bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of the Autoregulation of Simian Virus 40 Gene
A
Authors: Alwine James C. , Reed Steven I. , Stark George R.
Published: January 2020

Cells infected by
tsA
mutants of simian virus 40 (SV40) overproduce early RNA. Overproduction results from failure of the temperature-sensitive
A
protein (T antigen) to inhibit early transcription. The amount of early RNA in the cytoplasm, determined quantitatively from the kinetics of hybridization to labeled complementary SV40 DNA, was elevated at both permissive (32°C) and nonpermissive (41°C) temperatures in all the early mutants tested (
tsA
7, -30, -58, and -209), but not in the late mutant
tsB
4. The amount of early RNA in a culture maintained at 32°C for 72 h and then shifted to 41°C was maximum when each cell was infected initially with at least one plaque-forming unit of
tsA
58. Azidocytidine (2′-deoxy-2′-azidocytidine), which inhibits initiation of DNA synthesis, did not cause overproduction of early RNA in cells infected with wild-type SV40, showing that the effect seen with
tsA
mutants is not due to interference with initiation of DNA synthesis per se. In parallel infections at 41°C, the amount of early RNA per copy of viral DNA was as much as 2,000 times greater with
tsA
58 than with wild-type SV40, even though there was no replication of the
tsA
58 DNA. Synthesis of late RNA could not be detected during the first 20 h of an infection by either virus at 32°C, indicating that late and early transcription are under different control. In three cell lines transformed by
tsA
mutants, the amount of early RNA increased moderately after a shift from 32 to 41°C, whereas with homologous cells transformed by wild-type virus, the amount of early RNA decreased, indicating that the
A
protein may be able to repress transcription of integrated SV40 DNA. All the observations are consistent with a simple model in which the binding of
A
protein at the origin of replication blocks either binding of RNA polymerase to the early promoter or its progress through the early gene(s).
[125I]deoxycytidine used in a rapid, sensitive, and specific assay for herpes simplex virus type 1 thymidine kinase
Authors: Summers W C , Summers W P
Published: January 2020

[125I]deoxycytidine was a good substrate for herpes simplex virus type 1 thymidine kinase (TK), whereas [125I]deoxycytidine was a very poor substrate for cellular TK. Simple, sensitive, and specific assays for viral TK could be carried out in vivo and in vitro even in the presence of cell TK. Autoradiographic detection of incorporated [125I]deoxycytidine provided a rapid and simple method for detection of and screening for viral TK in infected as well as viral TK-transformed cells.
Defective Friend Spleen Focus-Forming Virus: Interfering Properties and Isolation Free from Standard Leukemia-Inducing Helper Virus
Authors: Eckner Robert J. , Hettrick Kristine L.
Published: January 2020

Defective Friend spleen focus-forming virus (SFFV) is able to interfere with the ability of its naturally occurring leukemia-inducing helper virus (LLV-F) to induce XC plaque formation in several different strains of mouse embryo cells. This interference has been observed by using two different SFFV preparations, one contained in an NB-tropic stock of Friend virus (FV) complex, and the second present in a C57BL-adapted strain of FV complex containing an associated B-tropic LLV-F helper. The LLV-F in NB-tropic FV complex effectively induced XC plaques in C57BL/6 (
Fv-1
bb
;
Fv-2
rr
) mouse embryo fibroblasts (MEF) only in the absence of coinfecting SFFV, indicating that
Fv-2
-associated resistance to SFFV-induced focus formation in vivo does not necessarily extend to the restriction of SFFV function(s) in vitro (i.e., in
Fv-2
rr
C57BL MEF). SFFV interference appears to be an intracellular event since LLV-F can adsorb onto, penetrate, and rescue defective murine sarcoma virus (MSV) from transformed 3T3FL S
+
L
−
cells with equal efficiency in the presence and absence of SFFV. However, significantly fewer LLV-infected S
+
L
−
cells released LLV-F progeny if SFFV was present. These observations suggest that Friend SFFV may be classified as a defective, interfering (DI) particle. Further support for this conclusion has come from studies designed to investigate two physical properties of defective SFFV particles. SFFV layered onto a 0 to 20% sucrose sedimentation gradient was recovered as a symmetrical band of virus that sedimented more slowly than standard LLV-F particles. Pooled SFFV-containing gradient samples contained visualizable type C virus particles and occasionally small amounts of detectable LLV-F. In an attempt to determine the buoyant density of sedimentation gradient-purified SFFV, pooled SFFV samples were layered onto a 25 to 50% sucrose equilibrium density gradient and were centrifuged to equilibrium. Greater than 50% of the infectious SFFV originally layered onto this gradient was recovered and seen as a narrow symmetrical band with peak SFFV infectivity at a sucrose density of 1.14 g/ml. The observed difference between SFFV and LLV-F buoyant densities appears to be related to an inherent physical property of each virus. Mixtures of these two viruses express the buoyant density of that virus population which is in excess in fabricated FV complexes probably due to the formation of SFFV-LLV aggregates. Finally, gradient-purified SFFV failed to induce XC plaques in MEF and did not function to rescue MSV as expected since SFFV itself is replication defective.
Prolongation of herpes simplex virus latency in cultured human cells by temperature elevation
Authors: O'Neill F J
Published: January 2020

