bluuebunny/crossref_metadata_2025 huggingface dataset:
Early region 1B of adenovirus 2 encodes two coterminal proteins of 495 and 155 amino acid residues
Authors: Anderson C W , Schmitt R C , Smart J E , Lewis J B
Published: January 2020

Partial sequence analysis of tryptic peptides has identified the E1B-495R (E1b-57K) (early transcription region 1B of 495 amino acid residues, with an approximate molecular weight of 57,000) protein of adenovirus 2 as encoded by the 495 amino acid open reading frame located in the adenovirus 2 DNA sequence between nucleotides 2016 and 3500. Additional proteins of 16,000 Mr and 18,000 Mr that are related to the E1B-495R protein were identified by cell-free translation of hybridization-selected mRNA. Analysis of [35S]methionine-containing amino terminal tryptic peptides by thin-layer chromatography showed that the E1B-495R, E1B-18K, and E1B-16K proteins all begin at the same initiation codon. The E1B-495R protein from 293 cells also has the same initial tryptic peptide, acetyl-methionyl-glutamyl-arginine. Sequence analysis of E1B-18K tryptic peptides indicated that this protein also has the same carboxy terminus as the E1B-495R protein and that it is derived from an mRNA that is spliced to remove sequences between nucleotides 2250 and 3269, resulting in a protein product of 155 amino acid residues. Analysis of E1B-16K tryptic peptides has not yet revealed the carboxy terminal structure of this protein. Both the E1B-495R and the E1B-155R (E1B-18K) proteins, as well as the E1B-16K protein, were precipitated from cell-free translations and from extracts of infected cells by antiserum against an amino terminal nonapeptide common to these proteins.
Antiviral and protein-inducing activities of recombinant human leukocyte interferons and their hybrids
Authors: Sen G C , Herz R , Davatelis V , Pestka S
Published: January 2020

The antiviral activities of recombinant human leukocyte interferons IFN-alpha A and IFN-alpha D as well as five hybrids of these interferons against retroviruses, vesicular stomatitis virus, and encephalomyocarditis virus were studied in feline, human, and murine cells. Although these interferon species had widely different potencies, their activities against these viruses were, in general, proportional. The IFN-alpha A/D (Bgl) hybrid was the most potent species, and the IFN-alpha D/A (Bgl) hybrid was the least potent. However, the latter species did not interfere with the action of the former species. Like natural human leukocyte interferon, each of the seven species of recombinant interferons induced the synthesis of at least five proteins in human fibroblasts, whereas induction of only one such protein was readily detected in a feline fibroblast line in which these interferon species inhibited the replication of all three viruses.
Monkey B-lymphotropic papovavirus DNA: nucleotide sequence of the region around the origin of replication
Authors: Furuno A , Miyamura T , Yoshiike K
Published: January 2020

We have determined the nucleotide sequence of the HindIII-B DNA segment of African green monkey B-lymphotropic papovavirus (LPV), which shows a highly restricted host range and whose genome is a 5.1-kilobase-long circular DNA. The segment, consisting of 1,123 base pairs, contained the origin of DNA replication, the putative control region for early transcription, and the region probably coding for the amino-terminal portion of T antigens. The symmetrical region at the center of replication origin, 5'-GAGGC CA GGGGCCCC TA GCCTC-3' (on the L strand), has diverged in the central portion from the corresponding regions of primate polyomaviruses simian virus 40, BK virus, and JC virus, but resembles that of mouse polyomavirus. The structure of the control region upstream of the replication origin was unique to LPV and contained several repeated sequences, the longest of which were two 60-base-pair tandem repeats. The amino-terminal region common to LPV small T and large T antigens showed some homology (41%) in the deduced amino acid sequence to that of both simian virus 40 and the mouse polyomavirus. Like other polyomaviruses, the probable carboxyl-terminal region unique to LPV small T antigen contained two sets of the Cys-x-Cys-x-x-Cys structure. These data show that, despite the unique structures in the control region, LPV is evolutionally related to the mouse polyomavirus and to simian virus 40.
Biochemical map of polypeptides specified by foot-and-mouth disease virus
Authors: Grubman M J , Robertson B H , Morgan D O , Moore D M , Dowbenko D
Published: January 2020

Pulse-chase labeling of foot-and-mouth disease virus-infected bovine kidney cells revealed stable and unstable viral-specific polypeptides. To identify precursor-product relationships among these polypeptides, antisera against a number of structural and nonstructural viral-specific polypeptides were used. Cell-free translations programmed with foot-and-mouth disease virion RNA or foot-and-mouth disease virus-infected bovine kidney cell lysates, which were shown to contain almost identical polypeptides, were immunoprecipitated with the various antisera. To further establish identity, some proteins were compared by partial protease digestion. Evidence for a membrane association of the polypeptides coded for by the middle genome region is also presented. A biochemical map of the foot-and-mouth disease virus genome was established from the above information.
Direction of transcription in bacteriophage T5 first-step transfer DNA
Authors: Davison J , Lafontaine D
Published: January 2020

The transcription of the T5 first-step transfer genes A2 and A3 has been shown to proceed from right to left (i.e., from A3 to A2) on the T5 DNA molecule.
Dexamethasone-stimulated expression of a proviral copy of mouse mammary tumor virus env mRNA
Authors: Robertson D L
Published: January 2020

