bluuebunny/crossref_metadata_2025 huggingface dataset:
Some biological properties of two new avian sarcoma viruses
Authors: Balduzzi P C , Notter M F , Morgan H R , Shibuya M
Published: January 2020

The new avian retroviruses UR1 and UR2 were isolated from spontaneous tumors of chickens by cocultivation of tumor material with susceptible chicken embryo fibroblasts. In vitro, UR1 induced formation of small foci of round and fusiform cells. On the other hand, cells infected by UR2 assumed an extremely elongated morphology. In vivo, both viruses induced fibrosarcomas and myxosarcomas with short latencies. Infectivity assays with and without mitomycin C showed that both viruses were defective for replication, but transformed nonproducing cell clones were obtained only with UR1. UR1-infected transformed nonproducing clones did not release particles detectable by reverse transcriptase assays, and fusion of transformed nonproducing cells with quail cells chronically infected with Rous sarcoma virus (a Bryan strain) failed to rescue infectious virus. This suggested that UR1 does not code for functional envelope glycoproteins. In this regard, UR1 appeared to be similar to Fujinami, PRCII, and Y73 viruses. The helper viruses of partially purified stocks of UR1 and UR2 appeared to belong to subgroup A, but these helper viruses were distinguishable from each other, as shown by host range experiments and neutralization tests. Hybridization studies with DNA complementary to the src gene of Rous sarcoma virus and RNAs extracted from both UR1 and UR2 showed no homology between the genomes of the new isolates and the transforming gene of Rous sarcoma virus.
Immunological characterization of a low oncogenic mouse mammary tumor virus BALB/cNIV mice
Authors: Vacquier J P , Cardiff R D , Blair P B
Published: January 2020

Antigenic determinants of mouse mammary tumor virus (MuMTV) from the low-mammary-tumor-incidence strain BALB/cNIV were compared by competition radioimmunoassay with those of MuMTV's isolated from several high- and low-mammary-tumor-incidence mouse strains, using rabbit hyperimmune sera against BALB/cNIV MuMTV and against MuMTV from the high-mammary-tumor-incidence strain BALB/cfC3H. Using anti-BALB/cfC3H serum in competition radioimmunoassay, BALB/cNIV MuMTV lacked antigenic determinants present on MuMTV's from the BALB/cfC3H, C3H, and GR strains. With anti-BALB/cNIV serum in competition radioimmunoassay, type-specific antigenicity was detected with BALB/cNIV MuMTV. We found class-specific antigenicities on BALB/cNIV MuMTV that were shared with RIII and C3Hf MuMTV's BALB/cNIV MuMTV was reportedly derived from the C3Hf strain by infection of BALB/c mouse mammary tissue after transplantation into a (BALB/c X C3Hf)F1 hybrid followed by retransplantation into the BALB/c strain (D. R. Pitelka, K. B. DeOme, and H. A. Bern. Proc. Am Assoc. Cancer Res. 6:51, 1965). However, BALB/cNIV MuMTV contained type-specific antigens not present on C3Hf MuMTV. The possible origin of these determinants is discussed.
Quantitative of murine mammary tumor virus-related RNA in mammary tissues of low- and high-mammary-tumor-incidence mouse strains
Authors: Marcus S L , Smith S W , Sarkar N H
Published: January 2020

Lactating mammary glands and hormonally induced mammary tumors of BALB/c mice from three geographically separated breeding colonies were examined by molecular hybridization, using murine mammary tumor virus (MuMTV) cDNA representing the entire viral genome to determine the amount of MuMTV-related RNA expressed in these tissues. The RNA extracted from these tissues by the classical sodium dodecyl sulfate-pronase, phenol-chloroform procedure (method 1) contained barely detectable levels of MuMTV-related sequences. In contrast, both normal lactating mammary glands and hormonally induced mammary tumors of these mice were found to contain approximately one to two copies of the MuMTV genome per cell by using a new procedure in which the RNA was extracted with guanidine derivatives (method 2). No significant differences in the MuMTV-related RNA content of the BALB/c mammary tissues were observed regardless of their colony of origin. Our results suggest that expression of MuMTV RNA does not change in BALB/c mammary glands during transformation to a malignant state and that MuMTV expression does not play a role in tumorigenesis in these mice. In view of the increased recovery of MuMTV-related RNA from BALB/c mice with method 2, we compared the level of MuMTV RNA expression in lactating mammary glands and mammary tumors of other mouse strains, including C57BL/6 and RIII, using both extraction methods. Yields of MuMTV-related RNA from mammary tissues increased by as much as 35- to 40-fold, using method 2 as compared with method 1. Therefore method 2, involving guanidine derivatives, appears to be method of choice for MuMTV-related RNA extraction from the mammary tissues of certain strains of mice, particularly those expressing relatively low levels of MuMTV RNA.
Viral protein synthesis in mouse hepatitis virus strain A59-infected cells: effect of tunicamycin
Authors: Rottier P J , Horzinek M C , van der Zeijst B A
Published: January 2020