Treatment of herpes simplex virus type 2 (HSV-2)-infected human fibroblast cells with cytosine arabinoside (ara-C) at 25 microgram/ml resulted in complete inhibition of virus replication. Removal of ara-C after 7 days of treatment ultimately resulted in renewed virus replication, but after a delay of at least 5 days. If however, the temperature was elevated from 37 degrees C to 39.5 to 40 degrees C at the time of ara-C reversal, infectious HSV-2 did not reappear. As long as the cultures were maintained at 39.5 to 40 degrees C (up to at least 128 days), HSV-2 was latent and infectious virus was undetectable. If the temperature was reduced to 37 degrees C at any time during the latent period, infectious virus was always reactivated, but only after a period of incubation at 37 degrees C of a least 11 days. Infectious-center assays performed with latent cultures indicated that only a very small fraction of cells could reactivate virus. The infectious-center titer did not show significant changes during much of the period of latency. This seemed to argue against the possibility that the latent cultures were synthesizing very small amounts of infectious virus. Additional studies were aimed at determining the minimum incubation period at 37 degrees C required to reactivate infectious HSV-2. Latent cultures reduced from 39.5 to 40 degrees C to 37 degrees C for less than 96 h did not yield infectious HSV-2, but those incubated at 37 degrees C for 96 h or more did.
Herpes simplex virus DNA polymerase as the site of phosphonoacetate sensitivity: temperature-sensitive mutants
Authors: Purifoy D J , Powell K L
Published: January 2020

Temperature-sensitive (ts) mutants in a number of complementation groups of herpes simplex virus type 1 (HSV-1) are deficient in DNA polymerase induction at the restrictive temperature. Twenty-two mutants in 15 complementation groups were tested for sensitivity to phosphonoacetate (PAA), a compound that inhibits HSV replication in vivo and the DNA polymerase in vitro. One mutant, tsD9, was resistant to PAA (Pr), whereas all others were sensitive. Revertants of tsD9 to the ts+ phenotype simultaneously lost PAA resistance. Additional Pr mutants were isolated from ts mutants belonging to several complementation groups of HSV-1. Double mutants (ts Pr phenotype) were used in three-factor recombination analyses to locate the PAA locus on the genetic map at a position indistinguishable from the ts lesion in tsD9. In all cases, resistance or sensitivity to PAA in vivo was correlated with resistance or sensitivity of DNA polymerase in vitro. These data are compatible with the temperature-sensitive lesion of tsD9 and the determinant of PAA sensitivity both residing in the structural gene for DNA polymerase.
Properties of deoxynucleoside 5'-monophosphatase induced by bacteriophage T5 after infection of Escherichia coli
Authors: Mozer T J , Warner H R
Published: January 2020

Bacteriophage T5 induced a deoxynucleoside 5'-monophosphatase during its infection of Escherichia coli. The enzyme was purified about 100-fold. It was clearly distinct from the host 5'-nucleotidase activity in its physical characteristics and substrate specificity. The enzyme was active on deoxynucleoside 5'-monophosphates but was not active as a phosphatase on ribonucleotides, deoxynucleoside 5'-triphosphates, deoxynucleoside 3'-monophosphates, or deoxyoligonucleotides. Furthermore, it did not have oligonucleotidase or exonuclease activity. The enzyme could exist in multimeric form but had a monomer molecular weight of about 25,000.
Isolation and characterization of a bacteriophage T5 mutant deficient in deoxynucleoside 5'-monophosphatase activity
Authors: Mozer T J , Thompson R B , Berget S M , Warner H R
Published: January 2020

A bacteriophage T5 mutant has been isolated that is completely deficient in the induction of deoxynucleoside 5'-monophosphatase activity during infection of Escherichia coli F. The mutant bacteriophage has been shown to be deficient in the excretion of the final products of DNA degradation during infection of E. coli F, and about 30% of the host DNA's thymine residues were reinocorporated into phage DNA. During infection with this mutant, host DNA degradation to trichloroacetic acid-soluble products was normal, host DNA synthesis was shut off normally, and second-step transfer was not delayed. However, induction of early phage enzymes and production of DNA and phage were delayed by 5 to 15 min but eventually reached normal levels. The mutant's phenotype strongly suggests that the enzyme's role is to act at the final stage in the T5-induced system of host DNA degradation by hydrolyzing deoxynucleoside 5'-monophosphates to deoxynucleosides and free phosphate; failure to do this may delay expression of the second-step-transfer DNA.
BALB/3T3 cells infected by the ts3 mutant of polyoma virus fail to accumulate virus-specific early RNA at the nonpermissive temperature
Authors: Cogen B , Eckhart W
Published: January 2020

We measured the accumulation of virus-specific early RNA in BALB/3T3 cells infected by the ts3 mutant of polyoma virus by annealing cytoplasmic RNA from infected cells to purified, radiolabeled, "early" strand of polyoma DNA. Cells infected by the ts3 mutant fail to accumulate virus-specific early RNA at the nonpermissive temperature.
Temperature-sensitive growth regulation in one type of transformed rat cells induced by the tsa mutant of polyoma virus
Authors: Seif R , Cuzin F
Published: January 2020

A fibroblast line of the 3T3 type with a low saturation density was established from Fisher rat embryo cells. After infection with either wild-type or tsa mutant polyoma virus, transformants were isolated and cloned at 33 degrees C on the basis of their ability either to grow as dense foci on plastic in liquid medium (type N) or to form colonies in soft agar (type A). Polyoma T antigen was detected in all of the transformed lines. The following growth characteristics were studied for both types at 33 and 41 degrees C: saturation density, growth in soft agar and at a low serum concentration, colony-forming ability, and generation time. tsa-N transformants behaved at 33 degrees C similarly to transformed cells, but reverted at 41 degrees C to the nontransformed phenotype for all of these characters. tsa-A transformants and all of the wild-type transformants exhibited the transformed phenotype at both low and high temperatures. These results led us to distinguish at least two types of virus-induced transformants. In one of them, the activity of the protein affected by the tsa mutation appears to be necessary for the expression of several of the characters defining the transformed state.
Peptide Map Comparison of Infectious Pancreatic Necrosis Virus-Specific Polypeptides
Authors: Dobos Peter , Rowe David
Published: January 2020

