Characterization of the Low-Molecular-Weight RNAs Associated with the 70S RNA of Rous Sarcoma Virus (original) (raw)

Structure and Molecular Weight of the 60-70S RNA and the 30-40S RNA of the Rous Sarcoma Virus

Proceedings of the National Academy of Sciences, 1974

The structure and molecular weight of the 60-70S RNA complex and the 30-40S RNA species of Rous sarcoma virus were analyzed in an electron microscope after treatment of the RNAs with the bacteriophage T4 gene-32 protein to stretch out the RNA strands. Although all RNA preparations treated with gene-32 protein showed considerable heterogeneity in length, a significant fraction

Evidence for 30-40S RNA as Precursor of the 60-70S RNA of Rous Sarcoma Virus

Proceedings of the National Academy of Sciences, 1973

Rous sarcoma virus harvested from cells at intervals of 3 min has the same density, sedimentation coefficient, and DNA polymerase as virus harvested at hourly intervals. The RNA of the Prague strain-C consists of a minor class of 60-70 S RNA, a major class 30-40 S RNA, and a 4-12 S class of RNA present at variable concentration. The RNA of the Schmidt-Ruppin strain-A contains more 60-70S than 30-40S RNA. Upon incubation of virus harvested at 3-min intervals at 400 in cell growth medium or Tris-

Analysis of the secondary and tertiary structures of Rous sarcoma virus RNA

Nucleic Acids Research, 1980

The secondary and tertiary structures of the 35S RNA of Rous Sarcoma virus have been investigated. T-, RNase partial digests have been first resolved into their components by gel electrophoresis under non denaturing conditions and then each component analyzed further by various techniques. More than one hundred structured fragments have thus been isolated and shown to consist of several individual nucleotide sequences located far apart on the genome. On the basis of the results,a cloverleaf model for the structure of RSV 35S RNA is proposed that has implications for the various biological functions of this RNA.

Molecular weight of RNA subunits of Rous sarcoma virus determined by electron microscopy

Journal of Virology, 1975

Secondary cultures of chicken embryo fibroblasts were infected with the Schmidt Ruppin strain of Rous sarcoma virus (RSV). Five days after infection, the medium was replaced at 2-h intervals with phosphate-free Eagle medium containing 50 muCi of [32P]orthophosphate per ml. Virus was collected by centrifugation, and the RNA was extracted and denatured with dimethyl sulfoxide, and the 33S subunit RNA was isolated by sucrose gradient centrifugation. The molecular weight of the RSV subunit RNA was determined by length measurement in the electron microscope, by using bacteriophage MS2 RNA as a length marker. Molecules of between 2.5 and 3.3 mum in length made up over 50% of the subunit RNA preparation. In this paper, we define RSV RNA subunits as that RNA released from the 70S RNA complex by dimethyl sulfoxide treatment, which sediments as a peak at 33S. Assuming the molecular weight of MS2 RNA to be 1.2 times 10-6, we calculate the molecular weight of RSV subunit RNA to be 3.12 times 10...

New procedure for isolation of Rous sarcoma virus-specific RNA from infected cells

Journal of Virology, 1979

The use of mercurated "strong stop" complementary DNA (complementary to the 5'-terminal 101 nucleotides of Rous sarcoma virus RNA) in the isolation of virus-specific RNA from infected chicken embryo fibroblasts is described. Strong stop Rous sarcoma virus complementary DNA was mercurated chemically, and, as a result of the low complexity of this DNA, short hybridization times (up to 15 min) and heating in the absence of formamide were found to be adequate conditions for the isolation of virus-specific RNA. The purity of the isolated RNA was demonstrated by analysis of labeled RNase T1-resistant oligonucleotides by two-dimensional polyacrylamide gel electrophoresis. The isolated RNA could be translated in the in vitro protein synthesis system derived from rabbit reticulocytes, and an analysis of polypeptides programmed by isolated RNA before and after immunoprecipitation further demonstrated both the purity of the isolated mRNA and the quantitative nature of the isolati...

The methionyl transfer RNAs of Rous sarcoma virus

Virology, 1975

Virions of ROW sarcoma virus contain the two major classes of methionyl-tRNA present in uninfected chick embryo fibroblasts. The formylated species, N-formyl methionyl tRNA, appears to be selectively associated with the viral 70 S RNA genome.

Physical properties of Rous Sarcoma Virus RNA

Proceedings of the National Academy of Sciences of the United States of America, 1968

The RNA's of several RNA tumor viruses have recently been isolated.1 Two major single-stranded RNA components were obtained, a fast-sedimenting RNA component with a sedimentation coefficient (820,w) of approximately 70S and a slowly sedimenting component of approximately 4S. Since the reported st9 as well as the corresponding molecular weights of the tumor virus RNA's are much larger than those of most other known viral RNA's, a determination of the structure and molecular weight of tumor virus RNA's is of great interest. Knowledge of the structure of tumor virus RNA's may also help to elucidate their replication, which is interrupted by inhibitors of DNA-dependent RNA synthesis such as actinomycin D;2 in addition, no virus-specific double-stranded RNA's have been isolated from infected or transformed cells.

Structure-function relationship of Rous sarcoma virus leader RNA

Nucleic Acids Research, 1982

Cells infected by RSV synthesize viral 35S RNA as well as subgenomic 28S and 22S RN'As coding for the Env and Src genes respectively. In addition, at least the 5' 101 nucleotides of the leader are also conserved and we have shown previously that this sequence contains a strong ribosome binding site (J.-L. Darlix et al., J. Virol. 29, 597). We now report the RNA sequence of Rous Sarcoma virus (RSV) leader RNA and propose a folding of this 5' untranslated region which brings the Cap, the initiation codon for Gag and the strong ribosome binding site close to each other. We also show that ribosomes protect a sequence just upstream from initiator AUG of Gag in vitro, and believed to interact with part of the strong ribosome binding site according to the folding proposed for the leader RNA. 0 I RL Pres Umited, Oxford, England.

Characterization of ribosome binding on Rous sarcoma virus RNA in vitro

Journal of Virology, 1985

We determined the sites at which ribosomes form initiation complexes on Rous sarcoma virus RNA in order to determine how initiation of Pr769'9 synthesis at the fourth AUG codon from the 5' end of Rous sarcoma virus strain SR-A RNA occurs. Ribosomes bind almost exclusively at the 5'-proximal AUG codon when chloride is present as the major anion added to the translational system. However, when chloride is replaced with acetate, ribosomes bind at the two 5'-proximal AUG codons, as well as at the initiation site for pFr76ga. We confirmed that the 5'-proximal AUG codon is part of a functional initiation site by identifying the seven-amino acid peptide encoded there. Our results suggest that (i) translation in vitro of Rous sarcoma virus virion RNA results in the synthesis of at least two polypeptides; (ii) the pattern of ribosome binding observed for Rous sarcoma virus RNA can be accounted for by the modified scanning hypothesis; and (iii) the interaction between 40S ribosomal subunits or 80S ribosomal complexes is stronger at the 5'-proximal AUG codon than at sites farther downstream, including the initiation site for the major viral proteins.