4 has a head and a long noncontractile tail. This paper describes its basic characters, and a quantitative study is made of its adsorption to exponential-phase cells of piliated and nonpiliated strains of
P. aeruginosa.
PO4 is found to contain double-stranded DNA and appears to be virulent towards its two host strains.
Viral Protein Synthesis in Bacteriophage φ29-Infected
Bacillus subtilis
Authors: Hawley Louise A. , Reilly Bernard E. , Hagen Elaine W. , Anderson Dwight L.
Published: January 2020

Twenty-three
14
C-labeled phage φ29-specific proteins in lysates of UV-irradiated
Bacillus subtilis
have been resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis and identified by autoradiography. Included in this group of proteins are the six major structural proteins of the virion. Analysis of the temporal sequence of viral protein synthesis indicates that three groups of proteins can be identified by time of appearance, beginning at 2 to 4, 4 to 6, or 8 to 10 min after infection, respectively. These proteins account for approximately 90% of the coding capacity of the φ29 genome.
DNA Polymerase Associated with Human Hepatitis B Antigen
Authors: Kaplan Paul M. , Greenman Richard L. , Gerin John L. , Purcell Robert H. , Robinson William S.
Published: January 2020

DNA polymerase activity was detected in each of eight preparations of concentrated human hepatitis B antigen (HBAg) rich in Dane particles prepared by high-speed centrifugation of antigen-positive human plasma and in none of seven control preparations prepared in the same way from HBAg-negative plasma. The incorporation of
3
H-thymidine-methyl-5′-triphosphate into DNA was dependent on four deoxyribonucleoside triphosphates and MgCl
2
. Treatment of the concentrated HBAg preparations with the nonionic detergent Nonidet P-40 (NP40) more than doubled the enzyme activity. Fractionation of the concentrated HBAg preparation in sucrose density gradients after treatment with NP40 revealed that the enzyme activity appeared within the density range of Dane core antigen but at a slightly higher density than the average for core antigen. The only particles observed by electron microscopy in this region of the gradient were typical 28-nm cores, suggesting that the DNA polymerase activity was associated with a subpopulation of cores. No DNA polymerase activity was found in purified 20-nm HBAg particles. The DNA product of the reaction remained associated with the 110
S
core and was not susceptible to DNase digestion when associated with the core. Inhibition of the reaction by actinomycin D and daunomycin suggested that the reaction was dependent on a DNA template associated with the core.
Analysis of Minimal Functions of Simian Virus 40 II. Enhancement of Oncogenic Transformation in Vitro by UV Irradiation
Authors: Seemayer N. H. , Defendi V.
Published: January 2020

After
light
UV irradiation (5,000 to 10,000 ergs/mm
2
) “complete” and “defective” simian virus 40 (SV40) showed an enhancement of oncogenic transformation capacity in Syrian hamster kidney cells in vitro up to 180 and 270% of the controls, respectively. Simultaneously with the enhancement of transformation, an increase in T-antigen induction was observed in CV-1 cells infected with light UV-irradiated SV40; infectivity, however, was correspondingly reduced by 1 log
10
. After strong UV irradiation (10,000 to 80,000 ergs/mm
2
) of “complete” and “defective” SV40, transformation capacity in vitro proved to be the most resistant viral function. It was only slightly reduced in comparison with a 4 to 5 log
10
reduction of infectivity. T-antigen induction of SV40 was also equally resistant to strong UV irradiation. We found no evidence of “multiplicity reactivation” involved in the high resistance of transformation capacity of SV40 after UV irradiation. Syrian hamster kidney cells transformed in vitro by UV-irradiated SV40 contained the SV40-specific T-antigen and showed the same morphology and growth characteristics as cells transformed by non-irradiated “complete” or “defective” SV40. They induced malignant tumors after subcutaneous inoculation into Syrian hamsters.
Cleavage of Circular, Superhelical Simian Virus 40 DNA to a Linear Duplex by S
1
Nuclease
Authors: Beard Peter , Morrow John F. , Berg Paul
Published: January 2020

