Homo sapiens
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ABSTRACT:
PROVIDER: PRJNA106865 | ENA |
REPOSITORIES: ENA
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Project description:Epigenetic gene regulation is a key determinant of heritable gene expression patterns and is critical for normal cellular function. Dysregulation of epigenetic transcriptional control is a fundamental feature of cancer, particularly manifesting as increased promoter DNA methylation with associated aberrant gene silencing which plays a significant role in tumor progression. We now globally map key chromatin parameters for genes with promoter CpG island DNA hypermethylation in colon cancer cells by combining microarray gene expression analyses with ChIP on chip technology. We first show that the silent state of such genes universally correlates with a broad distribution of a low, but distinct, level of the PcG mediated histone modification, methylation of lysine 27 of histone 3 (H3K27me) and a very low level of the active mark, H3K4me2. This chromatin pattern, and particularly H3K4me2 levels, crisply separates DNA hypermethylated genes from those where histone deacetylation is responsible for transcriptional silencing. Moreover, the chromatin pattern can markedly enhance identification of truly silent and DNA hypermethylated genes. We additionally find that when DNA hypermethylated genes are de-methylated and re-expressed, they adopt a “bivalent” chromatin pattern which is associated with the poised gene expression state of a large group of ES cell genes, and is characterized by an increase in levels of both the H3K27me3 and H3K4me2 marks. Our data have great relevance for the increasing interest in re-expression of DNA hypermethylated genes for the treatment of cancer. Keywords: DNA methylation, chromatin, histone modifications, cancer, epigenetic, ChIP-chip, expression microarray, hypermethylome, DNA-hypermethylation, DAC, TSA, colorectal cancer
2009-01-20 | GSE11173 | GEO
Project description:Many DNA-hypermethylated cancer genes are occupied by the polycomb (PcG) repressor complex in embryonic stem cells (ESCs). Their prevalence in the full spectrum of cancers, the exact context of chromatin involved, and their status in adult cell renewal systems are unknown. Using a genome-wide analysis, we demonstrate that approximately 75% of hypermethylated genes are marked by PcG in the context of bivalent chromatin in both ESC and adult stem/progenitor cells. A large number of these genes are key developmental regulators and a subset, which we call the "DNA hypermethylation module", comprise a portion of the PcG target genes that are downregulated in cancer. Genes with bivalent chromatin have a low, poised gene transcription state that has been shown to maintain stemness and self-renewal in normal stem cells. However, when DNA-hypermethylated in tumors, we find these genes are further repressed. We also show that the methylation status of these genes can cluster important subtypes of colon and breast cancers. By evaluating the subsets of genes that are methylated in different cancers with consideration of their chromatin status in ESCs, we provide evidence that DNA-hypermethylation preferentially targets the subset of PcG genes that are developmental regulators, and this may contribute to the stem-like state of cancer. Additionally, the capacity for global methylation profiling to cluster tumors by phenotype may have important implications for further refining tumor behavior patterns that may ultimately aid therapeutic interventions.
2012-03-02 | GSE35573 | GEO
Project description:CpG island promoter hypermethylation of tumor suppressor genes is a common hallmark of human cancer, and new large-scale epigenomic technologies might be useful in our attempts to define the complete DNA hypermethylome of tumor cells. Here we report a functional search for hypermethylated CpG islands using the colorectal cancer cell line HCT-116, in which two major DNA methyltransferases, DNMT1 and DNMT3b, have been genetically disrupted (DKOcells). Using methylated DNA immunoprecipitation (MeDIP) methodology in conjunction with promoter microarray analyses we found that DKO cells experience a significant loss of hypermethylated CpG islands. Further characterization of these candidate sequences demonstrates CpG island promoter hypermethylation and silencing of genes with potentially important roles in tumorigenesis, such as the Ras guanine-nucleotide release factor RASGRF2, the apoptosis-associated basic helixloop transcription factor BHLHB9, and the homeobox gene HOXD1. Hypermethylation of these genes occurs already in premalignant lesions and accumulates during tumorigenesis. Thus, our results demonstrate the usefulness of DNMT genetic disruption strategies combined with MeDIP in searching for unknown hypermethylated candidate genes in human cancer that might aid our understanding of the biology of the disease and be of potential translational use. Keywords: Human Proximal Promoter Array; DNA Methylation Comparative experiment: methylated DNA immunoprecipitated/INPUT(Total Genomic DNA) from the colon cancer cell line HCT116 wild type vs methylated DNA imonoprecipitated/INPUT from the colon cancer cell line HCT116 DNMT1/3b doubleknockout (DKO)
2010-05-27 | E-GEOD-9267 | biostudies-arrayexpress