We identified eight protein species in virions of mouse hepatitis virus strain A59. Based on their sizes, prosthetic groups, and locations in virions, these proteins were designated gp180/E2, gp90/E2, pp54/N, gp26.5/E1, gp25.5/E1, p24/E1, p22/X, and p14.5/Y. The positions of the last two proteins in virions are not known. Host protein synthesis in Sac(-) cells infected with mouse hepatitis virus strain A59 was inhibited, and the following novel proteins appeared: gp150, gp90, p54, gp26.5, gp25.5, p24, p22, and p14.5. Except for gp150, these polypeptides all co-electrophoresed with mouse hepatitis virus strain A59 structural proteins. In addition, all of these proteins could be immunoprecipitated with a convalescent mouse serum or a rabbit antiserum raised against purified disrupted virus. After a 15-min pulse of infected cells with radioactive amino acids at 7h postinfection, gp90 was not detected, whereas gp26.5 and gp25.5 were only labeled to a small extent. During a subsequent chase period gp150 was processed to gp90, whereas the radioactivity in gp26.5 and gp25.5 increased concomitantly with a reduction of label in p24. Tunicamycin, an antibiotic which inhibits the synthesis of glycopeptides bearing N glycosidically linked oligosaccharides, prevented the appearance of gp150 in mouse hepatitis virus strain A59-infected cells. Instead, a 110,000-dalton protein accumulated. In contrast, the syntheses of the smaller viral glycoproteins gp26.5 and gp25.5 were resistant to this drug, indicating that these glycosylations were of the O glycosidical type. Although the production of infectious virus in tunicamycin-treated cells was inhibited by more than 99%, release of noninfectious viral particles continued. An analysis of these particles revealed that they lacked the peplomeric glycoproteins gp90/E2 and gp180/E2. Obviously, although the surface projections were not essential for budding of virus particles from the cells, they were required for infectivity.
Genomic arrangement of an adenovirus-simian virus 40 hybrid virus, Ad2+ND4del
Authors: Westphal H
Published: January 2020

Ad2+ND4del is an adenovirus type 2-simian virus 40 hybrid virus nondefective for growth in human cells. The virus was first observed when stocks of Ad2+ND4, a hybrid isolated from primary monkey kidney cells, were propagated in human cells. This paper describes the DNA sequence at two sites of DNA recombination, the site of the left adenovirus type 2-simian virus 40 junction and the site of a deletion of internal simian virus 40 sequences. Since the deletion was observed when the virus was switched from monkey to human cells, an analysis of gene expression in the region of DNA rearrangement may prove useful for the elucidation of molecular events that accompany virus growth in different hosts.
Genetic mapping of endogenous mouse mammary tumor viruses: locus characterization, segregation, and chromosomal distribution
Authors: Traina V L , Taylor B A , Cohen J C
Published: January 2020

The restriction endonuclease EcoRI has been used to study the inheritance of strain difference in endogenous mouse mammary tumor virus DNA sequences. This enzyme, which cleaves at only one site within the nondefective viral genome, generates DNA fragments containing mouse mammary tumor virus sequences which vary in size according to the locations of EcoRI restriction sites in the flanking mouse sequences, thereby defining unique integration sites of the viral genome. Recombinant inbred strains of mice have been used to study the inheritance of these DNA fragments which hybridize to mouse mammary tumor virus cDNA sequences. The results define 11 segregating units consisting of 1 or 2 fragments. These units were shown to segregate among the recombinant inbred strains, and in some instances linkage was established. Two units were shown to be linked on chromosome 1. Another unit was mapped to chromosome 7, which is presumably identical to the previously defined genetic locus Mtv- 1. One other mouse mammary tumor virus locus was tentatively assigned to chromosome 6. The results are consistent with the view that integration of mouse mammary tumor virus can take place at numerous sites within the genome, and once inserted, these proviruses appear to be relatively stable genetic entities.
Gene products of McDonough feline sarcoma virus have an in vitro-associated protein kinase that phosphorylates tyrosine residues: lack of detection of this enzymatic activity in vivo
Authors: Barbacid M , Lauver A V
Published: January 2020

The primary translational product of the McDonough (SM) strain of feline sarcoma virus (FeSV) is a 180,000-dalton molecule, SM P180, that contains the p15-p12-p30 region of the FeLV gag gene-coded precursor protein and a sarcoma virus-specific polypeptide. In addition, cells transformed by SM-FeSV express a 120,000-dalton molecule, SM P120, that is highly related to the non-helper virus domain of SM P180. Both SM-FeSV gene products were found to be intimately associated with the membrane fraction of SM-FeSV-transformed cells. Immunoprecipitates containing SM P180 and SM P120 exhibited a protein kinase activity capable of phosphorylating tyrosine residues of both viral gene products but not immune immunoglobulin G molecules. By independently immunoprecipitating each of the two SM-FeSV proteins we found that most of the tyrosine-specific phosphorylating activity was associated with the SM P120 molecule. In vivo analysis of 32P-labeled SM P180 and SM P120 revealed their phosphoprotein nature; however, both molecules exhibited low levels of phosphorylation and did not contain phosphotyrosine residues. Finally, we did not detect any significant elevation in the levels of phosphotyrosine in the protein fraction of SM-FeSV transformants. Thus, if SM-FeSV were to induce malignant transformation by a mechanism involving phosphorylation of tyrosine residues, the viral gene products must interact with a small subset of cellular proteins that do not represent a significant fraction of the total cellular protein content.
Suppression of Vesicular Stomatitis Virus Defective Intefering Particle Generation by a Function(s) Associated with Human Chromosome 16
Authors: Kang C. Yong , Weide Lamont G. , Tischfield Jay A.
Published: January 2020