S
1
nuclease, the single-strand specific nuclease from
Aspergillus oryzae
can cleave both strands of circular covalently closed, superhelical simian virus 40 (SV40) DNA to generate unit length linear duplex molecules with intact single strands. But circular, covalently closed, nonsuperhelical DNA, as well as linear duplex molecules, are relatively resistant to attack by the enzyme. These findings indicate that unpaired or weakly hydrogen-bonded regions, sensitive to the single strand-specific nuclease, occur or can be induced in superhelical DNA. Nicked, circular SV40 DNA can be cleaved on the opposite strand at or near the nick to yield linear molecules. S
1
nuclease may be a useful reagent for cleaving DNAs at regions containing single-strand nicks. Unlike the restriction endonucleases, S
1
nuclease probably does not cleave SV40 DNA at a specific nucleotide sequence. Rather, the sites of cleavage occur within regions that are readily denaturable in a topologically constrained superhelical molecule. At moderate salt concentrations (75 mM) SV40 DNA is cleaved once, most often within either one of the two following regions: the segments defined as 0.15 to 0.25 and 0.45 to 0.55 SV40 fractional length, clockwise, from the EcoR
I
restriction endonuclease cleavage site (defined as the zero position on the SV40 DNA map). In higher salt (250 mM) cleavage occurs preferentially within the 0.45 to 0.55 segment of the map.
In Vitro Polyoma DNA Synthesis: Characterization of a System from Infected 3T3 Cells
Authors: Hunter Tony , Francke Bertold
Published: January 2020

A lysate from hypotonically swollen polyoma-infected BALB/3T3 cells incorporated labeled deoxynucleotide triphosphates into both viral and cellular DNAs. The incorporation was stimulated by the presence of ATP, deoxynucleotide triphosphates, thiols, and magnesium ions. Strong inhibition of incorporation was observed with thiol reagents and arabinosyl nucleotide triphosphates. The rate of in vitro synthesis increased with the temperature of incubation as expected. Incorporation into cellular DNA for up to 2 h was observed in lysates from virus-infected and serum-stimulated cells but not from resting cells. Synthesis in the system, therefore, appeared to reflect the physiological state of the cells before preparation of the lysate. Incorporation into viral DNA stopped far sooner than that into cellular DNA. During the initial phase of the in vitro incubation, incorporation occurred into viral replicative intermediates (RI). These RIs had identical properties to those isolated after in vivo pulse labeling and a substantial proportion of them was matured to form I DNA at later times in the incubation through all the stages known to occur in vivo. Density labeling of the in vitro product showed that practically all of the RIs pre-existing in the infected cell took part in the in vitro reaction. Analysis of DNA labeled in vitro in the presence of 5-bromodeoxyuridine triphosphate showed that synthesis occurred on RIs at all stages of replication and that the progeny strands were elongated by up to 80% of unit viral DNA length. Pre-existing RIs, pulse labeled in vivo, showed evidence of a pool at a late stage of replication which required elongation of their progeny strands by approximately 25% during conversion to form I molecules. From density-labeling experiments, we were also able to show that viral DNA synthesis in vitro was semiconservative. The major reason for cessation of viral DNA synthesis in vitro was the very limited ability of the lysate to initiate new rounds of viral DNA synthesis.
Interactions of Polyoma and Mouse DNAs I. Lytic Infection of Bromodeoxyuridine-Prelabeled Mouse Embryo Cells
Authors: Türler Hans
Published: January 2020

