J Neurooncol (2014) 116:429–435 DOI 10.1007/s11060-013-1328-7

TOPIC REVIEW

TET family proteins: new players in gliomas Er-Bao Bian • Gang Zong • Yong-Sheng Xie Xiao-Ming Meng • Cheng Huang • Jun Li • Bing Zhao

•

Received: 28 August 2013 / Accepted: 27 December 2013 / Published online: 7 January 2014 Ó Springer Science+Business Media New York 2014

Abstract DNA methylation at the 5-position of cytosine (5mC) in the mammalian genome has emerged as a pivotal epigenetic event that plays important roles in development, aging and disease. The three members of the TET protein family, which convert 5mC to 5-hydroxymethylcytosine, has provided a potential mechanism resulting in DNA demethylation and maintaining cellular identity. Recent studies have shown that epigenetic modifications play a key role in the regulation of the molecular pathogenesis of gliomas. In this review we focus on demonstrating the TET proteins in DNA demethylation and transcriptional regulation of different target genes. In addition, we address the role of TET proteins in gliomas. This review will provide valuable insights into the potential targets of gliomas, and may open the possibility of novel therapeutic approaches to this fatal disease. Keywords

Gliomas  TETs  5hmc  Demethylation

Abbreviations 5hmC 5-Hydroxymethylcytosine WHO World Health Organization Er-Bao Bian and Gang Zong have contributed equally to this study. E.-B. Bian  G. Zong  Y.-S. Xie  B. Zhao Department of Neurosurgery, The Second Affiliated Hospital of Anhui Medical University, Hefei 230601, China E.-B. Bian  G. Zong  Y.-S. Xie  B. Zhao (&) Cerebral Vascular Disease Research Center, Anhui Medical University, 678 Fu Rong Road, Hefei 230601, Anhui, China e-mail: [email protected] X.-M. Meng  C. Huang  J. Li School of Pharmacy, Anhui Medical University, Hefei 230032, China

GBM TET IDH1 Cys DSBH 2-OG DNMT1 MBDs MLL BER 5fC 5caC TDG TSSs 5mC a-KG OGT 2-HG CP4H TIMP

Glioblastoma multiforme Ten-eleven translocation Isocitrate dehydrogenase 1 Cysteine-rich Double-strandedb-helix 2-Oxoglutarate DNA methyltransferase-1 5-Methylcytosine binding proteins Mixed lineage leukemia Base excision repair 5-Formylcytosine 5-Carboxylcytosine Thymine DNA glycosylase Transcription start sites 5-Methylcytosine a-ketoglutarat O-GlcNAc transferase 2-Hydroxyglutarate Collagen prolyl-4-hydroxylases Tissue inhibitors of metalloproteinase

Introduction Human gliomas are the most common form of primary and aggressive brain tumor of the central nervous system [1]. Traditional pathological approaches classified gliomas as astrocytic, oligodendroglial, mixed oligoastrocytic, and ependymal tumors, which based on the predominant glial cell type (astrocytes or oligodendrocytes) present in tumor and the protein expression pattern as seen in immunohistochemistry [2, 3]. In addition, gliomas are classified according to the 2007 World Health Organization (WHO)

123

430

classification into grades I–IV depending on their extent of malignancy [4]. Patients with WHO grade II tumors which are usually well differentiated, slow-growing typically have a 5-year survival rate of *50 % [5]. Patients with Anaplastic astrocytomas (WHO grade III) exhibit three of the histological features usually nuclear pleomorphism, mitotic activity, and endothelial proliferation and survive 2–3 years [6, 7]. Grade IV gliomas, named glioblastoma multiforme (GBM), are characterized by diffuse invasiveness, immunosuppression, aggressive proliferation, vascularization, and resistance to conventional radiotherapy and chemotherapy [8]. GBM is thought of as the most malignant primary tumors of the brain with highly aggressive course and a median survival time of 9–12 months despite multiple therapeutic regimens designed to optimize surgery, radiation and chemotherapy [5, 9, 10]. Clearly, there is an urgent need for understanding the molecular mechanisms of gliomas recurrence and resistance to common treatment modalities. Recent works have provided novel insights through the identification of genetic alterations affecting genes directly functioning as epigenetic regulators such as ten-eleven translocation (TET) family proteins, or by linking mutations in genes encoding the isocitrate dehydrogenase 1 (IDH1) and 2 (IDH2) with specific changes that influence the function and activity of TETs, thereby resulting in changes in DNA demethylation. Here we review insights into the role of epigenetics modification by TETs in pathogenesis of gliomas.