An investigation of virus-specific protein synthesis in infectious pancreatic necrosis virus (IPNV)-infected rainbow trout gonad cells was undertaken to find a relationship between the coding capacity of the virus genome (two segments of double-stranded RNA of 2.5 × 10
6
and 2.3 × 10
6
molecular weight) and the sizes and relative amounts of virus-specific proteins. Using polyacrylamide slabgel electrophoresis and autoradiography, eight distinct virus-specific polypeptides were detected in infected, [
35
S]methionine-labeled cells. These proteins may be grouped into three size classes on the basis of molecular weight: (i) large, α (90,000); (ii) medium, β
1
(59,000), β
2
(58,000), and β
3
(57,000); and (iii) small, γ
1
(29,000), γ
1A
(28,000), γ
2
(27,000), and γ
3
(25,000). The combined molecular weight of these polypetides (373,000) is beyond the coding capacity of the virus genome. Purified IPNV contained polypeptides α, β
3
, γ
1
, and γ
1A
. Pulse-chase experiments and tryptic peptide map comparisons revealed that only four of the eight intracellular proteins were primary gene products, namely, α, β
1
, γ
1
, and β
2
, with a combined molecular weight of 205,000. Of these primary gene products only the α polypeptide was found to be stable, whereas the other three underwent intracellular proteolytic cleavage during virus morphogenesis. Polypeptide β
1
was cleaved to generate β
2
and β
3
; γ
1
was trimmed to produce γ
1A
, and the only nonstructural primary gene product, γ
2
, was found to be a precursor of γ
3
. These results suggest that IPNV possesses a unique mechanism to synthesize three size classes of proteins using mRNA transcripts from two high-molecular-weight double-stranded RNA genome segments.
Transformation-defective mutants of Rous sarcoma virus with src gene deletions of varying length
Authors: Kawai S , Duesberg P H , Hanafusa H
Published: January 2020

The RNAs of transformation-defective (td) deletion mutants of the Schmidt-Ruppin strain of Rous sarcoma virus were found to vary in size when compared by polyacrylamide gel electrophoresis. Three of seven td mutants appeared to recombine with a mutant of Rous sarcoma virus (Schmidt-Ruppin), which has a temperature-sensitive sarcoma (src) gene and is termed ts68, to give rise to recombinants with a reduced temperature sensitivity. The results suggested that different clones of td mutants exist: some in which the src gene appears to be deleted, and others in which the src gene is only partially deleted. A direct correlation between RNA size and the extent of src gene deletion measured by recombination was not obtained, possibly because the recombination assay could only detect src sequences homologous to the lesion(s) of ts68, whereas the electrophoretic analysis of the RNA measured src deletions as well as other possible alterations of the RNA.
DNA-binding proteins in the cytoplasm of vaccinia virus-infected mouse L-cells
Authors: Soloski M J , Esteban M , Holowczak J A
Published: January 2020

Mouse L cell fibroblasts were infected with vaccinia virus and labeled 2 to 3 h postinfection with [35S]methionine. Labeled proteins were fractionated on native and denatured DNA-cellulose columns and then analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Twenty-four 90,000 to 12,500, were detected. VDP-12A (molecular weight, 29,750) had affinity for denatured but not native DNA, and its synthesis was dependent on viral DNA replication. VDP-20 (molecular weight, 41,000) bound very tightly to native and denatured DNA and was displaced only after boiling the protein-DNA-cellulose matrix in 1% sodium dodecyl sulfate. VDP-8,-11,-12,-13, -and-14 behaved electrophoretically like the polypeptide species previously shown to be present in DNA-protein complexes prepared from infected cells. The molecular weights of VDP-10 (50,000), VDP-11 (36,000), and VDP-8 (67,000) were similar to the polypeptide subunits of polyadenylate polymerase and phosphohydrolase I, enzymes purified from virions which have also been shown to have affinity for DNA.
Presence of human chromosome 21 alone is sufficient for hybrid cell sensitivity to human interferon
Authors: Slate D L , Shulman L , Lawrence J B , Revel M , Ruddle F H
Published: January 2020

Human/mouse somatic cell hybrids with chromosome 21 as the only detectable human genetic material were sensitive to both human leukocyte and fibroblast interferons. The presence of additional human chromosomes decreased the amount of interferon needed to attain a given level of virus resistance. Decreased cytopathic effects, decreased virus yields, and the appearance of a specific phosphorylated protein associated with interferon treatment were all observed in hybrids maintaining only human chromosome 21. The phosphorylated protein found in extracts of these human interferon-treated hybrid cells was of mouse origin.
Anatomy of Bovine Mammillitis DNA I. Restriction Endonuclease Maps of Four Populations of Molecules That Differ in the Relative Orientation of Their Long and Short Components
Authors: Buchman Timothy G. , Roizman Bernard
Published: January 2020

