Robert R. Friis

Induction of avian tumor viruses in normal cells by physical and chemical carcinogens

Abstract Avian tumor viruses were induced in normal chicken cells treated with ionizing radiations or chemical carcinogens and mutagens. The induced leukosis viruses possess a buoyant density, DNA polymerase, polypeptides, and 70S RNA typical of avian tumor viruses. Induced leukosis viruses act as helper agents for the defective Bryan high titer strain of Rous sarcoma virus and with one exception belong to subgroup E as judged by host-range, interference, and neutralization patterns. Induction of leukosis viruses was successful in chicken cells lacking the “natural” group specific (gs) antigen of the avian tumor viruses as well as in cells carrying this antigen. This observation indicates that the viral genome is present in gs+ and in gs− cells. Therefore, the chromosomal locus which controls the presence of natural gs antigen in chicken cells does not represent the viral genome itself but regulates its expression in normal cells. These findings have implications for the origin of RNA tumor viruses and for theories of carcinogenesis.

An avian leukosis virus related to RSV(O): Properties and evidence for helper activity

Abstract The ability of Rous sarcoma virus type O [RSV(O)] to reproduce depends on the host cell. Most single infections in type C A chicken cells yield infectious progeny, whereas cells of Japanese quail embryos produce noninfectious progeny under such conditions. Quail embryos do not contain the congenitally transmitted group-specific ( gs ) antigen of the avian leukosis and sarcoma complex; all type C A chick embryos harbor this antigen. In addition, cultures from some C A embryos release a non-transforming avian RNA tumor virus which shares surface antigens with RSV(O) and which can act as helper virus for RSV(O) in quail cells. This agent is referred to as Rous-associated virus type O (RAV-O). RSV produced with the aid of RAV-O in quail cells has the envelope properties of parental RSV(O). Our observations suggest that RAV-O plays an essential role in the replication of RSV(O) in gs negative cells and that it is also involved in the reproduction of RSV(O) in gs positive cells. RAV-O differs from the common type of helper virus by its greater physical instability, its poor ability to interfere with related viruses, and its dependence on RSV(O) in the infection of quail cells.

Conditional lethal mutants of avian sarcoma viruses: I. Physiology of ts 75 and ts 149☆

Abstract Ts 75 and ts 149 are two temperature-sensitive mutants of avian sarcoma virus B77 which fail to reproduce and to induce neoplastic transformation at 41°. Both mutants are indistinguishable from wild type in somatic properties of the virion: host range, type-specific antigenicity, and rate of inactivation at 41°. The temperature-sensitive step of ts 75 occurs late in the infectious cycle allowing the synthesis of group-specific antigen in increased amounts and of viral RNA under nonpermissive conditions. Ts 149 has an early temperature sensitive phase and does not produce group-specific antigen at 41°. The maintenance of some neoplastic properties in cells transformed by either ts 75 or ts 149 is continuously dependent on a temperature sensitive viral function: Shift of transformed cells to 41° results in disappearance of neoplastic traits. Double infection of cells at 41° with wild-type avian sarcoma virus and ts 75 or ts 149 results in the rescue of 3 markers derived from the mutant virus: temperature sensitivity, host range, and morphology of the transformed cell. Rescue of ts 75 or ts 149 with wild-type avian leukosis viruses at 41° has not been accomplished.

RNA tumor viruses of pheasants: Characterization of avian leukosis subgroups F and G

Abstract Endogenous leukosis-like viruses of ring-necked pheasants ( Phasianus colchicus ) and golden pheasants ( Chrysolophus pictus ) have been isolated and characterized. The majority of the normal pheasant embryo cultures contain helper activity for the defective Bryan high titer strain of Rous sarcoma virus. The Rous sarcoma pseudotypes produced with endogenous helper activity from ring-necked pheasants belong to subgroup F. The pseudotypes from golden pheasant cells constitute subgroup G. Subgroup F and G pseudotypes can infect all known genetic types of chicken fibroblasts as well as pheasant and Japanese quail cells, but do not plate on goose cells. Duck cells are resistant to subgroup G but not to F. The subgroup F and G helper viruses isolated from Rous sarcoma viral pseudotypes show interference with their homologous subgroup. RAV-61, a standard of subgroup F, interferes with pseudotypes produced with endogenous helper activity from ring-necked pheasant cells but not with subgroup G pseudotypes. Subgroups F and G do not cross-react with subgroup A to E in neutralization tests. Some normal ring-necked pheasant sera have anti-F activity. Subgroup F and probably also G leukosis-like viruses can undergo genetic recombination with nondefective avian sarcoma viruses.

The reproductive and cell-transforming capacities of avian sarcoma virus B77: Inactivation with UV light

Abstract Avian sarcoma virus strain Bratislava 77 (B77) was irradiated with ultraviolet light, and virus survivors were studied for their ability to transform cells and to reproduce. Two classes of radiation-damaged particles were found. One can still transform but fails to reproduce; the other reproduces but cannot transform. The first class of particles also fails to induce the synthesis of noninfectious virus. It cannot be rescued by superinfection of the transformed cells with an avian leukosis virus, and it is unable to maintain the transformed state during prolonged culture. The second class of particles is antigenically indistinguishable from B77, interferes with avian sarcoma viruses of subgroup C and produces pseudotypes with RSV(0).

