Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey, United States of America.
PLoS One. 2010 Jan 5;5(1):e8579. doi: 10.1371/journal.pone.0008579.
Nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) is a transcription factor which regulates the expression of many cytoprotective genes. In the present study, we found that the expression of Nrf2 was suppressed in prostate tumor of the Transgenic Adenocarcinoma of Mouse Prostate (TRAMP) mice. Similarly, the expression of Nrf2 and the induction of NQO1 were also substantially suppressed in tumorigenic TRAMP C1 cells but not in non-tumorigenic TRAMP C3 cells. Examination of the promoter region of the mouse Nrf2 gene identified a CpG island, which was methylated at specific CpG sites in prostate TRAMP tumor and in TRAMP C1 cells but not in normal prostate or TRAMP C3 cells, as shown by bisulfite genomic sequencing. Reporter assays indicated that methylation of these CpG sites dramatically inhibited the transcriptional activity of the Nrf2 promoter. Chromatin immunopreceipitation (ChIP) assays revealed increased binding of the methyl-CpG-binding protein 2 (MBD2) and trimethyl-histone H3 (Lys9) proteins to these CpG sites in the TRAMP C1 cells as compared to TRAMP C3 cells. In contrast, the binding of RNA Pol II and acetylated histone H3 to the Nrf2 promoter was decreased. Furthermore, treatment of TRAMP C1 cells with DNA methyltransferase (DNMT) inhibitor 5-aza-2'-deoxycytidine (5-aza) and histone deacetylase (HDAC) inhibitor trichostatin A (TSA) restored the expression of Nrf2 as well as the induction of NQO1 in TRAMP C1 cells. Taken together, these results indicate that the expression of Nrf2 is suppressed epigenetically by promoter methylation associated with MBD2 and histone modifications in the prostate tumor of TRAMP mice. Our present findings reveal a novel mechanism by which Nrf2 expression is suppressed in TRAMP prostate tumor, shed new light on the role of Nrf2 in carcinogenesis and provide potential new directions for the detection and prevention of prostate cancer.
核因子-红细胞 2 相关因子 2（Nrf2）是一种转录因子，可调节许多细胞保护基因的表达。在本研究中，我们发现 Nrf2 的表达在 Transgenic Adenocarcinoma of Mouse Prostate（TRAMP）小鼠的前列腺肿瘤中受到抑制。同样，Nrf2 的表达和 NQO1 的诱导在致瘤性 TRAMP C1 细胞中也受到显著抑制，但在非致瘤性 TRAMP C3 细胞中则没有。对小鼠 Nrf2 基因启动子区域的检查发现了一个 CpG 岛，该岛在前列腺 TRAMP 肿瘤和 TRAMP C1 细胞中特定 CpG 位点发生甲基化，但在正常前列腺或 TRAMP C3 细胞中则没有，这一点通过亚硫酸氢盐基因组测序得到了证实。报告基因检测表明，这些 CpG 位点的甲基化极大地抑制了 Nrf2 启动子的转录活性。染色质免疫沉淀（ChIP）检测显示，与 TRAMP C3 细胞相比，甲基化 CpG 结合蛋白 2（MBD2）和三甲基组蛋白 H3（Lys9）蛋白在 TRAMP C1 细胞中与这些 CpG 位点的结合增加。相比之下，RNA Pol II 和乙酰化组蛋白 H3 与 Nrf2 启动子的结合减少。此外，用 DNA 甲基转移酶（DNMT）抑制剂 5-氮杂-2'-脱氧胞苷（5-aza）和组蛋白去乙酰化酶（HDAC）抑制剂曲古抑菌素 A（TSA）处理 TRAMP C1 细胞，可恢复 Nrf2 的表达以及 NQO1 在 TRAMP C1 细胞中的诱导。综上所述，这些结果表明，在 TRAMP 小鼠的前列腺肿瘤中，Nrf2 的表达受到启动子甲基化的表观遗传抑制，这种抑制与 MBD2 和组蛋白修饰有关。我们的研究结果揭示了 Nrf2 在 TRAMP 前列腺肿瘤中表达受抑制的一种新机制，为 Nrf2 在肿瘤发生中的作用提供了新的认识，并为前列腺癌的检测和预防提供了新的方向。
