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B Vennström

Publications and source records attributed to B Vennström.

At least 91 records · Page 5Linked to original sources
PubMed1986

Replication of the bovine papillomavirus type 1 genome; antisense transcripts prevent episomal replication.

A subgenomic fragment, representing 69% of the bovine papillomavirus type 1 (BPV-1) genome, has the capacity to transform mouse C127 cells in vitro and to replicate episomally in these cells. In the present study we have cloned this BPV-1 fragment between two retrovirus-derived long terminal repeats (LTRs) in the two possible orientations. The constructs were designated pMR and pML. The pMR construct contained the BPV-1 genome in the same transcriptional orientation as that of the LTRs whereas the pML construct contained the BPV-1 fragment in the opposite orientation. Both types of construct were capable of transforming mouse C127 cells with approximately the same efficiency. Analysis of the intracellular DNA from cells transformed with pMR and pML revealed that both cell types contained a large number of viral DNA copies. However, whereas the pMR-transformed cell clones contained episomally replicating BPV-1 DNA, the pML-transformed cell clones all contained integrated viral genomes. Analysis of RNA from the transformed cells revealed that the pML-transformed cells, unlike the pMR-transformed cells, contained large amounts of antisense BPV-1 transcripts which presumably interfere with the mRNAs that are required for episomal BPV-1 replication. The results are of importance for the future design of BPV-1 vector constructs.

Bovine papillomavirus 1↗
PubMed1986

Rous-associated virus 1-induced erythroleukemic cells exhibit a weakly transformed phenotype in vitro and release c-erbB-containing retroviruses unable to transform fibroblasts.

Avian leukosis viruses induce erythroblastosis in chicks by integrating into the c-erbB gene and thus activating c-erbB transcription. We characterized Rous-associated virus 1-induced leukemic erythroblasts in vitro and showed that they mostly resemble erythropoietin-independent but otherwise normal erythroid progenitors. Some leukemic cells, however, were able to both differentiate and proliferate extensively in vitro. All 14 leukemias studied expressed high levels of erbB-related proteins that were 5 to 10 kilodaltons larger but otherwise very similar to the gp74erbB protein of avian erythroblastosis virus ES4 with respect to biosynthesis, glycosylation, and cell surface expression. Two leukemias contained and released retroviruses that transduced erbB. Chicken embryo fibroblasts fully infected with these viruses expressed high levels of erbB RNA and protein but retained a normal phenotype. Our results suggest that certain forms of c-erbB, activated by long terminal repeat insertion or viral transduction, are capable of inducing erythroleukemia but unable to transform fibroblasts.

Animals↗
PubMed1985

How do retroviral oncogenes induce transformation in avian erythroid cells?

The v-erb B oncogene, as well as other oncogenes of the src-gene family transform immature erythroid cells from chick bone marrow in vivo and in vitro. The erb B-transformed erythroid cells differ from normal late erythroid precursors (CFU-E) in that they have acquired the capacity to undergo self-renewal as well as to differentiate terminally. They also do not require the normal erythroid differentiation hormone, erythropoietin, for either process. Cooperation of v-erb B with a second oncogene, v-erb A, results in a differentiation arrest of the transformed cells, which now only use the self-renewal pathway. Studies with conditional and non-conditional mutants in both v-erb B and v-erb A will be presented to elucidate further how the transforming proteins encoded by these oncogenes, gp74erb B and gp75gag-erb A, affect the differentiation programme of the infected erythroid precursor with the outcome of hormone-independent leukaemic cells arrested at an early stage of erythroid differentiation.

Animals↗
PubMed1985

Transfer of genes into embryonal carcinoma cells by retrovirus infection: efficient expression from an internal promoter.

