Purification of DNA complementary to the env gene of avian sarcoma virus and analysis of relationships among the env genes of avian leukosis-sarcoma viruses.
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The env gene of avian leukosis-sarcoma viruses encodes a glycoprotein that determines the host range and surface antigenicitiy of virions. We have purified radioactive DNA (cDNAgp) complementary to at least a portion of the env gene for viral subgroups A and C; complementary DNA was synthesized with purified virions of wild-type avian sarcoma virus, and RNA from a mutant with a deletion in env was used to select DNA specific to env by molecular hybridization. The genetic complexity of cDNAgp for subgroup A (ca. 2,000 nucleotides) was sufficient to represent the entire deletion and most or all of the env cistron. The deletions in env in two independently isolated strains of virus (Bryan and rdNY8SR) overlap, and cDNAgp represents nucleotide sequences common to both deletions. By contrast, we could detect no overlap between deletions in env and deletions in the adjacent viral gene src. Laboratory stocks of viral subgroups A, B, C, D and E do not contain detectable amounts of env deletions when tested by molecular hybridization; hence, segregation of deletions in env is a less frequent event that the segregation of deletions in the viral transforming gene src (Vogt, 1971). We found extensive homology among the nucleotide sequences encoding the env genes of virus strains indigenous to chickens (subgroups A, B, C, D, and E) although subgorups B, D and E appear to differ slightly from subgroups A and C at the env locus. By contrast, viruses obtained from pheasant cells (subgroups F and G) have env genes with little or no relationship to env genes of chikcen viruses. According to available data, viruses of subgroup F arose by recombination between an avarian sarcoma virus and viral genes in the genome of ring-necked pheasants, whereas subgroup G viruses may be entirely endogenous to golden pheasants. (+info)
Infectious DNA of spleen necrosis virus is integrated at a single site in the DNA of chronically infected chicken fibroblasts.
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The infectious DNAs of a number of avian leukosis-sarcoma and reticuloendotheliosis viruses were digested with six nucleotide-specific restriction endonucleases, and the digests were tested for infectivity. All of the enzymes inactivated the viral infectivities except for EcoRI, which did not inactivate the infectivity of the DNA of two of the reticuloendotheliosis viruses, spleen necrosis and chick syncytial viruses. The infectious DNA of spleen necrosis virus after digestion with EcoRI had a buoyant density in CsCl solution greater than the density of the high-molecular-weight infectious viral DNA. The infectious EcoRI-digested spleen necrosis virus DNA from chronically infected chicken cells was uniform in size, 10 megadaltons, which indicated a single site of integration. The infectious EcoRI-digested spleen necrosis virus DNA from acutely infected cells was heterogeneous in size, ranging from 8-14 megadaltons, which indicated multiple sites of integration. These results are consistent with the hypothesis that cells that integrate infectious spleen necrosis virus DNA at a single site survive and multiply, whereas cells that integrate infectious viral DNA at additional sites either die or selectively lose or inactivate the DNA in the additional sites. (+info)
Surface morphology of normal and neoplastic rat cells.
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Nontumorigenic rat cells and their tumorigenic counterparts were studied with scanning electron microscopy under controlled conditions in vitro and with transmission electron microscopy after replantation in vivo to discern if external morphology reflected the cell's neoplastic state or the etiology of transformation. Interphase cells in six of seven nontumorigenic lines were flat and monolayered under confluent conditions and exhibited smooth, nonactive cell surfaces. A nontumorigenic cell line morphologically transformed with human adenovirus-2 consisted of spherical cells with blebbed surfaces. Cells from six tumorigenic lines transformed with avian sarcoma virus had highly active surfaces with many surface projections. Cells from two chemical carcinogen-transformed rat embryo lines were flat with no surface projections in subconfluent culture and rounded with only a few microvilli at high densities, but cells from a sarcoma chemically induced in an adult rat were villous. When villous cells were syngeneically replanted in vivo, they lost most microvilli. The external morphology of cells was influenced by a number of factors simultaneously, with no universal pattern associated with tumorigenic capacity or transforming agent. (+info)
Type C viral gag gene expression in chicken embryo fibroblasts and avian sarcoma virus-transformed mammalian cells.
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Sensitive radioimmunoassays were developed for avian type C viral gag gene-coded proteins. These assays were used to examine the restriction to virus production by avian embryo cells and mammalian cells transformed by avian sarcoma viruses. The results indicate that although a high-molecular-weight primary translational product of the gag gene is expressed, its cleavage and processing are incomplete. Furthermore, analysis of intermediate cleavage products provided information regarding the order of sequences coding for the individual viral proteins within the avian type C viral gag gene. (+info)
Avian sarcoma and leukemia virus (ASLV) integration in vitro: mutation or deletion of integrase (IN) recognition sequences does not prevent but only reduces the efficiency and accuracy of DNA integration.
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A subpopulation of the avian erythroblastosis virus v-erbA protein, a member of the nuclear hormone receptor family, is glycosylated.
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The v-erbA oncogene of avian erythroblastosis virus is derived from a cellular gene for a thyroid hormone (T4/T3 thyronine) receptor and encodes a DNA-binding protein found principally in the nucleus of the infected cell. I report here that a subpopulation of the v-erbA protein is glycosylated. The v-erbA protein, therefore, is another member of the newly recognized family of eucaryotic transcription factors and related polypeptides which are glycoproteins. (+info)
Sequence-specific DNA binding by the v-erbA oncogene protein of avian erythroblastosis virus.
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The v-erbA oncogene, a transduced copy of a thyroid hormone receptor, plays an important role in establishment of the transformed cell phenotype induced by avian erythroblastosis virus. The ability of thyroid hormone receptors to bind to specific sites on chromatin and to thereby modify the expression of adjacent target genes is a crucial element in their mechanism of action in the normal cell. The v-erbA protein also bound at high affinity to a set of DNA fragments recognized by the rat thyroid hormone receptor, but the relative affinity of the v-erbA protein for the different binding sites was distinct from that previously reported for the thyroid hormone receptors. (+info)
The transforming gene product of avian erythroblastosis virus, v-erbB, is derived from the epidermal growth factor (EGF) receptor but has lost its extracellular ligand-binding domain and was mutated in its cytoplasmic portion, which is thought to be responsible for biological signal generation. We have repaired the deletion of extracellular EGF-binding sequences and investigated the functional consequences of cytoplasmic erbB mutations. Within the resulting EGF receptors, the autophosphorylation activities of the cytoplasmic domains of v-erbB-H and v-erbB-ES4 were fully ligand dependent in intact cells. However, the mitogenic and transforming signaling activities of an EGF receptor carrying v-erbB-ES4 (but not v-erbB-H) cytoplasmic sequences remained ligand independent, whereas those of a receptor with a v-erbB-H cytoplasmic domain were regulated by EGF or transforming growth factor alpha. Thus, structural alterations in the cytoplasmic domain of growth factor receptor tyrosine kinases may induce constitutive signaling activity without autophosphorylation. These findings provide new insight into the mechanism of receptor-mediated signal transduction and suggest a novel alternative for subversion of cellular control mechanisms and proto-oncogene activation. (+info)