Human-mouse somatic cell hybrids were made between adenine phosphoribosyltransferase-deficient mouse L cells and a strain of human primary fibroblasts and selected in medium containing alanosine and adenine (J. A. Tischfield and F. H. Ruddle, Proc. Natl. Acad. Sci. U.S.A.
71
:45-49, 1974). These hybrids were tested for the generation of defective interfering (DI) particles of vesicular stomatitis virus to determine whether or not a host gene controls the induction of DI particles. None of the seven independently arising hybrid clones tested generated detectable DI particles during 13 successive undiluted passages. In addition, the parental human cells also failed to generate DI particles. In contrast, the parental mouse cells generated a detectable level of DI particles during continuous passage. Thus, failure to generate DI particles appears to act in a dominant fashion in these hybrids. Human chromosome 16 and adenine phosphoribosyltransferase were present, as a direct consequence of the selection system, in all of the hybrid clones that failed to generate DI particles. It was the only human chromosome observed in the cells of every hybrid clone. This was verified by both isozyme and karyotype analyses. After hybrids were back-selected (with 2,6-diaminopurine) for loss of human adenine phosphoribosyltransferase and chromosome 16, they gained the ability to generate DI particles. Replication of DI particles already present in virus stocks, however, was normal in all of the hybrid clones and the parental human cells. This suggests that the induction, but not the replication, of DI particles is affected by the human genome and that a factor on human chromosome 16 seems to selectively suppress the mouse cell's ability to generate DI particles in the hybrids. These results support the idea that the induction of DI particles is controlled in part by host cell function(s), as suggested previously (C. Y. Kang and R. Allen, J. Virol.
25
:202-206, 1978).
Production and characterization of monoclonal antibodies against the Epstein-Barr virus membrane antigen
Authors: Strnad B C , Schuster T , Klein R , Hopkins R F , Witmer T , Neubauer R H , Rabin H
Published: January 2020

Five murine hybridoma lines that produce monoclonal antibodies against Epstein-Barr virus membrane antigen (MA) were established. Immunoprecipitation experiments demonstrated that three of the antibodies precipitated both the 236,000 (236K) MA and the 212K MA. The other two antibodies precipitated the 86K MA. Antibodies against the 236K-212K MA and the 86K MA mediated complement-dependent cytolysis of Epstein-Barr-virus-infected cells. The antibodies against the 86K MA neutralized both the B95-8 and P3HR-1 viruses.
Transformation of hamster kidney cells by fragments of BK virus DNA
Authors: Grossi M P , Corallini A , Valieri A , Balboni P G , Poli F , Caputo A , Milanesi G , Barbanti-Brodano G
Published: January 2020

Hamster kidney cells were transformed, with comparable efficiency, by circular or linear molecules of complete BK virus (BKV) genome and by agarose gel-purified fragments of BKV DNA obtained by single or double digestions with various restriction endonucleases. Only fragments containing the complete early region of BKV DNA displayed transforming activity. Analysis by blot hybridization of the arrangement of viral DNA sequences in a cloned cell line transformed by a 3.8-kilobase fragment, obtained after sequential digestion of BKV DNA with HhaI and BamHI, showed the presence of seven viral integrations into the cellular DNA. Apparently all of the integrated viral molecules contained the entire early region of BKV DNA. Large T antigen, small t antigen, and the 56,000-dalton nonviral Tau antigen were detected in transformed cells by immunoprecipitation. The pattern of integration of viral sequences in transformed cells was constant over many generations. Likewise, large T antigen was always detected in transformed cells at various passage levels. These results may suggest that all of the sequences of the early region coding for large T antigen are required for transformation by BKV. Alternatively, subgenomic segments of the BKV DNA early region may be unable to transform because the appropriate polyadenylation site, necessary to obtain a complete functional transcriptional unit, is removed by the restriction enzyme cleavage.
Effect of glycosylation inhibitors on the release of enveloped vaccinia virus
Authors: Payne L G , Kristensson K
Published: January 2020

Addition of 1 to 10 mM 2-deoxy-D-glucose (2-dg) or glucosamine (gln) to the growth medium of vaccinia virus-infected cells inhibited the release of extracellular enveloped vaccinia virus (EEV) without affecting the production of intracellular naked vaccinia virus (INV) particles. In contrast, INV infectivity (particles per PFU) was decreased sevenfold by 50 mM 2-dg. Treatment with 2-dg reduced but did not eliminate glycosylation of the INV 37,000-molecular-weight glycoprotein. The kinetics of sensitivity to inhibitor addition experiments and inhibitor reversal experiments indicated that EEV release was dependent on glycosylation before 8 h postinfection. This was supported by polyacrylamide gel electrophoretic analysis of the synthesis kinetics for cell membrane-associated vaccinia glycoproteins in 2-dg-inhibited infected cells. The dependence of vaccinia protein glycosylation before 8 h postinfection for efficient EEV release was observed in spite of the fact that the period of greatest glycoprotein synthesis was 8 to 12 h postinfection. The presence of 2-dg resulted in an incompletely glycosylated 89,000-molecular-weight glycoprotein, as indicated by a reduction in the apparent glycoprotein molecular weight. The morphological event affected by the inhibitors was the acquisition by INV of a double-membrane structure from the Golgi apparatus. This morphological intermediate is necessary for release of EEV.
Characterization of the major Epstein-Barr virus-specific RNA in Burkitt lymphoma-derived cells
Authors: Arrand J R , Rymo L
Published: January 2020

