CONTROL REGION FOR ADENOVIRUS VA RNA-TRANSCRIPTION
CONTROL REGION FOR ADENOVIRUS VA RNA-TRANSCRIPTION
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DOI：
10.1073/pnas.78.6.3378
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发表时间：
1981-01-01
期刊：
影响因子：
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通讯作者：
WEINMANN, R
中科院分区：
文献类型：
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作者：
GUILFOYLE, R;WEINMANN, R
含有腺病毒的VA RNA基因的质粒被含有DNA依赖性RNA聚合酶III的粗细胞质提取物忠实地转录[Wu，G. J.（1978）]。通过用假单胞菌的新酶对这些DNA模板进行体外定点诱变并在pBR 322中再克隆，构建了一系列有序的缺失，其影响主要RNA聚合酶III病毒产物VAI RNA的体外转录。可以定义特异性合成VAI RNA所需的三个区域。一个是在基因内部的+10到+76核苷酸，以全或无的方式影响转录。转录起始于替换基因5“末端下游最多10个核苷酸的质粒序列。缺失超过核苷酸+15的变体不支持VAI RNA的转录。去除+76下游的3“-末端序列允许正确起始。起始位点上游的第二个区域影响转录物的第一个核苷酸的精确比对（Thimmapaya等人，1979年）。+76下游的第三个区域编码转录终止的信号，并且新信号与其他病毒DNA序列一起引入。转录竞争实验表明，转录调节因子结合的主要位点位于核苷酸+55和+70之间，并表明控制区是双功能的。VAI RNA的内部对照区，60个碱基长和11个碱基下游的5“末端的基因，可以被定义。
Plasmids containing the VA RNA genes of adenovirus are faithfully transcribed by a crude cytoplasmic extract containing DNA-dependent RNA polymerase III [Wu, G.-J. (1978)]. By subjecting these DNA templates to in vitro site-directed mutagenesis with a novel enzyme of Pseudomonas and recloning in pBR322, an ordered series of deletions was constructed which affect the in vitro transcription of the major RNA polymerase III viral product, VAI RNA. Three regions that are required for specific synthesis of VAI RNA can be defined. One, inside the gene at nucleotides +10 to +76, affects the transcription in an all-or-none fashion. Transcription is initiated on plasmid sequences that replace up to 10 nucleotides downstream from the 5'' end of the gene. Variants with deletions past nucleotide +15 do not support the transcription of VAI RNA. Removal of 3''-end sequences downstream from +76 allows correct initiation. A 2nd region, upstream from the initiation site, affects the exact alignment of the 1st nucleotide of the transcript (Thimmapaya et al., 1979). A 3rd region, downstream from +76, encodes signals for termination of transcription, and new signals were brought in with other viral DNA sequences. Transcription competition experiments indicate that the primary site for binding of a transcriptional regulation factor is located between nucleotides +55 and +70 and suggest that the control region is bifunctional. An internal control region for VAI RNA, .apprx. 60 bases long and 11 bases downstream from the 5'' end of the gene, can be defined.