摘要
目的:探讨大鼠γ-谷氨酰半胱氨酸合成酶(γ-GCS)催化亚单位(GCLC)基因启动子(-745～-705)区段E-box元件对其转录调控是否存在组织细胞差异性。方法:分别将GCLC5’端上游序列驱动的荧光素酶报道基因载体GCLC-Luc、E-box缺失的delE-box-GCLC-Luc及pGL3-enhancer空载体瞬时转染大鼠不同组织来源的细胞,测得荧光素酶活性;并经蛋白定量及pSV-β-半乳糖苷酶的活性测定,获得校正荧光素酶活性值,通过比较校正荧光素酶活性值差异间接判断E-box元件对GCLC转录活性的影响。结果:在大鼠心肌细胞H9C2(2-1),转染delE-box-GCLC-Luc组荧光素酶活性值较转染GCLC-Luc组明显升高(P<0.001),说明E-box能使其GCLC转录活性下调;在大鼠肾小球细胞HBZY-1,转染delE-box-GCLC-Luc组荧光素酶活性值较GCLC-Luc组明显降低(P<0.02),提示E-box具有上调该细胞GCLC转录活性的作用;而在大鼠神经胶质瘤细胞C6和小肠隐窝上皮细胞IEC-6,组间差异无统计学意义(P>0.05),提示E-box对其GCLC转录活性无影响。结论:E-box元件能组织特异性的调控大鼠GCLC基因的转录,该元件可能是决定GCLC基因存在组织差异性表达的重要顺式作用元件。
Objective：To investigate the tissue specificity of E-box element in the regulation of rat γ-GCS catalytic subunit gene（GCLC gene）.Methods：GCLC-Luc and its E-box deletion mutant delE-box-GCLC-Luc were used to transfect rat cells from different tissues：rat myocardium myoblast cells H9C2（2-1）;kidney epithelial cells HBZY-1;glioma fibroblast cells C6 and intestina epithelial cells IEC-6.A SV40 promoter drived β-galactosidase reporter vector were cotransfected for calibrating the results.The luciferase activities of the transfected cells were determined by luminometry.Results：The luciferase activity of delE-box-GCLC-Luc transfected H9C2（2-1）was much higher than that of GCLC-Luc transfected cells（P〈0.001）;In contrast,the luciferase activity of delE-box-GCLC-Luc transfected HBZY-1was lower than that of the GCLC-Luc transfected cells（P〈0.05）;There was no difference in the luciferase activities of two groups of C6 cells（P=0.703）and IEC-6 cells（P=0.675）transfected with the delE-box-GCLC-Luc and GCLC-Luc vectors.Conclusion：The regulation of E-box element in rat GCLC gene promoter might be tissue-specific：it inhibits the transcriptional activity of GCLC promoter in H9C2（2-1）,as well as L2,but enhances the transcriptional activity in HBZY-1,and has no effects on C6 and IEC-6.
出处
《广州医学院学报》
2007年第1期1-5,共5页
Academic Journal of Guangzhou Medical College
基金
国家自然科学基金资助项目(30170401)
广东省自然科学基金团队项目(04205342)
作者简介
陈辉（1973．11-），女，主治医师，硕士。研究方向：慢性阻塞性肺疾病。
通讯作者：Tel：020—81340482，Email：pxran@vip．163．com
