Mars M, Beaud G
Institut Jacques Monod du C.N.R.S., Paris, France.
J Mol Biol. 1987 Dec 20;198(4):619-31. doi: 10.1016/0022-2836(87)90205-1.
We have reported the isolation of cis-acting regulatory DNA sequences promoting expression of the herpes virus thymidine kinase gene in vaccinia virus recombinants. In this work we show that each of the inserts from recombinants VpT25, 28, 36 and 56 contains a vaccinia virus early promoter. The position of each of the early RNA start sites in the nucleotide sequence of these four vaccinia virus inserts was precisely mapped by an S1 nuclease mapping procedure. Among the four recombinants analysed only VpT56-infected cells also contained a substantial amount of a transcript with the same 5' end at late period. The insert present in VpT25 contained a new late RNA start site 50 nucleotides upstream from that of the early RNA. The four inserts were mapped on the vaccinia virus genome. We also localized the 5' end of the mRNA of a vaccinia virus host-range gene, whose DNA nucleotide sequence has recently been established. The 45 nucleotides preceding the RNA start site from most of 19 known vaccinia virus early promoters were found to be A + T-rich (at least 80%) and contained shorter A-rich (at least 60%) regions, beginning approximately 25 nucleotides upstream from the RNA start site. The information content, as expressed by the parameter Rsequence, of early vaccinia virus promoters revealed ten bits of information in the sequence of 28 nucleotides upstream from the early RNA start sites. Most of the information needed to locate an early promoter is contained within the nucleotide sequence upstream from an RNA start site. A consensus sequence consists of two blocks: the sequence AA(A/T)N(T/A)N(A/G)AAAANAANA starting at position -27 and the sequence (T/A)(C/T)N(A/T)T(A/G) starting at position -5. It was concluded that vaccinia virus early promoters may be characterized by an A + T-rich region of approximately 45 nucleotides preceding the RNA start site and include a specific 3'-terminal sequence of 28 nucleotides containing at least ten bits of information. A procedure for localizing putative early RNA start sites in nucleotide sequences is proposed.
我们已报道了在痘苗病毒重组体中促进疱疹病毒胸苷激酶基因表达的顺式作用调控DNA序列的分离。在这项工作中，我们表明重组体VpT25、28、36和56的每个插入片段都包含一个痘苗病毒早期启动子。通过S1核酸酶图谱分析程序精确地绘制了这四个痘苗病毒插入片段核苷酸序列中每个早期RNA起始位点的位置。在分析的四个重组体中，只有感染VpT56的细胞在后期也含有大量具有相同5'末端的转录本。VpT25中存在的插入片段在早期RNA起始位点上游50个核苷酸处含有一个新的晚期RNA起始位点。这四个插入片段被定位在痘苗病毒基因组上。我们还定位了痘苗病毒宿主范围基因mRNA的5'末端，其DNA核苷酸序列最近已确定。发现19个已知痘苗病毒早期启动子中的大多数在RNA起始位点之前的45个核苷酸富含A + T（至少80%），并包含较短的富含A的区域（至少60%），从RNA起始位点上游约25个核苷酸处开始。以参数Rsequence表示的痘苗病毒早期启动子的信息含量在早期RNA起始位点上游28个核苷酸的序列中显示出10位信息。定位早期启动子所需的大部分信息包含在RNA起始位点上游的核苷酸序列中。共有序列由两个区域组成：从-27位开始的序列AA(A/T)N(T/A)N(A/G)AAAANAANA和从-5位开始的序列(T/A)(C/T)N(A/T)T(A/G)。得出的结论是，痘苗病毒早期启动子的特征可能是在RNA起始位点之前有一个约45个核苷酸的富含A + T的区域，并包括一个28个核苷酸的特定3'末端序列，该序列包含至少10位信息。提出了一种在核苷酸序列中定位假定的早期RNA起始位点的方法。
