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雌激素对载脂蛋白AⅠ基因启动子不同区域转录活性的影响

崔丽1,毛用敏2,赵莉莉2,崔让庄2   

  1. 1. 天津医科大学研究生院博士在读；天津市南开医院
    2. 天津市胸科医院
  • 收稿日期:2010-11-23 修回日期:2011-02-25 出版日期:2011-10-15 发布日期:2011-10-15
  • 通讯作者: 崔丽

The Effect of Estradiol on modulating transcription of different fractions of apoAⅠgene promoter

  • Received:2010-11-23 Revised:2011-02-25 Published:2011-10-15 Online:2011-10-15

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摘要： 目的：探讨雌激素对Apo AⅠ基因启动子不同区域转录活性的影响。方法：利用含荧光素酶报告基因的表达载体pGL2构建携带ApoAI基因启动子不同区域片段的重组质粒pGL2/-41AⅠ、pGL2/-256AⅠ和pGL2/-2500AⅠ，以及同时携带ApoCⅢ/AⅣ基因片段的重组质粒：pGL2/-41AⅠCⅢAⅣ、pGL2/-256AⅠ CⅢAⅣ 、pGL2/-2500AⅠCⅢAⅣ；采用阳离子脂质体法将重组质粒与pRL-null内参质粒共转染HepG2细胞，24h后加入10 μmol/L雌激素。雌激素刺激24h后检测细胞荧光素酶报告基因的荧光强度，以反映ApoAI的转录水平。结果 转染pGL2/-256AⅠ,pGL2/-2500AⅠ,pGL2/-256AⅠCⅢAⅣ及pGL2/-2500AⅠCⅢAⅣ 的HepG2细胞在雌激素刺激下荧光素酶相对活性明显高于对照组（P﹤0.05）；而pGL2/-41AⅠ及pGL2/-41AⅠCⅢAⅣ质粒转染细胞对雌激素刺激前后产生的荧光素酶活性差异无统计学意义（P﹥0.05）。结论：Apo AⅠ启动子-256bp~-41bp区域可能包含与雌激素作用相关的反应元件，受雌激素刺激转录活性增强；而ApoCⅢ/AⅣ基因调控区不受雌激素的影响。

关键词: 雌激素, 载脂蛋白AⅠ, 基因转录

Abstract: Objective: to analyze the modulating mechanism of estrogen on different fractions of apo A? gene promoter. Methods: Construct recombinants containing different length DNA fragments of apo AI gene (-41/+397, -256/+397, and -2500/+397) with or without a 7-kb region apo CIII/AIV intergenic region in basic pGL2 vector that containing lueiferase reporter gene. The final recombinants with the internal control vector pRL-null with renilla luciferase reporter gene were transfected to HepG2 cells by the cationic lipid method. After 24h, the cells were treated with estradiol at 10μmol/L concentration; After 48h, cells were collected and lysized. The activities of these luciferase enzymes were measured with Dual-GloTM Luciferase Assay System. Results: In estradiol induced experiments, exception of the recombinant plasmids containing only the basal promoter (-41/+397bp) with or without region of apo CIII/AIV, The all recombinants expression of luciferase enzymes was great increased. There was no different between pGL2/-41AⅠand pGL2/-41AⅠCⅢAⅣ in the presence of estradiol. Conclusion: Promoter area containing -256bp may be the smallest reactive element for estrogen. It can increase transcription activity after stimulating by estrogen. The apoCIII/AIV gene modulating was not affected by estrogen.

Key words: estrogen, apolipoprotein AⅠ, gene transcription modulation