In this paper, we report that the DNA of bovine mammillitis virus (BMV) consists of two covalently linked components that are 71.5 × 10
6
and 15.7 × 10
6
in molecular weight and designated L and S, respectively. We further report that the BMV DNA consists of four equimolar populations differing only in the orientation of the L and S components relative to each other. This conclusion is based on the following: (i) The sum molecular weight of fragments generated by digestion of BMV DNA with
Hsu
I,
Hpa
I,
Bgl
II, or
Xba
I significantly exceeds the established molecular weight of the intact DNA. (ii) In each digest, the fragments form three groups differing in molar concentration. In reference to the molar concentration of intact DNA, each enzyme digest contained a set of four fragments 0.25 M in concentration, a set of four fragments 0.5 M in concentration, and a variable size set, unique for each enzyme digest, 1.0 M in concentration. (iii) Experiments involving digestion of intact DNA by lambda exonuclease followed by restriction endonuclease digestion established that each of four 0.5 M fragments were positioned at the termini of the BMV DNA. (iv) Complete maps for the fragments generated by each enzyme established that the 0.25 M fragments arise by fusion of the sequences of the terminal fragments when these are juxtaposed as a consequence of the inversion of L and S components. The maps also established the dimensions of the L and S components. We conclude that the structure of BMV DNA is similar to that of HSV DNA previously shown to consist of two unequal size components that invert relative to each other.
Mutator and antimutator phenotypes of suppressed amber mutants in genes 32, 41, 44, 45, and 62 in bacteriophage T4
Authors: Watanabe S M , Goodman M F
Published: January 2020

Bacteriophage T4 genes 32, 41, 44, 45, 56, and 62 are essential to DNA replication. Amber mutants (suppressed by su+1, su+2, or su+3 bacteria) in these genes were examined for any mutator or antimutator effects on the reversion of a transition mutation. In every case except for mutations in gene 56, elevated or lowered error frequencies were observed. These results indicate the importance of all of the replicative proteins in the determination of error frequency.
Inhibitory Effect of Fatty Acids on the Entry of the Lipid-Containing Bacteriophage PR4 into
Escherichia coli
Authors: Reinhardt Albert , Cadden Stephen , Sands Jeffrey A.
Published: January 2020

Various unsaturated fatty acids (notably palmitoleic acid and oleic acid) interfered with plaque production by the lipid-containing bacteriophage PR4 on lawns of
Escherichia coli
. Addition of fatty acid to give 50 μg/ml (∼0.2 mM) at the time of infection prevented phage replication. If, however, the fatty acid was added after infection, normal amounts of phage were produced. If the fatty acid was added (to 50 μg/ml) to the host cell culture a long enough time before infection such that the fatty acid concentration in the growth medium at the time of infection was reduced to ≲5 μg/ml (due to fatty acid incorporation by the host cells), normal phage replication occurred also. Neither palmitoleic acid nor oleic acid prevented PR4 attachment to
E. coli
. Several types of experiments indicated that it is the entry process of the virus that is inhibited by these fatty acids. Specifically, if the fatty acid was added at the time of infection, the host cells were not killed by the virus and no detectable amounts of viral protein were synthesized. In addition, experiments using purified radioisotope-labeled virions showed directly that entry is inhibited. Mutants of PR4 that did replicate in the presence of oleic acid arose spontaneously at a frequency of 10
−6
. Three of these mutants that have been further characterized have protein and phospholipid compositions indistinguishable from those of wild-type PR4.
Roles of bacteriophage lambda gene products O and P during early and late phases of infection cycle
Authors: Klinkert J , Klein A
Published: January 2020

Ring-to-ring (early) replication of bacteriophage lambda DNA was blocked after heat inactivation of the P protein. Rolling circle (late) replication continued for several rounds at the rate reached when the temperature shift was carried out. The same differential effect was observed after inhibition of RNA or protein synthesis during the two different phases of replication. In contrast, inactivation of the O protein resulted in a fast stop of lambda DNA synthesis at early and late times after infection. The results were consistent with the following interpretations. (i) The lambda P gene product plays a role in the initiation of the ring-to-ring replication. (ii) Ring-to-ring replication continues parallel to rolling circle replication, possibly diminishing with time after infection. (iii) The O function is stable in and necessary for the structural integrity of an elongation complex. It is unstable in free form and probably released from such a replication complex after each round of replication at the ring-to-ring stage.
Biochemical transformation of mouse cells by herpes simplex virus type 2: enhancement by means of low-level photodynamic treatment
Authors: Verwoerd D W , Rapp F
Published: January 2020

The biochemical transformation of thymidine kinase-deficient cells by UV-inactivated herpes simplex virus is enhanced by low-level photodynamic treatment of the infected cells. At the concentration of proflavine used, the virus was not inactivated and both virus and cellular DNA syntheses were only marginally inhibited. The observed enhancement of the transfer of a virus gene to the cell genome suggests a possible cocarcinogenic role for photodynamically active dyes at very low concentrations.
Mechanism of poliovirus inactivation by ammonia
Authors: Ward R L
Published: January 2020

Poliovirus inactivation by ammonia causes a slight reduction in the sedimentation coefficients of viral particles, but has no detectable effect on either the electrophoretic pattern of viral capsid proteins or the isoelectric points of inactivated particles. These virions still attach to cells, but are unable to repress host translation or stimulate the synthesis of detectable amounts of viral RNA. Although ammonia has no detectable effect on naked poliovirus RNA, it causes cleavage of this RNA when still within viral particles. Therefore, the RNA genome appears to be the only component of poliovirus significantly affected by ammonia.
Anatomy of herpes simplex virus (HSV) DNA. X. Mapping of viral genes by analysis of polypeptides and functions specified by HSV-1 X HSV-2 recombinants
Authors: Morse L S , Pereira L , Roizman B , Schaffer P A
Published: January 2020