TET family proteins In mammals, three members of the TET gene family have been identified: TET1, TET2 and TET3 [11, 12]. All three TET proteins share a similar catalytic C-terminal CD domain, including a cysteine-rich (Cys) region and a double-strandedb-helix (DSBH) fold which exhibit 2-oxoglutarate (2-OG)- and Fe(II)-dependent dioxygenase activity [13, 14]. TET proteins were shown to be capable of converting 5-methylcytosine (5mC) to 5hmC by their CD domains in the presence of ATP and in an Fe(II) and aketoglutarate-dependent manner [15, 16]. Additionally, TET1 and TET3, but not Tet2, contain a CXXC zinc finger domain at their amino-terminus, which is a known DNAbinding domain that is frequently found in chromatin binding proteins, such as DNA methyltransferase1(DNMT1), 5mC binding proteins (MBDs) and mixed lineage leukemia (MLL) protein [17, 18]. Unlike the CXXC domains in other proteins closely involved in methylation and chromatin remodeling, which are known to bind unmethylated CpG dinucleotides, the function of this domain in TET1 and TET3 is still poorly known [19–

123

J Neurooncol (2014) 116:429–435

21]. Several recent studies suggest that the CXXC domain of TET1 recognizes CG-rich sequences of both methylated and unmethylated DNA, and it prefers to bind to regions in the genome of high CpG content [22, 23]. It is interesting to point out that many of the CXXC-proteins are also involved in DNA methylation and methylatin of histone. For example, CXXC9/DNMT1 is a DNA methyltransferase that appears to be mainly responsible for the maintenance of methylation pattern on the daughter strand after DNA methylation [24]. In addition, MLL(CXXC7) and MLL2 (CXXC12), known as histone methyltransferases, catalyze the formation of the transcription promoting H3K4 methyl modification [25]. Apart from the function in epigenetic modification, CXXC proteins have also been found to mediate signal transduction. IDAX (CXXC4) functions as a negative regulator of the Wnt signaling pathway that play a key role in regulating cell proliferation, invasion and survival [26, 27].

Role of TET proteins in DNA demethylation DNA methylation is catalyzed by DNA methyltransferases (DNMTs) via covalent addition of a methyl group at the 50 carbon of the cytosine ring, resulting in 5mC [28, 29]. In theory, decreasing levels of 5mC in genomic DNA can be generated through a passive or an active DNA demethylation pathway [13]. More and more studies suggest that there might be multiple pathways or mechanisms by which TET family proteins regulate DNA demethylation (Fig. 1). TET hydroxylases may catalyze active DNA demethylation induced by oxidation of 5mC to 5hmC and its replacement with unmethylated cytosine through a sequential deamination and the base excision repair (BER) DNA repair pathway [30]. In addition, TET proteins were reported to be capable of further converting 5hmC to 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), which can subsequently be recognized and excised by thymine DNA glycosylase (TDG) [15, 31, 32]. Cortellino et al. [30] have demonstrated that demethylation is an active process that requires TDG catalytic activity immediately downstream of the deaminase-catalyzed conversion of 5mC into thymine and/or 5hmC into 5hmU. Deamination of hmC to hmU by an AID/APOBEC enzyme followed by BER-mediated conversion of hmU to cytosine, which is assicated with demethylation [33, 34]. Furthermore, depletion of TDG in mouse ES or iPS cells caused significant 5caC accumulation [15]. In addition to TDG, the methyl-CpG-binding protein MBD4 has glycosylase activity, which is implicated in the regulation of dynamic DNA methylation patterns coupled with deamination and/or oxidation of 5mC [35]. These studies suggest that the role of the TET proteins may be to prevent hypermethylation and promote demethylation by

J Neurooncol (2014) 116:429–435

431

AID/APOBEC

TETs 5mc

5hmc

5hmu

TETs

5cac

5fc DNMTs

BER TDG

cytosine

Fig. 1 Overview of multiple pathways or mechanisms by which TET family proteins mediate DNA demethylation

Target genes

Transcriptional activation

5hm

HCF1

DNMTs

TETs PRC Target genes

OGT Sin3A Transcriptional repression

Fig. 2 TETs function as transcriptional repression or activation of target expression by different pathways

the deamination of the BER repair pathway or TDG pathway.

