人NFBD1启动子的鉴定与初步分析  

Cloning and Identification of Human NFBD1 Promoter

作　　者：卜友泉[1] 宋方洲[1] 易发平[1] 马永平[1] 

机构地区：[1]重庆医科大学生物化学与分子生物学教研室,重庆400016

出　　处：《中国生物化学与分子生物学报》2007年第8期631-637,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金(No.30371479);重庆医科大学青年基金(2004-12);国家公派留学基金(No.2004850010)资助项目~~

摘　　要：NFBD1,也称MDC1,是一个参与细胞内DNA损伤后细胞应答反应的重要分子.为了进一步深入研究其转录调控机制,本研究克隆鉴定了NFBD1的启动子.首先应用5′RACE技术鉴定了NFBD1的转录起始位点,首次发现NFBD1至少存在3种丰度和转录起始位点不同的转录变异体.然后,通过PCR定向克隆和酶切亚克隆策略,构建了覆盖NFBD1基因5′侧翼区起始密码子ATG上游5kb区域的一系列NFBD1启动子荧光素酶报告基因重组体.启动子活性分析表明,NFBD1启动子区域定位于主要转录起始位点区域附近1.5kb的区域内.采用转录因子结合位点预测分析软件分析表明,NFBD1启动子缺乏TATA盒,但含有典型的CCAAT盒和GC盒以及其它潜在的转录因子结合位点,提示Sp1和NF-Y等转录因子可能参与NFBD1的转录调控.NFBD1 （a nuclear factor with BRCT domain 1 ）, also known as MDC1 （mediator of DNA damage checkpoint protein 1）, is a large nuclear protein that mainly participates in DNA damage response. To further investigate its transcriptional regulation, NFBD1 promoter has been cloned and identified. At first, the transcriptional start sites for NFBD1 gene have been identified by 5＇ RACE （rapid amplification of cDNA ends） and bioinformation analysis. Results indicated that human NFBD 1 gene has at least three transcriptional variants with distinct transcriptional start sites. Several overlapping genomic fragments from the 5＇- flanking region of NFBD 1 gene have been then cloned into pGL3-basic vector to construct NFBD 1 promoter reporters. Luciferase reporter assay indicated that NFBD 1 promoter region was mainly located in a 1.5 kb region nearby the major transcriptional start site. Transcriptional factor binding analysis indicated that NFBD1 promoter lacked TATA box, but contained classical CCAAT box and GC box as well as other putative transcriptional factor binding sites. These results suggested that transcriptional factors such as Spl and NF-Y might be involved in the transcriptional regulation of NFBD 1 gene.

关 键 词：NFBDl RACE 启动子 转录调控 

分 类 号：Q78[生物学—分子生物学]