In an earlier paper (Morse et al., J. Virol 24:231--248, 1977) we reported on the provenance of the DNA sequences in 26 herpes simplex virus type 1 (HSV-1) X HSV-2 recombinants as determined from analyses of their DNAs with at least five restriction endonucleases. This report deals with the polypeptides specified by the recombinants and by their HSV-1 and HSV-2 parents. We have identified (i) the corresponding HSV-1 and HSV-2 polypeptides with molecular weights ranging from 20,000 to more than 200,000, (ii) the polypeptides that undergo rapid post-translational processing, and (iii) polypeptides that vary intratypically in apparent molecular weight. By comparing the segregation patterns of the polypeptides with those of the DNA sequence of the recombinants, we have mapped the templates specifying 26 polypeptides and several viral functions on the physical map of HSV DNA. The data show the following: (i) alpha polypeptides map at the termini of the L and S components of the HSV DNA. Although alpha ICP 27 maps entirely within the reiterated region of the L component, the template for alpha ICP 4 may lie only in part within the reiterated sequences of the S component. Of note is the finding that cells infected with a recombinant that contains both HSV-1 and HSV-2 DNA sequences in the S component produced alpha ICP 4 of both HSV-1 and HSV-2. (ii) Templates specifying beta and gamma polypeptides map in the L component and appear to be randomly distributed. (iii) Thymidine kinase and resistance to phosphonoacetic acid mapped in the L component. In addition, we have taken advantage of the rapid inhibition of host protein synthesis characteristic of HSV-2 infections and syncytial plaque morphology to also map the template(s) responsible for these functions in the L component. The implications of the template arrangement in HSV DNA are discussed.
Herpesvirus infection modifies adenovirus RNA metabolism in adenovirus type 5-transformed cells
Authors: Spector D , Pizer L I
Published: January 2020

The effect of herpes simplex virus (HSV) infection of mRNA metabolism was examined in a system where the fate of specific RNA sequence can be assayed. Adenovirus type 5-transformed rat embryo cell line 107 synthesizes adenovirus-specific RNA (ad-RNA), which functions in the cytoplasm as mRNA. We have utilized ad-RNA as a model for mRNA metabolism, and in a preliminiary study we characterized ad-RNA in the nucleus and cytoplasm by hybridization to filter-bound adenovirus DNA. The results indicated the as-RNA accumulates in the nucleus and that cytoplasmic polyadenylic acid [poly(A)]-containing ad-RNA turns over with a half-life of a few hours. Pulse-chase experiments confirmed these observations and a half-life of about h was determined for the poly(A)-containing cytoplasmic ad-RNA. A second class of ad-RNA remains in the nucleus, where it turns over with a longer hlaf-life (about 24 h). The infection of 107 cells by HSV was restricted at 37 degree C, giving a burst size of 5 PFU per cell and allowing continued host DNA synthesis. Protein synthesis was inhibited greater than 50% by 7 h after infection, and total RNA synthesis was 50% inhibited by 4 h after infection. During the first 8 h after infection, HSV has little effect on the rate of synthesis of ad-RNA as determined by hybridization of nuclear RNA samples, but,during the same period, HSV inhibits the accumulation of poly(A)-containing ad-RNA in the cytoplasm. The degree of this inhibition increases steadily throughout this period and reaches 60% by 6.5 to 8 h after infection. Nosignificant effect was seen on the accumulation of total cellular poly(A)-containing RNA. It was concluded from these experiments that HSV infection alters the metabolism of ad-RNA so as to prevent the normal appearance of the poly(A)-containing mRNA in the cytoplasm. The result for ad-RNA may not represent the behavior of total cellular poly(A)-containing RNA under conditions where infection is restricted.
Structure of native and chloroform-methanol-treated mycobacteriophage R1
Authors: Fay D , Bowman B U
Published: January 2020

The morphologies of native and chloroform-methanol-treated mycobacteriophage R1 were compared by electron microscopy, utilizing three negative stains. R1 was determined to be a complex phage. The head appears as an elongated cylinder with a pointed end (93 +/- 3 by 42 +/- 3 nm) constructed from an orderly arrangement of capsomeres. The phage tail measures 209 +/- 11 by 11 +/- 1 nm and possesses a striated surface with two base plates at its distal end. Treatment of R1 with chloroform-methanol resulted in disruption of both the head and tail structures and was accompanied by loss of infectivity. However, because no likely lipid-containing structure was observed in native phages, there is the possibility that the mechanism of chloroform-methanol inactivation is something other than lipid extraction.
Chromatographic separation of the polyoma virus proteins and renaturation of the isolated VP1 major capsid protein
Authors: Brady J N , Consigli R A
Published: January 2020

Treatment of purified polyoma virions with 6 M guanidine-hydrochloride and 0.01 M beta-mercaptoethanol resulted in the immediate loss of both hemagglutinating and plaque-forming ability. Gel filtration through Sepharose CL-6B beads allowed separation of the dimer, VP1, VP2, VP3, and histone proteins VP4-7 in highly purified form. Renaturation of the purified VP1 protein resulted in the formation of subunits that were morphologically, biophysically, and immunologically similar to native virion capsomeres.
Isolation of a bacterial host selective for bacteriophage T4 containing cytosine in its DNA
Authors: Runnels J , Snyder L
Published: January 2020