Mechanisms of gene expression control by TETs proteins The enzymatic activity of TET proteins catalysing the methylation of the 5-position of cytosine (mC) have essential roles in regulating gene expression and maintaining cellular identity. Interestingly, it was found that TETs regulate gene transcription by binding sites located at transcription start sites (TSSs) of CpG-rich promoters [36]. Gene-expression profiling analysis showed that \10 % of the expression alteration of TET1 target genes in TET1depleted ESCs. Unexpectedly, it was also found that an even higher fraction of the upregulated genes were related to TET1 knockdown [36, 37]. These results indicate that TETs could act as transcriptional repression or activation of target expression (Fig. 2). Recent studies regarding the effects of TET proteins on gene expression showed that TET1 plays a repressive role, independent of its enzymatic activity as a hydroxylase [36, 38]. TET1 promotes the silencing of many developmental regulators and somatic lineage differentiation genes that

are associated with PRC2 [36]. Wu et al. [39] further demonstrated that the chromatin-binding ability of PRC2 members was decreased in TET1-depleted cells, indicating that TET1 can directly or indirectly facilitate the recruitment of PRC2 to many TET1 target genes. Interestingly, recent studies found extensive overlap between TET1 and polycomb repressor complex target genes, and despite a requirement of TET1 for the transcriptional activity of PRC2, there was no direct interaction between these proteins [36, 38]. Another mechanism by which transcriptional repression of target genes are regulated by TET1 is via the interaction TET1 with the Sin3A co-repressor complex [36]. A significant target overlap is observed between TET1 and Sin3A. In addition, a subset of TET1 target genes are de-repressed by TET1 or Sin3A knockdown was observed, suggesting that transcriptional repression by TET1 is regulated via the recruitment of Sin3A [36, 39]. Recent studies indicate that TET1 could also interact with the O-GlcNAc transferase (OGT), an enzyme that catalyses O-GlcNAcylation [40, 41]. OGT preferentially associates with TET1 to genes promoters in close proximity of CpG-rich TSSs characterized by low levels of DNA modification, suggesting a link between TET1 and OGT activities in regulating CpG island methylation [42]. OGT is important for TET1-mediated gene repression, where depletion of OGT led to decreased TET1 localization and 5hmC enrichment on TET1-target genes. Mutation of the putative O-GlcNAcylation site on TET1 led to decreased O-GlcNAcylation and level of the TET1 protein [41]. TET, a novel DNA-binding transcriptional regulator, contributes to gene activation likely through alteration of the gene de-repression epigenetic mark, 5hmC. 5hmC may promote transcriptional activation in a context-dependent manner. Moreover, enforced expression of TET1 in cells not only leads to the accumulation of 5hmC, but also contributes to DNA demethylation and reactivates DNA methylation-silenced reporter genes [23]. These results suggest that TET proteins mediate gene transcriptional activation through regulation of 5hmC. In addition, knockdown of TET1 in mouse J1 ES cells also resulted in down-regulation of Nanog expression at both the RNA and protein level [11]. TET10 s regulation of DNA methylation might also involve its negative regulation, direct or indirect, of de novo DNA methyltransferase Dnmt3b [43]. DNMTs methylate the Nanog promoter, leading to the down-regulation of Nanog expression and consequent loss of ES cell identity [11]. These dates indicate that TET1 regulates Nanog expression by preventing the Nanog promoter from hypermethylation mediated by DNMTs. Recently, one published study from Deplus et al. [44] demonstrate that TET2 and TET3 interacted directly with the OGT glycosyltransferase regulate transcriptional activation. TET2/3–OGT co-localize on chromatin at active

123

432

J Neurooncol (2014) 116:429–435

promoters enriched for H3K4me3 and reduction of either TET2/3 or OGT activity results in a direct decrease in H3K4me3 and concomitant decreased transcription. TET2/ 3–OGT mediated transcriptional activation of host cell factor 1(HCF1), a component of the H3K4 methyltransferase SET1/COMPASS complex. Additionally, several key regulators of haematopoiesis, including the Cebpa, Tal1, Runx1, and several Mllgenes, showed decrease in H3K4me3 in a TET2 knockout mouse. These findings suggest that TET2/3–OGT may induce transcriptional activation through the recruitment of H3K4me3 to an overlapping specific gene promoter.

