Chen K H, Wood T, He F, Narayan S, Wilson S H
Sealy Center For Molecular Science, University of Texas Medical Branch, Galveston 77555-1068, USA.
Gene. 1995 Oct 27;164(2):323-7. doi: 10.1016/0378-1119(95)00498-u.
The core promoter of the human DNA polymerase beta (beta Pol)-encoding gene (POL beta) is regulated through cis-elements for the ATF/CREB protein(s), and GC box-binding and initiation-site-binding proteins. The mechanism of promoter regulation has been studied using a nuclear extract transcription system from HeLa cells [Narayan et al., J. Biol. Chem. 269 (1994) 12755-12763]. To study the homologous promoter (ppol beta) in a bovine system, we cloned and characterized the 5'-flanking region of the bovine gene (pol beta). A 15.3-kb fragment of bovine genomic DNA containing the first two exons and 11 kb of 5'-flanking region was isolated from a testis library in bacteriophage lambda EMBL3. S1 nuclease mapping and primer extension analysis of the 5'-end of the pol beta mRNA identified the major transcription start point (tsp), which is located 142-bp 5' of the translational start codon. In transient expression assays using a bovine cell line, analysis of various 5'-deletion mutants demonstrated that a fragment of only 91-bp 5' of the tsp had promoter activity similar to that of a 1.37-kb fragment, so that cis-elements for basal transcription are located within this approx. 100-bp core promoter, as in the human promoter (pPOL beta). Comparison of the core promoters from the bovine and human genes revealed striking similarity, including an almost precise match of the tsp, the ATF/CREB-binding and Sp1-binding sites, and the spacing separating them.
人类DNA聚合酶β（βPol）编码基因（POLβ）的核心启动子是通过与ATF/CREB蛋白、GC盒结合蛋白和起始位点结合蛋白的顺式元件来调控的。启动子调控机制已通过HeLa细胞核提取物转录系统进行了研究[纳拉扬等人，《生物化学杂志》269（1994）12755 - 12763]。为了在牛系统中研究同源启动子（ppolβ），我们克隆并鉴定了牛基因（polβ）的5'侧翼区域。从噬菌体λEMBL3中的睾丸文库中分离出一个15.3 kb的牛基因组DNA片段，该片段包含前两个外显子和11 kb的5'侧翼区域。对polβ mRNA的5'末端进行S1核酸酶图谱分析和引物延伸分析，确定了主要转录起始点（tsp），其位于翻译起始密码子上游142 bp处。在使用牛细胞系的瞬时表达试验中，对各种5'缺失突变体的分析表明，tsp上游仅91 bp的片段具有与1.37 kb片段相似的启动子活性，因此基础转录的顺式元件位于这个约100 bp的核心启动子内，就像人类启动子（pPOLβ）一样。牛和人类基因核心启动子的比较显示出惊人的相似性，包括tsp、ATF/CREB结合位点和Sp1结合位点几乎精确匹配，以及它们之间的间隔。
