Avian sarcoma virus-transformed quail clones defective in the production of focus-forming virus

The cellular sites of integration of avian sarcoma virus (ASV) have been examined in clones of duck embryo cells infected with the Bratislava 77 strain of ASV using restriction endonuclease digestion, agarose gel electrophoresis, Southern blotting, and hybridization with labeled ASV complementary DNA probes. DNA prepared from 11 clones of duck embryo cells infected with the Bratislava 77 strain of ASV was digested with the restriction enzymes HpaI, which cleaves once within the viral genome, and Hind III, which cleaves twice within the viral genome, and the virus-cell DNA juncture fragments were resolved by agarose gel electrophoresis. Analysis of the virus-cell junctures present in individual ASVinfected duck embryo clones revealed that all clones contain at least one copy of nondefective proviral DNA with some clones containing as many as 5 to 6 copies of proviral DNA. A comparison of the virus-cell juncture fragments present in different ASV-infected clones showed that each clone contains a unique set of virus-cell junctures. These data suggest that ASV DNA can integrate at multiple sites within the duck embryo cell genome and that these sites appear to be different as defined by digestion with the restriction enzymes HpaI and HindIII.

Section: Resultsmentioning

confidence: 86%

“…In two clones, we observed proviral DNA containing internal deletions of 1.0 Md (clone 9) and 2.2 Md (clones 4 and 9). Although these deletions have not been mapped within the proviral DNA, a number of examples of deletions occurring in viral sequences other than the src gene were recently described (6,10,11). Fig.…”

Section: Asv-infected De Cells (Lane U)mentioning

confidence: 99%

Analysis of cellular integration sites in avian sarcoma virus infected duck embryo cells

Gilmer¹,

Parsons²

1979

“…Since the transformation of mammalian cells by RSV only requires the transforming gene of the virus to be expressed and does not depend upon the ability of a virus to replicate, the frequency of generation of nonvirogenic cells may reflect the proportion of replication-defective viruses in a particular virus stock. Previous reports that nonproducer avian cells are generated after single infection by nondefective strains of RSV would tend to support this conclusion (19,23). It is also possible that there is something peculiar about the establishment of infection in nonpermissive mammalian cells that generates a high proportion of defective transforming proviruses.…”

Section: Discussionmentioning

confidence: 85%

Cell fusion for genetic analysis of two nonconditional Rous sarcoma virus replication mutants

Steimer¹,

Boettiger²

1979

Procedures for characterizing replication-defective viruses in nonpermissive mammalian cells were developed and applied to three nonvirogenic Rous sarcoma virus (RSV)-transformed mammalian cell lines-B4, a line of Bryan virus-transformed hamster cells, and two SRD-RSV transformed rat cell lines, LR3/1 and LR3/2. Cell fusion was used to study virus complementation. The three cell lines (i) fused with helper virus-infected chicken cells and the host range of the rescued virus examined, (ii) tested for complementation by fusion with chicken cells exhibiting various patterns of endogenous virus expression, (iii) fused with chicken cells infected with the temperature-sensitive replication mutant LA334 and assayed for complementation at permissive and nonpermissive temperatures, and (iv) tested for complementation of defective viruses in other RSV-transformed mammalian cell lines by fusing pairs of nonvirogenic cell lines and permissive chicken cells. Based upon these complementation studies, we concluded that B4 virus is defective only in the env gene, LR3/1 virus is an absolute mutant in the gag and/or pol genes, and LR3/2 virus is a leaky env mutant. Clones of LR3/1 and LR3/2 virus-infected chicken cells were established, and the results obtained from the characterization of these viruses in permissive avian cells substantiates the conclusions reached in the fusion-rescue studies.

“…The genomes of a few of these avian and mammalian retrovirus nonconditional mutants have been analyzed to identify the types of lesions involved (4,15,19,24). For the mammalian system the analysis of the genomes of nonconditional mutants has revealed that a few of these mutants have smaller virion RNAs, indicating that deletions are responsible for their defectiveness.…”

mentioning

confidence: 99%

“…Other types of avian nonconditional mutants include those with insertions (19, 20a) or substitutions (9) in the wildtype genome which account for the mutant phenotype. Other genomes, however, such as a few Rous sarcoma virus proviruses of nonproducer quail cells, resemble wild-type virus at least in size and by restriction endonuclease analysis (19).…”

mentioning

confidence: 99%

Four Moloney murine leukemia virus-infected rat cell clones producing replication-defective particles: protein and nucleic acid analyses

Yoshimura¹,

Yamamura²

1981

Four cloned rat cell lines (NX-1 to -4) infected with Moloney murine leukemia virus and defective in virus replication were found to be all different by viral protein and nucleic acid analyses. All four clones produced noninfectious particles and, except for NX-2, at about the same level as wild type. Compared with wild-type virions these defective particles contained larger amounts of gag precursor proteins and very little or no p30 or p15. Analysis of intracellular precursor proteins revealed that NX-2 to -4 synthesized normal Pr65gag, whereas NX-1 produced a slightly smaller precursor. Both NX-1 and NX-4 synthesized an intracellular polyprotein with a size similar to that of wild-type Pr180 gag-pol. Restriction endonuclease analysis of NX-1 to -4 cellular DNA showed that each clone contained a single integrated provirus which possessed large terminal repeat sequences at both the 5' and 3' ends. The proviruses of NX-1 to -3 appeared normal by restriction endonuclease analysis, but NX-4 provirus had a deletion of 1,700 base pairs comprising part of the polymerase region. The noninfectious particles produced by all four clones packaged Moloney viral RNAs and rat RNAs of two different sizes.