GADD45A does not promote DNA demethylation.
GADD45A does not promote DNA demethylation.
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DOI：
10.1371/journal.pgen.1000013
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发表时间：
2008-03-07
期刊：
影响因子：
4.5
通讯作者：
Pfeifer GP
中科院分区：
文献类型：
--
作者：
Jin SG;Guo C;Pfeifer GP
虽然体细胞中的DNA甲基化模式被认为是相对稳定的，但它们在胚胎发育、配子发生和恶性转化期间发生了巨大变化。DNA甲基转移酶的酶学已经被很好地理解，但是从DNA中去除甲基化胞嘧啶的机制（活性DNA去甲基化）仍然是个谜。最近，已经报道了生长停滞和DNA损伤诱导蛋白GADD 45 A在DNA去甲基化中的作用。我们已经研究了GADD 45 A在DNA去甲基化的功能，更详细地使用基因再激活和DNA甲基化检测。与以前的报道相反，我们无法证实GADD 45 A在DNA去甲基化中的功能作用。哺乳动物细胞中主动DNA去甲基化的机制仍然未知。在哺乳动物发育过程中，全基因组DNA去甲基化发生在发育中的生殖细胞和受精卵母细胞中。这种快速的DNA去甲基化是一个活跃的过程，在没有DNA复制的情况下发生。主动DNA去甲基化的机制对该领域的研究人员来说是一个难题，即碳-碳键的断裂以从DNA胞嘧啶环中去除甲基似乎在能量上是不利的，并且在短时间窗口内从两条DNA链中消除约3000万个5-甲基胞嘧啶碱基引起了关于在此过程中维持基因组稳定性的问题。最近，据报道，GADD 45 A蛋白，一种与DNA损伤反应有关的小的酸性蛋白，在几种哺乳动物细胞系中促进活性DNA去甲基化中起着至关重要的作用。我们注意到GADD 45 A不满足哺乳动物DNA去甲基化酶因子的一个可能的要求，因为它不在卵母细胞或受精卵中表达。然后，我们研究了GADD 45 A在DNA去甲基化中的作用，使用甲基化的报告质粒和DNA甲基化分析的几个内源性基因在细胞系中过表达GADD 45 A。与之前的报道相反，我们无法证明GADD 45 A在DNA去甲基化中的作用。在哺乳动物中，促进DNA去甲基化在全基因组水平上的活性仍有待鉴定。
Although DNA methylation patterns in somatic cells are thought to be relatively stable, they undergo dramatic changes during embryonic development, gametogenesis, and during malignant transformation. The enzymology of DNA methyltransferases is well understood, but the mechanism that removes methylated cytosines from DNA (active DNA demethylation) has remained enigmatic. Recently, a role of the growth arrest and DNA damage inducible protein GADD45A in DNA demethylation has been reported. We have investigated the function of GADD45A in DNA demethylation in more detail using gene reactivation and DNA methylation assays. Contrary to the previous report, we were unable to substantiate a functional role of GADD45A in DNA demethylation. The mechanism of active DNA demethylation in mammalian cells remains unknown. During mammalian development, genome-wide DNA demethylation occurs both in developing germ cells and in fertilized oocytes. This rapid DNA demethylation is an active process that occurs in the absence of DNA replication. The mechanism of active DNA demethylation represents a conundrum for researchers in this field, i.e. the breakage of a carbon-carbon bond to remove a methyl group from the DNA cytosine ring appears energetically unfavorable, and the elimination of approximately 30 million 5-methylcytosine bases from both DNA strands within a short time window raises questions about the maintenance of genome stability during this process. Recently, it has been reported that the protein GADD45A, a small acidic protein that has been implicated in the DNA damage response, plays a crucial role in promoting active DNA demethylation in several mammalian cell lines. We noticed that GADD45A does not fulfill one likely requirement for a mammalian DNA demethylase factor in that it is not expressed in oocytes or zygotes. We then investigated the role of GADD45A in DNA demethylation using methylated reporter plasmids and DNA methylation analysis of several endogenous genes in cell lines overexpressing GADD45A. Contrary to the previous report, we were not able to demonstrate a role of GADD45A in DNA demethylation. The activity that promotes DNA demethylation at a genome-wide level in mammals remains to be identified.
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