Cytoplasmic RNA prepared from five lymphoid cell lines and a Burkitt lymphoma biopsy was radioactively labeled in vitro and hybridized to cloned EcoRI restriction endonuclease fragments of B95-8 Epstein-Barr virus DNA. The results confirmed that the most abundant cytoplasmic RNA species in such cells is specified by a small region of the genome defined by the EcoRI J fragment. Detailed mapping experiments precisely localized these transcripts within the sequence of the rightmost one-third of the EcoRI J fragment. DNA sequencing suggested that this region of the Epstein-Barr virus genome is unable to code for protein. The major early transcripts consisted of two non-polyadenylated RNA species, each about 170 nucleotides in length. They were both transcribed off the same strand of the DNA and showed significant sequence homology with each other. The coding sequences of the two small RNAs contained potential intragenic control regions for RNA polymerase III.
Temporal Patterns of Human Cytomegalovirus Transcription: Mapping the Viral RNAs Synthesized at Immediate Early, Early, and Late Times After Infection
Authors: Wathen Michael W. , Stinski Mark F.
Published: January 2020

The transcription of the human cytomegalovirus genome was investigated at immediate early, early, and late times after infection. Viral RNAs associated with either the whole cell, the nucleus, the cytoplasm, or the polyribosomes were analyzed. At immediate early times, i.e., in the absence of de novo viral protein synthesis, the viral RNA in high abundance originated from a region of the long unique section of the prototype arrangement of the viral genome (0.660 to 0.770 map units). The viral RNA in low abundance originated from the long repeat sequences (0.010 to 0.035 and 0.795 to 0.825 map units) and a region in the long unique section (0.201 to 0.260 map units). Viral RNAs associated with the polyribosomes as polyadenylated RNA were mapped to these restricted regions of the viral genome and characterized according to size class in kilobases. At 24 h after infection in the presence of an inhibitor of viral DNA replication, i.e., at early times, the stable viral RNAs in highest abundance mapped in the long repeat sequences. Viral RNAs at intermediate abundance under these conditions mapped in two regions of the long unique section of the viral genome (0.325 to 0.460 and 0.685 to 0.770 map units). Stable viral RNAs that were associated with the polyribosomes in high abundance as polyadenylated RNA orginated from the long repeat sequences, but not from the long unique section of the viral genome. An analysis of whole-cell RNA at late times (72 h) indicated that the abundant transcription was in the regions of the long unique sequences (0.325 to 0.460 and 0.660 to 0.685 map units), and transcription of intermediate abundance was from the long repeat sequences. However, stable viral mRNA's derived from the long repeat sequences were associated with the polyribosomes at late times after infection. In addition, mRNA's originating from the long and short unique sequences were found associated with the polyribosomes at higher relative concentration than at early times after infection. It is proposed that expression of the immediate early viral genes is required to transcribe the early viral genes in the long repeat and adjacent sequences. These sequences are also transcribed at late times after infection while viral DNA synthesis continues. The expression of viral genes in most of the long and short unique sequences appears to require viral DNA replication.
Isolation of vesicular stomatitis virus defective interfering genomes with different amounts of 5'-terminal complementarity
Authors: Kolakofsky D
Published: January 2020

I isolated at least 30 different vesicular stomatitis virus defective interfering (DI) genomes, distinguished by chain length, by five independent undiluted passages of a repeatedly cloned virus plaque. Labeling of the 3' hydroxyl ends of these DI genomes and RNase digestion studies demonstrated that the ends of these DI genomes were terminally complementary to different extents (approximately 46 to 200 nucleotides). Mapping studies showed that the complementary ends of all of the DI genomes were derived from the 5' ends of the nondefective minus-strand genome. Regardless of the extent of terminal complementarity, all of the DI genomes synthesized the same 46-nucleotide minus-strand leader RNA.
Identification of virus-coded nonstructural polypeptides in bunyavirus-infected cells
Authors: Fuller F , Bishop D H
Published: January 2020

Analyses of bunyavirus-infected cell extracts identified at least two virus-induced nonstructural polypeptides. With snowshoe hare (SSH), La Crosse (LAC), and six SSH-LAC reassortant viruses, it was shown that one of these nonstructural polypeptides (NSs, approximate molecular weight, 7.4 X 10(3)) is coded by the SSH small (S)-size viral RNA species. This nonstructural polypeptide was not detected (at least in the same relative abundancies) in LAC virus-infected cells or in cells infected with reassortants having LAC S RNA. For SSH virus, tryptic peptide analyses of either [3H]leucine- or [3H]arginine-labeled NSs indicated that it contains unique sequences not present in the SSH nucleocapsid (N) polypeptide (also coded by the S RNA; J. R. Gentsch and D. H. L. Bishop, J. Virol. 28:417-419, 1978). Analyses of SSH virus-infected cell extracts and extracts of cells infected with SSH-LAC reassortants having SSH medium (M)-size RNA species indicated that a nonstructural polypeptide (NSM; approximate molecular weight, 12 X 10(3)) is coded by the SSH M RNA species. In extracts of LAC virus-infected cells (or cells infected with SSH-LAC reassortants having LAC M RNA), a polypeptide with an electrophoretic mobility slightly faster than that of the SSH NSM polypeptide was observed (approximate molecular weight, 11 X 10(3)); it has been designated LAC NSM. The relationships of the NSM polypeptides to the other M RNA-coded polypeptides (G1 and G2; J. R. Gentsch and D. H. L. Bishop, J. Virol. 30;767-770, 1979) have not been determined. Two additional polypeptides present in both LAC- and SSH-infected cell extracts also appear to be virus induced (one with an approximate molecular weight of 10 X 10(3), p10; the other with an approximate molecular weight of 18 X 10(3), p18). Whether these polypeptides are virus coded has not been determined.
Identification of the polyprotein termination site on encephalomyocarditis viral RNA
Authors: Drake N L , Palmenberg A C , Ghosh A , Omilianowski D R , Kaesberg P
Published: January 2020

