微核糖核酸506在人结肠癌细胞中的靶基因预测和鉴定

目的 预测并验证微核糖核酸506(miR-506)的真实靶基因.方法 通过多个生物信息学软件对miR-506可能调控的靶基因进行预测,结合其生物学功能和前期基因芯片结果,确定候选靶基因.根据miR-506结合候选靶基因的3'UTR位点,构建相应的报告基因载体及突变体.将所构建的报告基因载体或突变体、β半乳糖苷酶对照报告酶载体和miR-506前体共转染至SW1116细胞中,24 h后检测荧光报告酶变化.结果 过氧化物酶体增殖物激活受体(PPAR)α和维甲酸受体(RXR)α皆为has-miR-506的可能靶基因,miR-506作用于PPARα基因3'UTR区可能存在3个位点,RXRa基因3'UTR区可能存在2个位点.针对于PPARα基因的三组报告基因载体中,位点7930-7936组变化最为明显[(2.68±0.55)倍],位点3547-3553组和位点8335-8341组的荧光报告酶表达变化为其突变体的(1.43±0.22)倍和(1.34±0.20)倍.针对于RXRa基因的两组报告基因载体的荧光报告酶变化则并不显著.结论 PPARα为miR-506的真实靶基因,而RXRα非miR-506的靶基因.

Target prediction and validation of microRNA 506 in human colon cancer cell

Objective To predict and verify the true target genes of microRNA 506 (miR-506).Methods Some bioinformatics softwares were used to predict the possible target genes of miR-506,then determined candidates by combining their biological function and results of gene chip.According to 3'UTR binding sites of target genes for miR-506,corresponding report vectors and mutants were constructed.Report construction or mutant,beta-gal reporter control vector and miR-506 precusor were co-transfected into SW1116 cells,then the changes of luciferase were assayed after 24 hours.Results Peroxisome-prdiferaton activated receptor (PPAR)α and retinoid X receptor (RXR)α mightbinding sites for miR-506,2 binding sites for RXRα.For 3 report vectors of PPARa.the changes of 7936,position 3547-3553 and position 8335-8341,respectively.However,for 2 report vectors of RXRα,the changes of luciferase were all insignificant.Conclusion PPARα is a true target gene of miR-506,not for RXRα.