An Escherichia coli B strain, B834 galU56, has been isolated which supports growth of bacteriophage T4 with cytosine in its DNA while restricting growth of T4 with hydroxymethylcytosine. This host is partially deficient in uridine diphosphoglucose as determined by the ability of DNA isolated from T4 grown on it to accept glucose in an in vitro assay. In this mutant an intact rgl restriction system recognizes unglucosylated hydroxymethylcytosine residues in phage DNA, while the absence of a functional rB restriction function prevents degradation of unmodified DNA containing cytosine.
Isolation and characterization of a virus-specific ribonucleoprotein complex from reticuloendotheliosis virus-transformed chicken bone marrow cells
Authors: Wong T C , Kang C Y
Published: January 2020

Chicken bone marrow cells transformed by reticuloendotheliosis virus (REV) produce in the cytoplasm a ribonucleoprotein (RNP) complex which has a sedimentation value of approximately 80 to 100S and a density of 1.23 g/cm3. This RNP complex is not derived from the mature virion. An endogenous RNA-directed DNA polymerase activity is associated with the RNP complex. The enzyme activity was completely neutralized by anti-REV DNA polymerase antibody but not by anti-avian myeloblastosis virus DNA polymerase antibody. The DNA product from the endogenous RNA-directed DNA polymerase reaction of the RNP complex hybridized to REV RNA but not to avian leukosis virus RNA. The RNA extracted from the RNP hybridized only to REV-specific complementary DNA synthesized from an endogenous DNA polymerase reaction of purified REV. The size of the RNA in the RNP is 30 to 35S, which represents the subunit size of the genomic RNA. No 60S mature genomic RNA was found within the RNP complex. The significance of finding the endogenous DNA polymerase activity in the viral RNP in infected cells and the maturation process of 60S virion RNA of REV are discussed.
Absence of Detectable Capping and Methylating Enzymes in Influenza Virions
Authors: Plotch Stephen J. , Tomasz Jeno , Krug Robert M.
Published: January 2020

In the presence of Mg
2+
and a specific dinucleotide primer (ApG or GpG), the influenza virion transcriptase synthesizes the eight discrete segments of complementary RNA (cRNA) containing polyadenylic acid (Plotch and Krug, J. Virol.
21
:24-34, 1977). Virions were examined for their ability to cap and methylate cRNA containing di- or triphosphorylated 5′ termini. By using the primers ppApG, pppApG, or ppGpG, viral cRNA was synthesized in vitro with [α-
32
P]-GTP and
S
-[
methyl
-
3
H]adenosylmethionine as labeled precursors. DEAE-Sephadex chromatography of the RNase T2 digest of the cRNA product demonstrated no
3
H incorporation at all and the absence of a
32
P-labeled cap structure. The 5′ terminus of ppApG-primed cRNA could be capped and methylated by enzymes from vaccinia virus, indicating that the two 5′-terminal phosphates derived from the primer were preserved in the product cRNA. The cap structure formed by the vaccinia enzymes and released by RNase T2 digestion as m
7
GpppA
m
pGp was radioactively labeled at its 3′-terminal phosphate only when [α-
32
P]CTP was used as the labeled precursor during transcription. This indicates that the 5′-terminal sequence of the cRNA is ppApGpC and that, therefore, ppApG most probably initiates transcription exactly at the 3′ GpCpU
OH
terminus of the virion RNA templates. Virions were also tested for their ability to cap and methylate ppApG in the absence of transcription. No such activities were detected, whereas under the same conditions the vaccinia virus enzymes successfully capped and methylated this compound. Consequently, these experiments, together with those reported earlier, have not detected in influenza virions any capping and methylating enzymes active on the 5′-initiated termini of viral cRNA chains synthesized in vitro, whether these termini possess one, two, or three phosphates. Some mechanism for capping and methylation of viral cRNA must, however, exist, because the viral mRNA (cRNA) synthesized in the infected cell contains 5′-terminal methylated cap structures (Krug et al., J. Virol.
20
:45-53, 1976). Possible mechanisms are discussed.
Restriction fragment analysis of bacteriophage SPP1 in vitro transcription by host RNA polymerase
Authors: Chenciner N , Milanesi G
Published: January 2020

In vitro transcription of SPP1 DNA occurred on only one of the two strands, the same which is predominantly transcribed in SPP1-infected cells. Transcripts were distributed in several size classes. Analysis of elongation kinetics and of size distribution, coupled with hybridization to DNA restriction fragments, showed that some regions of the template have more initiation sites than others; some have none. Some regions were transcribed directly, some were transcribed from initiation sites located in other regions, and one was never transcribed. Several transcription initiation sites on SPP1 DNA are located on EcoRI fragment 1; four to five others are distributed among other fragments. Cutting the DNA with EcoRI did not introduce artifactual initiation sites. In vitro transcription units can be localized and oriented with respect to the EcoRI restriction map of SPP1 DNA.
Detergent-inhibited, heat-labile nucleoside triphosphatase in cores of avian myeloblastosis virus
Authors: Jensen K F
Published: January 2020

Endogenous DNA synthesis was studied in isolated core particles of avian myeloblastosis virus. It was found that cores contained an enzymatic activity which rapidly converted the added nucleoside triphosphates to diphosphates (but not further) at 0 degrees C, thus inhibiting DNA synthesis. This triphosphatase probably originates from the viral membranes. In the cores the enzyme is completely inactivated by low concentrations (0.02%) of Nonident P-40. Also, the enzyme is very thermolabile and denatures rapidly at 38 degrees C.
Transcriptional mapping of rabies virus in vivo
Authors: Flamand A , Delagneau J F
Published: January 2020