We show by sequence analysis of a 420-base-long region adjacent to the 3' polyadenylic acid of encephalomyocarditis viral RNA and by carboxy terminus analysis of protein E that the termination site of encephalomyocarditis virus polyprotein translation consists of two successive UAG codons located at positions 121 to 126 from the 3' polyadenylic acid.
Antibodies Against the Genome-Linked Protein VPg of Cowpea Mosaic Virus Recognize a 60,000-Dalton Precursor Polypeptide
Authors: Zabel Pim , Moerman Marja , van Straaten Flip , Goldbach Rob , van Kammen Albert
Published: January 2020

We have prepared a rabbit antiserum specifically directed against the genome-linked protein (VPg) of cowpea mosaic virus by injecting an hydrolysate of purified virion RNA. Using this antiserum as a probe in combination with “Western” (protein) blots of subcellular fractions of cowpea mosaic virus-infected cowpea (
Vigna unguiculata
) cells, we have detected a bottom component RNA-encoded, 60,000-dalton polypeptide which is membrane bound and presumably represents the immediate precursor of VPg.
Isolation and biochemical characterization of partially transformation-defective mutants of avian myelocytomatosis virus strain MC29: localization of the mutation to the myc domain of the 110,000-dalton gag-myc polyprotein
Authors: Ramsay G M , Hayman M J
Published: January 2020

Recently, we isolated three mutants of MC29 virus which, although able to transform fibroblasts with the same efficiency as wild-type MC29, were 100-fold less efficient at transforming macrophages. In this study we found that MC29-transformed quail producer cell line Q10 was able to generate these partially transformation defective mutants at a high frequency. Using tryptic peptide mapping, we determined that the smaller gag-myc polyproteins encoded by the transformation-defective viruses had lost myc-specific tryptic peptides. This suggested that the mutations which resulted in the transformation-defective viruses being inefficient at transforming macrophages were located in the v-myc sequence and thus directly implicated v-myc and the gag-myc polyprotein in transformation by MC29.
Deletions Within the Transformation-Specific RNA Sequences of Acute Leukemia Virus MC29 Give Rise to Partially Transformation-Defective Mutants
Authors: Bister Klaus , Ramsay Gary M. , Hayman Michael J.
Published: January 2020

The viral RNAs of three nonconditional mutants of avian myelocytomatosis virus MC29 were analyzed. These mutants, which were originally isolated from the quail producer line Q10 and were designated 10A, 10C, and 10H, have lost most of the ability to transform hematopoietic cells in vitro and to induce tumors in vivo, but they still transform cultured fibroblasts with the same efficiency as wild-type (
wt
) MC29. Electrophoretic analyses showed that the mutant genomic RNAs were smaller than the 5.7-kilobase genome of
wt
MC29; the genomes of mutants 10A, 10C, and 10H were about 5.5, 5.3, and 5.1 kilobases long, respectively. Analyses of the transformation-specific sequences of these mutant RNAs by a combination of T
1
oligonucleotide fingerprinting and hybridization with cDNA from the transformation-specific sequences
myc
of
wt
MC29 or competition hybridization including
wt
MC29 RNA revealed that deletions of
myc
-specific sequences had occurred. The deletions in all three mutants overlapped, since they all had lost one particular
myc
-specific oligonucleotide. In agreement with the size of the genomic RNAs, mutants 10C and 10H had lost two additional
myc
oligonucleotides, and mutant 10A contained a modified
myc
oligonucleotide. The locations of the deletions were deduced from comparisons with previously established oligonucleotide maps of several members of the MC29 subgroup of acute leukemia viruses and by hybridization of
wt
and mutant RNAs to molecularly cloned subgenomic fragments of
wt
MC29 proviral DNA, representing the 5′ and 3′ domains of the
myc
sequence. We found that the deleted sequences represented overlapping internal segments of the
myc
sequence and that the borders of
myc
with the partial complements of the virion genes
gag
and
env
appeared to be conserved in mutant and
wt
MC29 RNAs. The correlation between the altered transforming potential for hematopoietic cells and the partial deletion of
myc
in the mutant RNAs provided direct genetic evidence for the involvement of
myc
in oncogenesis. However, the unaffected efficiency of these mutants in fibroblast transformation suggested that the deleted sequences are not essential for the fibroblast-transforming potential of the
onc
gene of MC29.
Phosphorylation and metabolism of the transforming protein of Rous sarcoma virus
Authors: Sefton B M , Patschinsky T , Berdot C , Hunter T , Elliott T
Published: January 2020

p60src, the transforming protein of Rous sarcoma virus, was found to contain 0.5 to 0.9 mol of total phosphate per mol of polypeptide. The protein is known to be phosphorylated at two sites, a serine in the amino-terminal domain and a tyrosine in the carboxy-terminal domain. Because our indirect analysis suggests that the serine is phosphorylated to approximately twice the extent of the tyrosine, we estimate that p60src contains approximately 0.3 to 0.6 mol of phosphoserine and 0.2 to 0.3 mol of phosphotyrosine per mol of polypeptide. p60src was found to represent approximately 0.02% of the total incorporated radioactivity in Rous sarcoma virus-transformed chick cells labeled with [35S]methionine for 48 h. This corresponds to approximately 500,000 molecules of p60src per cell. Pulse-chase experiments revealed that the half-life of p60src ranged from 2 to 7 h, depending on the strain of virus examined. The P60src of the Schmidt-Ruppin strain was significantly more stable than that of the Prague strain.
Molecular Cloning and Physical Mapping of Restriction Endonuclease Fragments of
Autographa californica
Nuclear Polyhedrosis Virus DNA
Authors: Cochran Mark A. , Carstens Eric B. , Eaton Bryan T. , Faulkner Peter
Published: January 2020