On CsCl isopycnic centrifugation of the DNA extracted from secondary mouse embryo (ME) cultures grown in the presence of 5-bromodeoxyuridine (BUdR) and 5-fluorodeoxyuridine (FUdR) for 40 h, 10 to 25% of the DNA was found to be unsubstituted, 70 to 80% was bromouracil-hybrid DNA, and 5 to 10% was heavy DNA. These results together with cell number determinations, autoradiography, and Feulgen microspectrophotometry revealed three types of cells in these cultures: (i) 60 to 80% of the cells replicated their DNA once, divided, and then stopped mitotic activity, (ii) 5 to 10% were going through a second round of DNA replication; whereas (iii) 10 to 30% did not replicate DNA during the BUdR-FUdR exposure. After the transfer of these cultures to normal medium (without BUdR-FUdR), up to 20% of the cells resumed DNA synthesis asynchronously within 60 h, but no increase in cell number was observed. BUdR-FUdR-treated cultures, which were infected with polyoma virus in the absence of the thymidine analogues, supported a lytic infection to the same extent as did untreated ME cultures. This was concluded from the similar number of cells, which were induced to synthesize DNA, from the similar replication rate of the viral DNA, from the similar number of cells containing polyoma capsid proteins, and from the similar yields of progeny virus determined by hemagglutination and plaque formation. Thus, BUdR-prelabeled ME cultures are suitable for the investigation of interactions of the polyoma and mouse genomes during the lytic infection.
Virus Replication in Enucleate Cells: Vesicular Stomatitis Virus and Influenza Virus
Authors: Follett E. A. C. , Pringle C. R. , Wunner W. H. , Skehel J. J.
Published: January 2020

The requirement of the presence of a nucleus for the replication of vesicular stomatitis virus and influenza virus has been examined by following the growth and development of these viruses in enucleate BS-C-1 cells. Vesicular stomatitis virus replicates normally in enucleate cells with the rate of production of infectious virus, the amount of virus-specific protein synthesis, and the type of proteins produced being essentially the same in nucleate and enucleate cells. Influenza virus does not replicate in enucleate cells, no virus gene products can be detected, and there is no inhibition of cellular protein synthesis.
Mutation to Overproduction of Bacteriophage T4 Gene Products
Authors: Karam J. D. , Bowles M. G.
Published: January 2020

R9 was isolated as one of several mutations that enhanced the growth of a leaky amber (
am
) mutant of bacteriophage T4 gene 62 (product required for phage DNA synthesis) under conditions of partial suppression by ribosomal ambiguity. R9 also enhanced the growth of leaky
am
mutants of some, but not all, other T4 “early” gene functions. R9 mapped between mutations in genes 43 and 62. By using assays involving polyacrylamide slab gel electrophoresis in the presence of sodium dodecyl sulfate, we observed the following. (i) R9 resulted in an overproduction of many T4 “early” proteins in infected cells. The most pronounced effects of R9 were observed when phage DNA synthesis and/or the functions of maturation genes 55 and 33 were not expressed. (ii) In rifampintreated infected cells, the capacity to synthesize T4 “early” proteins decayed more slowly in the presence of the R9 mutation than in the presence of the wild-type counterpart of R9. R9 appeared to have no effect on the rates of RNA synthesis either during early or late times after infection. The results suggest that the R9 mutation leads to increased functional stability of T4 “early” messengers.
Antiserum Inactivation of Electrophoretically Purified M13 Diploid Virions: Model for the F-Specific Filamentous Bacteriophages
Authors: Beaudoin Jacques , Pratt David
Published: January 2020

Antiserum inactivation experiments were carried out on electrophoretically purified diploid virions from a cross between two complementing amber mutants of phage M13. The total (homozygous plus heterozygous) diploid population, assayed on a permissive host where only one genome is needed for plaque formation, was inactivated at the same rate as haploids. Heterozygous diploids, assayed on a nonpermissive host, where both genomes are needed for plaque formation, were twice as sensitive as haploids and the total diploid population. These results have led us to propose a model for serum inactivation of the F-specific filamentous phages. According to this model, phage-neutralizing antibodies attach anywhere along the length of the phage and allow the phage to penetrate only up to the first bound antibody molecule.
Structural Studies of Avian Myeloblastosis Virus: Comparison of Polypeptides in Virion and Core Component by Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis
Authors: Stromberg Kurt , Hurley Nancy E. , Davis Nancy L. , Rueckert Roland R. , Fleissner Erwin
Published: January 2020