Synthesis of the proteins of rabies virus was studied in hamster cells infected with UV-irradiated virus. The UV target size of genes L, N, M1, and M2 was measured during primary transcription. Except for N, the target size of the remaining genes was considerably larger than that of their physical sizes. The data fit the hypothesis that four genes occupy a single transcriptional unit and that transcription of rabies virus proceeds in the order N, M1, M2, and L.
Identification of nucleotide sequences which may encode the oncogenic capacity of avian retrovirus MC29
Authors: Sheiness D , Fanshier L , Bishop J M
Published: January 2020

The retrovirus strain MC29 induces a variety of tumors in chickens, including myelocytomatosis and carcinomas of the kidney and liver. In addition, the virus can transform cultures of embryonic avian macrophages and fibroblasts. We have characterized the genome of MC29 virus and have identified nucleotide sequences that may encode the oncogenic potential ofthe virus. MC29 virus can replicate only with the assistance of a related helper virus. The defect in replication is apparently a consequence of a deletion in one or more viral genes: the haploid genome of the MC29 virus has a molecular weight of ca. 1.7 X 10(6), whereas the genome of the helper virus MCAV has a molecular weight of ca. 3.1 X 10(6). Although MC29 virus transforms fibroblasts in culture, its genome has no detectable homology with the gene src that is responsible for transformation of fibroblasts by avian sarcoma viruses. We prepared radioactive single-stranded DNA complementary to nucleotide sequences present in the genome of MC29 virus but not in the genome of MCAV (cDNA(MC29)). If they are contiguous, these sequences (ca. 1,500 nucleotides) are sufficiently complex to encode at least one protein. Homologous sequences were not detectable in several strains of avian sarcoma viruses or in an endogenous virus of chickens. Our findings confirm and extend recent reports from other laboratories and lead to the conclusion that MC29 virus may contain a previously unidentified gene(s) that is capable of transforming several distinct target cells. The evolutionary origins of this putative gene and its location on the viral genome can be explored with cDNA(MC29).
Physical mapping of herpes simplex virus-induced polypeptides
Authors: Marsden H S , Stow N D , Preston V G , Timbury M C , Wilkie N M
Published: January 2020

Analysis of the polypeptides induced by 29 herpes simplex virus type 1/type 2 intertypic recombinants and correlation of the data with the crossover points in the recombinant DNAs have enabled the map positions of many polypeptides to be deduced. These include 25 polypeptides which label with [35S]methionine, 11 which label with [32P]orthophosphate, and 4 which label with [14C]glucosamine. Together with the data of Preston et al. (J. Virol., in press) on the mapping of five immediate-early polypeptides, the results show that representatives of four groups of proteins--immediate-early, late, phosphorylated, and glycosylated--map in both long and short regions. The functional organization of the herpes simplex virus genome does not therefore restrict any of these four groups to either the long or the short region.
Model for DNA packaging into bacteriophage T4 heads
Authors: Black L W , Silverman D J
Published: January 2020

The mechanism of DNA packaging into bacteriophage T4 heads in vivo was investigated by glucosylation of hydroxymethylcytosine residues in a conditionally glucose-deficient host. Cytoplasmic DNA associated with partially packaged ts49 heads can be fully glucosylated, whereas DNA already packaged into these heads is shown to be resistant to glucosylation. After temperature shift and completion of arrested packaging into the reversible temperature-sensitive ts49 head, the structure of the DNA in the mature ts49 phage was investigated by restriction enzyme digestion, autoradiography, and other techniques. Such mature DNA appears to be fully glucosylated along part of its length and nonglucosylated on the remainder. Its structure suggests that the DNA is run into the head linearly and unidirectionally from one mature end and that there is little sequence specificity in that portion of the T4 DNA which first enters the capsid. This technique should be useful in investigation of the three-dimensional structure of first- and last-packaged DNA within the head; preliminary studies including autoradiography of osmotically shocked phage suggest that the DNA which first enters the head is deposited toward the center of the capsid and that the end of the DNA which first enters the head exits first upon injection. In conjunction with studies of the structure of condensed DNA, the positions and functions of T4 capsid proteins in DNA packaging, and the order of T4 packaging functions [Earnshaw and Harrison, Nature (London) 268:598-602, 1977; Hsiao and Black, Proc. Natl. Acad. Sci. U.S.A. 74:3652-3656, 1977; Müller-Salamin et al., J. Virol. 24:121-134, 1977; Richards et al., J. Mol. Biol. 78:255-259, 1973], the features described above suggest the following model: the first DNA end is fixed to the proximal apex of the head at p20 and the DNA is then pumped into the head enzymatically by proteins (p20 + p17) which induce torsion in the DNA molecule.
Chromatin pattern consisting of repeating bipartite structures in WI-38 cells infected with human cytomegalovirus
Authors: Kierszenbaum A L , Huang E S
Published: January 2020

A novel chromatin structural pattern displaying bipartite and oblate ellipsoid structures orderly arranged along a fiber axis has been observed in WI-38 cells infected with human cytomegalovirus. This chromatin type coexists with chromatin fibers showing conventional nucleosomes. Each bipartite-oblate structure is 40 nm in length, about four times as long as an ordinary nucleosome, and the number of these structures per micrometer (11/micrometer) is clearly less than that of typical cellular nucleosomes (32/micrometer).
Bacteriophage lambda mutants (lambdatp) that overproduce repressor
Authors: Truitt C L , Chu H , Walker J R
Published: January 2020

