Human DNA polymerase alpha gene: sequences controlling expression in cycling and serum-stimulated cells.

B E Pearson; H P Nasheuer; T S Wang

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Human DNA polymerase alpha gene: sequences controlling expression in cycling and serum-stimulated cells.

机译：人DNA聚合酶α基因：控制自行车和血清刺激细胞中表达的序列。

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We have investigated the DNA polymerase alpha promoter sequence requirements for the expression of a heterologous gene in actively cycling cells and following serum addition to serum-deprived cells. An 11.4-kb genomic clone that spans the 5' end of this gene and includes 1.62 kb of sequence upstream from the translation start site was isolated. The transcription start site was mapped at 46 +/- 1 nucleotides upstream from the translation start site. The upstream sequence is GC rich and lacks a TATA sequence but has a CCAAT sequence on the opposite strand. Analysis of a set of deletion constructs in transient transfection assays demonstrated that efficient expression of the reporter in cycling cells requires 248 bp of sequence upstream from the cap site. Clustered within these 248 nucleotides are sequences similar to consensus sequences for Sp1-, Ap1-, Ap2-, and E2F-binding sites. The CCAAT sequence and the potential E2F- and Ap1-binding sites are shown to be protected from DNase I digestion by partially purified nuclear proteins. The DNA polymerase alpha promoter can confer upon the reporter an appropriate, late response to serum addition. No single sequence element could be shown to confer serum inducibility. Rather, multiple sequence elements appear to mediate the full serum response.

机译：我们已经研究了DNA聚合酶α启动子序列的要求，在主动循环的细胞中以及在血清添加到血清剥夺的细胞中后表达异源基因。分离了一个跨越该基因5'端并包含翻译起始位点上游1.62 kb序列的11.4 kb基因组克隆。转录起始位点位于翻译起始位点上游46 +/- 1个核苷酸处。上游序列富含GC，缺乏TATA序列，但在相反链上具有CCAAT序列。在瞬时转染测定中对一组缺失构建体的分析表明，报告子在循环细胞中的有效表达需要在帽位点上游248 bp的序列。聚集在这248个核苷酸中的序列类似于Sp1-，Ap1-，Ap2-和E2F结合位点的共有序列。 CCAAT序列和潜在的E2F-和Ap1结合位点显示被部分纯化的核蛋白保护免受DNase I消化。 DNA聚合酶α启动子可以赋予报告分子对血清添加的适当的晚期反应。没有显示出单一序列元件可赋予血清诱导性。而是，多个序列元件似乎介导了完整的血清反应。

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《Molecular and Cellular Biology》 |1991年第4期|共15页
作者
B E Pearson; H P Nasheuer; T S Wang;
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中图分类细胞生物学;
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Human DNA polymerase alpha gene: sequences controlling expression in cycling and serum-stimulated cells.

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