Two different systems of dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in separate laboratories detected analogous patterns of dye bands in virions of avian myeloblastosis virus (AMV). At least 11 of the dye bands co-migrated with the major polypeptides reported in Rous sarcoma virus. Particles with the morphology of the AMV core component, obtained after exposure of AMV to the nonionic surfactant Sterox SL, contained major polypeptides p12, p27, p60, p64, p91, and p98. The polypeptide p12 has been previously shown to be the major constituent of the inner ribonucleoprotein (RNP) of the AMV core, and has been designated p12(N). Two RNP polypeptides, p64 and p91, co-electrophoresed with purified AMV DNA polymerase and have now been designated p64(P) and p91(P). The polypeptide p27 has been identified as a probable constituent of the core shell, and has accordingly now been designated p27(C). In comparison to virions of AMV, the AMV core component contained a greatly reduced amount of polypeptide p15 and appeared to lack a major polypeptide, p19. Consequently, these polypeptides may be associated either with the exterior of the core shell or the interior of the viral envelope. Glycopeptides were not detected in AMV cores, in agreement with earlier reports that they reside in external projections from the viral envelope.
Translation of mRNA from Simian Virus 40-Infected Cells into Simian Virus 40 Capsid Protein by Cell-Free Extracts
Authors: Lodish Harvey F. , Weinberg Robert , Ozer Harvey L.
Published: January 2020

Messenger RNA was isolated from simian virus 40 (SV40)-infected and mock-infected cells by chromatography on poly(U) sepharose. When added to cell-free extracts from Chinese hamster ovary cells or rabbit reticulocytes, RNA from the infected cells, but not from mock-infected cells, stimulated synthesis of the major SV40 capsid protein. Identification of this species was done by sodium dodecyl sulfate gel electrophoresis, peptide mapping, and immunoprecipitation. The in vitro synthesized capsid protein was slightly different from virion assembled capsid protein, as shown by separation upon chromatography on hydroxylapatite and by minor differences in the peptide map.
Interference in Bacteriophage Growth by a Resident Plasmid λdv II. Role of the Promoter-Operator
Authors: Matsubara Kenichi
Published: January 2020

The susceptibilities to interference by λdv of a set of λ phages differing in the genetic structure of their right promoter-operator (pRoR) were compared. For this purpose mutant phages were added to λdv-carrier cells at various multiplicities, and abilities to escape from interference, as represented by percentages of infected complexes to produce progeny phages, were compared. It was observed that phages that carry strongly constitutive pRoR were able to escape from interference at a low multiplicity of infection, whereas phages with weakly constitutive pRoR were able to escape only when a large number of these genomes entered into a cell. Several mutations in the pRoR were arranged in order of their constitutivity, or ability to escape from interference. Next, the abilities of a set of λdv plasmids differing in their pRoR to cause interference were compared. The results showed that interference increased with increase in constitutivity of the pRoR in the plasmid genome. These observations are thought to reflect a regulatory system of the plasmid replicon, and a possible mechanism for the system is discussed.
Synthesis of Reovirus Oligo Adenylic Acid In Vivo and In Vitro
Authors: Silverstein Samuel C. , Astell Caroline , Christman Judith , Klett Hannah , Acs George
Published: January 2020

