Adenovirus EIIA early promoter: transcriptional control elements and induction by the viral pre-early EIA gene, which appears to be sequence independent.
Adenovirus EIIA early promoter: transcriptional control elements and induction by the viral pre-early EIA gene, which appears to be sequence independent.
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腺病毒 EIIA 早期启动子：转录控制元件和病毒前早期 EIA 基因的诱导，该基因似乎与序列无关。
DOI：
10.1073/pnas.82.8.2230
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发表时间：
1985
影响因子：
11.1
通讯作者：
Thimmappaya,B
中科院分区：
文献类型：
--
作者：
Murthy,SC;Bhat,GP;Thimmappaya,B
对腺病毒EIIA早期(E)启动子进行了分子解剖，以研究转录所需的序列元件，并检测病毒早期EIA基因产物反式激活其特异的核苷酸序列。将EIIA-E启动子区域与细菌氯霉素乙酰转移酶(CAT)基因的编码序列融合的嵌合基因用于瞬时检测，以确定转录控制区。缺失定位研究表明，-86的上游DNA序列对于HeLa细胞的最佳基础转录和EIA诱导的转录都是足够的。构建了一系列连接子扫描(LS)突变体，以精确识别控制转录的核苷酸序列。对这些LS突变体的分析使我们能够确定启动子上对EIIA-E转录至关重要的两个区域。这些区域位于-29和-21之间(区域I)和-82和-66之间(区域II)。I区的突变会影响启动子的启动，并且在功能上与通常研究的启动子的“TATA”序列相似。为了检测EIIA-E启动子是否含有EIA反式激活的特异性DNA序列，对LS突变体进行了含有携带EIA基因的质粒的共转染实验。在本实验中，所有LS突变体的CAT活性都是由EIA基因诱导的，表明EIA基因对EIIA-E启动子的转录诱导不是序列特异性的。
A molecular dissection of the adenovirus EIIA early (E) promoter was undertaken to study the sequence elements required for transcription and to examine the nucleotide sequences, if any, specific for its trans-activation by the viral pre-early EIA gene product. A chimeric gene in which the EIIA-E promoter region fused to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene was used in transient assays to identify the transcriptional control regions. Deletion mapping studies revealed that the upstream DNA sequences up to -86 were sufficient for the optimal basal level transcription in HeLa cells and also for the EIA-induced transcription. A series of linker-scanning (LS) mutants were constructed to precisely identify the nucleotide sequences that control transcription. Analysis of these LS mutants allowed us to identify two regions of the promoter that are critical for the EIIA-E transcription. These regions are located between -29 and -21 (region I) and between -82 and -66 (region II). Mutations in region I affected initiation and appeared functionally similar to the "TATA" sequence of the commonly studied promoters. To examine whether or not the EIIA-E promoter contained DNA sequences specific for the trans-activation by the EIA, the LS mutants were analyzed in a cotransfection assay containing a plasmid carrying the EIA gene. CAT activity of all of the LS mutants was induced by the EIA gene in this assay, suggesting that the induction of transcription of the EIIA-E promoter by the EIA gene is not sequence-specific.