The kinetics of dexamethasone induction of mouse mammary tumor virus RNAs in a cell line (designated line 8) which contains a deleted proviral DNA has been examined. This DNA is an apparent reverse transcription product of the env mRNA. A detectable increase in the 3.8-kilobase RNA (mouse mammary tumor virus env mRNA) was present within 15 min after hormone addition. The 2.5-kilobase RNA detected previously during hormone induction studies was also observed in these cells. Both RNAs increased in concentration up to about 4 h after hormone addition and thereafter attained the steady-state level. Pr73env was detected by immunoprecipitation and appeared to be identical to bona fide Pr73env isolated from wild-type mouse mammary tumor virus-infected cells. An additional 5.7 kilobases of RNA was detected in augmented quantities after hormone addition, but the structure of this RNA is not known.
Evidence for secondary structure within the virion RNA of echovirus 22
Authors: Seal L A , Jamison R M
Published: January 2020

Phenol-extracted echovirus 22 virion RNA is infectious, but unlike poliovirus virion RNA, it resists digestion with pancreatic RNase and nuclease P-1, a 3' exonuclease selective for single-stranded RNA. These data indicate the presence of an enzyme-resistant portion somewhere in the RNA molecule and suggest that it is a double-stranded or base-paired region distant from the unblocked 3' terminus. Equilibrium density gradient centrifugation of native echovirus 22 virion RNA results in a single peak with a density of 1.63 g/cm3. When sheared before centrifugation, the molecule is resolved into two RNA species: one with an approximate density of 1.70 to 1.71 g/cm3, as is observed also for single-stranded poliovirus virion RNA, and the other with a density of 1.58 to 1.59 g/cm3. Data obtained from rate zonal centrifugation may be used to calculate an approximate sedimentation coefficient corrected to water at 20 degrees C of 34 and a molecular weight of 2.4 X 10(6) for the virion RNA. We propose a model for echovirus 22 RNA composed of a linear RNA molecule with a 5' hairpin.
Molecular cloning and characterization of a leukemia-inducing myeloproliferative sarcoma virus and two of its temperature-sensitive mutants
Authors: Kollek R , Stocking C , Smadja-Joffe F , Ostertag W
Published: January 2020

The myeloproliferative sarcoma virus (MPSV) induces extensive hematopoietic changes, including spleen foci in adult mice, and transforms fibroblasts in vitro. NRK nonproducer cell lines of MPSV and ts temperature-sensitive mutants were analyzed by restriction enzyme digestion and Southern blotting. EcoRI fragments containing the proviral DNAs of MPSV and two temperature-sensitive mutants and rat cellular sequences homologous to c-mos were molecularly cloned. By comparing restriction enzyme cleavage sites, it was shown that the MPSV genome consists only of sequences related either to Moloney murine leukemia virus or to the c-mos mouse oncogenic sequences. Two regions of fragment heterogeneity were observed: (i) in the defective pol gene, where MPSV and the two cloned temperature-sensitive mutants were different from Moloney murine sarcoma virus and from each other, although MPSV wild-type retained more of the pol gene than any of the Moloney murine sarcoma virus isolates; (ii) in the area 3' to the mos gene, which was identical in MPSV and its temperature-sensitive mutants but different from other Moloney murine sarcoma virus variants. Transfection of cloned MPSV DNA in RAT4 cells and virus rescue on infection with Friend murine leukemia virus yielded MPSV which transformed fibroblasts in vitro and also induced spleen foci in adult mice, thus proving that both properties are coded by the same viral genome.
Effects of deletions on expression of the herpes simplex virus thymidine kinase gene from the intact viral genome: the amino terminus of the enzyme is dispensable for catalytic activity
Authors: Halpern M E , Smiley J R
Published: January 2020

We have transferred two deletions affecting the 5' end of the herpes simplex virus thymidine kinase (TK) gene into the intact viral genome. One, extending from -12 to +189, had no effect on TK mRNA synthesis and only a small effect on TK activity, although the first 27 codons of the TK polypeptide were deleted. The other, extending from -85 to +85, severely impaired TK mRNA synthesis. We conclude that the amino terminus of the TK polypeptide is dispensable for catalytic activity, and that expression of TK in viral infections requires some of the same promoter elements used in uninfected cells.
Fluidity of a retrovirus genome
Authors: Clark S P , Mak T W
Published: January 2020

Comparison of the genomic sequences of the Friend spleen focus-forming virus with other murine retroviral sequences indicated that the spleen focus-forming virus was derived from at least three retroviruses. The 5' end of the virus, from the primer binding site through most of gag, was derived from AKV. The rest of gag and all of pol were of uncertain origin, but were probably derived from the same xenotropic virus that gave rise to the 5' half of env. The remainder was derived from Friend murine leukemia virus. The positions of a 585-base deletion, a 6-base duplication, and a point insertion that leads to a frame shift and premature protein termination in the ecotropic 3' end of env were invariant between three spleen focus-forming virus strains, indicating that they had a single common ancestor. However, the point of crossover between xenotropic viral sequences and Friend murine leukemia virus was different in each strain, and two strains were much more closely related to each other than to the third in the xenotropic region, indicating that these strains had diverged by multiple recombinations. Furthermore, a different nucleotide comprised the single point insertion near the 3' end of env, suggesting that these viruses have an extremely high transition and transversion rate.
Bacteriophage phi 29 proteins required for in vitro DNA-gp3 packaging
Authors: Bjornsti M A , Reilly B E , Anderson D L
Published: January 2020

