Epigenetic Silencing of ALDH1L1, a Metabolic Regulator of Cellular Proliferation, in Cancers.
Epigenetic Silencing of ALDH1L1, a Metabolic Regulator of Cellular Proliferation, in Cancers.
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DOI：
10.1177/1947601911405841
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发表时间：
2011-02-01
期刊：
影响因子：
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通讯作者：
Krupenko, Sergey A
中科院分区：
文献类型：
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作者：
Oleinik, Natalia V;Krupenko, Natalia I;Krupenko, Sergey A
FDH（10-甲酰四氢叶酸脱氢酶，ALDH 1 L1基因的产物）是细胞溶质中的主要叶酸代谢酶，参与细胞增殖的调节。我们以前已经证明，FDH是强烈和普遍下调恶性人类肿瘤和癌细胞系。在这里，我们报告说，启动子甲基化是一个主要的机制控制FDH水平在人类癌症。计算分析已经确定了ALDH 1 L1启动子区域中广泛的CpG岛。它含有96个CpG对，覆盖ALDH 1 L1基因的-525和+918 bp之间的区域，包括启动子、整个外显子1和紧邻外显子下游的内含子1的一部分。亚硫酸氢盐测序分析揭示了癌细胞系中岛的广泛甲基化（76%-95%的CpG）。与这些发现一致，用甲基转移酶抑制剂5-氮杂-2 '-脱氧胞苷处理缺乏FDH的A549细胞恢复了FDH表达。对来自肺腺癌患者的样品的分析证明了肿瘤样品中ALDH 1 L1 CpG岛的甲基化，并且在相应的正常组织中完全缺乏甲基化。在肝组织中观察到相同的现象：CpG岛在从正常肝细胞提取的DNA中是无甲基化的，但在肝细胞癌中是广泛甲基化的。ALDH 1 L1 mRNA和蛋白水平与岛的甲基化状态相关，肿瘤样本显示表达下调甚至基因完全沉默。我们的研究还表明，外显子1显着增加ALDH 1 L1启动子的转录活性，在荧光素酶报告分析。有趣的是，外显子在具有强烈下调或沉默的ALDH 1 L1基因的样品中广泛甲基化。
FDH (10-formyltetrahydrofolate dehydrogenase, the product of the ALDH1L1 gene), a major folate-metabolizing enzyme in the cytosol, is involved in the regulation of cellular proliferation. We have previously demonstrated that FDH is strongly and ubiquitously down-regulated in malignant human tumors and cancer cell lines. Here, we report that promoter methylation is a major mechanism controlling FDH levels in human cancers. A computational analysis has identified an extensive CpG island in the ALDH1L1 promoter region. It contains 96 CpG pairs and covers the region between -525 and +918 bp of the ALDH1L1 gene including the promoter, the entire exon 1, and a part of intron 1 immediately downstream of the exon. Bisulfite sequencing analysis revealed extensive methylation of the island (76%-95% of CpGs) in cancer cell lines. In agreement with these findings, treatment of FDH-deficient A549 cells with the methyltransferase inhibitor 5-aza-2'-deoxycytidine restored FDH expression. Analysis of the samples from patients with lung adenocarcinomas demonstrated methylation of the ALDH1L1 CpG island in tumor samples and a total lack of methylation in respective normal tissues. The same phenomenon was observed in liver tissues: the CpG island was methylation free in DNA extracted from normal hepatocytes but was extensively methylated in a hepatocellular carcinoma. Levels of ALDH1L1 mRNA and protein correlated with the methylation status of the island, with tumor samples demonstrating down-regulation of expression or even complete silencing of the gene. Our studies have also revealed that exon 1 significantly increases transcriptional activity of ALDH1L1 promoter in a luciferase reporter assay. Interestingly, the exon is extensively methylated in samples with a strongly down-regulated or silenced ALDH1L1 gene.