中文摘要：

目的对人HAS3基因的核心启动子区域进行初步鉴定和分析,为深入研究HAS3基因的转录调控奠定基础。方法以前期构建的人HAS3基因启动子荧光素酶报告基因重组体P（-761/-305）为模板,采用PCR介导的基因定点突变技术构建4个不同的系列删除体,并对HAS3启动子区域中的Sp1结合位点和核心启动子元件进行定点突变,构建相应的定点突变重组体;采用荧光素酶双报告基因分析技术检测各重组体的启动子活性。结果构建了P（-761/-569）、P（-563/-305）、P（-490/-305）和P（-433/-305）4个系列删除体和5个定点突变体;启动子活性分析结果表明,P（-761/-569）无启动子活性,P（-563/-305）、P（-490/-305）和P（-433/-305）均具有较强的启动子活性,Sp1结合位点和核心启动子元件的定点突变可导致HAS3启动子活性的降低。结论 HAS3基因的核心启动子区域主要位于其转录起始位点上游附近-433～-305 bp内,Sp1可能在HAS3的转录调控中起重要作用。

英文摘要：

Objective To preliminarily identify and characterize the core promoter region for human hyaluronan synthase 3（HAS3） gene.Methods The previous constructed HAS3 promoter reporter,P（-761/-305）,was used as template to make four different deletion mutants and site-directed mutants by using PCR based site-directed mutagenesis.Dual luciferase reporter assay was used to determine promoter reporter activity.Results Four deletion mutants were constructed,including P（-761/-569）,P（-563/-305）,P（-490/-305） and P（-433/-305）.Among the four deletion mutants,P（-761/-569） had no promoter activity whereas P（-563/-305）,P（-490/-305） and P（-433/-305） showed significant promoter activity.Site-directed mutagenesis of Sp1-binding site and core promoter element resulted in a decrease in HAS3 promoter activity.Conclusion The core promoter region of HAS3 gene is mainly located in a 129-bp region（-433 bp to-305 bp） nearby the major transcriptional start site.Sp1 might play an important role in the transcriptional regulation of HAS3 gene.