Glioma cells

Normal cells

IDH1/2

IDH1/2 mutation

α-KG

α-KG

2-HG

2-HG

TETs

5hmc

TIMP3

5hmc

TIMP3

The role of TET family proteins in gliomas Epigenetic modification of DNA by cytosine methylation to produce 5mC has become well known as a pivotal epigenetic event in gliomas. 5mCs, especially when accumulated, are important transcriptional silencers at many specific gene promoters and endogenous retrotransposons in the genome [45]. TETs, converting 5mCs to 5-hydroxymethylcytosines (5hmCs), contributes to DNA demethylation in mammalian cells. Recent studies suggest that DNA demethylation mediated by TETs play an important role in gliomas (Fig. 3). One published paper by Parsons et al. [46] demonstrated the existence of a novel gliomaassociated mutation in isocitrate dehydrogenase-1 (IDH1) in 12 % of GBM patients through a genome wide mutation analysis. As a number of subsequent studies have confirmed the findings, mutations in either IDH1 or its mitochondrial counterpart IDH2 have even higher frequencies (up to 70 %) in grade II–III gliomas and secondary glioblastomas [47]. IDH1 and IDH2 catalyze the oxidative decarboxylation of isocitrate to a-ketoglutarat (a-KG) and reduce NADP to NADPH [48]. IDH1 and IDH2 mutations occur frequently in gliomas, resulting in loss of native enzymatic activities and confer novel activity of converting a-KG to 2-hydroxyglutarate (2-HG) [49]. A recent study from Xu et al. [50] demonstrated that 2-HG is a competitive inhibitor of multiple a-KG dioxygenases, including the collagen prolyl-4-hydroxylases (CP4H), histone demethylases, and the TET family 5mC hydroxylases. The expression of mutant IDH1/2 and D-2-HG inhibited the activity of TET2 in catalyzing the 5mC-to-5hmC conversion, which provide a biochemical basis for the mutual exclusivity of IDH1/2 and TET2 gene mutations. Given that mutant TET2 in myeloid malignancies, mutant IDH1/2 in gliomas and both mutations in myeloid malignancies, it may be possible that TET2 mutation is also present in malignant gliomas [51–53]. Loss at the TET2 locus (4q24) has been reported in a small fraction of glioblastomas (2 %) [54]. Depletion of TET2 in immortalized human

123

Prevention of gliomas

Gliomas

Fig. 3 Overview of the role of TETs in normal cells and gliomas cells

astrocytes phenocopies the transforming effects of mutant IDH expression, but TET2 mutations have not been described in brain tumors [55]. In support of this, reduction in TET2 enzyme levels by promoter methylation, while no TET2 mutation, was recently found in low-grade diffuse astrocytomas, indicating that this might be an alternative pathomechanism in low-grade gliomas lacking IDH1 and IDH2 mutation [56]. Mutations TET2 gene displayed uniformly low levels of 5hmC in genomic DNA and altered gene expression patterns in myeloid cancers [57, 58]. The specific type of TET2 mutation in high-grade gliomas remains to be identified. It is likely that mutant TET2 exists in high-grade glioma, though TET2 mutations have not been observed in low-grade glioma. It is interesting to point out that five primary glioblastomas and one secondary glioblastoma investigated did not show expression of TET1, suggesting that gene transcriptional silencing might also be involved in the regulation of TET1 activity in gliomas [59]. Strikingly, most gliomas with wild-type IDH1 status show nuclear exclusion of TET1 that is significantly associated with the loss of 5hmC in glioma [59]. A recent study demonstrated that with mRNA levels of the TET enzymes upregulated in the less aggressive proneural subgroup compared to the mesenchymal subgroup, whereas low levels of TET1 and TET3 were also associated with reduced survival in glioblastoma [60]. 5hmC is generated from 5mC by the action of the TET enzymes and may be an intermediate in a pathway for active DNA demethylation [32, 61]. One recent study reported that in brain tumors, 5hmC levels were high in low grade tumors and reduced in malignant glioma, but did