The formation of reovirus double-stranded (ds) RNA and of oligo adenylic acid (oligo A) is inhibited by 5 μg of actinomycin D per ml added at the time of viral infection. Viral proteins are synthesized and assembled into dsRNA-deficient particles under these conditions. The addition of cycloheximide to infected cells during the mid-logarithmic phase of viral replication terminates protein and dsRNA synthesis, but allows continued oligo A synthesis for about 1 h. The
3
H-labeled oligo A formed in the presence of cycloheximide is incorporated into particles whose density in CsCl is identical to that of reovirions. Using the large particulate or virus factory-containing cytoplasmic fraction of infected L-cells, we have established an in vitro system for the synthesis of oligo A. The in vitro product migrates slightly faster in sodium dodecyl sulfate acrylamide gels than marker oligo A. Oligo A synthesis in vitro continues for about 1 h, requires, the presence of only one ribonucleoside triphosphate (ATP), is not inhibited by DNase or RNase, but is abruptly terminated by the addition of chymotrypsin to the reaction mixture. Oligo A formed both in vivo and in vitro is released from the factory fraction by chymotrypsin digestion. The enzymes which catalyze the synthesis of oligo A, dsRNA, and single-stranded RNA all exhibit a similar temperature dependence with an optimum of ∼45 C. These results indicate that oligo A is formed within the core of the nascent virion after the completion of dsRNA synthesis; they suggest that the oligo A polymerase is an alternative activity of the virion-bound transcriptase and that it is regulated by outer capsomere proteins.
Bacteriophage Interference in
Bacillus subtilis
168
Authors: Yasbin R. E. , Ganesan A. T. , Young F. E.
Published: January 2020

Strains of
Bacillus subtilis
lysogenic for temperate bacteriophage SPO2 inhibit the development of bacteriophage φ1. After infection by bacteriophage φ1, DNA and RNA synthesis in the lysogenic host terminates, culminating in cell death. Bacteriophage SPO2 also prevents the production of bacteriophage φ105. Mechanisms for these two types of bacteriophage interference are discussed.
Comparison of 6/94 Virus and Sendai Virus RNA by RNA-RNA Hybridization
Authors: Kolakofsky Daniel , Spahr Pierre-Francois , Koprowski Hilary
Published: January 2020

The genomic RNA of 6/94 virus, an agent isolated from the brains of multiple sclerosis patients, was studied for sequence homology by RNA-RNA hybridization with closely related Sendai virus and another paramyxovirus virus, Newcastle disease virus. It was found that the genomic RNA of 6/94 virus hybridizes equally as well to the virus-specific 18
S
RNA found in Sendai-infected cells as that of Sendai virus.
Transcription Unit Mapping in Bacteriophage T7 II. Proportionality of Number of Gene Copies, mRNA, and Gene Product
Authors: Bräutigam Alan R. , Sauerbier Walter
Published: January 2020

The effect of UV irradiation of bacteriophage T7 on in vivo early RNA synthesis has been studied by direct quantitation of the gene-specific RNA transcripts. The results show that the early region of phage T7 is transcribed from left to right as a single unit. Furthermore, gene inactivation, the UV sensitivity of synthesis of gene-specific RNA, and the UV sensitivity of synthesis of the corresponding proteins all follow pseudo first-order kinetics in multiply infected cells, demonstrating a random statistical correlation between both transcriptional sampling of gene copies and translational sampling of the resultant RNA transcripts. In addition, these simple kinetics imply an absence of positive feedback mechanisms compensating for the differential decline of individual early gene products in cells multiply infected with phage T7.
Host Cell Regulation of Induction of Epstein-Barr Virus
Authors: Glaser Ronald , Nonoyama Meihan
Published: January 2020

When Epstein-Barr virus (EBV) negative cells (Raji) were treated with iododeoxyuridine, only the early antigen (EA) component was induced. There was no significant increase in EBV DNA and no virus particles were observed. Somatic-cell hybrids were prepared from the fusion of Raji and D98 cells (D98/Raji). When these cells were treated with iododeoxyuridine, early antigen EBV DNA, and virus particles were synthesized. These data suggest cellular control over the expression of the EBV genome.
Mechanism of Sindbis Virus-Induced Intrinsic Interference with Vesicular Stomatitis Virus Replication
Authors: Hunt John M. , Marcus Philip I.
Published: January 2020

