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首页> 外文期刊>Journal of Virology >Induction of endogenous guinea pig retrovirus by 5-bromodeoxyuridine: amplification of virus-specific RNA.
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Induction of endogenous guinea pig retrovirus by 5-bromodeoxyuridine: amplification of virus-specific RNA.

机译：用5-溴脱氧尿苷的内源性豚鼠逆转录病毒诱导：病毒特异性RNA的扩增。

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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.
机译：用5-溴脱氧（BUDR）治疗正常豚鼠胚胎细胞（BUDR）激活内源性豚鼠逆转录病毒。在本报告中，通过使用病毒互补DNA（cDNA）与细胞RNA的杂交测定了Budr处理对正常豚鼠胚胎细胞中内源性逆转录病毒RNA水平的影响。我们发现，未处理细胞的总RNA的0.0075％（每个细胞/每个细胞拷贝）是特异性的，而在Budr处理后，从细胞48小时获得的总细胞RNA的0.32％（每个细胞/每个细胞）是病毒特异性的。因此，在治疗48小时后，BudR引起病毒特异性RNA的大约40倍扩增。几种证据支持假设，使得在Budr治疗后观察到的病毒特异性RNA的扩增涉及转录而不是转录后处理的改变。在不同时期在Budr治疗后，在核和细胞质中发生类似的病毒特异性RNA浓度的增加。在48小时后，核病毒特异性RNA增加99倍，每种细胞每种细胞29倍至2,880份，而细胞质病毒特异性RNA从每种细胞85倍的增加47倍至每种细胞4,000份。在表现或不存在BuDR的情况下，放线霉素D存在的病毒特异性RNA的衰减率类似，表明Budr不稳定病毒特异性RNA。在Budr处理的细胞中，病毒特异性RNA的T1 / 2在武器的继续存在下为170分钟，或在除去后150分钟后，在未处理的细胞中150分钟。核病毒特异性RNA序列的尺寸分布在与二甲基亚甲醚变性之后，在未处理的和BUDR处理的细胞中相似，表明在未处理和BUDR处理细胞中的病毒RNA类似的核处理。在未处理的或BUDR处理的细胞中，未观察到核前体对38s病毒特异性RNA的累积。在未处理和BudR处理细胞的总细胞RNA中存在类似的病毒特异性RNA（14秒，38秒和70s）。另外，病毒特异性RNA存在于未处理的细胞的纯化的下变异物中。最后，通过Coffin等人的方法进行核RNA在核RNA脉冲脉冲脉冲脉冲脉冲脉冲脉冲脉冲脉冲脉冲的特异性病毒特异性RNA的量的直接分析。 （J.Mol.Biol.86：373-396,1977）用于定量测定脉冲标记的病毒特异性RNA。发现标记的病毒特异性RNA包含0.0035至0.004％的细胞总脉冲标记的核RNA，用BUDR处理48小时。放射性标记的病毒特异性RNA的50倍增加可能会令人满意的是在48小时后观察到的BUDR处理后观察到的病毒特异性RNA的40倍。

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