The expression of retroviral genomes after infection of embryonal carcinoma (EC) cells is usually restricted. We have constructed a retroviral vector which has the potential to express efficiently in EC cells an inserted sequence, the selectable neomycin (neo) gene, from an internal thymidine kinase promoter. A second retrovirus vector contained in addition a v-myc oncogene, which in NIH 3T3 cells could be expressed from the viral long terminal repeat (LTR) through a subgenomic mRNA. 10-100% of the EC cells infected with recombinant virus gave rise to neo-resistant colonies following drug selection. The neo gene was expressed in both the non-permissive EC and the permissive NIH 3T3 cells at similar levels, and transcription of neo-specific RNA was initiated at the proper site in the thymidine kinase promoter. In contrast, v-myc mRNA was produced only in the NIH 3T3 cells and not in EC cells, even after induction of differentiation. Thus, genes inserted into retrovirus constructs can easily be introduced into EC cells by virus infection, even without drug selection, and they can be efficiently expressed when transcribed from promoters other than the viral LTRs.

Animals↗
PubMed1984

Transformation of mammalian fibroblasts and macrophages in vitro by a murine retrovirus encoding an avian v-myc oncogene.

A murine retrovirus which expresses the avain v-myc OK10 oncogene was constructed. The virus, denoted MMCV, readily transforms fibroblasts of established lines, such as mouse NIH/3T3 and rat 208F cells, to anchorage-independent growth in agarose. The virus also transforms primary mouse cells: (i) virus-infected macrophages are induced to form large colonies in semi-solid media, and can easily be expanded into mass cultures; (ii) MMCV-infected fibroblastic cells from mouse limb buds undergo morphological transformation and grow in semi-solid medium. MMCV thus transforms both mouse fibroblastic cells and macrophages in vitro, in a fashion similar to the v-myc-containing avian viruses in chicken cells. The possibility of introducing a transforming myc gene into mammalian cells by virus infection provides a novel approach for studying the mechanism of myc transformation in cells from many lineages.

Animals↗
PubMed1983

Isolation and characterization of multiple human genes homologous to the oncogenes of avian erythroblastosis virus.

Human DNA sequences complementary to the oncogenes v-erbA and v-erbB of avian erythroblastosis virus have been isolated from a genomic DNA library. Two clones, lambda he-A1 and lambda he-A2, were related to the erbA gene and one to the erbB gene (lambda he-B). The two erbA genes were only distantly related to each other as judged from hybridization analysis. Furthermore, human chromosomal DNA appears to contain one or two additional genes analogous to the lambda he-A2 sequence, whereas the mouse genome contained only two genes complementary to lambda he-A1 and lambda he-A2, respectively. Polyadenylated RNA species, 5.0 kb in size, were found in the human HeLa and the human hematopoietic K562 cell lines, suggesting that at least some of the erb-related genes are active and do not represent pseudogenes. Taken together, the data demonstrate that two distantly related classes of erbA genes exist in human and mouse DNA, and that multiple copies of genes belonging to one of these two classes exist in the human genome.

Alpharetrovirus↗
PubMed1983

Transforming capacities of avian erythroblastosis virus mutants deleted in the erbA or erbB oncogenes.

Mutants of avian erythroblastosis virus (AEV) were constructed by deleting large nucleotide segments in each of the viral oncogenes termed v-erbA and v-erbB. Mutants in erbA (erbA -B +) retained the ability to transform fibroblasts in vitro, and these cells exhibited most of the transformation characteristics that typify wild-type AEV-transformed fibroblasts. In addition, the mutants induced small erythroid colonies upon infection of bone marrow cells in culture. Chickens inoculated with erbA -B + virus or with erbA -B +-transformed cells developed sarcomas or atypical erythroid leukemias. The erythroid cells transformed in vivo or in vitro by the erbA -B + viruses appeared not to be as tightly blocked in differentiation as wild-type transformed cells. In contrast, fibroblasts infected with the erbA +B - mutant resembled normal cells in all transformation parameters tested, and no bone marrow cell transformation was observed with the mutant. The results indicate that the main transforming properties of AEV are encoded in erbB and that its effects are enhanced by erbA.

Alpharetrovirus↗
PubMed1983

Avian acute leukemia virus OK 10: analysis of its myc oncogene by molecular cloning.

