CAJ | 학술논문

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HepG2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CYP3 A4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CYP3 A4*1 G增强CYP3 A4基因表达起负调控作用。
목적：탐토CYP3A4*1G증강CYP3A4기인표체적부조공작용。방법：설계합성포함CYP3A4*1G G혹A등위기인적일계렬기인편단（외현자10여내함자10교계처287화181 bp），구건CYP3A4*1G정향위우CYP3A4계동자상유적형광소매보고기인재체，여내삼질립pRL-TK공전염HepG2세포，통과쌍형광소매보고기인계통검측형광소매활성。결과：여CYP3 A4계동자상비교， G여A등위기인재287 bp DNA편단중균강저료형광소매표체（F＝795．575，P＜0．001），차G여A등위기인대CYP3A4계동자활성적억제정도비교차이무통계학의의（P＞0．05）。여CYP3A4계동자상비교，G 여A 등위기인재181 bp DNA 편단중균강저료형광소매표체（F＝23．218，P＜0．001），이G여A등위기인대CYP3A4계동자활성적억제정도비교차이무통계학의의（P＞0．05）。결론：재CYP3A4내함자10여외현자10교계처CYP3A4*1G존재여CYP3A4*1G변이무관적CYP3A4계동자적억제원건，해억제원건대CYP3 A4*1 G증강CYP3 A4기인표체기부조공작용。
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.