摘要
目的:通过比较小鼠不同细胞类型之间Pdx-1基因转录起始区的表观遗传修饰差异,探讨表观遗传修饰对Pdx-1基因转录表达的作用。方法:采用免疫共沉淀-实时定量PCR法检测小鼠胚胎干细胞(mES)、小鼠成纤维细胞株NIH3T3细胞和小鼠β细胞株NIT-1细胞Pdx-1和MLH1基因转录起始区DNA甲基化和组蛋白修饰(H3K4m3、H3K9m3和H3乙酰化)的状况。同时采用实时定量RT-PCR检测上述3种细胞各基因mRNA表达水平。分析基因的DNA甲基化水平、H3K4m3、H3K9m3和H3乙酰化修饰与基因表达之间的相互关系。结果:(1)以mES细胞为对照,NIT-1细胞的Pdx-1基因转录起始区呈低DNA甲基化和高H3K4m3修饰(P<0.05),NIH3T3细胞的Pdx-1基因的转录起始区的DNA甲基化、H3乙酰化、H3K4m3和H3K9m3修饰水平明显增高(P<0.05);(2)Pdx-1基因仅在NIT-1细胞表达,其表达与DNA甲基化存在等级负相关(r=-0.802,P<0.01),与H3K4m3修饰存在直线相关(r=0.997,P<0.01),与H3K9m3修饰存在等级负相关(r=-0.879,P<0.01);(3)管家基因MLH1的表达与所检测的表观遗传修饰无相关性。结论:DNA甲基化、H3K9m3与H3K4m3修饰能相互协调,共同调控Pdx-1基因的表达,对胚胎干细胞向β细胞分化具有重要意义。
AIM：To investigate the role of epigenetic modification in Pdx-1 gene transcription and expression,and to compare the differences between epigenetic modifications of Pdx-1 gene promoter in various cell types of mice. METHODS： The promoter DNA methylation and histone modification status of Pdx-1 and MLH1 genes in NIT-1 cells,NIH3T3 cells and mouse embryonic stem cells were measured by chromatin immunoprecipitation-real time PCR method. The expression levels of these genes in the three cell lines were measured by real time RT-PCR. The relation between epigenetic modifications and gene expression was analyzed. RESULTS： （1） Compared to mES cells,there was lower DNA methylation and higher H3K4m3 modification levels in the promoter of Pdx-1 gene in NIT-1 cells （P〈0.05）. DNA methylation,H3 acetylation,H3K4m3 and H3K9m3 modification levels in the promoter of Pdx-1 gene in NIH3T3 cells were distinctly increased （P〈0.05）. （2） Pdx-1 gene transcription expressed only in NIT-1 cells. The Spearman＇s rho between Pdx-1 gene expression and DNA methylation （r=-0.802,P〈0.01） was observed. The Pearson correlation between Pdx-1 gene expression and H3K4m3 modification （r=0.997,P〈0.01） was also found. The Spearman’s rho between Pdx-1 gene expression and H3K9m3 modification （r=-0.879,P〈0.01） was observed. （3） No correlation between housekeeper MLH1 gene expression and epigenetic modification was found. CONCLUSION： DNA methylation,H3K4m3 and H3K9m3 modification coordinated participate to regulate and control the expression of Pdx-1 gene. It is of great significance to the differentiation of β cells from ES cells.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2010年第4期786-790,共5页
Chinese Journal of Pathophysiology