In vitro assembly of bacteriophage phi 29 in crude extracts involves efficient packaging of a DNA-protein complex (DNA- gp3 ) into a prohead with the aid of the gene 16 product ( gp16 ) and subsequent assembly of neck and tail proteins ( Bjornsti et al., J. Virol. 41:508-517, 1982; Bjornsti et al., J. Virol. 45:383-396, 1983; Bjornsti et al., Proc. Natl. Acad. Sci. U.S.A. 78:5861-5865, 1981). To define the viral proteins required for the DNA- gp3 encapsidation phase, we purified biologically active proheads and DNA- gp3 and constructed a chimeric plasmid, pUM101 , which contained and expressed gene 16 of phi 29 and no other viral genes. The plasmid-specified gp16 was both necessary and sufficient to package 24% of the DNA- gp3 added to the purified proheads , and the DNA-filled heads so produced were efficiently complemented to infectious phage by the addition of neck and tail proteins. Purified proheads and DNA- gp3 gave linear dose-response curves with slopes of approximately 1; in contrast, a 4-fold dilution of gp16 resulted in a 1,000-fold reduction of phi 29, suggesting a requirement for multiple copies of this protein.
Immunogenicity of herpes simplex virus glycoproteins gC and gB and their role in protective immunity
Authors: Glorioso J , Schröder C H , Kumel G , Szczesiul M , Levine M
Published: January 2020

The relative antigenicity of the individual herpes simplex virus type 1 (KOS) glycoproteins gC and gB was analyzed in BALB/c mice by using KOS mutants altered in their ability to present these antigens on cell surface membranes during infection. The mutants employed were as follows: syn LD70 , a non-temperature-sensitive mutant defective in the synthesis of cell surface membrane gC; tsF13 , a temperature-sensitive mutant defective in the processing of the precursor form of gB to the mature cell surface form at 39 degrees C; and ts606 , an immediate early temperature-sensitive mutant defective in the production of all early and late proteins including the glycoproteins. By comparing the relative susceptibility to immunolysis of mouse 3T3 cells infected at 39 degrees C with wild-type virus, presenting the full complement of the glycoprotein antigens, gC, gB, and gD, with target cells infected with mutants presenting only subsets of these antigens, we determined that a major portion of cytolytic antibody contained in hyperimmune anti-herpes simplex virus type 1 (KOS) mouse antiserum was directed against glycoproteins gC and gB. The relative immunogenicity of wild-type and mutant virus-infected cells also was compared in BALB/c mice. Immunogen lacking the mature form of gB induced a cytolytic antibody titer comparable to that of the wild-type virus, whereas that lacking the mature form of gC showed a 70% reduction in titer. The absence of the mature cell surface forms of gB and gC in immunogen preparations resulted in a 4- to 15-fold reduction in in virus neutralizing titer. Animals immunized with ts606 -infected cells (39 degrees C) induced relatively little virus-specific cytolytic and neutralizing antibody. Analysis of the glycoprotein specificities of these antisera by radioimmunoprecipitation showed that the antigens immunoprecipitated reflected the viral plasma membrane glycoprotein profiles of the immunogens. The absence of the mature forms of gC or gB in the immunizing preparation did not appreciably affect the immunoprecipitating antibody response to other antigens. Mice immunized with wild-type and mutant virus-infected cells were tested for their resistance to intracranial and intraperitoneal challenge with the highly virulent WAL strain of herpes simplex virus type 1. Despite the observed alterations in serum virus-specific antibody induced with the individual immunogens, all animals survived an intraperitoneal challenge of 10 50% lethal doses. However, differences in the survival of animals were obtained upon intracranial challenge.(ABSTRACT TRUNCATED AT 400 WORDS)
Characterization of the env gene and long terminal repeat of molecularly cloned Friend mink cell focus-inducing virus DNA
Authors: Adachi A , Sakai K , Kitamura N , Nakanishi S , Niwa O , Matsuyama M , Ishimoto A
Published: January 2020

The highly oncogenic erythroleukemia-inducing Friend mink cell focus-inducing (MCF) virus was molecularly cloned in phage lambda gtWES.lambda B, and the DNA sequences of the env gene and the long terminal repeat were determined. The nucleotide sequences of Friend MCF virus and Friend spleen focus-forming virus were quite homologous, supporting the hypothesis that Friend spleen focus-forming virus might be generated via Friend MCF virus from an ecotropic Friend virus mainly by some deletions. Despite their different pathogenicity, the nucleotide sequences of the env gene of Friend MCF virus and Moloney MCF virus were quite homologous, suggesting that the putative parent sequence for the generation of both MCF viruses and the recombinational mechanism for their generation might be the same. We compare the amino acid sequences in lymphoid leukemia-inducing ecotropic Moloney virus and Moloney MCF virus, and erythroblastic leukemia-inducing ecotropic Friend virus, Friend-MCF virus, and Friend spleen focus-forming virus. The Friend MCF virus long terminal repeat was found to be 550 base pairs long. This contained two copies of the 39-base-pair tandem repeat, whereas the spleen focus-forming virus genome contained a single copy of the same sequence.
Expression in COS-1 cells of Epstein-Barr virus nuclear antigen from a complete gene and a deleted gene
Authors: Robert M F , Shedd D , Weigel R J , Fischer D K , Miller G
Published: January 2020

