Jones P C, Roizman B
J Virol. 1979 Aug;31(2):299-314. doi: 10.1128/JVI.31.2.299-314.1979.
This report concerns the stable viral RNA sequences that accumulate in HEp-2 cells infected with herpes simplex virus type 1. By hybridizing labeled total DNA and restriction endonuclease DNA fragments with excess unlabeled total nuclear and cytoplasmic RNA, we determined the genetic complexity of the RNA and we mapped the regions on the physical map of herpes simplex virus type 1 DNA that are homologous to the RNA. Our results show the following. (i) The viral RNAs accumulating in the nucleus and cytoplasm of cells infected and maintained in the presence of inhibitory concentrations of either cycloheximide or emetine were homologous to 33 and 12% of viral DNA, respectively. All of the fragments tested contained sequences homologous to nuclear RNA. However, only the fragments mapping between 0.00 and 0.18, and 0.53 and 1.00 map units contained sequences homologous to cytoplasmic RNA. (ii) The viral RNAs that accumulate in the nucleus and cytoplasm of cells infected and maintained in the presence of inhibitory concentrations of phoaphonoacetic acid were homologous to 39 and 26% of viral DNA, respectively. In this instance all of the fragments except those mapping between 0.42 and 0.53 map units contained sequences homologous to cytoplasmic RNA. (iii) The viral RNAs that accumulate in the nucleus and cytoplasm 8 h after infection were homologous to greater than 50 and 41%, respectively. All of the fragments tested contained sequences homologous to cytoplasmic RNA. (iv) The viral RNAs that accumulate in the nucleus and cytoplasm of cells infected and maintained in the presence of canavanine are homologous to 33 and 19% of viral DNA, respectively. All of the fragments tested contained sequences homologous to both nuclear and cytoplasmic RNAs. Our results indicate the following. First, there are at least three phases of transcription of viral DNA. Phase 1 does not require the synthesis of host cell or viral proteins. Phase 2 requires the synthesis of viral proteins made before the initiation of viral DNA synthesis. Phase 3 appears to be related to the initiation of viral DNA synthesis. Second, both the extent of transcription and the accumulation of viral RNA in the cytoplasm are tightly regulated. The genetic complexity of total RNA accumulating in infected cells increased in each successive phase. Moreover, the genetic complexity of nuclear RNA was invariably higher than that of cytoplasmic RNA in each phase. Lastly, the results of the studies on viral RNA accumulating in canavanine-treated cells reinforce the hypothesis made previously that more than one polypeptide in each of the alpha and beta polypeptide groups is involved in the transcription preceding the transitions from alpha to beta and beta to gamma polypeptide synthesis, respectively, and that canavanine selectively inactivated subsets of these polypeptides permitting only partial transitions from alpha to beta and beta to gamma to occur.
本报告涉及在感染单纯疱疹病毒1型的HEp - 2细胞中积累的稳定病毒RNA序列。通过将标记的总DNA和限制性内切酶DNA片段与过量未标记的总核RNA和细胞质RNA杂交，我们确定了RNA的遗传复杂性，并在单纯疱疹病毒1型DNA的物理图谱上绘制了与RNA同源的区域。我们的结果如下所示。（i）在存在抑制浓度的放线菌酮或依米丁的情况下感染并维持的细胞的细胞核和细胞质中积累的病毒RNA分别与33%和12%的病毒DNA同源。所有测试片段都含有与核RNA同源的序列。然而，只有位于0.00至0.18以及0.53至1.00图谱单位之间的片段含有与细胞质RNA同源的序列。（ii）在存在抑制浓度的磷乙酸的情况下感染并维持的细胞的细胞核和细胞质中积累的病毒RNA分别与39%和26%的病毒DNA同源。在这种情况下，除了位于0.42至0.53图谱单位之间的片段外，所有片段都含有与细胞质RNA同源的序列。（iii）感染后8小时在细胞核和细胞质中积累的病毒RNA分别与超过50%和41%的病毒DNA同源。所有测试片段都含有与细胞质RNA同源的序列。（iv）在存在刀豆氨酸的情况下感染并维持的细胞的细胞核和细胞质中积累的病毒RNA分别与33%和19%的病毒DNA同源。所有测试片段都含有与核RNA和细胞质RNA同源的序列。我们的结果表明以下几点。首先，病毒DNA的转录至少有三个阶段。阶段1不需要宿主细胞或病毒蛋白的合成。阶段2需要在病毒DNA合成起始之前合成的病毒蛋白。阶段3似乎与病毒DNA合成的起始有关。其次，转录程度和病毒RNA在细胞质中的积累都受到严格调控。在感染细胞中积累的总RNA的遗传复杂性在每个连续阶段都增加。此外，在每个阶段，核RNA的遗传复杂性总是高于细胞质RNA。最后，对在刀豆氨酸处理的细胞中积累的病毒RNA的研究结果强化了先前提出的假设，即α和β多肽组中的每种多肽不止一种分别参与从α到β以及从β到γ多肽合成转变之前的转录，并且刀豆氨酸选择性地使这些多肽的亚群失活，仅允许从α到β以及从β到γ的部分转变发生。
