人HAS3启动子的结构与功能初步分析  

Structural and Functional Analysis of Human HAS3 Promoter

作　　者：镇磊[1,2] 卜友泉[1,2] 杜刚[1,2] 杨正梅[3] 兰欢[1,2] 崔涛[1,2] 刘革力[1,2] 宋方洲[1,2] 

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 [2]重庆医科大学分子医学与肿瘤研究中心,重庆400016 [3]重庆医科大学实验动物中心,重庆400016

出　　处：《中国生物化学与分子生物学报》2009年第6期534-541,共8页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金(No.30801356;30871237;30872758);重庆市科委自然科学基金(No.2007BB5302);重庆医科大学重点课题(No.XBZD200707)资助项目~~

摘　　要：透明质酸合酶3(hyaluronan synthase3,HAS3),是一个参与透明质酸合成的酶分子,在上皮形成和肿瘤转移等过程中起重要作用.为了进一步研究HAS3基因的转录调控机制,本研究克隆鉴定了HAS3基因的启动子.首先应用5′RACE(rapid amplification of cDNA ends,cDNA末端快速扩增)技术鉴定了HAS3基因的转录起始位点,发现了丰度和转录起始位点不同的两种新的HAS3基因剪接变异体.通过PCR定向克隆策略,构建了覆盖HAS3基因5′端侧翼区起始密码子ATG上游约4.3kb区域的一系列HAS3基因启动子荧光素酶报告基因重组体.启动子活性分析表明,HAS3基因启动子定位于转录起始位点附近约450bp的区域内.转录因子结合位点分析表明,HAS3基因启动子缺乏典型的TATA盒,但含有典型的GC盒以及C/EBP等其它潜在的转录因子结合位点.结果提示,Sp1和C/EBP等转录因子可能参与HAS3基因的转录调控.Hyaluronan synthase 3, HAS3, is a hyaluronan-synthesizing enzyme that plays a critical role in the epidermis tissue and tumor development. To further investigate its transcriptional regulatory meehanism, the promoter of HAS3 gene has been cloned and identified in the present study. The 5′ RACE （rapid amplification of cDNA ends） assay and bioinformatic analysis also revealed two novel transcriptional HAS3 variants with distinct transcriptional start sites. A series of overlapped genomic fragments in the 5＇-flanking region of HAS3 gene were cloned into pGL3-basic reporting vectors. Luciferase assays indicated a 450-bp region from the transcriptional start site exerted a core activity, containing classical GC boxes and putative binding sites for transcriptional factors such as C/EBP, but lacking typiacl TATA boxes. These results suggested that the transcriptional factors such as Spl and C/EBP might be invovled in the transcriptional regulation of HAS3 gene.

关 键 词：透明质酸合酶3 CDNA末端快速扩增 启动子 转录调控 

分 类 号：Q78[生物学—分子生物学]