Stringer J R, Holland L E, Swanstrom R I, Pivo K, Wagner E K
J Virol. 1977 Mar;21(3):889-901. doi: 10.1128/JVI.21.3.889-901.1977.
We have quantitatively analyzed the size and amount of herpes simplex virus (HSV)-specific RNA synthesized in HeLa cells using DNA and RNA excess hybridization. At 2 h after infection (early), transcripts from 20% of the total HSV DNA are present on polyribosomes as poly(A+) RNA. At this time, viral poly(A+) RNA comprises 60 to 75% of the newly synthesized poly(a+) mRNA on polyribosomes. By 6 h after infection (late), poly(A+) HSV RNA transcribed from 35 to 40% of the viral DNA is found on polyribosomes. These viral poly(A+) transcripts comprised as much as 90% of newly synthesized poly(A+) mRNA and are measurably larger than viral poly(A+) transcripts isolated early. Some but not all of this size difference is due to the fact that the poly(A) tails on early transcripts are shorter than those found on transcripts made late. Even late after infection, a small but readily measurable amount of cellular poly(A+) RNA is still being made and entering polyribosome complexes. In the nucleus, late after infection, poly(A+) HSV RNA is complementary to 50% of the total HSV DNA. Both early and late after infection, total nuclear viral transcripts are, on the average, larger than viral transcripts found on polyribosomes; however, nuclear HSV poly(A+) RNA is not measureably larger than the corresponding cytoplasmic viral poly(A+) sequences at either time. A major portion (30 to 40%) of the polyribosomal HSV RNA made either early or late after infection is not polyadenylated. This HSV poly (A-) RNA is transcribed from the same sequences as HSV poly(A+) RNA but, when labeled and isolated either early or late after infection, both nuclear and polyribosomal viral poly(A-) RNA molecules sediment faster in sucrose-formaldehyde gradients than their polyadenylated counterparts.
我们利用DNA和RNA过量杂交技术，对在HeLa细胞中合成的单纯疱疹病毒（HSV）特异性RNA的大小和数量进行了定量分析。感染后2小时（早期），占HSV总DNA 20%的转录本以多聚腺苷酸（poly(A+)）RNA的形式存在于多核糖体上。此时，病毒多聚腺苷酸（poly(A+)）RNA占多核糖体上新合成的多聚腺苷酸（poly(A+)）mRNA的60%至75%。感染后6小时（晚期），在多核糖体上发现了由35%至40%的病毒DNA转录而来的多聚腺苷酸（poly(A+)）HSV RNA。这些病毒多聚腺苷酸（poly(A+)）转录本占新合成的多聚腺苷酸（poly(A+)）mRNA的比例高达90%，且明显大于早期分离的病毒多聚腺苷酸（poly(A+)）转录本。这种大小差异部分但并非全部是由于早期转录本上的多聚腺苷酸（poly(A)）尾巴比晚期转录本上的短。即使在感染后期，仍有少量但易于测量的细胞多聚腺苷酸（poly(A+)）RNA在合成并进入多核糖体复合物。在细胞核中，感染后期，多聚腺苷酸（poly(A+)）HSV RNA与50%的HSV总DNA互补。感染后的早期和晚期，总的核病毒转录本平均都比多核糖体上的病毒转录本大；然而，在这两个时期，核HSV多聚腺苷酸（poly(A+)）RNA与相应的细胞质病毒多聚腺苷酸（poly(A+)）序列相比，并没有明显更大。感染后早期或晚期产生的多核糖体HSV RNA中，很大一部分（30%至40%）没有多聚腺苷酸化。这种HSV多聚腺苷酸（poly(A-)）RNA与HSV多聚腺苷酸（poly(A+)）RNA转录自相同的序列，但在感染后的早期或晚期进行标记和分离时，核和多核糖体病毒多聚腺苷酸（poly(A-)）RNA分子在蔗糖-甲醛梯度中的沉降速度都比它们的多聚腺苷酸化对应物快。
