DNA methylation presents distinct binding sites for human transcription factors.
DNA methylation presents distinct binding sites for human transcription factors.
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作者：
Hu S;Wan J;Su Y;Song Q;Zeng Y;Nguyen HN;Shin J;Cox E;Rho HS;Woodard C;Xia S;Liu S;Lyu H;Ming GL;Wade H;Song H;Qian J;Zhu H
DNA甲基化，特别是启动子区域的CpG甲基化，通常被认为是一种有效的表观遗传修饰，其抑制转录因子（TF）的募集，导致转录抑制。在这里，我们使用蛋白质芯片为基础的方法，系统地调查整个人类TF家族，并发现许多纯化的TF与甲基化CpG（mCpG）依赖的DNA结合活性。有趣的是，一些TF对不同序列的甲基化和未甲基化DNA基序表现出特异性结合活性。为了阐明潜在的机制，我们专注于Kruppel样因子4（KLF 4），并通过定点突变解耦其mCpG和CpG结合活性。此外，KLF 4在体内结合人胚胎干细胞中特异性甲基化或未甲基化的基序。我们的研究表明，mCpG依赖的TF结合活性是一种普遍的现象，并提供了一个新的框架，以了解在基因转录的表观遗传调控的TF的作用和机制。DNA甲基化--http://dx.doi.org/10.7554/eLife.00726.001 DNA中胞嘧啶或腺嘌呤碱基上添加一个甲基--在调节基因作为蛋白质的表达中起着关键作用。它有助于诸如X失活的过程，其中一个X染色体拷贝在女性中被沉默，以及基因组印记，其中基因的表达取决于它是从哪个亲本遗传的。DNA甲基化也与癌症的发展有关。然而，它产生这些效应的分子机制尚未完全了解。在哺乳动物中，CpG位点的甲基化（由鸟嘌呤碱基旁边的胞嘧啶碱基组成）通常被认为是通过阻止称为转录因子的蛋白质与称为启动子的DNA区域结合来减少基因表达。如果甲基化破坏了DNA和转录因子之间的相互作用，这可以直接发生，或者如果与甲基化DNA结合的其他蛋白质与转录因子竞争结合位点，这可以间接发生。然而，到目前为止，只有少数与甲基化DNA结合的蛋白质被鉴定出来。现在，Hu等人已经筛选了大约1300种人类转录因子和210种辅因子（与转录因子相互作用的蛋白质）的整个家族，以确定它们与大约150种不同的甲基化DNA片段结合的能力。他们发现，47种蛋白质可以与甲基化的CpG位点结合，大多数蛋白质对特定的DNA序列表现出偏好。此外，一些转录因子和辅因子与具有不同序列的甲基化和非甲基化DNA靶结合。这两种类型的结合在很大程度上是独立的，如阻止称为KLF 4的转录因子与甲基化DNA结合的突变并不阻止其与未甲基化DNA结合的事实所示，反之亦然。Hu等人的工作表明，甲基化胞嘧啶可以有效地作为“第五碱基”-除了腺嘌呤，胞嘧啶，鸟嘌呤和胸腺嘧啶-并强调DNA甲基化对调节基因表达的重要性。DOI：http://dx.doi.org/10.7554/eLife.00726.002网站
DNA methylation, especially CpG methylation at promoter regions, has been generally considered as a potent epigenetic modification that prohibits transcription factor (TF) recruitment, resulting in transcription suppression. Here, we used a protein microarray-based approach to systematically survey the entire human TF family and found numerous purified TFs with methylated CpG (mCpG)-dependent DNA-binding activities. Interestingly, some TFs exhibit specific binding activity to methylated and unmethylated DNA motifs of distinct sequences. To elucidate the underlying mechanism, we focused on Kruppel-like factor 4 (KLF4), and decoupled its mCpG- and CpG-binding activities via site-directed mutagenesis. Furthermore, KLF4 binds specific methylated or unmethylated motifs in human embryonic stem cells in vivo. Our study suggests that mCpG-dependent TF binding activity is a widespread phenomenon and provides a new framework to understand the role and mechanism of TFs in epigenetic regulation of gene transcription. DOI: http://dx.doi.org/10.7554/eLife.00726.001 DNA methylation—the addition of a methyl group to a cytosine or adenine base within DNA—has a key role in regulating the expression of genes as proteins. It contributes to processes such as X-inactivation, in which one copy of the X chromosome is silenced in females, and genomic imprinting, in which the expression of a gene depends upon which parent it was inherited from. DNA methylation has also been implicated in the development of cancer. However, the molecular mechanisms by which it produces these effects are not fully understood. In mammals, the methylation of CpG sites—which consist of a cytosine base next to a guanine base—is typically thought to reduce gene expression by preventing proteins called transcription factors from binding to regions of DNA called promoters. This can occur directly if methylation disrupts interactions between the DNA and the transcription factors, or indirectly if other proteins that bind to the methylated DNA compete with the transcription factors for binding sites. However, only a small number of proteins that bind to methylated DNA have so far been identified. Now, Hu et al. have screened the entire family of roughly 1300 human transcription factors and 210 co-factors (proteins that interact with transcription factors) for their ability to bind to some 150 different stretches of methylated DNA. They found that 47 of the proteins could bind to methylated CpG sites, with the majority showing a preference for specific DNA sequences. Moreover, some transcription factors and co-factors bind to methylated and non-methylated DNA targets with distinct sequences. These two types of binding are largely independent, as illustrated by the fact that mutations that prevent a transcription factor called KLF4 from binding to methylated DNA do not prevent it binding to unmethylated DNA, and vice versa. The work of Hu et al. suggests that methylated cytosine can effectively act as a ‘fifth base’—in addition to adenine, cytosine, guanine and thymine—and emphasizes the importance of DNA methylation for regulating gene expression. DOI: http://dx.doi.org/10.7554/eLife.00726.002