Benbrahim-Tallaa Lamia, Waterland Robert A, Dill Anna L, Webber Mukta M, Waalkes Michael P
Laboratory of Comparative Carcinogenesis, National Cancer Institute at National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina 27709, USA.
Environ Health Perspect. 2007 Oct;115(10):1454-9. doi: 10.1289/ehp.10207.
Aberrant DNA methylation is common in carcinogenesis. The typical pattern appears to involve reduced expression of maintenance DNA methyltransferase, DNMT1, inducing genomic hypomethylation, whereas increased expression of de novo DNMT3a or 3b causes gene-specific hypermethylation.
During cadmium-induced malignant transformation, an unusual pattern of genomic hypermethylation occurred that we studied to provide insight into the roles of specific DNMTs in oncogenesis.
Gene expression and DNA methylation were assessed in control and chronic cadmium-transformed prostate epithelial cells (CTPE) using reverse transcription-polymerase chain reaction (RT-PCR), Western blot analysis, methylation-specific PCR, and methyl acceptance assay.
During the 10-weeks of cadmium exposure that induced malignant transformation, progressive increases in generalized DNMT enzymatic activity occurred that were associated with over-expression of DNMT3b without changes in DNMT1 expression. Increased DNMT3b expression preceded increased DNMT enzymatic activity. Procainamide, a specific DNMT1 inhibitor, reversed cadmium-induced genomic DNA hypermethylation. Reduced expression of the tumor suppressor genes, RASSF1A and p16, began about the time DNMT3b overexpression first occurred and progressively decreased thereafter. RASSF1A and p16 promoter regions were heavily methylated in CTPE cells, indicating silencing by hypermethylation, while the DNA demethylating agent, 5-aza-2'-deoxycytidine, reversed this silencing. DNMT1 inhibition only modestly increased RASSF1A and p16 expression in CTPE cells and did not completely reverse silencing.
These data indicate that DNMT3b overexpression can result in generalized DNA hypermethylation and gene silencing but that DNMT1 is required to maintain these effects. The pattern of genomic DNA hypermethylation together with up-regulation of DNMT3b may provide a unique set of biomarkers to specifically identify cadmium-induced human prostate cancers.
异常DNA甲基化在致癌过程中很常见。典型模式似乎涉及维持性DNA甲基转移酶DNMT1的表达降低,诱导基因组低甲基化,而从头甲基化酶DNMT3a或3b的表达增加导致基因特异性高甲基化。
在镉诱导的恶性转化过程中,发生了一种不寻常的基因组高甲基化模式,我们对此进行研究以深入了解特定DNA甲基转移酶在肿瘤发生中的作用。
使用逆转录聚合酶链反应(RT-PCR)、蛋白质印迹分析、甲基化特异性PCR和甲基接受试验,评估对照和慢性镉转化前列腺上皮细胞(CTPE)中的基因表达和DNA甲基化。
在诱导恶性转化的10周镉暴露期间,发生了普遍DNMT酶活性的逐渐增加,这与DNMT3b的过表达相关,而DNMT1的表达没有变化。DNMT3b表达的增加先于DNMT酶活性的增加。普鲁卡因胺,一种特异性DNMT1抑制剂,可逆转镉诱导的基因组DNA高甲基化。肿瘤抑制基因RASSF1A和p16的表达降低大约在DNMT3b首次过表达时开始,此后逐渐下降。RASSF1A和p16启动子区域在CTPE细胞中高度甲基化,表明通过高甲基化沉默,而DNA去甲基化剂5-氮杂-2'-脱氧胞苷可逆转这种沉默。DNMT1抑制仅适度增加CTPE细胞中RASSF1A和p16的表达,并未完全逆转沉默。
这些数据表明,DNMT3b过表达可导致普遍的DNA高甲基化和基因沉默,但DNMT1是维持这些效应所必需的。基因组DNA高甲基化模式以及DNMT3b的上调可能提供一组独特的生物标志物,以特异性识别镉诱导的人类前列腺癌。