Several DNAs representing the genome of the avian acute leukemia virus OK 10 were isolated by molecular cloning from a transformed quail cell line, 9C, which contained at least six OK 10 proviruses. Recombinant lambda phages harboring the OK 10 genome and additional flanking cellular DNA sequences were studied by restriction endonuclease mapping and hybridization to viral cDNA probes. Six of the clones represented complete proviruses with similar, if not identical, viral sequences integrated at different positions in the host DNA. The organization of the OK 10 genome was determined by electron-microscopic analysis of heteroduplexes formed between the cloned OK 10 DNA and DNAs representing the c-myc gene and the genomes of two other avian retroviruses, Rous-associated virus-1 and MC29. The results indicated that the OK 10 proviral DNA is about 7.5 kilobases in size with the following structure: 5'-LTR-gag-delta polmyc-delta env-LTR-3', where LTR indicates a long terminal repeat. The oncogene of OK 10, v-mycOK 10, forms a continuous DNA segment of around 1.7 kilobases between pol and env. It is similar in structure and length to the v-myc gene of MC29, as demonstrated by restriction endonuclease and heteroduplex analyses. Two of the OK 10 proviruses were tested in transfection experiments: both DNAs gave rise to virus with the transforming capacities of OK 10 when Rous-associated virus-1 was used to provide helper virus functions.

Animals↗
PubMed1982

Isolation and characterization of chicken DNA homologous to the two putative oncogenes of avian erythroblastosis virus.

The genome of avian erythroblastosis virus contains two independently expressed genetic loci (v-erbA and v-erbB) whose activities are probably responsible for oncogenesis by the virus. Both loci are closely related to nucleotide sequences found in the DNA and RNA of chickens and other vertebrates. We have isolated and characterized chicken DNA homologous to v-erbA and v-erbB. The two viral genes are represented by separate domains within chicken DNA (c-erbA and c-erbB), which are separated by a minimum of 12 kilobases (kb) of DNA and may not be linked at all. The nucleotide sequences shared by the viral and cellular erb loci are colinear, but the cellular loci are interrupted by multiple intervening sequences of various lengths. Polyribosomes prepared from normal chicken embryos contain two polyadenylated RNAs transcribed from c-erbA and two transcribed from c-erbB. The evident coding regions of these RNAs represent an unusually small fraction of the lengths of the RNAs, as if the 3' untranslated domains of the RNAs might be exceptionally large (3-11 kb). These findings indicate that the c-erb loci are normal vertebrate genes rather than genes of cryptic endogenous retroviruses, and that they may have a role in the metabolism of normal cells. It appears that the viral erb genes, like most other retrovirus oncogenes, have been copied from cellular genes. In the viral genome, the two genes are devoid of introns, but they remain independently expressed loci, and they remain colinear with the coding domains of their cellular progenitors.

Alpharetrovirus↗
PubMed1981

Molecular cloning of the avian myelocytomatosis virus genome and recovery of infectious virus by transfection of chicken cells.

The avian retrovirus myelocytomatosis virus 19 (MCV) possesses an interesting diversity of oncogenic potentials, but the virus has proven difficult to study because of its inability to replicate without the assistance of a helper virus. We have therefore isolated and amplified the genome of MCV by molecular cloning in a procaryotic vector. The topography of the cloned DNA was explored by the use of restriction endonucleases and radioactive complementary DNAs representing specific domains in avian retrovirus genomes. The cloned DNA appeared to be an authentic representation of the MCV genome: the size and genetic topography of the DNA were comparable to those of MCV, and transfection of the cloned DNA into chicken cells (in company with the DNA of a suitable helper virus) gave rise to virus with the genome and transforming potentials of MCV. The availability of cloned MCV DNA should facilitate a variety of genetic and biochemical manipulations directed at elucidating the mechanism of oncogenesis by MCV.

Animals↗
PubMed1980

Structure of genes for virus-associated RNAI and RNAII of adenovirus type 2.