Lambda tp mutants, selected for their ability to form turbid plaques on lon hosts, overproduce repressor. The tp1 and tp2 mutations have been located within (or adjacent to) the cIII gene. The tp1 mutation reduced late gene expression, as measured by endolysin synthesis (in the absence of functional cI repressor) and progeny phage yield. The tp4 mutation was mapped in the cY-cII region, and complementation tests indicated that tp4 affects the diffusible product of the cII gene. The tp4 mutation also reduced progeny production, but did not markedly affect endolysin synthesis.
Cell-free translation of purified virion-associated high-molecular-weight RNA synthesized in vitro by vaccinia virus
Authors: Bossart W , Paoletti E , Nuss D L
Published: January 2020

Virion-associated high-molecular-weight (HMW) RNA synthesized in vitro by purified vaccinia virus particles has been translated in a wheat germ cell-free protein synthesizing system. Purified HMW RNA directs the synthesis of translation products which are identical to the translation products made in response to in vitro-synthesized, virion-released 8 to 12S mRNA. The translation of HMW RNA proceeds exclusively through a 5'-terminal cap-mediated initiation step. Furthermore, only one coding sequence is translated per HMW RNA molecule, and that sequence is probably located near the 5' end of the molecule. These conclusions are based on the following results. (i) Sodium dodecyl sulfate--polyacrylamide gel electrophoresis patterns of translation products synthesized in response to HMW RNA and in response to 8 to 12S mRNA were qualitatively identical. (ii) On an equal weight basis, HMW RNA was 25 to 30% as active as 8 to 12S mRNA in stimulating in vitro protein synthesis. (iii) Unmethylated HMW RNA was translated at 10% the efficiency of the methylated form of this RNA. (iv) m7pG inhibited the translation of fully methylated HMW RNA by 90%. (v) After the initiation step of translation was blocked by aurintricarboxylic acid, the rate with which amino acids were incorporated into individual polypeptides decreased in a similar manner for the translation of both HMW RNA and 8 to 12S mRNA. Virion-released 8 to 12S mRNA derived from virion-associated HMW RNA during a chase in the presence of ATP, GTP, and S-adenosylmethionine was also translated. At low RNA concentrations, the derived RNA appeared to stimulate amino acid incorporation more efficiently than the HMW RNA precursor. However, at higher concentrations of this RNA, protein synthesis was severely inhibited.
Membrane fusion as a mechanism of simian virus 40 entry into different cellular compartments
Authors: Maul G G , Rovera G , Vorbrodt A , Abramczuk J
Published: January 2020

Permissive and nonpermissive simian virus 40 (SV40)-infected cells were ultrastructurally analyzed. Viral particles were found in the cytoplasm, rough endoplasmic reticulum, nuclear envelope, lysosomes, and mitochondria. Upon entering the cell the virion obtains a tight membrane envelope. It seems to be either released from the envelope upon fusion with other membranes of the cell or aggregated into tubular membrane specializations upon fusion with other membrane-enveloped particles. Reconstructed morphological sequences and the finding of SV40 in different spaces of the cell suggest that entry of SV40 into the different compartments and eventually into the site of replication is facilitated by its capacity for being enveloped by a variety of membranes (notably the cell membrane and the nuclear membrane) and the sequential fusion and fission of these membranes.
Comparison of the expression of the src gene of Rous sarcoma virus in vitro and in vivo
Authors: Sefton B M , Beemon K , Hunter T
Published: January 2020

We have compared the polypeptide products of the src gene of several strains of Rous sarcoma virus produced by in vitro translation of heat-denatured 70S virion RNA in the nuclease-treated reticulocyte lysate with those present in chick cells transformed by these viruses. We have done this by immunoprecipitation, using sera from rabbits injected at birth with Schmidt-Ruppin Rous sarcoma virus. In vitro translation results in the synthesis of at least nine polypeptides which appear to be encoded by the src gene. These range in size from 17,000 to 60,000 daltons. The sera from tumor-bearing rabbits precipitated these polypeptides arising from the in vitro translation of RNA from Schmidt-Ruppin Rous sarcoma virus of both subgroup A and subgroup D and from one stock of Prague Rous sarcoma virus of subgroup C. In each case, all of this family of related polypeptides could be precipitated except the smallest, the 17,000-dalton polypeptide. No precipitation of analogous polypeptides resulting from the translation of RNA from other strains of Rous sarcoma virus was observed. Cells transformed by these three strains of Rous sarcoma virus contain easily detectable amounts of a polypeptide, p60src, essentially identical to the 60,000-dalton in vitro product. With one exception, they do not contain significant amounts of polypeptides analogous to the smaller in vitro products which can be precipitated by these sera. Cells transformed by one stock of Schmidt-Ruppin Rous sarcoma virus of subgroup A did contain a 39,000-dalton polypeptide, which was related, by peptide mapping, to the 60,000-dalton polypeptide and was similar in size to a precipitable in vitro product. The 60,000-dalton polypeptide present in transformed cells appeared to be phosphorylated 10 to 25 min after its synthesis, metabolically very stable, and not derived from a precursor polypeptide. All immunoprecipitates from transformed cells which contained p60src also contained an 80,000-dalton phosphoprotein. This polypeptide is unrelated to p60src, as determined by peptide mapping, and may well be a host cell polypeptide which is specifically associated with p60src.