In a previous study the BamHI-K fragment of Epstein-Barr virus DNA was shown to induce a nuclear antigen, Epstein-Barr virus nuclear antigen (EBNA), when cotransfected with the herpes simplex virus thymidine kinase gene into mouse LTK- cells. We have now inserted the BamHI-K fragment and a BamHI/HindIII subfragment, I1f , into shuttle vectors containing the origin of replication of simian virus 40. These plasmids have been introduced into COS-1, which are monkey kidney cells transformed by an origin-defective simian virus 40 genome. This expression system permitted rapid characterization of antigens, mRNAs, and proteins related to EBNA. The same-sized EBNA protein (approximately 78,000) was made after transfection with BamHI-K (5.2 kilobase pairs [kbp]) or the I1f subfragment (2.9 kbp). A deletion of about 600 bp occurred when the I1f fragment was propagated on the pSV2 plasmid in Escherichia coli. The deleted fragment gave rise to a smaller protein (approximately 52,000). These data provide evidence that EBNA is encoded by the 2.9-kbp I1f and is not an induced cellular protein. Nuclear antigen and polypeptide expression occurred equally well when the Epstein-Barr virus DNA was cloned on PSV2 -gpt or pSVOd . The latter plasmid lacks sequences allowing for efficient early gene transcription as well as splicing and polyadenylation signals which are present in pSV2 . Preliminary mapping of the EBNA gene transcripts demonstrated that two mRNAs (2.9 and 2.4 kilobases [kb]) are homologous to the I1f fragment. Taken together, the data suggest that the 2.9-kbp I1f fragment contains the structural gene for EBNA synthesis. COS-1 cells will thus provide a valuable system in which to analyze functional domains of the EBNA gene.
Role of mink cell focus-inducing virus in leukemias induced by Friend ecotropic virus
Authors: Silver J
Published: January 2020

Recombinant viruses have been implicated in the pathogenesis of murine leukemias induced by a variety of long-latency retroviruses. Neonatal mice of several strains were inoculated with Friend ecotropic virus (F-Eco) and analyzed for the presence of mink cell focus-inducing (MCF) virus or DNA restriction enzyme fragments which were specific for Friend MCF virus (F-MCF). MCF virus was detected within 2 weeks of inoculation in NFS /N mice and at about 2 months after inoculation in BALB/c mice. Both of these strains developed erythroblastosis after inoculation with F-Eco. In contrast, MCF virus was not detected in F-Eco-inoculated C57BL mice. These mice were resistant to erythroblastosis but developed lymphoma or myelogenous leukemia or both at about 5 months after inoculation. Thus, although MCF viruses were associated with F-Eco erythroblastosis in NFS /N and BALB/c mice, they were not necessary for F-Eco-induced lymphoid or myeloid leukemias in C57BL mice. To investigate the association between resistance to erythroblastosis and absence of MCF virus, C57BL mice were inoculated with pseudotypic mixtures of F-Eco plus F-MCF; MCF virus replicated well in these mice, but the mice remained resistant to erythroblastosis. Furthermore, in genetic crosses between C57BL and NFS /N or BALB/c, some mice inherited resistance to F-Eco erythroblastosis without inheriting the C57BL resistance to the generation of MCF viruses. These results indicate that C57BL mice carry a gene for resistance to F-Eco erythroblastosis which is distinct from the C57BL genes which interfere with the generation of MCF viruses.
Production of a monospecific antiserum against the early region 1A proteins of adenovirus 12 and adenovirus 5 by an adenovirus 12 early region 1A-beta-galactosidase fusion protein antigen expressed in bacteria
Authors: Scott M O , Kimelman D , Norris D , Ricciardi R P
Published: January 2020

Antisera were prepared against the amino acid sequences encoded within the N-terminal half of the adenovirus 12 (Ad12) early region 1A (E1A) gene. This was accomplished by construction of a plasmid vector which encoded the N-terminal 131 amino acids of Ad12 E1A joined in frame to the coding sequence of beta-galactosidase. After induced synthesis in Escherichia coli, the Ad12 E1A-beta-galactosidase fusion protein (12-1A-FP) was extracted with urea and used to raise antibodies in rabbits. The 12-1A-FP antisera immunoprecipitated major phosphoproteins of 39,000 and 37,000 apparent molecular weights from Ad12-transformed and infected cells. The 12-1A-FP antisera also immunoprecipitated E1A phosphoproteins from Ad5-transformed and infected cells. Immunospecificity of the 12-1A-FP antisera was demonstrated by the ability of 12-1A-FP antigen to block immunoprecipitation of E1A proteins. Furthermore, E1A proteins immunoprecipitated from in vivo-labeled cells comigrated with those translated in vitro by RNA that had been hybridization selected to E1A DNA.
Induction of tumors and generation of recovered sarcoma viruses by, and mapping of deletions in, two molecularly cloned src deletion mutants
Authors: Wang L H , Edelstein B , Mayer B J
Published: January 2020

td108 , a transformation-defective (td) deletion mutant of the Schmidt-Ruppin strain of Rous sarcoma virus of subgroup A (SR-A), was molecularly cloned. Two isolates of td viruses, td108 -3b and td108 -4a, obtained by transfection of the molecularly cloned td108 DNAs into chicken embryo fibroblasts, were tested for their ability to induce tumors and generate recovered avian sarcoma viruses ( rASVs ) in chickens. Both td viruses were able to induce tumors with a latency and frequency similar to those observed previously with biologically purified td mutants of SR-A. rASVs were isolated from most of the tumors examined. The genomic RNAs of those newly obtained rASVs were analyzed by RNase T1 oligonucleotide fingerprinting. The results showed that they had regained the deleted src sequences and contained the same set of marker src oligonucleotides as those of rASVs analyzed previously. The src oligonucleotides of rASVs are distinguishable from those present in SR-A. We conclude that those rASVs must have been generated by recombination between the molecularly cloned td mutants and the c-src sequence. The deletions in the td mutants were mapped by restriction enzyme analysis and nucleotide sequencing. td108 -3b was found to contain an internal src deletion of 1,416 nucleotides and to retain 57 and 105 nucleotides of the 5' and 3' src coding sequences, respectively. td108 -4a contained a src deletion of 1,174 nucleotides and retained 180 and 225 nucleotides of the 5' and 3' src sequences, respectively. Comparison of sequences in the 5' src and its upstream region of td108 -3b with those of SR-A, rASV1441 (a td108 -derived rASV analyzed previously), and c-src suggested that the 5' recombination between td108 and c-src occurred from 7 to 20 nucleotides upstream from the beginning of the src coding sequence.
DNA binding properties of a mutant T antigen from the simian virus 40-transformed human cell line simian virus 80
Authors: Gruss C , Baumann E , Knippers R
Published: January 2020