J Neurooncol (2014) 116:429–435

not exhibit any correlation with IDH1 mutation status [59, 60]. However, other study demonstrated that in gliomas, IDH1 mutations are associated with decreased 5hmC [50]. Recent study from mass spectrometry studies demonstrated a reduction in 5hmC levels in a small series of astrocytomas and glioblastomas compared to normal brain [62, 63]. Additionally, a significant relationship between low levels of 5hmC and reduced survival in malignant glioma was observed [60]. Taken together, these findings that the loss of TETs caused by the accumulation of 2-HG produced by mutant IDH1 and IDH2 proteins, which result in the lower level of 5hmc in gliomas [32, 49, 50, 60]. Tumor suppressor gene silencing through epigenetic modification contributes to cancer formation. Epigenetic events include aberrant DNA methylation such as localized CpG island hypermethylation that leads to inactivation of specific tumor-suppressor genes [64, 65]. Recent studies suggest that TET-catalyzed DNA hydroxymethylation may be a novel regulatory mechanism of DNA methylation and demethylation. Loss of TETs have reported in diverse solid tumors including human breast, liver, prostate cancers and brain tumor [59, 66, 67]. In addition, expression of both TET1 and TET2 but not TET3 was dramatically downregulated in Ischemia reperfusion-injured kidney [68]. Very recently, one study from Hsu et al. [66] demonstrated that TET1 depletion facilitates cell invasion, tumor growth, and cancer metastasis in prostate xenograft models and correlates with poor survival rates in breast cancer patients. However, overexpression of TET1 reduces cell invasion and breast xenograft tumor formation. Mechanistically, TET1 suppresses cell invasion through its dioxygenase and DNA binding activities. Furthermore, TET1 maintains the expression of tissue inhibitors of metalloproteinase (TIMP) family proteins 2 and 3 through its dioxygenase and DNA binding activities inhibiting their DNA methylation. Methylation of TIMP3 was observed in medulloblastoma, the activation of an apoptotic pathway via the enforce expression of TIMP-3 induced apoptosis of transduced human glioma cells and the growth inhibition of human glioma tumor xenografts in immunodeficient mice [69, 70]. Low levels of TET1 was associated with reduced survival in glioblastoma [60]. These results suggest that the loss of TETs protein lead to methylation of TIMP3 following the decrease of TIMP3 in gliomas.

Conclusion and prospective As outlined in this review, extensive studies on TETs have shed light on their functional role in regulating DNA demethylation and gene transcription. TETs regulate target genes depending on the catalytic activity-dependent and independent functions of TETs proteins. Repression or

433

activation of the targeted genes would depend on the interaction of TETs with molecular complexes.TET enzymes appear to have epigenetic functions that go beyond their catalytic role, and further investigations are needed to investigate these mechanisms and determine how they coevolved with the catalytic role. In the future, it is anticipated that TETs will be used for promising novel targets for innovative epigenetic therapy approaches. Acknowledgments This project was supported by the National Science Foundation of China (No. 81072066). Conflict of interest

None.

References 1. Williams SR, Joos BW, Parker JC, Parker JR (2008) Papillary glioneuronal tumor: a case report and review of the literature. Ann Clin Lab Sci 38:287–292 2. Joseph JV, Balasubramaniyan V, Walenkamp A, Kruyt FA (2013) TGF-beta as a therapeutic target in high grade gliomas: promises and challenges. Biochem Pharmacol 85:478–485 3. Louis DN (2006) Molecular pathology of malignant gliomas. Annu Rev Pathol 1:97–117 4. Rychly B, Sidlova H, Danis D (2008) The 2007 World Health Organisation classification of tumours of the central nervous system, comparison with 2000 classification. Cesk Patol 44:35–36 5. Dolecek TA, Propp JM, Stroup NE, Kruchko C (2012) CBTRUS statistical report: primary brain and central nervous system tumors diagnosed in the United States in 2005–2009. Neuro Oncol 14(Suppl 5):v1–v49 6. Goodenberger ML, Jenkins RB (2012) Genetics of adult glioma. Cancer Genet 205:613–621 7. Louis DN, Ohgaki H, Wiestler OD, Cavenee WK, Burger PC, Jouvet A et al (2007) The 2007 WHO classification of tumours of the central nervous system. Acta Neuropathol 114:97–109 8. Zhu VF, Yang J, Lebrun DG, Li M (2012) Understanding the role of cytokines in glioblastoma multiforme pathogenesis. Cancer Lett 316:139–150 9. Daumas-Duport C, Scheithauer B, O’Fallon J, Kelly P (1988) Grading of astrocytomas. A simple and reproducible method. Cancer 62:2152–2165 10. Yao XH, Liu Y, Chen K, Gong W, Liu MY, Bian XW et al (2011) Chemoattractant receptors as pharmacological targets for elimination of glioma stem-like cells. Int Immunopharmacol 11:1961–1966 11. Ito S, D’Alessio AC, Taranova OV, Hong K, Sowers LC, Zhang Y (2010) Role of Tet proteins in 5mC to 5hmC conversion, EScell self-renewal and inner cell mass specification. Nature 466:1129–1133 12. Tahiliani M, Koh KP, Shen Y, Pastor WA, Bandukwala H, Brudno Y et al (2009) Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science 324:930–935 13. Kriukiene E, Liutkeviciute Z, Klimasauskas S (2012) 5-Hydroxymethylcytosine the elusive epigenetic mark in mammalian DNA. Chem Soc Rev 41:6916–6930 14. Tan L, Shi YG (2012) Tet family proteins and 5-hydroxymethylcytosine in development and disease. Development 139:1895–1902 15. He YF, Li BZ, Li Z, Liu P, Wang Y, Tang Q et al (2011) Tetmediated formation of 5-carboxylcytosine and its excision by TDG in mammalian DNA. Science 333:1303–1307