A DNA sequence, 552 base pairs in length, encoding the two "virus-associated" (VA) RNAs of adenovirus type 2 is presented. Comparison of the oligonucleotide maps of VA RNAI and VA RNII with the established sequence permits identification of the genes for these RNAs. VA RNAI is 157-160 nucleotides long and VA RNAII 158-163 nucleotides long, depending on the exact length of their heterogeneous 3' end. The genes are separated by a spacer of about 98 nucleotides. The RNAs exhibit scattered regions of primary sequence homology and can adopt secondary structures which resemble each other closely in their configuration and stability. VA RNAII is also capable of assuming a different configuration that is energetically more favorable. The data suggest that the two RNA genes may have arisen by duplication of an ancestral gene and that the folding of the RNA chain may be of importance for the function of VA RNAs. Hypothetical RNA polymerase III recognition sequences and the coding potential of the region are discussed.

Adenoviruses, Human↗
PubMed1980

Molecular cloning of the avian erythroblastosis virus genome and recovery of oncogenic virus by transfection of chicken cells.

Avian erythroblastosis virus (AEV) causes erythroblastosis and sarcomas in birds and transforms both erythroblasts and fibroblasts to neoplastic phenotypes in culture. The viral genetic locus required for oncogenesis by AEV is at present poorly defined; moreover, we know very little of the mechanism of tumorigenesis by the virus. To facilitate further analysis of these problems, we used molecular cloning to isolate the genome of AEV as recombinant DNA in a procaryotic vector. The identity of the isolated DNA was verified by mapping with restriction endonucleases and by tests for biological activity. The circular form of unintegrated AEV DNA was purified from synchronously infected quail cells and cloned into the EcoRI site of lambda gtWES x B. A restriction endonuclease cleavage map was established. By hybridization with complementary DNA probes representing specific parts of avian retrovirus genomes, the restriction map of the cloned AEV DNAs was correlated with a genetic map. These data show that nucleotide sequences unique to AEV comprise at least 50% of the genome and are located approximately in the middle of the AEV genome. Our data confirm and extend previous descriptions of the AEV genome obtained by other procedures. We studied in detail two recombinant clones containing AEV DNA: the topography of the viral DNA in the two clones was virtually identical, except that one clone apparently contained two copies of the terminal redundancy that occurs in linear viral DNA isolated from infected cells; the other clone probably contained only one copy of the redundant sequence. To recover infectious virus from the cloned DNA, we developed a procedure for transfection that compensated for the defectiveness of AEV in replication. We accomplished this by ligating cloned AEV DNA to the cloned DNA of a retrovirus (Rous-associated virus type 1) whose genome could complement the deficiencies of AEV. Ligation of the two viral DNAs was facilitated by using a neutral fragment of DNA as linker between otherwise noncompatible termini. Cloned AEV DNA gave rise to infectious AEV capable of transforming fibroblasts and bone marrow cells in culture and of inducing both sarcomas and erythroleukemia in chickens. We conclude that the cloned DNAs represent the authentic genome of AEV undisturbed by the cloning procedure. Molecular cloning offers a powerful approach to the identification and characterization of retrovirus genomes.

Alpharetrovirus↗
PubMed1979

A DRB (5,6 dichloro-beta-D-ribofuranosylbenzimidazole)-resistant adenovirus mRNA.

5,6-Dichloro-beta-D ribofuranosyl benzimidazole (DRB) inhibits transcription from the major late adenovirus promoter, located on the r-strand at map position 16.3 on the viral genome. 100-500 nucleotides long RNA chains with capped 5'-termini are transcribed from this promoter in the presence of 70 muM DRB. Synthesis of the mRNA for polypeptide IX which is unspliced and maps on the r-strand between positions 9.7 and 10.9 appears, however, to be DRB-resistant although it is transcribed by polymerase II and capped. Translatable mRNA for polypeptide IX is synthesized in the presence of DRB while the mRNAs for the other viral structural proteins are not synthesized. This differential DRB-inhibition of polymerase II transcription suggests either that short mRNAs may escape inhibition or that unspliced mRNAs are insensitive to the drug.