We investigated whether the T antigen of the simian virus 40-transformed human cell line simian virus 80 ( SV80 ) specifically recognizes DNA sequences of its own template, i.e, the viral sequences integrated in the SV80 cellular genome. In vitro DNA binding experiments clearly indicated that, in contrast to wild-type T antigen, SV80 T antigen does not specifically bind to sites on the integrated viral DNA in SV80 cells.
Characterization of a Rous sarcoma virus mutant defective in packaging its own genomic RNA: biochemical properties of mutant TK15 and mutant-induced transformants
Authors: Koyama T , Harada F , Kawai S
Published: January 2020

The accompanying paper (S. Kawai and T. Koyama , J. Virol. 51:147-153, 1984) describes the isolation and biological properties of a mutant, TK15 , derived from a Rous sarcoma virus mutant, tsNY68 . The cis-acting defect of the mutant is analyzed biochemically in this paper. TK15 virions released from virus-producing 15c (+) cells were deficient in viral genomic 39S RNA, although comparable amounts of viral RNAs were transcribed in 15c (+) and tsNY68 -infected cells. Analysis of provirus DNA occurring in 15c (+) cells suggested that the mutant genome had a deletion of ca. 250 bases near the 5' end of the genome somewhere between the primer binding site and the 5' end of the gag-coding region. These findings indicate that at least part of the sequence lost in the TK15 genome is indispensable for packaging viral genomic RNA into virions. TK15 induces nonvirus -producing 15c (-) transformants at high frequency. Southern blot analysis of DNAs from those 15c (-) clone cells revealed that TK15 -derived proviruses contained various extents of internal deletions. Many 15c (-) clones had a provirus carrying only the src gene with long terminal repeat sequences at both ends. The mechanism for the segregation of 15c (-) cells is discussed.
Only site-directed antibodies reactive with the highly conserved src-homologous region of the v-abl protein neutralize kinase activity
Authors: Konopka J B , Davis R L , Watanabe S M , Ponticelli A S , Schiff-Maker L , Rosenberg N , Witte O N
Published: January 2020

Antisera specific for six regions of the v- abl protein were used to serologically characterize the Abelson murine leukemia virus tyrosine kinase. Chemically synthesized peptides corresponding to the predicted v- abl protein sequence and larger regions of the v- abl protein expressed as fusion proteins in bacteria were used as immunogens. The specificity of each antiserum was confirmed by immunoprecipitation analysis with defined deletion mutants of Abelson murine leukemia virus. Several of these v- abl -specific antisera display much higher titers and avidities than serum harvested from mice bearing Abelson murine leukemia virus-induced tumors, previously the only source of anti- abl -specific serum. Two antisera were found to block the in vitro autophosphorylation of the v- abl protein as well as its ability to phosphorylate a peptide substrate. Examination of the sites against which the kinase-blocking antisera were prepared revealed that both are in close proximity to the in vivo sites of tyrosine phosphorylation, which fall within the region of high homology with v-src and other tyrosine kinases. Antisera directed against other regions of v- abl did not inhibit kinase activity.
Ion etching of human adenovirus 2: structure of the core
Authors: Newcomb W W , Boring J W , Brown J C
Published: January 2020

The surface of human adenovirus 2 was etched by irradiating intact virions with low-energy (1-keV) Ar+ ions in a Technics Hummer V sputter coater . Viral structures exposed by the etching process were shadowed and then examined in the electron microscope. Periods of etching that were sufficient to reduce the viral diameter by 20 to 30 nm revealed distinct substructural elements in the virion core. Cores were found to consist of a cluster of 12 large, uniformly size spheres which abutted one another in the intact virion. The spheres, for which we suggest the name " adenosomes ," had a diameter of 23.0 +/- 2.3 nm, and they were related to each other by two-, three-, and fivefold axes of rotational symmetry. The results support the view, originally suggested by Brown et al. (J. Virol. 16:366-387, 1975) that the adenovirus 2 core is composed of 12 large spheres packed tightly together in such a way that each is directed toward the vertex of an icosahedron . Such a structure, constructed of 23.0-nm-diameter spheres, would have an outside diameter (vertex-to-vertex distance) of 67.0 nm and a face-to-face distance of 58.2 nm. It could be accommodated inside the icosahedral adenovirus capsid if each large sphere were located beneath a capsid vertex.
Organization of RNA transcripts from a vaccinia virus early gene cluster
Authors: Morgan J R , Roberts B E
Published: January 2020