123

434 16. Loenarz C, Schofield CJ (2009) Oxygenase catalyzed 5-methylcytosine hydroxylation. Chem Biol 16:580–583 17. Frauer C, Rottach A, Meilinger D, Bultmann S, Fellinger K, Hasenoder S et al (2011) Different binding properties and function of CXXC zinc finger domains in Dnmt1 and Tet1. PLoS One 6:e16627 18. Iyer LM, Tahiliani M, Rao A, Aravind L (2009) Prediction of novel families of enzymes involved in oxidative and other complex modifications of bases in nucleic acids. Cell Cycle 8:1698–1710 19. Cierpicki T, Risner LE, Grembecka J, Lukasik SM, Popovic R, Omonkowska M et al (2010) Structure of the MLL CXXC domain-DNA complex and its functional role in MLL-AF9 leukemia. Nat Struct Mol Biol 17:62–68 20. Song J, Rechkoblit O, Bestor TH, Patel DJ (2011) Structure of DNMT1-DNA complex reveals a role for autoinhibition in maintenance DNA methylation. Science 331:1036–1040 21. Xu C, Bian C, Lam R, Dong A, Min J (2011) The structural basis for selective binding of non-methylated CpG islands by the CFP1 CXXC domain. Nat Commun 2:227 22. Xu Y, Wu F, Tan L, Kong L, Xiong L, Deng J et al (2011) Genome-wide regulation of 5hmC, 5mC, and gene expression by Tet1 hydroxylase in mouse embryonic stem cells. Mol Cell 42:451–464 23. Zhang H, Zhang X, Clark E, Mulcahey M, Huang S, Shi YG (2010) TET1 is a DNA-binding protein that modulates DNA methylation and gene transcription via hydroxylation of 5-methylcytosine. Cell Res 20:1390–1393 24. Kim JK, Samaranayake M, Pradhan S (2009) Epigenetic mechanisms in mammals. Cell Mol Life Sci 66:596–612 25. Langemeijer SM, Aslanyan MG, Jansen JH (2009) TET proteins in malignant hematopoiesis. Cell Cycle 8:4044–4048 26. Hino S, Kishida S, Michiue T, Fukui A, Sakamoto I, Takada S et al (2001) Inhibition of the Wnt signaling pathway by Idax, a novel Dvl-binding protein. Mol Cell Biol 21:330–342 27. Kojima T, Shimazui T, Hinotsu S, Joraku A, Oikawa T, Kawai K et al (2009) Decreased expression of CXXC4 promotes a malignant phenotype in renal cell carcinoma by activating Wnt signaling. Oncogene 28:297–305 28. Baylin SB (2005) DNA methylation and gene silencing in cancer. Nat Clin Pract Oncol 2(Suppl 1):S4–S11 29. Jones PA, Baylin SB (2002) The fundamental role of epigenetic events in cancer. Nat Rev Genet 3:415–428 30. Cortellino S, Xu J, Sannai M, Moore R, Caretti E, Cigliano A et al (2011) Thymine DNA glycosylase is essential for active DNA demethylation by linked deamination-base excision repair. Cell 146:67–79 31. Inoue A, Zhang Y (2011) Replication-dependent loss of 5-hydroxymethylcytosine in mouse preimplantation embryos. Science 334:194 32. Ito S, Shen L, Dai Q, Wu SC, Collins LB, Swenberg JA et al (2011) Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine. Science 333:1300–1303 33. Guo JU, Su Y, Zhong C, Ming GL, Song H (2011) Hydroxylation of 5-methylcytosine by TET1 promotes active DNA demethylation in the adult brain. Cell 145:423–434 34. Wu SC, Zhang Y (2010) Active DNA demethylation: many roads lead to Rome. Nat Rev Mol Cell Biol 11:607–620 35. Otani J, Arita K, Kato T, Kinoshita M, Kimura H, Suetake I et al (2013) Structural basis of the versatile DNA recognition ability of the methyl-CpG binding domain of methyl-CpG binding domain protein 4. J Biol Chem 288:6351–6362 36. Williams K, Christensen J, Pedersen MT, Johansen JV, Cloos PA, Rappsilber J et al (2011) TET1 and hydroxymethylcytosine in transcription and DNA methylation fidelity. Nature 473:343–348