Heterologous viral interference is induced by Sindbis virus against vesicular stomatitis virus (VSV) in a manner analogous to intrinsic interference with Newcastle disease virus replication. Interference in both systems (i) depends upon early expression of the inducing virus genome, (ii) shows similar kinetics of induction, (iii) does not involve interferon action, and (iv) appears to be manifest as an all-or-none effect. VSV can be added to the list of viruses blocked by intrinsic interference. Sindbis virus-induced intrinsic interference with VSV replication is not mediated through homotypic interference by defective interfering particles; rather, T-particle and B-particle synthesis is inhibited. Significantly, intrinsic interference has no effect on primary transcription directed by the virion-associated transcriptase in VSV-challenged cells. However, Sindbis virus appears to induce interference with the VSV-RNA synthesized subsequent to primary transcription, namely, that which is dependent on protein synthesis. Thus, the target of intrinsic interference appears to be a reaction subsequent to primary transcription but prior to the appearance of protein synthesis-dependent VSV RNA, secondary transcription.
Sialoglycoprotein of Vesicular Stomatitis Virus: Role of the Neuraminic Acid in Infection
Authors: Schloemer Robert H. , Wagner Robert R.
Published: January 2020

Neuraminidase free of proteolytic activity substantially reduced the infectivity of vesicular stomatitis (VS) virus but less effectively than trypsin. The only sugar residue hydrolyzed by neuraminidase was
N
-acetyl neuraminic acid, ∼89% of which was liberated from virion glycoprotein and the rest from virion glycolipid. Desialylation of virion glycoprotein but not of glycolipid resulted in progressive loss of infectivity. Sialyl transferase prepared and partially purified from BHK-21 cells catalyzed resialylation by CMP-[
14
C]sialic acid of the glycoprotein of neuraminidase-treated VS virions and supersialylation of unhydrolyzed VS viral glycoprotein. Resialylation of desialylated VS virions resulted in substantial (26-fold) restoration of their infectivity. We conclude that terminal neuraminic acids of VS viral sialoglycoprotein play an important role in initiation of infection with this virus.
Characterization of the Large Picornaviral Polypeptides Produced in the Presence of Zinc Ion
Authors: Butterworth Byron E. , Korant Bruce D.
Published: January 2020

Zinc ion inhibits the posttranslational cleavages of human rhinovirus-1A, encephalomyocarditis virus, and poliovirus polypeptides. Each virus displayed a different susceptibility to zinc. However, in each case the cleavages of the capsid precursor and the cleavages analogous to the C → D → E conversion in encephalomyocarditis virus were most sensitive to zinc. Higher concentrations of zinc resulted in the buildup of even larger precursor polypeptides of a size between 106,000 and 214,000 daltons. The sizes of these polypeptides and the relative position of their gene loci on the viral RNA were determined. These data were used to place these polypeptides in the over-all scheme of viral protein processing.
Transcriptional Regulation of T4 Bacteriophage-Specific Enzymes Synthesized In Vitro
Authors: Cohen Paul S. , Natale Peter J. , Buchanan John M.
Published: January 2020