The detailed organization of the RNAs transcribed from an early gene cluster encoded by vaccinia virus has been determined from the information derived from several complementary techniques. These include hybrid selection coupled with cell-free translation to locate DNA sequences complementary to mRNAs encoding specific polypeptides; RNA filter hybridization to size and locate on the DNA mature RNAs as well as higher-molecular-weight RNAs; S1 nuclease mapping to precisely locate the 5' and 3' ends of the RNAs; S1 nuclease mapping to precisely locate the 5' and 3' ends of the RNAs; and fractionation of hybrid-selected mRNAs in an agarose gel containing methyl mercury hydroxide followed by the cell-free translation of these mRNAs to definitively ascertain the size of the mRNA encoding each polypeptide. The early gene cluster is located between 21 and 26 kilobases from the left end of the vaccinia virus genome and is encoded by a 5.0-kilobase EcoRI fragment which spans the HindIII-N, -M, and -K fragments. Transcribed towards the left terminus are four mature mRNAs, 1,450, 950, 780, and 400 nucleotides in size, encoding polypeptides of 55, 30, 20, and 10 kilodaltons, respectively. These mRNAs are colinear with the DNA template and are closely spaced such that the 5' terminus of one mRNA is within 50 base pairs of the 3' terminus of the adjacent RNA. In addition to the mature size mRNAs, there are higher-molecular-weight RNAs, 5,000, 3,300, 2,350, 2,300, 1,800, 1,700, and 1,350 nucleotides in size. The 5' and 3' termini of the high-molecular-weight RNAs are coterminal with the 5' and 3' termini of the mature size mRNA. The implications of this arrangement and the biogenesis of these early mRNAs are discussed.
Many rhinovirus serotypes share the same cellular receptor
Authors: Abraham G , Colonno R J
Published: January 2020

Twenty-four human rhinovirus serotypes were grown and purified by centrifugation in metrizamide density gradients. These preparations had a lower buoyant density (1.24 g/cm3) and higher specific infectivities (1:24 to 1:240) than did rhinoviruses described previously (E. J. Stott and R. J. Killington, Annu. Rev. Microbiol. 26:503-524, 1972). Binding conditions in which the unique cellular receptors for virus attachment were saturated were determined for each serotype. Competition binding assays between pairs of serotypes allowed 20 of the 24 serotypes to be assigned to the same cellular receptor. The remaining four serotypes appeared to attach to a different cellular receptor. Since most serotypes were chosen for study at random, it seems likely that many of the yet unstudied rhinoviruses will share this common cellular receptor.
Nucleotide sequence of an mRNA transcribed in latent growth-transforming virus infection indicates that it may encode a membrane protein
Authors: Fennewald S , van Santen V , Kieff E
Published: January 2020

The most abundant Epstein-Barr virus mRNA in a latently infected cell line, IB4, established by in vitro growth transformation with virus, was a 2,8-kilobase RNA encoded by largely unique DNA near the right end of the genome. The RNA was transcribed from right to left, and two introns were spliced out. This region of the genome was sequenced, and the exons of the RNA were identified by S1 analysis of DNA-RNA hybrids and primer extension. The first start codon in the RNA was 40 nucleotides from its 5' end. Beginning with the start codon, there was a 1,158-nucleotide open reading frame which crossed both introns. The important characteristics of the translated protein were as follows. (i) The amino terminus was highly charged and not suggestive of a leader sequence. (ii) There were six markedly hydrophobic alpha-helical domains, each having 21 amino acids and connected by 5 to 7 amino acid segments predicted to be reverse turns. (iii) The carboxy-terminal 200 amino acids were markedly acidic, containing 6 glutamic and 37 aspartic acids. The hydrophobic region is predicted to form six membrane-spanning regions, leaving the short charged amino terminus and long acidic carboxy terminus on the inside of the membrane. This protein could be responsible for the new antigen detected in the plasma membrane of Epstein-Barr virus-transformed cells, lymphocyte-determined membrane antigen. There were two other open reading frames in the RNA.
Human polyomavirus JC virus genome
Authors: Frisque R J , Bream G L , Cannella M T
Published: January 2020

The complete DNA sequence of the human JC virus, which was found to consist of 5,130 nucleotide pairs, is presented. The amino acid sequence of six proteins could be deduced: the early, nonstructural proteins, large T and small t antigens; the late capsid proteins, VP1, VP2, and VP3; and the agnogene product encoded within the late leader sequence, called the agnoprotein in simian virus 40. The extent of homology between JC virus DNA and the genomes of simian virus 40 (69%) and BK virus (75%) confirmed the close evolutionary relationship of these three polyomaviruses. The sequences showing the greatest divergence in these viral DNAs occurred within the tandem repeats located to the late side of the replication origins.
A bacteriophage T5 mutant with an increased frequency of single-chain interruptions
Authors: Rhoades M
Published: January 2020

A T5 mutant whose DNA has two sites where single-chain interruptions occur with a higher frequency than in wild-type DNA was isolated. Both sites occurred within the same strand as do the natural interruptions in T5 DNA, and both were due to alteration of the mechanism that generates the interruptions.
Spontaneous and induced leukemias of myeloid origin in recombinant inbred BXH mice
Authors: Bedigian H G , Johnson D A , Jenkins N A , Copeland N G , Evans R
Published: January 2020