A restriction fragment library containing
Autographa californica
nuclear polyhedrosis virus (AcNPV) DNA was constructed by using the pBR322 plasmid as a vector. The library, which is representative of more than 95% of the viral genome, consists of 2 of the 7
Bam
HI fragments, 12 of the 24
Hind
III fragments, and 23 of the 24
Eco
RI fragments. The cloned fragments were characterized and used to generate physical maps of the genome by hybridizing nick-translated recombinant plasmid to Southern blots of AcNPV DNA digested with
Sma
I,
Bam
HI,
Xho
I,
Pst
I,
Hind
III, and
Eco
RI restriction endonucleases. This information was used to define our strain of AcNPV (HR3) with respect to other strains for which physical maps have been previously published. The hybridization data also indicate that reiteration of DNA sequences occurs at the
Hind
III-L and -Q regions of the genome.
DNA-processing activities associated with the purified alpha, beta 2, and alpha beta molecular forms of avian sarcoma virus RNA-dependent DNA polymerase
Authors: Hizi A , Gazit A , Guthmann D , Yaniv A
Published: January 2020

The RNA-dependent DNA polymerase purified from B77 avian sarcoma virus exhibited two distinct DNA-processing activities. The alpha and beta 2 isoenzymes possessed an endodeoxyribonuclease activity capable of nicking simian virus 40 superhelical DNA, whereas the alpha beta isoenzyme performed as an untwisting topoisomerase. Both activities associated with the three molecular forms of the retroviral DNA polymerase were dependent on the presence of either Mn2+ or Mg2+ ions. From analysis of the denaturated DNA products, it is apparent that the alpha and beta 2 isoenzymes introduced two nicks, one per each strand in the superhelical simian virus 40 DNA molecules, whereas the alpha beta polymerase converted these supercoiled molecules to the relaxed covalently closed circular form. The notion that the DNA-processing activities are located on the DNA polymerase molecules was supported by the following: (i) the three isoenzymes were of a high purity; (ii) the activities cosedimented in glycerol gradients with the DNA polymerase activities of the alpha, beta 2, and alpha beta molecular forms; and (iii) immunoglobulin directed against the purified polymerase immunoprecipitated the DNA-processing activities. Chemical treatments of the DNA polymerase molecules (with pyridoxalphosphate, iodoacetamide, and sulfhydryl reagents), which inhibited the polymerase activity, also suppressed the endonucleolytic and topoisomerase activities, suggesting that cystein and amino groups play an important role in the active sites of the DNA-processing activities as well.
Transformation-deficient adenovirus mutant defective in expression of region 1A but not region 1B
Authors: Solnick D , Anderson M A
Published: January 2020

A adenovirus type 5 host range mutant (hr440) has been isolated which is defective in a splicing event required to generate the middle-sized mRNA from early region 1A. This defect has been ascribed to two adjacent nucleotide changes which lie five and six nucleotides from the 5' splice site for this mRNA (Solnick, Nature 291:508-510, 1981). One of these changes introduces an amber codon into the reading frame of the largest region 1A mRNA, resulting in the production of a truncated polypeptide. Like other region 1A mutants, hr440 is defective in the production of mRNA from early regions 2 and 3, but hr440 is unusual in that transcription from regions 1B and 4 is normal. Furthermore, although region 1B expression is unaffected, hr440 does not transform baby rat kidney cells. Therefore, expression of early region 1B is insufficient for transformation, eliminating the possibility that region 1A is required only to induce such expression.
Late events in T4 bacteriophage DNA replication. III. Specificity of DNA reinitiation as revealed by hybridization to cloned genetic fragments
Authors: Halpern M , Mattson T , Kozinski A W
Published: January 2020

Through the use of the technique of hybridization to cloned genes, the site specificity of the reinitiation of T4 DNA replication was examined at late times after infection, when a large amount of DNA had accumulated in the infected cell. Replication was examined under two conditions; (i) when there was recombination but the repair of the recombinants was inhibited, and (ii) when recombination was followed by covalent joining. When no covalent repair of recombinant was allowed, reinitiation occurred in the areas known to be also involved in the initiation of replication of the parental molecule: thus late reinitiation, if covalent joining is prevented, is site specific. When there was covalent joining, reinitiation displayed no apparent site specificity. The results are discussed in light of the possibility that at late times after infection recombinant intersections act as primers. The similarity of the model proposed to the "break-and-copy" model for lambda phage and the fitness of the proposed model to the genetic phenomena described by others are emphasized.
Identification of proteins encoded by a fragment of herpes simplex virus type 2 DNA that has transforming activity
Authors: Galloway D A , Goldstein L C , Lewis J B
Published: January 2020

