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Induction of endogenous guinea pig retrovirus by 5-bromodeoxyuridine: amplification of virus-specific RNA.

  • A R Davis
  • D P Nayak
  • J Lofgren
Publication Date
Jun 01, 1978


Treatment of normal guinea pig embryo cells with 5-bromodeoxyuridine (BUdR) activates endogenous guinea pig retrovirus. In this report the effect of BUdR treatment upon the level of endogenous retroviral RNA in normal guinea pig embryo cells was determined by using hybridization of viral complementary DNA (cDNA) to cellular RNA. We found that 0.0075% (120 copies per cell) of total RNA of untreated cells was virus-specific, whereas 0.32% (5,120 copies per cell) of total cellular RNA obtained from cells 48 h after BUdR treatment was virus-specific. Thus, BUdR causes an approximately 40-fold amplification of virus-specific RNA after 48 h of treatment. Several lines of evidence favor the hypothesis that the amplification of virus-specific RNA observed after BUdR treatment involves enhancement of transcription rather than an alteration of post-transcriptional processing. At different times after BUdR treatment, similar increases in virus-specific RNA concentration occur in both nucleus and cytoplasm. After 48 h of BUdR treatment, nuclear virus-specific RNA increased 99-fold, from 29 copies per cell to 2,880 copies per cell, whereas cytoplasmic virus-specific RNA increased 47-fold from 85 copies per cell to 4,000 copies per cell. Decay rates of virus-specific RNA in the presence of actinomycin D were similar in the presence or absence of BUdR, indicating that BUdR does not stabilize virus-specific RNA. In BUdR-treated cells the t1/2 of virus-specific RNA was 170 min either in the continued presence of BUdR or after the removal of BUdR, and 150 min in untreated cells. The size distribution of nuclear virus-specific RNA sequences, after denaturation with dimethyl sulfoxide, was similar in untreated and BUdR-treated cells, suggesting similar nuclear processing of viral RNA in both untreated and BUdR-treated cells. The accumulation of nuclear precursors to 38S virus-specific RNA was not observed at steady-state levels in untreated or BUdR-treated cells. Similar species of virus-specific RNA (14S 24S, 38S, and 70S) were present in the total cellular RNA of untreated and BUdR-treated cells. Additionally, virus-specific RNA was present in purified polyribosomes of untreated cells. Finally, direct analysis of the amount of radiolabeled virus-specific RNA in nuclear RNA pulse-labeled for 30 min with [3H]uridine was performed by the method of Coffin et al. (J. Mol. Biol. 86:373-396, 1977) for quantitative determination of pulse-labeled virus-specific RNA. It was found that labeled virus-specific RNA comprised 0.0035 to 0.004% of the total pulse-labeled nuclear RNA of cells treated for 48 h with BUdR. This 50-fold increase in radiolabeled virus-specific RNA may full- account for the 40-fold increase in steady-state levels of virus-specific RNA observed after 48 h of BUdR treatment.

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