BXH-2 recombinant inbred (RI) mice produce high titers of B-ecotropic murine leukemia virus beginning early in life and have a high incidence of non-T-cell leukemias that occur before 1 year of age. The leukemias that develop are in some cases associated with hind limb paralysis. In addition, a dualtropic mink cell focus-forming virus has been isolated from leukemic cells of BXH-2 mice. Immunological and cytochemical characterization of the BXH-2 leukemias showed that they are of the myeloid lineage. To assess the oncogenicity of the BXH-2 viruses, newborn mice of several BXH RI strains were inoculated at birth with biologically cloned B-ecotropic or mink cell focus-forming murine leukemia virus. These studies demonstrated that the B-ecotropic virus can induce myeloid leukemias in other BXH RI strains, whereas the dualtropic mink cell focus-forming isolates were nononcogenic in the strains tested. DNA-DNA reassociation analysis indicated that the organotropism of the B-ecotropic murine leukemia virus is confined to lymphoid tissues. Southern analysis of tumor DNAs showed that there was amplification of ecotropic virus-specific sequences in BXH-2 myeloid tumors and in all leukemias induced in other BXH RI strains by inoculation of the BXH-2 B-ecotropic virus. Although B-ecotropic virus is expressed in central nervous tissues of paralyzed BXH-2 mice, we were unable to induce the disorder in several BXH RI strains inoculated intracranially at birth with either the B-ecotropic or dualtropic virus. These results suggest that the paralysis that occurs in BXH-2 mice is due to the infiltration of leukemic cells into the central nervous system.
A membrane-associated, carbohydrate-modified form of the v-abl protein that cannot be phosphorylated in vivo or in vitro
Authors: Watanabe S M , Rosenberg N E , Witte O N
Published: January 2020

Abelson murine leukemia virus encodes a transforming protein which contains tyrosine kinase activity and is phosphorylated in vivo and in vitro. We found that P160 and P160-derived virus strains expressed an additional, altered v-abl protein which could not be phosphorylated. The altered v-abl protein (L-v-abl) differed from the phosphorylated form (K-v-abl) in that it was glycosylated and localized exclusively to the membrane fraction. Tunicamycin inhibition of N-linked carbohydrate addition did not restore phosphorylation. It did, however, reveal that L-v-abl had additional sequences relative to K-v-abl. The coding sequences required for this region and for the expression of L-v-abl were identified by replacing sequences in the P120 virus genome, which did not express L-v-abl, with sequences from the P160 virus genome. The necessary sequences were localized to the Moloney murine leukemia virus-derived gag gene. Comparison between the in vitro altered P120 and wild-type P120 virus strains indicated that expression of L-v-abl did not increase the efficiency of lymphoid transformation. Although the biological role of L-v-abl is not clear, our analyses have revealed that a specific amino terminal gag sequence can prevent v-abl from acting as a kinase substrate and can alter the cellular localization and modification of v-abl. These properties distinguish L-v-abl from previously reported v-abl proteins.
Regulation of expression and nucleotide sequence of a late vaccinia virus gene
Authors: Weir J P , Moss B
Published: January 2020

A subset of vaccinia virus genes are expressed only after DNA replication. To investigate the regulation of such transcriptional units, a representative gene encoding a major late polypeptide (Mr, 28,000) was mapped and sequenced. Translatable mRNAs were heterogeneous in length and overlapped several early genes downstream. The 5' end of the message was located, and the DNA segment upstream was excised and ligated to the coding sequence of the easily assayable procaryotic chloramphenicol acetyltransferase gene. The resulting chimeric gene was recombined into the thymidine kinase locus of the vaccinia virus genome, and infectious recombinant virus was isolated. Both the time of chloramphenicol acetyltransferase synthesis in infected cells and the requirement for DNA replication indicate that the sequence upstream of the late gene contains cis-acting transcriptional regulatory signals.
Immunoprecipitation of some forms of simian virus 40 large-T antigen by antibodies to synthetic peptides
Authors: Paucha E , Harvey R , Smith A E
Published: January 2020

Antibodies were raised against six synthetic peptides corresponding to overlapping amino acid sequences (106 through 145) from a putative DNA binding domain in simian virus 40 (SV40) large-T antigens. All six antipeptide sera immunoprecipitated large-T from crude extracts of SV40-transformed cells, but the efficiency varied widely; in general, antibodies to the longer peptides produced the strongest anti-large-T activity. Antisera were purified by immunoaffinity chromatography on immobilized peptide. The purified antisera recognized only some forms of large-T; full-sized large-T from transformed cells, super-T from SV3T3 C120 cells, and 70,000-dalton T-antigen from Taq-BamHI cells were immunoprecipitated, whereas large-T from productively infected cells reacted irreproducibly, and the full-sized protein, synthesized in vitro or eluted from sodium dodecyl sulfate-containing gels, and the 33,000- and 22,000-dalton truncated large-Ts from Swiss SV3T3 and MES2006 cells, respectively, were not immunoprecipitated. This pattern of reactivity was explained when extracts were fractionated by sucrose density centrifugation, and it was found that only rapidly sedimenting forms of large-T were immunoprecipitated by the antipeptide sera; that is, large-T complexed with nonviral T antigen was detected, whereas lighter forms were not detected. Cascade immunoprecipitations did not support the view that this result was caused by the low affinity of the peptide antisera for large-T, and Western blotting experiments confirmed that the peptide antisera react directly with immobilized, monomeric large-T but not with nonviral T antigen. Immunoprecipitation assays to detect large-T:nonviral T antigen complexes bound specifically to fragments of SV40 DNA showed that under conditions of apparent antibody excess, DNA still bound to the complex.
Involvement of retrovirus reverse transcriptase-associated RNase H in the initiation of strong-stop (+) DNA synthesis and the generation of the long terminal repeat
Authors: Resnick R , Omer C A , Faras A J
Published: January 2020

