转录因子JunB和JunD调控成釉细胞MMP-20基因转录活性的研究  

Study of Transcription Factor JunB and JunD on Regulating the Expression of Gene MMP-20

作　　者：郝佳[1] 张新新[1] 袁杰[1] 张娟娟[1] 刘晓影[2] 孙岩[1] 高玉光[3] 

机构地区：[1]潍坊医学院口腔医学研究所,山东潍坊261053 [2]潍坊医学院细胞生物学教研室 [3]滨州医学院附属医院口腔科

出　　处：《潍坊医学院学报》2013年第1期14-17,25,共4页Acta Academiae Medicinae Weifang

基　　金：国家自然科学基金资助课题(课题编号:30973327);山东省自然科学基金资助课题(课题编号:ZR2010HM076)

摘　　要：目的通过研究转录因子JunB,JunD对成釉细胞中MMP-20基因表达的调控作用,从而进一步明确Jun家族在牙釉质形成中的影响。方法首先构建JunB和JunD真核表达载体重组质粒并瞬时转染重组质粒进入成釉细胞中;利用双荧光素酶基因检测报告系统分析不同浓度JunB,JunD对MMP-20启动子特征性序列区域的转录活性的影响;确定有明显作用的启动子区段,利用基因定点突变和双荧光素酶基因检测报告系统分析JunB对MMP-20基因启动子转录活性的影响;最后再观察JunB与JunD共转染时对MMP-20基因活性表达的改变。结果成功构建JunB,JunD真核重组表达载体,顺利将重组质粒转染入成釉细胞;双荧光素酶基因检测报告系统分析显示JunB对MMP20启动子活性表达上调,而JunD对MMP20启动子活性表达无作用;突变MMP20启动子AP1的两个结合位点后,MMP20启动子的转录活性下降,同时JunB也失去了上调MMP20启动子转录活性的作用。最后将JunB和JunD共转染时,在JunB存在的前提下,随着JunD转染量的增加,小鼠成釉细胞MMP20启动子转录活性明显减弱。结论该研究表明转录因子JunB与成釉细胞内MMP-20启动子的特征性序列相互作用,从而调控MMP20的表达水平;而JunD对MMP20启动子特征性序列无明显作用。所以为进一步研究Jun家族成员在釉质发育过程中的作用建立了重要的生物学基础。Objective Through the research of transcription factor JunB and JunD on regulating the expression of gene MMP20 in ameloblast,lhus further clearing the influence of Jun family in enamel development. Methods Firstly recombinant plasmid of eukaryotic expression vector of JunB and JunD was constructed and recombinant plasmid was ｜ransfected into ameloblast instantly. Double luciferase genetic testing report system were performed to measure the effect of different concentration of JunB and JunD to transcription activity of MMP20 promoter. To make sure the promoter section of apparent effect,and to analyse the influence of JunB/D to MMP20 gene promoter transcription activity by using the gent, point mutation and double luciferase genetic testing report system. Finally the changes of MMP20 gene expres- sion were observed when the JunB and JunD commonly transfect. Results Eukaryotic recombination expression vector of JtmB and JunD were constructed successfully,and the recombinant plasmid was transfect into ameloblast smoothly. Double luciferase genetic testing report system analysis showed that JunB can rise the activity expression of MMP20 promoter, bur JunD have no action to MMP20 promoter activity express. After mutating MMP20 promoter AP1 two binding site, transcription activity of MMP20 promoter declined, and at the same time JunB had lost its action to rise MMP20 promoter transcription activity. Finally When JunB and JunD commonly transfect into ameloblast, JunB existed in the premise,with the increased dose of JunD transfection, mice into glaze cell transcription activity of MMP20 promoter decreased signifi- cantly in mice ameloblast. Conclusion The study suggests that transcription factor JunB were interacted with the characteristic sequence of MMP20 promoter in ameloblast, so as to control MMP20 expression level. But transcription factor JunD have no influence on the characteristic sequence of MMP20 promoter. Further study the function of the Jun family in the process of enamel development can establish the i

关 键 词：转录因子JunB JUND MMP20 双荧光素酶基因检测报告 基因定点突变 

分 类 号：R392.13[医药卫生—免疫学]