{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"omics_type":["Unknown","Transcriptomics","Genomics","Proteomics"],"submitter":["John Thomson"],"study_type":["methylation profiling by array"],"organism":["Mus musculus"],"species":["Mus musculus"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-GEOD-77727"],"description":["Through the analysis of mouse liver tumours promoted by distinct routes (DEN exposure alone, DEN exposure plus non-genotoxic insult with phenobarbital and non-alcoholic fatty liver disease); we report that the cancer associated hyper-methylated CGI events in mice are also predicated by silent promoters that are enriched for both the DNA modification 5-hydroxymethylcytosine (5hmC) and the histone modification H3K27me3 in normal liver. During cancer progression these CGIs undergo hypo-hydroxymethylation, prior to subsequent hyper-methylation; whilst retaining H3K27me3. A similar loss of promoter-core 5hmC is observed in Tet1 deficient mouse livers indicating that reduced Tet1 binding at CGIs may be responsible for the epigenetic dysregulation observed during hepatocarcinogenesis. Consistent with this reduced Tet1 protein levels are observed in mouse liver tumour lesions. As in human, DNA methylation changes at CGIs do not appear to be direct drivers of hepatocellular carcinoma progression in mice. Instead dynamic changes in H3K27me3 promoter deposition are strongly associated with tumour-specific activation and repression of transcription. Our data suggests that loss of promoter associated 5hmC in diverse liver tumours licences DNA methylation reprogramming at silent CGIs during cancer progression. 5-mc is a well establisehd epigenetic mark typically related to gene silencing events. Phenobarbital (PB) is a well studied non-genotoxic carcinogen with roles in epigenetic perturbation.  We profile 5mC in both control mouse livers as well as in the livers of 12 week PB treated mice. We also profile 5mC in liver tumours arising in the presence of long term PB exposure (35 week: resulting in Ctnnb1 mutated tumours) to a Ha-Ras liver tumour which arose without PB.  Samples: 2 control and 2 PB exposed mouse livers, 3 liver tumours resulting from long term PB exposrue and 1 liver tumour arising without PB"],"repository":["biostudies-arrayexpress"],"sample_protocol":["Nucleic Acid Extraction - Genomic DNA (5mC-IP - 2.5 µg in 450 µl TE), sonicated to yield a fragment distribution of approximately 300-1000 bp, was denatured by incubation at 100°C for 10 min. Samples were rapidly chilled on wet-ice. At this point, 45 µl (10%) of denatured sample was removed and saved as input, and 45 µl of 10X IP buffer (100 mM Na-Phosphate pH 7.0 (mono and dibasic), 1.4 M NaCl, 0.5 % Triton X-100) and 1 µg of 5mC (ActiveMotif; #39769) antibody were added to the remaining sample. Samples were incubated at 4C with gentle agitation overnight. Then, 40 µl of magnetic beads (Dynabeads® Protein G, Invitrogen, UK) in 1X IP buffer were added to each sample to allow magnetic separation of the antibody from the unbound DNA using a magnetic tube rack. Samples were incubated at 4C for 1 hr with gentle agitation. Beads were collected with a magnetic rack and washed with 1000 µl of 1X IP buffer at RT for 10 min with gentle agitation; washing was repeated three times. Beads were collected with a magnetic rack and re-suspended in 250 µl of digestion buffer (50 mM Tris pH 8.0, 10 mM EDTA , 0.5 % SDS) followed by addition of 10 µl of proteinase K (20 mg/ml; Roche Applied Science, Mannheim, Germany) and incubation at 52C for 1.5 hr with constant shaking (≥800 rpm). Finally, beads were removed using a magnetic rack and DNA was purified from the remaining sample using a QIAquick PCR Purification Kit (QIAGEN,CA, USA), eluting in a final volume of 40 µl dH2O. Inputs were also purified using a QIAquick PCR Purification Kit and eluted in 40 µl dH2O. Subsequently, 10 ng of input and IP DNA was subjected to whole genome amplification (WGA) using the GenomePlex® Complete Whole Genome Amplification Kit (Sigma-Aldrich, UK) as per the manufacturer's instructions. Amplified DNA was run on a 1.2% agarose gel to confirm consistency of fragment size between samples.","Hybridization - Labelled samples were applied commercially to a "Nimblegen 2.1M Deluxe promoter array" by Nimblegen, Iceland","Scaning - Arrays were scanned commercially by Nimblegen, Iceland","Labeling - Amplification of DNA samples was carried out commercially by NimbleGen, Iceland (Cy5- (IP) or Cy3- (Input) labelled)"],"figure_sub":["MIAME Score","Raw Data","Organization","Assays and Data","MAGE-TAB Files","Array Designs"],"pubmed_authors":["Richard Meehan","John Thomson","Jonathan Moggs","Remi Terranova","Harri Lempiäinen","Michael Schwarz","Arne Muller"],"additional_accession":[]},"is_claimable":false,"name":"IP of 5-methylcytosine (5-mc) enriched DNA fragments from control, PB treated mouse livers, resulting Ctnnb1 mutated PB liver tumours and PB minus Ha-Ras mutate liver tumour","description":"Through the analysis of mouse liver tumours promoted by distinct routes (DEN exposure alone, DEN exposure plus non-genotoxic insult with phenobarbital and non-alcoholic fatty liver disease); we report that the cancer associated hyper-methylated CGI events in mice are also predicated by silent promoters that are enriched for both the DNA modification 5-hydroxymethylcytosine (5hmC) and the histone modification H3K27me3 in normal liver. During cancer progression these CGIs undergo hypo-hydroxymethylation, prior to subsequent hyper-methylation; whilst retaining H3K27me3. A similar loss of promoter-core 5hmC is observed in Tet1 deficient mouse livers indicating that reduced Tet1 binding at CGIs may be responsible for the epigenetic dysregulation observed during hepatocarcinogenesis. Consistent with this reduced Tet1 protein levels are observed in mouse liver tumour lesions. As in human, DNA methylation changes at CGIs do not appear to be direct drivers of hepatocellular carcinoma progression in mice. Instead dynamic changes in H3K27me3 promoter deposition are strongly associated with tumour-specific activation and repression of transcription. Our data suggests that loss of promoter associated 5hmC in diverse liver tumours licences DNA methylation reprogramming at silent CGIs during cancer progression. 5-mc is a well establisehd epigenetic mark typically related to gene silencing events. Phenobarbital (PB) is a well studied non-genotoxic carcinogen with roles in epigenetic perturbation.  We profile 5mC in both control mouse livers as well as in the livers of 12 week PB treated mice. We also profile 5mC in liver tumours arising in the presence of long term PB exposure (35 week: resulting in Ctnnb1 mutated tumours) to a Ha-Ras liver tumour which arose without PB.  Samples: 2 control and 2 PB exposed mouse livers, 3 liver tumours resulting from long term PB exposrue and 1 liver tumour arising without PB","dates":{"release":"2016-05-17T00:00:00Z","modification":"2023-09-18T04:01:14.832Z","creation":"2022-03-07T00:16:10.915Z"},"accession":"E-GEOD-77727","cross_references":{"GEO":["GSE77727"],"EFO":["EFO_0002759"]}}