Lack of Sequence Homology Among RNAs of Avian Leukosis-Sarcoma Viruses, Reticuloendotheliosis Viruses, and Chicken Endogenous RNA-Directed DNA Polymerase Activity

Chil-Yong Kang, Howard M. Temin
1973 Journal of Virology  
The relatedness of the RNAs of the three avian systems, including six avian leukosis-sarcoma viruses, four reticuloendotheliosis viruses, and the microsome fraction of normal uninfected chicken embryo cells, containing RNA and a DNA polymerase have been studied by nucleic acid hybridization. All six avian leukosis-sarcoma viruses have closely related nucleotide sequences; and all four reticuloendotheliosis viruses have closely related nucleotide sequences. But, almost no similarities were
more » ... ed between the RNAs of avian leukosis-sarcoma viruses ald reticuloendotheliosis viruses. The RNA template of the endogenous RNA-directed DNA polymerase activity of normal uninfected chicken cells had no detectable relationship to RNAs of avian leukosis-sarcoma and reticuloendotheliosis viruses. There are three avian systems with RNA and a DNA polymerase: avian leukosis-sarcoma viruses (ALV), reticuloendotheliosis viruses (REV), and a fraction from uninfected chicken cells and embryos. Virions of both ALV and REV contain 60-70S RNA and a DNA polymerase and have C-type morphology (2, 18, 19, 22, 24, 29) . However, there are no serological relationships between the structural proteins or the native DNA polymerases of ALV and REV virions (11, 18, 20) . In addition, there is no complementation, phenotypic mixing, or recombination between ALV and REV (11). A microsome fraction isolated from normal, uninfected chicken cells and embryos contained endogenous RNA-directed DNA polymerase activity (14). This endogenous DNA polymerase activity was both RNase sensitive and partially resistant to actinomycin D. The early DNA product of this activity was isolated as a complex with RNA, and the product DNA hybridized to RNA from the same fraction (14, 15) . The experiments reported in this paper were performed to determine if there were any relationships between the RNAs of these three avian systems. In a preliminary study, we did not find a relationship among the three avian systems with RNA and a DNA polymerase (14) . These experiments have now been repeated with better methods of hybridization, with a larger number of viruses, and with DNA products made in the presence of actinomycin D which contain more of the sequences in the RNAs (9). Two different methods for determining nucleic acid sequence homologies were used. In one method, virion 32P-labeled RNA was annealed with an excess of DNA product, and the extent of protection of the RNA from digestion by RNase was determined. Previous reports (8, 9) have shown that excess product DNA from the endogenous DNA polymerase reaction of Rous sarcoma virus (RSV) virions protected from RNase digestion all of the homologous virion 32P-labeled 60-70S RNA. The presence of actinomycin D in the reaction mixture decreased the ratio of product DNA to viral RNA needed for protection from RNase digestion (9). In the other method, 'H-labeled product DNA was annealed with excess virion or cell RNA, and the extent of protection of the DNA from digestion by the single-strand-specific nuclease, Si (from Aspergillus oryzae), was determined as a function of RNA concentration and time (13, 16) . Association kinetics, developed by 1314 on May 9, 2020 by guest
doi:10.1128/jvi.12.6.1314-1324.1973 fatcat:f5qo72f42jhdzif76qdjf3uvke