Reconstructed enzymatic reactions containing purified reverse transcriptase and defined analog substrates which mimic those purported to be natural substances for reverse transcription in vivo were employed to delineate the mechanism of strong-stop (+) DNA synthesis. Our analysis of this system has indicated that strong-stop (+) DNA synthesis is initiated after the introduction of a nick in the viral RNA genome between a polypurine sequence and an inverted repeat that represents the end of the long terminal repeat. Since inhibitors of the reverse transcriptase-associated RNase H activity prevent the introduction of the nick and the synthesis of strong-stop (+) DNA synthesis, it appears that this particular reverse transcriptase-associated enzymatic activity is responsible for the initiation of strong-stop (+) DNA. Our data also indicated that the RNase H activity creates a second nick in the viral RNA genome 11 nucleotides upstream from the strong-stop (+) DNA initiation site since the strong-stop (+) DNA synthesized in these reactions is covalently linked to an oligoribonucleotide 11 residues in length. Nucleotide sequence analysis of the oligoribonucleotide primer molecule indicated that a single homogenous oligomer was associated with strong-stop (+) DNA exhibiting the sequence rArGrGrGrArGrGrGrGrGrA. The oligoribonucleotide primer can be removed from strong-stop (+) DNA by the purified reverse transcriptase, which creates a nick at the junction between the primer and strong-stop (+) DNA. These data demonstrate that the initiation of strong-stop (+) DNA synthesis is mediated by RNase H and that the site of initiation is exactly at the end of the long terminal repeat, providing evidence for yet another function of this reverse transcriptase-associated enzymatic activity in the synthesis of retrovirus DNA.
Identification of the DNA sequences encoding the large subunit of the mRNA-capping enzyme of vaccinia virus
Authors: Morgan J R , Cohen L K , Roberts B E
Published: January 2020

The DNA sequences encoding the large subunit of the mRNA-capping enzyme of vaccinia virus were located on the viral genome. The formation of an enzyme-guanylate covalent intermediate labeled with [alpha-32P]GTP allowed the identification of the large subunit of the capping enzyme and was used to monitor the appearance of the enzyme during the infectious cycle. This assay confirmed that after vaccinia infection, a novel 84,000-molecular-weight polypeptide corresponding to the large subunit was rapidly synthesized before viral DNA replication. Hybrid-selected cell-free translation of early viral mRNA established that vaccinia virus encoded a polypeptide identical in molecular weight with the 32P-labeled 84,000-molecular-weight polypeptide found in vaccinia virions. Like the authentic capping enzyme, this virus-encoded cell-free translation product bound specifically to DNA-cellulose. A comparison of the partial proteolytic digestion fragments generated by V8 protease, chymotrypsin, and trypsin demonstrated that the 32P-labeled large subunit and the [35S]methionine-labeled cell-free translation product were identical. The mRNA encoding the large subunit of the capping enzyme was located 3.1 kilobase pairs to the left of the HindIII D restriction fragment of the vaccinia genome. Furthermore, the mRNA was determined to be 3.0 kilobases in size, and its 5' and 3' termini were precisely located by S1 nuclease analysis.
Antigenic differences between two viruses, isolated in Japan and Korea, that cause hemorrhagic fever with renal syndrome
Authors: Yamanishi K , Dantas J R , Takahashi M , Yamanouchi T , Domae K , Takahashi Y , Tanishita O
Published: January 2020

Hantaan virus (HV) 76-118, isolated from Apodemus agrarius coreae in Korea, and hemorrhagic fever with renal syndrome (HFRS) virus B-1, isolated from a rat in Japan, were examined for polypeptide compositions and for differences in immune responses in rats. In immunoprecipitation experiments, a major polypeptide of ca. 50 kilodaltons (K) was detected with antisera against HV 76-118 in cell extracts from Vero E6 cells infected with HFRS virus B-1, whereas three major polypeptides of 74 K (glycosylated), 57 K (glycosylated), and 50 K were detected with antisera against HFRS virus B-1. On the other hand, two polypeptides with molecular weights of 55,000 (glycosylated) and 50,000 were detected with either antiserum in cell extracts infected with HV 76-118. In neutralizing antibody tests with antisera prepared in rats, a remarkable difference in antibody titer (5 to 30 times higher to the homologous virus than to the heterologous virus) was observed between the two viruses. However, this difference was not so marked (1 to 4 times higher to the homologous virus) in the immunofluorescent antibody test. Twenty hybrid cell lines producing mouse monoclonal antibodies against HV 76-118 were isolated by fusion of spleen cells from BALB/c mice immunized against HV (strain 76-118) with mouse myeloma cells. The specificity of these monoclonal antibodies was established by immunofluorescent antibody, neutralizing antibody, and fluorescent antibody to membrane antigen tests and by analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These hybrid cell lines were classified into three groups based principally on the IF staining pattern of the HV-infected cells: (i) antibodies which showed a discrete patch pattern in the cytoplasm by the immunofluorescent antibody test, reacting with the membrane antigen of infected cells and immunoprecipitating a 55-K glycoprotein from HV 76-118-infected cell lysates and a 57-K glycoprotein from the heterologous (strain B-1) HFRS virus-infected cell lysates. Among these, depending on the neutralizing antibody activity and the reaction with the heterologous antigen, three subgroups designated I-A, I-B, and I-C were established; (ii) antibodies which showed large granular dots in the cytoplasm, neither having neutralizing antibody activity nor immunoprecipitating any antigen; (iii) antibodies which showed defined granular dots throughout the cytoplasm, reacting with a 50-K polypeptide of both virus strains. These antibodies also classified into two subgroups based on the reactivity with the B-1 strain.(ABSTRACT TRUNCATED AT 400 WORDS)