123

J Neurooncol (2014) 116:429–435 37. Wu H, Zhang Y (2011) Mechanisms and functions of Tet proteinmediated 5-methylcytosine oxidation. Genes Dev 25:2436–2452 38. Wu H, D’Alessio AC, Ito S, Xia K, Wang Z, Cui K et al (2011) Dual functions of Tet1 in transcriptional regulation in mouse embryonic stem cells. Nature 473:389–393 39. Wu H, D’Alessio AC, Ito S, Wang Z, Cui K, Zhao K et al (2011) Genome-wide analysis of 5-hydroxymethylcytosine distribution reveals its dual function in transcriptional regulation in mouse embryonic stem cells. Genes Dev 25:679–684 40. Kreppel LK, Blomberg MA, Hart GW (1997) Dynamic glycosylation of nuclear and cytosolic proteins. Cloning and characterization of a unique O-GlcNAc transferase with multiple tetratricopeptide repeats. J Biol Chem 272:9308–9315 41. Shi FT, Kim H, Lu W, He Q, Liu D, Goodell MA et al (2013) Ten-eleven translocation 1 (Tet1) is regulated by O-linked Nacetylglucosamine transferase (Ogt) for target gene repression in mouse embryonic stem cells. J Biol Chem 288:20776–20784 42. Vella P, Scelfo A, Jammula S, Chiacchiera F, Williams K, Cuomo A et al (2013) Tet proteins connect the O-linked N-acetylglucosamine transferase Ogt to chromatin in embryonic stem cells. Mol Cell 49:645–656 43. Freudenberg JM, Ghosh S, Lackford BL, Yellaboina S, Zheng X, Li R et al (2012) Acute depletion of Tet1-dependent 5-hydroxymethylcytosine levels impairs LIF/Stat3 signaling and results in loss of embryonic stem cell identity. Nucleic Acids Res 40:3364–3377 44. Deplus R, Delatte B, Schwinn MK, Defrance M, Mendez J, Murphy N et al (2013) TET2 and TET3 regulate GlcNAcylation and H3K4 methylation through OGT and SET1/COMPASS. EMBO J 32:645–655 45. Jaenisch R, Bird A (2003) Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nat Genet 33(Suppl):245–254 46. Parsons DW, Jones S, Zhang X, Lin JC, Leary RJ, Angenendt P et al (2008) An integrated genomic analysis of human glioblastoma multiforme. Science 321:1807–1812 47. Kanu OO, Hughes B, Di C, Lin N, Fu J, Bigner DD et al (2009) Glioblastoma multiforme oncogenomics and signaling pathways. Clin Med Oncol 3:39–52 48. Guo C, Pirozzi CJ, Lopez GY, Yan H (2011) Isocitrate dehydrogenase mutations in gliomas: mechanisms, biomarkers and therapeutic target. Curr Opin Neurol 24:648–652 49. Yen KE, Bittinger MA, Su SM, Fantin VR (2010) Cancer-associated IDH mutations: biomarker and therapeutic opportunities. Oncogene 29:6409–6417 50. Xu W, Yang H, Liu Y, Yang Y, Wang P, Kim SH et al (2011) Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of alpha-ketoglutarate-dependent dioxygenases. Cancer Cell 19:17–30 51. Figueroa ME, Abdel-Wahab O, Lu C, Ward PS, Patel J, Shih A et al (2010) Leukemic IDH1 and IDH2 mutations result in a hypermethylation phenotype, disrupt TET2 function, and impair hematopoietic differentiation. Cancer Cell 18:553–567 52. Guilhamon P, Eskandarpour M, Halai D, Wilson GA, Feber A, Teschendorff AE et al (2013) Meta-analysis of IDH-mutant cancers identifies EBF1 as an interaction partner for TET2. Nat Commun 4:2166 53. Tefferi A, Lim KH, Abdel-Wahab O, Lasho TL, Patel J, Patnaik MM et al (2009) Detection of mutant TET2 in myeloid malignancies other than myeloproliferative neoplasms: CMML, MDS, MDS/MPN and AML. Leukemia 23:1343–1345 54. Parsons DW, Jones S, Zhang X et al (2008) Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature 455:1061–1068 55. Koivunen P, Lee S, Duncan CG, Lopez G, Lu G, Ramkissoon S et al (2012) Transformation by the (R)-enantiomer of 2-hydroxyglutarate linked to EGLN activation. Nature 483:484–488

