Analysis of TET expression/activity and 5mC oxidation during normal and malignant germ cell development.
Analysis of TET expression/activity and 5mC oxidation during normal and malignant germ cell development.
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DOI：
10.1371/journal.pone.0082881
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Schorle H
中科院分区：
文献类型：
--
作者：
Nettersheim D;Heukamp LC;Fronhoffs F;Grewe MJ;Haas N;Waha A;Honecker F;Waha A;Kristiansen G;Schorle H
在哺乳动物发育过程中，受精卵和原始生殖细胞在一个细胞周期内失去DNA甲基化，从而产生了主动DNA去甲基化的概念。最近的研究发现，Tet羟基酶是DNA去甲基化的关键酶，催化5-甲基胞嘧啶氧化为5-羟甲基胞嘧啶。碱基切除修复机制的进一步氧化和激活导致用未修饰的胞嘧啶取代修饰的胞嘧啶。在这项研究中，我们分析了TET1-3的表达/活性，并筛选出5mC氧化产物在成人睾丸和生殖细胞癌中的存在。通过对人类睾丸切片的分析，我们发现随着精子发生的进行，5-羟甲基胞嘧啶、5-甲酰胞嘧啶和5-羧基胞嘧啶的水平呈下降趋势，而5-甲基胞嘧啶的水平保持不变。这些数据表明，在精子发生过程中，活跃的DNA去甲基化被下调，导致成熟精子中的甲基化标志保持不变。我们证明，与非精原细胞瘤相比，所有原位癌和大多数精原细胞瘤都是低甲基化和低羟甲基化的。有趣的是，在分析的所有生殖细胞癌实体中都能检测到5-甲酰胞嘧啶和5-羧基胞嘧啶，但水平与5-甲基胞嘧啶或5-羟甲基胞嘧啶状态无关。对生殖细胞癌组织和相应细胞系的基因表达数据的荟萃分析表明，TET1和DNA糖基酶TDG高表达，表明生殖细胞癌利用氧化途径进行活性DNA去甲基化。在异种移植实验中，精原细胞瘤样Tcam-2细胞转移到胚胎样癌状态DNMT3b和DNMT3L，其中DNMT3b和DNMT3L强烈上调，这与5-甲基胞嘧啶水平的增加相关。此外，5-羟甲基胞嘧啶水平升高，表明从头甲基化和主动去甲基化伴随着这一转变过程。最后，没有检测到IDH1(IDH1 R132)和IDH2(IDH2 R172)突变导致生殖细胞癌细胞系产生Tet抑制肿瘤代谢产物2-羟基戊二酸。
During mammalian development the fertilized zygote and primordial germ cells lose their DNA methylation within one cell cycle leading to the concept of active DNA demethylation. Recent studies identified the TET hydroxylases as key enzymes responsible for active DNA demethylation, catalyzing the oxidation of 5-methylcytosine to 5-hydroxymethylcytosine. Further oxidation and activation of the base excision repair mechanism leads to replacement of a modified cytosine by an unmodified one. In this study, we analyzed the expression/activity of TET1-3 and screened for the presence of 5mC oxidation products in adult human testis and in germ cell cancers. By analyzing human testis sections, we show that levels of 5-hydroxymethylcytosine, 5-formylcytosine and 5-carboxylcytosine are decreasing as spermatogenesis proceeds, while 5-methylcytosine levels remain constant. These data indicate that during spermatogenesis active DNA demethylation becomes downregulated leading to a conservation of the methylation marks in mature sperm. We demonstrate that all carcinoma in situ and the majority of seminomas are hypomethylated and hypohydroxymethylated compared to non-seminomas. Interestingly, 5-formylcytosine and 5-carboxylcytosine were detectable in all germ cell cancer entities analyzed, but levels did not correlate to the 5-methylcytosine or 5-hydroxymethylcytosine status. A meta-analysis of gene expression data of germ cell cancer tissues and corresponding cell lines demonstrates high expression of TET1 and the DNA glycosylase TDG, suggesting that germ cell cancers utilize the oxidation pathway for active DNA demethylation. During xenograft experiments, where seminoma-like TCam-2 cells transit to an embryonal carcinoma-like state DNMT3B and DNMT3L where strongly upregulated, which correlated to increasing 5-methylcytosine levels. Additionally, 5-hydroxymethylcytosine levels were elevated, demonstrating that de novo methylation and active demethylation accompanies this transition process. Finally, mutations of IDH1 (IDH1 R132) and IDH2 (IDH2 R172) leading to production of the TET inhibiting oncometabolite 2-hydroxyglutarate in germ cell cancer cell lines were not detected.
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影响因子：
4.5
作者：
Jin SG;Guo C;Pfeifer GP
通讯作者：
Pfeifer GP
影响因子：
46.9
作者：
Deng, Jie;Shoemaker, Robert;Xie, Bin;Gore, Athurva;LeProust, Emily M.;Antosiewicz-Bourget, Jessica;Egli, Dieter;Maherali, Nimet;Park, In-Hyun;Yu, Junying;Daley, George Q.;Eggan, Kevin;Hochedlinger, Konrad;Thomson, James;Wang, Wei;Gao, Yuan;Zhang, Kun
通讯作者：
Zhang, Kun
影响因子：
6.1
作者：
Benchaib, M;Braun, V;Guérin, JF
通讯作者：
Guérin, JF
影响因子：
11.2
作者：
Jin SG;Jiang Y;Qiu R;Rauch TA;Wang Y;Schackert G;Krex D;Lu Q;Pfeifer GP
通讯作者：
Pfeifer GP
影响因子：
64.8
作者：
Barreto, Guillermo;Schaefer, Andrea;Niehrs, Christof
通讯作者：
Niehrs, Christof