Determination of the defective function in two mutants of Rous sarcoma virus

Abstract LA 335 and LA 337 are two temperature-sensitive mutants of the Prague strain of Rous sarcoma virus (subgroup C) which are unable to initiate infection at the nonpermissive temperature (41°). The virions of LA 335 and LA 337 are shown to possess a reverse transcriptase activity which is more heat labile than that of the wild type. Genetic studies are described which indicate that the inability of LA 335 or LA 337 to initiate infection at 41° is due to this temperature-sensitive enzyme. The genetic markers for reverse transcriptase of LA 335 and LA 337 undergo high-frequency recombinations with markers for host range, and possibly transformation.

A study of the relationship of reticuloendotheliosis virus to the avion leukosis-sarcoma complex of viruses

Abstract Reticuloendotheliosis virus (REV) was compared on biological and biochemical grounds with members of the avian leukosis-sarcoma complex of viruses (ALSV). Stocks of REV contained no virus which produced interference with any subgroup of ALSV. REV was unable to complement various strains of Rous sarcoma virus (RSV) as measured by the absence of broadening of host range, “helper” virus activity with the defective Bryan high titer strain of RSV, or the induction of release of RAV-60 from chick embryo cells containing the chick helper factor. Hence, no biological interaction has been detected between REV and the avian leukosis-sarcoma complex of viruses. The RNA structure of the REV was studied using glycerol velocity gradients and polyacrylamide gel electrophoresis. The RNA of REV was distinctly like that of RAV-49, a subgroup C avian leukosis virus. The native RNA of REV recovered after phenol extraction cosedimented with the RNA of RAV-49, and after heat denaturation, both RNAs showed the reduced sedimentation velocity and increased rate of migration in polyacrylamide gel which is characteristic of the tumor virus RNAs. Proteins of REV were examined on SDS discontinuous gel electrophoresis. Five major proteins were detected, two of which were glycoproteins. Coelectrophoresis of REV with avian sarcoma virus B 77 showed no polypeptides that were identical in electrophoretic mobilities. In addition, lactoperoxidase catalyzed iodination was employed to selectively label the surface proteins of REV. Two surface proteins were detected; these corresponded to the two viral glycoproteins.

A replication defective mutant of Rous sarcoma virus which fails to make a functional reverse transcriptase

Abstract ts 672 is a replication defective temperature sensitive mutant of Rous sarcoma virus which produces large yields of noninfectious viral particles (NI 672) during growth in cells at the nonpermissive temperature. This study has been directed to identifying the temperature sensitive function of ts 672 by analysis of the NI 672 particles and by comparing this mutation to other known mutations in functions affecting replication. The only structural defect observed with NI 672 was the absence of the virion-associated RNA dependent DNA polymerase activity characteristic of all infectious RNA tumor viruses. Although there are other viral mutants which exhibit an altered polymerase activity, ts 672 is unique in that the DNA polymerase is only temperature sensitive prior to or during the assembly of the virus particle. The DNA polymerase activity of the ts 672 virions produced at the permissive temperature is just as temperature stable as the wild type enzyme. Mixed infection tests performed with ts 672 and with two other DNA polymerase mutants have failed to show a complementation, thus supporting the idea that ts 672 is defective in the DNA polymerase function. Recombination studies with ts 672 and an avian leukosis virus have shown that there is a very high degree of genetic linkage between the temperature sensitive DNA polymerase function of ts 672 and the determinants for the host range of these viruses, the viral envelope proteins.

Abortive infection of Japanese quail cells with avian sarcoma viruses

Abstract The replication of Prague strains of Rous sarcoma virus (PR RSV) and avian sarcoma virus B77 in Japanese quail cells is abortive in a quantitative sense. Although efficient infection of quail cells, as evidenced by transformation, was obtained with these viruses, the yield of progeny was suppressed by 102-fold or more relative to the chick cell host, and for many clones of transformed quail cells tested, no yield could be detected. Infectious center assays, however, indicated that each transformed cell produces a small yield of progeny. The yield of progeny can be enhanced by prior infection of quail cells with avian leukosis viruses or rescued by superinfection with avian sarcoma viruses inoculated at higher multiplicity than the initial infecting virus. B77-transformed, abortively infected quail cells synthesize viral envelope proteins as evidenced by the complementation of the defective Bryan high titer strain of Rous sarcoma virus (BH RSV); the dominant progeny species of such complementation is, however, genotypically BH RSV. The possibility that the replication of the B77 genome in quail cells is inefficient is suggested.

Inhibition of avion sarcoma virus replication by glucosamine

Abstract The effect of glucosamine on the replication of avian sarcoma viruses has been studied. Six hours after addition of 20 μmoles/ml of glucosamine to the culture medium replication of infectious virus was inhibited by 99.9%. The production of viral particles detectable by physical techniques ceased in parallel with the synthesis of focus forming virus. Immunofluorescence studies indicate that the glucosamine block affects the production of viral envelope antigens. Accumulation of such antigens in the cytoplasm or at the cell surface was not observed in glucosamine-treated cultures, although the accumulation of a nonglycosylated polypeptide precursor cannot be excluded. Synthesis of group specific ( gs1 ) protein was also inhibited by glucosamine, but to a lesser degree than that of viral envelope antigens.