Selective association of the methyl-CpG binding protein MBD2 with the silent p14/p16 locus in human neoplasia
Selective association of the methyl-CpG binding protein MBD2 with the silent p14/p16 locus in human neoplasia
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DOI：
10.1073/pnas.101617298
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发表时间：
2001-04-24
影响因子：
11.1
通讯作者：
Wolffe, AP
中科院分区：
文献类型：
--
作者：
Magdinier, F;Wolffe, AP
抑癌基因的DNA甲基化是人类癌症的一个共同特征。细胞周期蛋白依赖性激酶抑制剂基因 p16/Ink4A 在多种恶性组织中高度甲基化，位于 9p21 染色体上游 20 kb 的 p14/ARF 基因在癌症中也甲基化。 p14/ARF（ARF，替代阅读框）不抑制细胞周期蛋白或细胞周期蛋白依赖性激酶复合物的活性；然而，这两种基因产物在癌症病因学中的重要性在于它们分别参与两个主要的细胞周期调节途径：p53 和视网膜母细胞瘤蛋白 Rb。由单独的启动子驱动的不同的第一外显子被剪接到共同的外显子2和3上，并且所得的蛋白质在不同的阅读框中被翻译。这两个基因均在正常细胞中表达，但当它们的 CpG 岛高度甲基化时，它们可以选择性或协同沉默。在此，我们检查了与异常甲基化启动子相关的甲基-CpG 结合蛋白的存在、通过染色质免疫沉淀测定乙酰化组蛋白 H3 和 H4 的分布，以及通过定量逆转录酶 PCR 检测 5-aza-2'-脱氧胞苷 (5aza-dC) 和曲古抑菌素 A 化学处理对结肠细胞系中基因诱导的影响。我们观察到甲基-CpG 结合蛋白 MBD2 靶向甲基化调节区并排除乙酰化组蛋白 H3 和 H4，导致局部失活染色质构型。当甲基化时，这些基因可以被 5aza-dC 诱导，但 5aza-dC 和曲古抑菌素 A 的联合作用会导致强劲的基因表达。因此，甲基-CpG 结合蛋白和组蛋白脱乙酰酶似乎在体内协同作用，DNA 甲基化对组蛋白乙酰化具有主导作用，并抑制癌症中高甲基化的肿瘤抑制基因的表达。
DNA methylation of tumor suppressor genes is a common feature of human cancer. The cyclin-dependent kinase inhibitor gene p16/Ink4A is hypermethylated in a wide range of malignant tissues and the p14/ARF gene located 20 kb upstream on chromosome 9p21 is also methylated in carcinomas. p14/ARF (ARF, alternative reading frame) does not inhibit the activities of cyclins or cyclin-dependent kinase complexes; however, the importance of the two gene products in the etiology of cancer resides in their involvement in two major cell cycle regulatory pathways: p53 and the retinoblastoma protein, Rb, respectively. Distinct first exons driven from separate promoters are spliced onto the common exons 2 and 3 and the resulting proteins are translated in different reading frames. Both genes are expressed in normal cells but can be alternatively or coordinately silenced when their CpG islands are hypermethylated. Herein, we examined the presence of methyl-CpG binding proteins associated with aberrantly methylated promoters, the distribution of acetylated histones H3 and H4 by chromatin immunoprecipitation assays, and the effect of chemical treatment with 5-aza-2 ' -deoxycytidine (5aza-dC) and trichostatin A on gene induction in colon cell lines by quantitative reverse transcriptase-PCR. We observed that the methyl-CpG binding protein MBD2 is targeted to methylated regulatory regions and excludes the acetylated histones H3 and H4, resulting in a localized inactive chromatin configuration. When methylated, the genes can be induced by 5aza-dC but the combined action of 5aza-dC and trichostatin A results in robust gene expression. Thus, methyl-CpG binding proteins and histone deacetylases appear to cooperate in vivo, with a dominant effect of DNA methylation toward histone acetylation, and repress expression of tumor suppressor genes hypermethylated in cancers.