In contrast to dihydrofolate reductase and four other phage-specific enzymes, the initiation of deoxynucleotide kinase is essentially prevented if rifampin is added to a culture of
Escherichia coli
B cells within 1.5 min after infection with T4. Deoxynucleotide kinase thus belongs to a group of so-called delayed-early enzymes that is not initiated from an immediate-early promoter site. We prepared crude extracts from infected cells in a manner designed to maintain the integrity of the complexes of native, endogenous T4 DNA with bacterial structural and enzymatic units concerned with RNA synthesis. The initiation of the synthesis of the mRNA for dihydrofolate reductase, an example of an immediate-early enzyme, and deoxynucleotide kinase, a special type of delayed-early enzyme, was studied with these extracts prepared from cells infected in the absence or presence of chloramphenicol. Initiation of transcription of the dihydrofolate reductase gene is immediate when programmed by extracts made either from cells treated with chloramphenicol prior to infection (CM extracts) or from cells 3 min into the normal infection cycle (3-min extracts). However, initiation of transcription of the deoxynucleotide kinase gene programmed by CM extracts is delayed 2 min relative to the immediate initiation of transcription of the deoxynucleotide kinase gene programmed by 3-min extracts. These experiments duplicated in vitro effects of the antibiotics on the synthesis of phage-specific mRNA previously noted only in vivo.
Inhibition of Herpes Virus-Induced Cell Fusion by Concanavalin A, Antisera, and 2-Deoxy-
d
-Glucose
Authors: Ludwig H. , Becht H. , Rott R.
Published: January 2020

Pseudorabies virus-induced cell fusion in rabbit kidney cells can be prevented by Concanavalin A added early after infection. The infected cells are not agglutinated and the infectivity of cell-free virus is not reduced. Sera from productively infected animals also inhibit polykaryocytosis, whereas a hyperimmune serum directed against virus structural components has no effect. 2-Deoxy-
d
-glucose reversibly disturbs virus-induced fusion and reduces significantly the virus infectivity.
DNA of a Human Hepatitis B Virus Candidate
Authors: Robinson William S. , Clayton David A. , Greenman Richard L.
Published: January 2020

Particles containing DNA polymerase (Dane particles) were purified from the plasma of chronic carriers of hepatitis B antigen. After a DNA polymerase reaction with purified Dane particle preparations treated with Nonidet P-40 detergent, Dane particle core structures containing radioactive DNA product were isolated by sedimentation in a sucrose density gradient. The radioactive DNA was extracted with sodium dodecyl sulfate and isolated by band sedimentation in a preformed CsCl gradient. Examination of the radioactive DNA band by electron microscopy revealed exclusively circular double-stranded DNA molecules approximately 0.78 μm in length. Identical circular molecules were observed when DNA was isolated by a similar procedure from particles that had not undergone a DNA polymerase reaction. The molecules were completely degraded by DNase 1. When Dane particle core structures were treated with DNase 1 before DNA extraction, only 0.78-μm circular DNA molecules were detected. Without DNase treatment of core structures, linear molecules with lengths between 0.5 and 12 μm, in addition to the 0.78-μm circles were found. These results suggest that the 0.78-μm circular molecules were in a protected position within Dane particle cores and the linear molecules were not within core structures. Length measurements on 225 circular molecules revealed a mean length of 0.78 ± 0.09 μm which would correspond to a molecular weight of around 1.6 × 10
6
. The circular molecules probably serve as primer-template for the DNA polymerase reaction carried out by Dane particle cores. Thermal denaturation and buoyant density measurements on the Dane particle DNA polymerase reaction product revealed a guanosine plus cytosine content of 48 to 49%.
Mechanism of Adsorption and Eclipse of Bacteriophage φX174 III. Comparison of the Activation Parameters for the In Vitro and In Vivo Eclipse Reactions With Mutant and Wild-Type Virus
Authors: Incardona Nino L.
Published: January 2020

