title: Medulloblastoma and ependymoma cells display increased levels of 5-carboxylcytosine and elevatedTET1expression.
abstract: Alteration of DNA methylation (5-methylcytosine, 5mC) patterns represents one of the causes of tumorigenesis and cancer progression. Tet proteins can oxidise 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine and 5-carboxylcytosine (5caC). Although the roles of these oxidised forms of 5mC (oxi-mCs) in cancer pathogenesis are still largely unknown, there are indications that they may be involved in the mechanisms of malignant transformation. Thus, reduction of 5hmC content represents an epigenetic hallmark of human tumours, and according to our recent report, 5caC is enriched in a proportion of breast cancers and gliomas. Nevertheless, the distribution of oxi-mCs in paediatric brain tumours has not been assessed.
  
  
  
  Here, we analyse the global levels and spatial distribution of 5hmC and 5caC in four brain tumour cell lines derived from paediatric sonic hedgehog (SHH) pathway-activated medulloblastomas (Daoy and UW228-3) and ependymomas (BXD-1425EPN and DKFZ-EP1NS). We show that, unlike HeLa cells, the paediatric tumour cell lines possess both 5hmC and 5caC at immunochemically detectable levels and demonstrate that both modifications display high degrees of spatial overlap in the nuclei of medulloblastomas and ependymomas. Moreover, although 5hmC levels are comparable in the four brain tumour cell lines, 5caC staining intensities differ dramatically between them with highest levels of this mark in a subpopulation of DKFZ-EP1NS cells. Remarkably, the 5caC enrichment does not correlate with 5hmC levels and is not associated with alterations in thymine DNA glycosylase (TDG) expression in SHH medulloblastoma and ependymoma cell lines but corresponds to elevated levels ofTET1transcript in UW228-3 and DKFZ-EP1NS cells.
  
  
  
  We demonstrate that both 5caC enrichment and elevatedTET1expression are observed in SHH medulloblastomas and ependymomas. Our results suggest that increased Tet-dependent 5mC oxidation may represent one of the epigenetic signatures of cancers with neural stem cell origin and, thus, may contribute to development of novel approaches for diagnosis and therapy of the brain tumours.
authors.affiliation: Division of Cancer and Stem Cells, School of Medicine, Centre for Biomolecular Sciences, University of Nottingham, University Park, Nottingham, NG7 2RD UK.
authors.forename: Ashley
authors.initials: A
authors.lastname: Ramsawhook
authors.affiliation: Division of Cancer and Stem Cells, School of Medicine, Centre for Biomolecular Sciences, University of Nottingham, University Park, Nottingham, NG7 2RD UK.
authors.forename: Lara
authors.initials: L
authors.lastname: Lewis
authors.affiliation: Children's Brain Tumour Research Centre, School of Medicine, QMC, University of Nottingham, Nottingham, NG7 2UH UK.
authors.forename: Beth
authors.initials: B
authors.lastname: Coyle
authors.affiliation: Division of Cancer and Stem Cells, School of Medicine, Centre for Biomolecular Sciences, University of Nottingham, University Park, Nottingham, NG7 2RD UK.
authors.forename: Alexey
authors.identifier: 0000-0002-1247-6634
authors.initials: A
authors.lastname: Ruzov
mesh_terms: D044503:5-Methylcytosine / Q000378:metabolism; D001932:Brain Neoplasms / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002528:Cerebellar Neoplasms / Q000235:genetics / Q000378:metabolism*; D002648:Child; D003596:Cytosine / Q000031:analogs & derivatives* / Q000378:metabolism; D004806:Ependymoma / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D008527:Medulloblastoma / Q000235:genetics / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000096:biosynthesis* / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015854:Up-Regulation
chemical_list: C560974:5-carboxylcytosine; D011518:Proto-Oncogene Proteins; D044503:5-Methylcytosine; D003596:Cytosine; D006899:Mixed Function Oxygenases; C462539:TET1 protein, human
keywords: 5-carboxylcytosine; 5-hydroxymethylcytosine; DNA methylation; Ependymoma; Immunohistochemistry; Medulloblastoma; Paediatric brain tumours