Selective DNA demethylation by fusion of TDG with a sequence-specific DNA-binding domain
Selective DNA demethylation by fusion of TDG with a sequence-specific DNA-binding domain
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DOI：
10.4161/epi.19509
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发表时间：
2012-04-01
期刊：
影响因子：
3.7
通讯作者：
Fedulov, Alexey V.
中科院分区：
文献类型：
--
作者：
Gregory, David J.;Mikhaylova, Lyudmila;Fedulov, Alexey V.
我们通过表观遗传手段选择性地操纵基因表达的能力是有限的，因为与可用于选择性基因沉默的方法相比，没有用于靶向重新激活表观遗传沉默的基因的方法。我们的目标是开发一种通过DNA去甲基化来选择性激活转录的工具。在这里，我们提出的证据表明，直接靶向胸腺嘧啶-DNA-糖基化酶（TDG）的DNA中的特定序列可以导致在潜在的调控序列的局部DNA去甲基化，并导致增强的基因诱导。当TDG融合到一个充分表征的DNA结合结构域[NF κ B B的Rel同源结构域（RHD）]时，我们观察到DNA甲基化降低，对诱导型一氧化氮合酶（NOS 2）的无关刺激的转录反应增加。这种效应在缺乏RHD结合位点或高水平甲基化的对照基因中没有观察到，在对照模拟转导细胞中也没有观察到。因此，通过这种方法可以实现表观遗传沉默基因的特异性重新激活，这为进一步探索生物学机制和改善对表观基因组的控制提供了广泛有用的策略。
Our ability to selectively manipulate gene expression by epigenetic means is limited, as there is no approach for targeted reactivation of epigenetically silenced genes, in contrast to what is available for selective gene silencing. We aimed to develop a tool for selective transcriptional activation by DNA demethylation. Here we present evidence that direct targeting of thymine-DNA-glycosylase (TDG) to specific sequences in the DNA can result in local DNA demethylation at potential regulatory sequences and lead to enhanced gene induction. When TDG was fused to a well-characterized DNA-binding domain [the Rel-homology domain (RHD) of NF kappa B], we observed decreased DNA methylation and increased transcriptional response to unrelated stimulus of inducible nitric oxide synthase (NOS2). The effect was not seen for control genes lacking either RHD-binding sites or high levels of methylation, nor in control mock-transduced cells. Specific reactivation of epigenetically silenced genes may thus be achievable by this approach, which provides a broadly useful strategy to further our exploration of biological mechanisms and to improve control over the epigenome.