Cloned BglII fragment N (map units 0.58 to 0.625) of herpes simplex virus type 2 DNA has been shown to transform rodent cells to an oncogenic phenotype (Galloway and McDougall, J. Virol. 38: 749-760, 1981). RNA homologous to this fragment directs the synthesis of five polypeptides in a cell-free translation system. The approximate molecular weights of these proteins are 140,000, 61,000, 56,000, 35,000, and 23,500. The 35,000-dalton protein is the major species late in infection and is the only species detected before the onset of viral DNA replication. The arrangement of the sequences encoding these proteins along the herpes simplex virus type 2 genome was determined by hybridization of the RNA to cloned PstI fragment of BglII-N and to single-stranded DNA segments cloned into M13mp7. Both the hybridization experiments and immunoprecipitation with monoclonal antibodies suggested that the 140,000- and 35,000-dalton proteins are at least partially colinear and share antigenic determinants.
Demonstration of biological activity and nucleotide sequence of an in vitro synthesized clone of the Moloney murine sarcoma virus mos gene
Authors: Donoghue D J
Published: January 2020

A clone of the Moloney murine sarcoma virus mos gene derived by in vitro reverse transcription was characterized. When assayed for focus formation by DNA transfection on NIH/3T3 cells, this clone was biologically inactive, presumably due to the absence of a long terminal repeat sequence. Therefore, a long terminal repeat was inserted into the clone by in vitro recombination, after which the most gene was able to transform NIH/3T3 cells efficiently. The nucleotide sequence encompassing the transforming region of this clone was determined. A single long open reading frame was observed, which potentially encodes a polypeptide of 41,000 daltons. This open reading frame initiates with the first five amino acids of the murine leukemia virus env gene, after which it enters the mos sequence, where it terminates. The nucleotide sequence described in this paper was compared with other sequences of mos in an effort to resolve discrepancies in the position of the long open reading frame. Although Moloney murine sarcoma virus retains the 3' splicing site of the murine leukemia virus env gene, a mos-specific mRNA which corresponds structurally to the murine leukemia virus env mRNA was not identified. The sequence described here revealed a single nucleotide change in the proposed env gene 3' splicing site which was retained in Moloney murine sarcoma virus. This deviation from the consensus 3' splicing sequence may underlie the observed absence of mos expression via the env gene splicing pathway.
Cellular proteins reactive with monoclonal antibodies directed against simian virus 40 T-antigen
Authors: Crawford L , Leppard K , Lane D , Harlow E
Published: January 2020

Several recently isolated monoclonal antibodies which reacted with simian virus 40 T antigens also reacted with proteins found in uninfected and untransformed cells. The proteins were different from each other, PAb419 reacting with a 35,000-molecular-weight protein, PAb427 reacting with a 75,000-molecular-weight phosphoprotein, PAb405 reacting with a 150,000-molecular-weight phosphoprotein, and PAb204 reacting with a 68,000-molecular-weight protein. It is suggested that although some of these cross-reactions may be fortuitous, they may, as an alternative, reflect similarities of shape and perhaps function between domains of the viral T antigen and the relevant host proteins.
Mutation affecting late gene expression in polyoma virus maps in the late region
Authors: Thomas E K , Hare J D
Published: January 2020

A mutation in polyoma virus strain 3049 which results in the overproduction of capsid proteins has been mapped to the late region of the genome between the HindIII site at 45.0 map units and the BamHI site at 58.6 map units. This region contains the coding sequence for VP3 and a portion of VP2, but does not include the late promoters or the coding sequence for the late leaders. The possible role of VP2 or VP3 in the regulation of genetic expression in polyoma virus is discussed.
Synthesis of coronavirus mRNAs: kinetics of inactivation of infectious bronchitis virus RNA synthesis by UV light
Authors: Stern D F , Sefton B M
Published: January 2020

Infection of cells with the avian coronavirus infectious bronchitis virus results in the synthesis of five major subgenomic RNAs. These RNAs and the viral genome form a 3' coterminal nested set. We found that the rates of inactivation of synthesis of the RNAs by UV light were different and increased with the length of the transcript. These results show that each RNA is transcribed from a unique promoter and that extensive processing of the primary transcripts probably does not occur.
Precursor Polypeptides to Structural Proteins of Visna Virus
Authors: Vigne Robert , Filippi Pierre , Quérat Gilles , Sauze Nicole , Vitu Christian , Russo Pierre , Delori Pierre
Published: January 2020

Visna virus is a retrovirus which replicates in fibroblast-like cells of the sheep choroid plexus through a lytic cycle. Visna virions contain three major low-molecular-weight proteins (p30, p16, and p14) which, together with the genomic RNA and several molecules of reverse transcriptase, constitute the core structure of the virions. The core is surrounded by an envelope containing a major glycoprotein (gp135). By analogy with the oncoviruses, these three groups of structural proteins (i.e., the internal proteins, the envelope glycoprotein, and the reverse transcriptase) are probably encoded by the
gag, env
, and
pol
genes, respectively. To elucidate the genetic organization of the visna virus genome and its expression, we studied the synthesis of viral proteins in infected sheep choroid plexus cells. Intracellular viral proteins were detected by immunoprecipitation of pulse-labeled cell extracts with monospecific sera raised against p30, p16, and gp135 and resolution of the proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Immunoprecipitation with anti-p30 and anti-p16 sera allowed the characterization of the 55,000-dalton polypeptide precursor to internal virion proteins p30, p16, and p14 (Pr55

gag

). Tryptic peptide mapping confirmed the precursor-product relationship between Pr55

gag

and the three internal proteins. In addition, a
gag
-related polypeptide of 150,000 daltons was also detected. This polypeptide, which was less abundant than Pr55

gag

, is a likely precursor to the viral reverse transcriptase (Pr150

gag-pol

). Pr55

gag

and Pr150

gag-pol

are not glycosylated. The precursor related to major envelope protein gp135 is a glycosylated polypeptide with an average molecular weight of 150,000 (gPr150

env

). Pulse-chase experiments indicated that gPr150

env

matures into glycoprotein gp135 intracellularly; however, gp135 was never preponderant in cell extracts. The non-glycosylated from of gPr150

env

, which accumulated in the presence of 2-deoxy-
d
-glucose, appeared as a polypeptide of about 100,000 daltons. These results indicated that visna virus codes for the largest non-glycosylated
env
-related precursor among all of the retroviruses and therefore probably contains the largest
env
gene.
Accumulation of Spliced Avian Retrovirus mRNA Is Inhibited in
S
-Adenosylmethionine-Depleted Chicken Embryo Fibroblasts
Authors: Stoltzfus C. Martin , Dane Richard W.
Published: January 2020

