Cochran M A, Puckett C, Moss B
J Virol. 1985 Apr;54(1):30-7. doi: 10.1128/JVI.54.1.30-37.1985.
A vaccinia virus gene that is expressed throughout the reproductive cycle was found to have two sets of RNA start sites approximately 55 nucleotides apart. The site nearest to the coding segment is used early in infection and the one further upstream is used after DNA replication. A series of 5' to 3' deletions were made in the promoter region, and the truncated DNA segments were then ligated to the coding portion of the procaryotic chloramphenicol acetyltransferase gene to measure expression. The effects of these mutations on chloramphenicol acetyltransferase synthesis were determined in a vaccinia virus helper-dependent transient expression system and by forming infectious vaccinia virus recombinants that contain the chimeric genes. Deletions extending up to 31 nucleotides before the late RNA start site had no effect on either early or late expression. Removal of an additional 15 nucleotides produced a dramatic decrease in late expression but had no effect on early expression. The latter was not diminished until the deletion was extended from 31 to 24 nucleotides before the early RNA start site. These results were confirmed by transcriptional analyses. We concluded that this vaccinia virus gene has two promoters and that the regulatory signals for each are located within 31 nucleotides of their sites of transcription.
发现一个在整个复制周期中都表达的痘苗病毒基因有两组RNA起始位点，彼此相距约55个核苷酸。最靠近编码区的位点在感染早期被使用，而另一个位于更上游的位点在DNA复制后被使用。在启动子区域进行了一系列从5'到3'的缺失，然后将截短的DNA片段连接到原核氯霉素乙酰转移酶基因的编码部分以测量表达。在痘苗病毒辅助依赖型瞬时表达系统中，并通过形成含有嵌合基因的感染性痘苗病毒重组体，确定了这些突变对氯霉素乙酰转移酶合成的影响。在晚期RNA起始位点之前长达31个核苷酸的缺失对早期或晚期表达均无影响。再去除15个核苷酸会导致晚期表达急剧下降，但对早期表达没有影响。直到缺失延伸至早期RNA起始位点之前的31个核苷酸至24个核苷酸时，早期表达才会减弱。这些结果通过转录分析得到了证实。我们得出结论，该痘苗病毒基因有两个启动子，并且每个启动子的调控信号位于其转录位点的31个核苷酸范围内。
