摘要
目的:探讨CYP3A4*1G增强CYP3A4基因表达的负调控作用。方法:设计合成包含CYP3A4*1G G或A等位基因的一系列基因片段(外显子10与内含子10交界处287和181 bp),构建CYP3A4*1G正向位于CYP3A4启动子上游的荧光素酶报告基因载体,与内参质粒pRL-TK共转染Hep G2细胞,通过双荧光素酶报告基因系统检测荧光素酶活性。结果:与CYP3A4启动子相比较,G与A等位基因在287 bp DNA片段中均降低了荧光素酶表达(F=795.575,P<0.001),且G与A等位基因对CYP3A4启动子活性的抑制程度比较差异无统计学意义(P>0.05)。与CYP3A4启动子相比较,G与A等位基因在181 bp DNA片段中均降低了荧光素酶表达(F=23.218,P<0.001),而G与A等位基因对CYP3A4启动子活性的抑制程度比较差异无统计学意义(P>0.05)。结论:在CYP3A4内含子10与外显子10交界处CYP3A4*1G存在与CYP3A4*1G变异无关的CYP3A4启动子的抑制元件,该抑制元件对CYP3A4*1G增强CYP3A4基因表达起负调控作用。
Aim：To study the negative regulation of CYP3A4＊1G enhancing CYP3A4 gene expression.Methods：A series of reporter gene vectors carrying CYP3A4＊1G G or A allele in different length DNA fragments（exon-intron junction,287 or 181 bp） located upstream of the CYP3A4 promoter in the forward direction were constructed .The above recombinant plasmids were co-transfected with internal control plasmid pRL-TK into HepG2 cells by LipofectamineTM 2000, and lucifer-ase activity was measured with dual luciferase reporter gene system .Results：Compared with CYP3A4 promoter, the G al-lele and the A allele in 287 bp DNA fragments both decreased the expression of luciferase （F=795.575,P〈0.001）,while there was no significant difference in the degree of inhibition to CYP 3A4 promoter by the both alleles（P〉0.05）.Com-pared with CYP3A4 promoter, the G allele and the A allele in 181 bp DNA fragments both reduced the expression of lucif-erase（F=23.218, P〈0.001）, while there was no difference in the extent of inhibition to CYP 3A4 promoter by the both alleles（P〉0.05）.Conclusion： There is a certain inhibition element independent on CYP 3A4＊1G variation of the CYP3A4 promoter,lying exon-intron 10 junction of CYP3A4,and it has negative regulation to CYP3A4＊1G enhancing CYP3A4 expression.
出处
《郑州大学学报（医学版）》
CAS
北大核心
2015年第6期765-768,共4页
Journal of Zhengzhou University(Medical Sciences)
基金
国家自然科学基金项目81173127
81171060
河南省杰出青年基金项目074100510020
河南省教育厅基础研究项目13A310663
河南省科技厅基础与前沿技术研究计划项目142300410206
