人支气管上皮细胞γ-谷氨酰半胱氨酸合酶催化亚单位基因转录的调控  

Transcriptional regulation of GCLC gene in human bronchus epithelial cells

作　　者：邹国明[1] 李冰[2] 冉丕鑫[1] 

机构地区：[1]广州医学院第一附属医院广州呼吸疾病研究所,广东广州510120 [2]广州医学院基础学院实验中心,广东广州510182

出　　处：《中国病理生理杂志》2008年第8期1543-1548,共6页Chinese Journal of Pathophysiology

基　　金：广东省自然基金团队资助项目(No.05200239)

摘　　要：目的:研究人支气管上皮细胞谷胱甘肽(GSH)合成的限速酶γ-谷氨酰半胱氨酸合酶(γ-GCS)催化亚单位(GCLC)基因转录调控。方法:通过PCR的方法克隆GCLC基因的部分转录调控区基因,构建表达虫荧光素酶报告基因的质粒;采用外切核酸酶Ⅲ结合S1核酸酶的方法构建了GCLC基因部分转录调控区基因的嵌套缺失体;基因转染上述质粒,通过虫荧光素酶报告基因的表达分析揭示嵌套缺失体突变基因的转录调控功能。结果:人GCLC基因转录起始位点上游-2515～-2236bp、-864～-838bp、-769～-538bp、-421～-341bp区间的转录调控区为正性转录调控区,为新发现的转录调控区域;而-810～-769bp区间特别是-782～-769bp区间为负性转录调控区,它的定位范围较原来更精确。结论:本研究有助于进一步深入探讨人支气管上皮细胞GCLC基因的转录调控,为γ-GCS及GSH的合成调控研究提供了良好的前期实验基础。AIM ： To investigate the transcriptional regulating function of the catalytic subunit （GCLC） gene of γ- glutamylcysteine synthase （γ- GCS）, the rate -limiting enzyme of glutathione （GSH） synthetic enzymes. METHODS ： The regulating region of the human GCLC gene was cloned by PCR method to construct the wild type plasmid, which expressed luciferase reporting gene. Many mutated plasmids expressing luciferase reporting gene were also constructed by the method of exonuclease In with S1 nuclease. The transcription regulation of GCLC gene was investigated by transient transfection of the wild type and the mutated plasmids through the observation of relative activities of luciferase. RESULTS： We found that the -2515- -2236bp, -864- -838bp, -769~ -538bp, -421 ~ -341 bpregionsupside from the transcriptional start site up -regulated the gene transcription are the new positive transcription -regulating regions and the - 810 - - 769 bp （ especial the - 782 - - 769 bp） region is the negative transcription - regulating region, which is more precise than ever. CONCLUSION ： The results of this experiment provide one good foundation to study the transcription management of the GCLC gene and GSH synthesis.

关 键 词：基因 转录调控 谷胱甘肽 

分 类 号：R56[医药卫生—呼吸系统]