Adenoviruses, Human↗
PubMed1978

Two initiation sites for adenovirus 5.5S RNA.

Adenovirus specific 5.5S RNA is heterogeneous at its 5' terminus. Complete pancreatic RNase digests of the RNA reveal a 5' terminal oligonucleotide (pp)pApGpCp in addition to the major 5' terminal (pp)pGpGpGpCp (1). Both 5' termini are detected early as well as late after adenovirus infection. In isolated nuclei, alpha-amanitin inhibits all 5.5S RNA transcription at a concentration of 200 microgram/ml, indicating that both initiation sites are recognized by RNA polymerase III.

Adenine Nucleotides↗
PubMed1978

Initiation of transcription in nuclei isolated from adenovirus infected cells.

Initiation of adenovirus transcription was analyzed by incubation of isolated nuclei from virus infected cells in the presence of beta-32P GTP or beta-32P ATP. Nucleotide analysis of RNA from nuclei incubated with beta-32P GTP shows that the label is incorporated exclusively into pppGp and ppGp. Under similar incubation conditions, the label from beta-32P ATP was incorporated primarily into the 5' phosphate of 5', 3' mononucleoside diphosphates, but label was also detected in pppAp, pppGp and in the 3' nucleoside monophosphates. Analysis of RNA, synthesized in the presence of different concentrations of alpha-amanitin, shows that only RNA polymerase III initiates virus specific transcription in isolated nuclei. The virus specific transcripts containing pppAp and pppGp in their 5' termini were identified as the 5.5S and 5.2S viral RNA species by hybridization and finger printing.

Adenoviruses, Human↗
PubMed1977

Fidelity of adenovirus RNA transcription in isolated HeLa cell nuclei.

An in vitro nuclear system from adenovirus type 2-infected cells was developed to study transcription of viral RNA. Nuclei isolated from adenovirus-infected HeLa cells late in the infectious cycle synthesized in vitro only RNA from the r-strand of adenovirus DNA. Around 15% of the virus-specific RNA in isolated nuclei was polyadenylated. Short pulse labeling of nascent RNA followbd by hybrization of size-fractionated RNA to specific restriction endonuclease fragments of the genome suggested that the origin(s) for transcription is located on the r-strand in the left 30% of the adenovirus 2 genome at late times in the infectious cycle. Pulse-chase experiments were used to estimate the elongation rate of adenovirus high-molecular-weight RNA in isolated nuclei. An elongation of a least six nucleotides per second was observed in vitro. Viral RNA synthesis in the vitro nuclei showed several similarities to the in vivo system late in the infectious cycle.

Adenoviruses, Human↗
PubMed1976

A new species of virus-coded low molecular weight RNA from cells infected with adenovirus type 2.

A virus-coded low molecular weight RNA (5.2S), which migrates slightly faster on polyacrylamide gels than the well characterized adenovirus-specific 5.5S RNA, has been isolated from cells infected with adenovirus type 2. Hybridization-competition experiments and RNA fingerprints indicate that the two virus-associated (VA) RNAs differ in their primary structures. The gene for 5.2S RNA is located to the right of the gene for 5.5S RNA, on the I strand of a DNA segment which extends between positions 30.3 and 32.2 on the map of adenovirus type 2 DNA. Both 5.5S and 5.2S RNA can be detected early after infection and also in the presence of cytosine-arabinoside or cycloheximide. After the onset of viral DNA replication, the synthesis of 5.2S RNA levels off, whereas 5.5S RNA is synthesized in increasing amounts. Both 5.2S and 5.5S RNAs are synthesized in isolated nuclei by an enzyme which resembles RNA polymerase III in its sensitivity to alpha-amanitin. In isolated nuclei, both RNA species are labeled with beta-32P-labeled GTP, which suggests that they are initiated at separate promotor sites.

Adenoviridae↗