The structure of the Rous sarcoma virus glycoprotein complex:In vitro phenotypic mixing of Rous sarcoma virus

Summary Gp85 the major envelope glycoprotein of Rous sarcoma virus can be released from intact by virus treatment with 2-mercaptoethanol. Attempts were made to reassociate released gp85 to 2-mercaptoethanol treated virus. In order to study the biological activity and to analyse the reaction product with biochemical methods gp85 from a subgroup different from the recipient virus was used. Successfulin vitro phenotypic mixing could be shown. The biochemical analysis, however, revealed that no reassociation of gp85 and gp35 could be obtained. The biological activity was due to a strong association of whole gp85-gp35 molecules derived from the gp85 preparation to the recipient virus. These molecules were present as a minor contamination in the gp85 preparation.With 3 Figures     

Inhibition of Rous sarcoma virus production by formycin

The effect of formycin, an adenosine analog, on the growth of chick embryo fibroblasts and on Rous sarcoma virus (RSV) production was studied. An adverse effect on cell proliferation was observed in the presence of 10 microM formycin. Treatment with 5 microM formycin for 24 hr reduced by a factor of about 1000 the yield of infections progeny whereas the cell growth remained unaltered. Moreover the few particles released in the presence of formycin showed a markedly decreased ability to synthesize viral cDNA. This impairment was shown to be related to a nonfunctional primer tRNA.     

Presence of complementary RNA in chicken sarcoma cells and Rous sarcoma virus

The subcellular localization of nucleotide sequences in Rous chicken sarcoma complementary to RNA of Rous sarcoma virus was investigated by RNA-RNA molecular hybridization. Samples of virion RNA labeled with radioactive iodine (¹²⁵I) were annealed with RNA from different fractions (nuclei, mitochondria, free and membrane-bound polysomes) isolated from Rous chicken sarcoma cells. The formation of RNase-resistant hybrids between virus RNA-¹²⁵I and RNA from mitochondria and membrane-bound polysomes was demonstrated; the relative frequency of occurrence of sequences complementary to virion RNA in the latter, moreover, was 446 times greater than in the former. The role of complementary ribonucleotide sequences is discussed.Laboratory of Biochemistry, N. N. Petrov Research Institute of Oncology, Ministry of Health of the USSR, Leningrad. (Presented by Academician of the Academy of Medical Sciences of the USSR A. N. Klimov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 85, No. 3, pp. 332–334, March, 1978.     

The methionyl transfer RNAs of Rous sarcoma virus.

An average volume of 1.11 pl of a standard TMV suspension was microinjected into single cultured tobacco cells using a glass capillary microneedle. The volume of virus suspension injected was calculated from 20 microneedles. The average number of characteristic TMV particles (290–310 nm) delivered was determined by direct count of the particles on slotted EM grids. All cells produced crystalline virus inclusions when injected with 620 or more TMV particles. One of two cells was infected after injection with 310 TMV particles and only one of four cells became infected after injection of 72 particles. Cell injections at lower concentrations produced no signs infection. Injected cells producing crystalline inclusions (the criterion for infection) were infectious when assayed on Nicotiana glutinosa L. Uninoculated controls and cells bioassayed immediately after injection with the standard and lowest infectious concentration gave a negative assay.     

Isolation and characterization of proteins from Rous sarcoma virus

When the Bryan high-titer strain of Rous sarcoma virus RSV (RAV-1) labeled with a mixture of ¹⁴C amino acids was dissociated with a neutral detergent (Brij 35), mercaptoethanol, and urea and analyzed by isoelectric focusing in urea, seven radioactive peaks with pIs between 3.5 and 9.9 were found. The peaks were further analyzed by polyacrylamide gel electrophoresis in SDS to determine the number and molecular weight of the individual protein components in each peak. A total of eight amino acid-¹⁴C-labeled proteins were identified in RSV (RAV-1) with molecular weights between 14,000 and 96,000 daltons. When ³H-glucosamine labeled RSV (RAV-1) was dissociated with SDS and the viral components separated by SDS-gel electrophoresis, four radioactive components were observed. Analysis of glucosamine-³H-labeled virus together with amino acid-¹⁴C-labeled virus by dissociation with Brij 35, urea, and mercaptoethanol and separation of components by isoelectric focusing followed by electrophoresis in SDS-containing gels revealed that three of the glucosamine labeled components were glycoproteins corresponding to the three largest amino acid-labeled proteins. The fourth glucosamine-labeled component did did not contain radioactive amino acids suggesting that it was protein-free carbohydrate. Separation of virion components on Bio-Gel columns revealed that the glycoproteins were released from virus with Brij 35, urea, and mercaptoethanol in a large complex which was further dissociated into smaller units by SDS. Two of the eight protein components had high complement fixation titers and formed crossing precipitin lines in agar gel diffusion with hamster antiserum to the avian tumor virus group-specific antigen.     

Specific inhibition of Rous sarcoma virus byα-amanitin

Summary The inhibition of the Rous sarcoma virus replication by -amanitin suggests that the cellular RNA polymerase II may play a part at some stage of the viral replication.     

The structure of the Rous sarcoma virus glycoprotein complex

Summary The viral envelope glycoprotein gp85 is released from purified Rous sarcoma virus by treatment with 2-mercaptoethanol, while the second surface antigen gp35 remains associated with the membrane of intact virus particles. The data represented substantiate and extend previous observations on the Rous sarcoma virus glycoprotein complex.With 2 Figures