The synthesis and processing of B77 avian sarcoma virus RNA in infected chicken embryo fibroblasts was followed in the presence and absence of cycloleucine, a competitive inhibitor of the synthesis of
S
-adenosylmethionine and thus an inhibitor of RNA methylations. An increase in the steady-state levels of genome-length RNA and a decrease in the steady-state levels of subgenomic RNA molecules were obtained in the
S
-adenosylmethionine-depleted avian sarcoma virus-infected cells after 24 h of treatment with the inhibitor. The total number of virus-specific RNA molecules per cell, however, remained relatively constant under either condition. The production of newly synthesized virus-specific RNA in cycloleucine-treated and untreated cells infected with a transformation-defective strain of B77 avian sarcoma virus was followed as a function of [
3
H]uridine labeling time. The accumulation of radioactive genome-length 8.4-kilobase (kb) RNA continued in cycloleucine-treated cells, and virus particle production proceeded at normal rates as previously shown by incorporation of labeled nucleoside precursors or amino acids. In contrast, newly synthesized 3.5-kb subgenomic mRNA, the putative mRNA for the envelope protein precursor, failed to accumulate in the treated cells. The extent of the inhibition in the appearance of the radioactive 3.5-kb RNA was correlated with the extent of the inhibition of viral genomic and cellular mRNA methylations and was a function of the cycloleucine concentration. Under conditions in which the accumulation of 3.5-kb envelope protein mRNA was blocked by the cycloleucine treatment, there were significant increases in the rate of synthesis of the polypeptide products of the genome-length RNA, the precursors to the non-glycosylated
gag
proteins (Pr76

gag

), and the reverse transcriptase (Pr 180

gag pol

) relative to the rate of synthesis of the envelope protein precursor (gPr 92

env

). These results suggest that there is an
S
-adenosylmethionine requirement for the splicing, but not for the synthesis, packaging, or messenger function, of avian retrovirus genome-length RNA. Possible reasons for this requirement are discussed.
Monoclonal antibodies to the p21 products of the transforming gene of Harvey murine sarcoma virus and of the cellular ras gene family
Authors: Furth M E , Davis L J , Fleurdelys B , Scolnick E M
Published: January 2020

We have isolated eight rat lymphocyte-myeloma hybrid cell lines producing monoclonal antibodies that react with the 21,000-dalton transforming protein (p21) encoded by the v-ras gene of Harvey murine sarcoma virus (Ha-MuSV). These antibodies specifically immunoprecipitate both phosphorylated and non-phosphorylated forms of p21 from lysates of cells transformed by Ha-MuSV. All eight react with the products of closely related ras genes expressed in cells transformed by two additional sarcoma viruses (rat sarcoma virus and BALB sarcoma virus) or by a cellular Harvey-ras gene placed under the control of a viral promoter. Three of the antibodies also react strongly with the p21 encoded by the v-ras gene of Kirsten MuSV. These same three antibodies immunoprecipitate the predominant p21 species synthesized normally in a variety of rodent cell lines, including the p21 produced at high levels in 416B murine hemopoietic cells. This suggests that an endogenous gene closely related to Kirsten-ras is expressed in these cells. The monoclonal antibodies have been used to confirm two properties associated with p21; localization at the inner surface of the membrane of Ha-MuSV-transformed cells, assayed by immunofluorescence microscopy, and binding of guanine nucleotides.
Fatty acid modification of Newcastle disease virus glycoproteins
Authors: Chatis P A , Morrison T G
Published: January 2020

The fatty acid acylation of Newcastle disease virus hemagglutininin-neuraminidase and fusion glycoproteins was assayed. [3H]palmitate label was associated with cytoplasmic fusion proteins (F0 and F1) and virion-associated F1. In contrast, there was no detectable [3H]palmitate label associated with the hemagglutin-neuraminidase protein in Newcastle disease virus-infected Chinese hamster ovary cells or chicken embryo cells or in virions released from these cells. Thus, fatty acid modification may not be important for the maturation of some glycoproteins.
Identification of a DNA fragment from a molecularly cloned mink cell focus-inducing murine leukemia virus specific for xenotropic virus-related sequences
Authors: Yoshimura F K
Published: January 2020

Molecular clones of closed circular DNA molecules of a mink cell focus-inducing murine leukemia virus (MCF-13 MuLV) were generated. Closed circular DNA molecules isolated from a Hirt extraction of recently infected NIH/3T3 cells were inserted at their unique EcoRI site into lambda gtWES.lambda B. Restriction endonuclease analysis of inserts of two clones indicated that they represented intact MCF-13 MuLV genomes. One viral insert contained two large terminal repeat sequences, and the other contained only one. A 300-base-pair DNA fragment located in the envelope region of the MCF-13 MuLV genome was determined to be related to xenotropic MuLV sequences.