白血病细胞系中WT1基因启动子区域DNA甲基化的研究

目的应用基于聚合酶链反应(PCR)的实验方法,研究白血病细胞系中WT1基因启动子区域的DNA甲基化水平及其与WT1基因表达的关系.方法采用RT-PCR技术及甲基化特异性PCR(methylation-specific PCR,MSP)技术,检测HL-60、Jurkat及KG-1等白血病细胞系中WT1基因mRNA表达水平及其启动子区域的DNA甲基化状态;以5-杂氮脱氧胞嘧啶(5-aza-CdR)对启动子区域DNA高甲基化的U937细胞系进行去甲基化处理后,观察WT1基因表达水平的改变.结果 HL-60、K562、KG-1、NB4及SHI-1细胞系中WT1水平高表达,而Jurkat和U937细胞表达水平极低,同时检测到这两个细胞系存在WT1基因启动子区域DNA高甲基化;经去甲基化处理后,U937细胞系的WT1基因表达水平较未处理者升高.结论 WT1基因启动子区域DNA高甲基化是抑制其表达的机制之一.

Study on DNA Methylation Status of WT1 Gene in its Promoter Gegion in Leukemia Cell Lines

Objective To study the DNA methylation pattern of WT1 gene promoter region within leukemia cells and correlation between that and WT1 gene expression by using the PCR-based methods. Methods RT-PCR and Methylation-specific PCR(MSP) performed to study WT1 gene expression in HL-60,Jurkat,KG-1,K562,NB4,SHI-1, and U937 cell lines and the DNA methylation status in promoter region of WT1 gene.After treatment of U937 cell line by 5-aza-CdR,a demethylation inducing agent,the changes of WT1 gene expression level in U937 cells was determined. Results HL-60,K562,KG-1,NB4,SHI-1 cell lines demonstrated higher WT1 expression,while that are extremely low in Jurkat and U937.The DNA hypermethylation in WT1 gene promoter region were identified in those two cell lines. The WT1 gene expression in U937 was enhanced after demethylation. Conclusion DNA hypermethylation in WT1 promoter region is one of the mechanisms to suppress its expression.