J Neurooncol (2014) 116:429–435 56. Kim YH, Pierscianek D, Mittelbronn M, Vital A, Mariani L, Hasselblatt M et al (2011) TET2 promoter methylation in lowgrade diffuse gliomas lacking IDH1/2 mutations. J Clin Pathol 64:850–852 57. Ko M, Huang Y, Jankowska AM, Pape UJ, Tahiliani M, Bandukwala HS et al (2010) Impaired hydroxylation of 5-methylcytosine in myeloid cancers with mutant TET2. Nature 468:839–843 58. Konstandin N, Bultmann S, Szwagierczak A, Dufour A, Ksienzyk B, Schneider F et al (2011) Genomic 5-hydroxymethylcytosine levels correlate with TET2 mutations and a distinct global gene expression pattern in secondary acute myeloid leukemia. Leukemia 25:1649–1652 59. Muller T, Gessi M, Waha A, Isselstein LJ, Luxen D, Freihoff D et al (2012) Nuclear exclusion of TET1 is associated with loss of 5-hydroxymethylcytosine in IDH1 wild-type gliomas. Am J Pathol 181:675–683 60. Orr BA, Haffner MC, Nelson WG, Yegnasubramanian S, Eberhart CG (2012) Decreased 5-hydroxymethylcytosine is associated with neural progenitor phenotype in normal brain and shorter survival in malignant glioma. PLoS One 7:e41036 61. Pfaffeneder T, Hackner B, Truss M, Munzel M, Muller M, Deiml CA et al (2011) The discovery of 5-formylcytosine in embryonic stem cell DNA. Angew Chem Int Ed Engl 50:7008–7012 62. Jin SG, Jiang Y, Qiu R, Rauch TA, Wang Y, Schackert G et al (2011) 5-Hydroxymethylcytosine is strongly depleted in human cancers but its levels do not correlate with IDH1 mutations. Cancer Res 71:7360–7365

435 63. Kraus TF, Globisch D, Wagner M, Eigenbrod S, Widmann D, Munzel M et al (2012) Low values of 5-hydroxymethylcytosine (5hmC), the ‘‘sixth base’’, are associated with anaplasia in human brain tumors. Int J Cancer 131:1577–1590 64. Kulis M, Esteller M (2010) DNA methylation and cancer. Adv Genet 70:27–56 65. Taberlay PC, Jones PA (2011) DNA methylation and cancer. Prog Drug Res 67:1–23 66. Hsu CH, Peng KL, Kang ML, Chen YR, Yang YC, Tsai CH et al (2012) TET1 suppresses cancer invasion by activating the tissue inhibitors of metalloproteinases. Cell Rep 2:568–579 67. Yang H, Liu Y, Bai F, Zhang JY, Ma SH, Liu J et al (2013) Tumor development is associated with decrease of TET gene expression and 5-methylcytosine hydroxylation. Oncogene 32:663–669 68. Huang N, Tan L, Xue Z, Cang J, Wang H (2012) Reduction of DNA hydroxymethylation in the mouse kidney insulted by ischemia reperfusion. Biochem Biophys Res Commun 422:697–702 69. Lam P, Sian Lim K, Mei Wang S, Hui KM (2005) A microarray study to characterize the molecular mechanism of TIMP-3mediated tumor rejection. Mol Ther 12:144–152 70. Muhlisch J, Bajanowski T, Rickert CH, Roggendorf W, Wurthwein G, Jurgens H et al (2007) Frequent but borderline methylation of p16 (INK4a) and TIMP3 in medulloblastoma and sPNET revealed by quantitative analyses. J Neurooncol 83:17–29

123