In a starvation buffer containing 10
−3
M divalent cations, φX174 undergoes viral eclipse above 20 C when attached to intact host cells. An in vitro structural transition that is similar to that observed in this in vivo eclipse reaction occurs over the same temperature range in 0.1 M CaCl
2
(pH 7.2). Since both reactions result in a loss of infectivity, their kinetics have been compared in this report. Both exhibit a biphasic first-order loss in PFU that is a result of two competing first-order processes. However, a single type of heterogeneity in the population of virions is not the basis for both competing slower reactions. The Arrhenius plots of the faster components show that the in vitro eclipse reaction has the same activation energy of 35 kcal/mol (ca. 1.47 × 10
5
J/mol) as the in vivo reaction but a 10-fold lower Arrhenius preexponential factor. This is further evidence that certain features of the in vivo mechanism are retained in the in vitro reaction. In the case of the slower components, the in vitro reaction has an activation energy of 37 kcal/mol (1.55 × 10
5
J/mol), whereas that of the in vivo reaction is only 5 kcal/mol (2.1 × 10
4
J/mol). A similar analysis has been performed on a cold-sensitive eclipse mutant of φX174. In vivo, the mutation is expressed by a two- to three-fold lower Arrhenius preexponential factor for both components of the eclipse reaction when compared to wt virus. The activation energies for both components are the same as wt virus. These results suggest that the mechanism of the eclipse reaction can be operationally divided into two aspects, each subject to mutational alteration.
In Vitro Synthesis of Proteins by Membrane-Bound Polyribosomes from Vesicular Stomatitis Virus-Infected HeLa Cells
Authors: Grubman Marvin J. , Ehrenfeld Ellie , Summers Donald F.
Published: January 2020

Membrane-bound polysomes from vesicular stomatitis virus (VSV)-infected HeLa cells synthesize predominantly three proteins in an in vitro protein synthesizing system. These three proteins have different molecular weights than the viral structural proteins, i.e., 115,000, 88,000, and 72,000. Addition of preincubated L or HeLa cell S10 or HeLa cell crude initiation factors stimulates amino acid incorporation and, furthermore, alters the pattern of proteins synthesized. Stimulated membrane-bound polysomes synthesize predominantly viral protein G and lesser amounts of N, NS, and M. In vitro synthesized proteins G and N are very similar to virion proteins G and N based on analysis of tryptic methionine-labeled peptides. Most methionine-labeled tryptic peptides of virion G protein contain no carbohydrate moieties, since about 90% of sugar-labeled peptides co-chromatograph with only about 10% of methionine-labeled peptides. Sucrose gradient analysis of the labeled RNA present in VSV-infected membrane-bound polysomes reveals a relative enrichment in a class of viral RNA sedimenting slightly faster than the total population of the 13 to 15
S
mRNA, as compared to a VSV-infected crude cytoplasmic extract. A number of proteins, other than the viral structural proteins, are synthesized in the cytoplasm of five lines of VSV-infected cells. One of these proteins has the same molecular weight as the major in vitro synthesized protein, P
88
. In vitro synthesized protein P
88
does not appear to be a precursor of viral structural proteins G, N, or M based on pulse-chase experiments and tryptic peptide mapping. Nonstimulated membrane-bound polysomes from uninfected HeLa cells synthesize the same size distribution of proteins as nonstimulated VSV-infected membrane-bound polysomes.
Characteristics of PRD1, a Plasmid-Dependent Broad Host Range DNA Bacteriophage
Authors: Olsen Ronald H. , Siak June-Sang , Gray Robert H.
Published: January 2020

Several distinctive properties of PRD1, an icosahedral plasmid-dependent phage, are described. The drug-resistance plasmid-dependent host range of PRD1 extends beyond the P incompatibility group and includes gram-negative bacteria containing plasmids of incompatibility groups N and W. PRD1 phage will infect pseudomonads and
Enterobacteriaceae
containing either a P or W incompatibility group plasmid. PRD1 adsorbs to the cell wall of R
+
bacteria and thus its infectivity indicates cell wall alterations by these drug-resistance plasmid groups. PRD1 nucleic acid is duplex DNA with an estimated molecular weight of 24 × 10
6
. The appearance of PRD1 in electron micrographs is suggestive of lipid content in addition to its buoyant density of 1.348 in CsCl and its sensitivity to chloroform. The latent period of PRD1 varies with the R
+
host bacterial strain used for growth of the phage.