Project description:Altered gene expression is a hallmark of human cancers and arises in part through abnormal epigenetic regulation of gene transcription. The best characterized epigenetic alteration involves tumor suppressor gene inactivation via transcriptional repression associated with aberrant DNA hypermethylation of promoter region CpG islands1. Despite characterization of a growing number of such genes, the majority have yet to be identified. We now describe a genome wide microarray gene expression approach for human colorectal cancer cells, which can efficiently identify hundreds of hypermethylated genes for any cancer type. We compared isogenic cells altered pharmacologically versus genetically to induce genomic demethylation, to pinpoint genes activated by DNA demethylation, but not by inhibition of class I and II histone deacetylases (HDACs). We achieve an 82% success rate in predicting genes with densely hypermethylated CpG islands and complete gene silencing. The genes are similarly hypermethylated in primary tumors and have previously undetected tumor suppressor functions. Our approach provides the first highly efficient, comprehensive, platform for defining the cancer “DNA hypermethylome" Keywords: gene expression profiling of pharmacological and genetic human cancer cells
2007-08-08 | GSE4763 | GEO
Project description:Chromatin immunoprecipitation with DNMT3B specific antibody followed by CGI microarray identified genes with or without CGIs, repeat elements and genomic contigs in RKO cells. ChIP-Chop analysis showed that majority of the target genes including P16, DCC, DISC1, SLIT1, CAVEOLIN1, TBX5, TBX18, HOXB13 and some histone variants, that harbor CGI in their promoters, were methylated in multiple colon cancer cell lines but not in normal colon epithelial cells. Further, these genes were reactivated in RKO cells after treatment with 5-aza-2â-deoxycytidine, a DNA hypomethylating agent. COBRA and bisulfite genomic sequencing showed that the CGIs encompassing the transcription start sites (TSS) of DCC, TBX5, TBX18, SLIT1 were also methylated in primary colorectal tumors compared to matching normal tissues whereas GNA11 was methylated in both. MassARRAY analysis demonstrated that the CGI located ~4.5 kb upstream of HOXB13 +1 site was tumor-specifically hypermethylated in primary colorectal cancers and cancer cell lines. Analysis of tumor suppressor properties of two aberrantly methylated transcription factors, HOXB13 and TBX18, revealed that both inhibited growth and clonogenic survival of colon cancer cells in vitro, but only HOXB13 abolished tumor growth in nude mice. Chromatin immunoprecipitation with DNMT3B specific antibody in RKO cells was compared to control antibody.
2010-05-06 | E-GEOD-18929 | biostudies-arrayexpress
Project description:Aberrant CpG methylation is a universal trait of cancer cell genomes and can result in epigenetic modulation of gene activity; however, at which stages tumour-specific epigenetic patterns arise is unknown. Here, we analyse the methylome of APCM in mouse intestinal adenoma as a model of intestinal cancer initiation, and inventory a map of over 13,000 adenoma-specific recurrent differentially methylated regions (DMRs). We find that multiple genes coding for Polycomb proteins are upregulated in adenoma, and concomitantly, hypermethylated DMRs form preferentially at Polycomb target sites. We establish that DMRs are absent from proliferating intestinal epithelial cells or intestinal stem cells, and thus arise de novo after loss of APC. Importantly, a core set of DMRs is conserved in human colon cancer, defining a class of early epigenetic alterations that are distinct from known sets of epigenetically silenced tumour suppressors. The data presented suggests a sequence of events that leads to an altered methylome of colon cancer cells, and may allow more specific selection of clinical epigenetic biomarkers. Analysis of the methylome and RNA expression in adenoma of Apc-Min/+ mutant mice and of normal intestine in Apc-Min/+ and Apc-+/+ wild type mice.
2013-04-03 | E-GEOD-38983 | biostudies-arrayexpress
Project description:Chromatin immunoprecipitation with DNMT3B specific antibody followed by CGI microarray identified genes with or without CGIs, repeat elements and genomic contigs in RKO cells. ChIP-Chop analysis showed that majority of the target genes including P16, DCC, DISC1, SLIT1, CAVEOLIN1, TBX5, TBX18, HOXB13 and some histone variants, that harbor CGI in their promoters, were methylated in multiple colon cancer cell lines but not in normal colon epithelial cells. Further, these genes were reactivated in RKO cells after treatment with 5-aza-2’-deoxycytidine, a DNA hypomethylating agent. COBRA and bisulfite genomic sequencing showed that the CGIs encompassing the transcription start sites (TSS) of DCC, TBX5, TBX18, SLIT1 were also methylated in primary colorectal tumors compared to matching normal tissues whereas GNA11 was methylated in both. MassARRAY analysis demonstrated that the CGI located ~4.5 kb upstream of HOXB13 +1 site was tumor-specifically hypermethylated in primary colorectal cancers and cancer cell lines. Analysis of tumor suppressor properties of two aberrantly methylated transcription factors, HOXB13 and TBX18, revealed that both inhibited growth and clonogenic survival of colon cancer cells in vitro, but only HOXB13 abolished tumor growth in nude mice.
2010-03-01 | GSE18929 | GEO
Project description:Aberrant CpG methylation is a universal trait of cancer cell genomes and can result in epigenetic modulation of gene activity; however, at which stages tumour-specific epigenetic patterns arise is unknown. Here, we analyse the methylome of APCM in mouse intestinal adenoma as a model of intestinal cancer initiation, and inventory a map of over 13,000 adenoma-specific recurrent differentially methylated regions (DMRs). We find that multiple genes coding for Polycomb proteins are upregulated in adenoma, and concomitantly, hypermethylated DMRs form preferentially at Polycomb target sites. We establish that DMRs are absent from proliferating intestinal epithelial cells or intestinal stem cells, and thus arise de novo after loss of APC. Importantly, a core set of DMRs is conserved in human colon cancer, defining a class of early epigenetic alterations that are distinct from known sets of epigenetically silenced tumour suppressors. The data presented suggests a sequence of events that leads to an altered methylome of colon cancer cells, and may allow more specific selection of clinical epigenetic biomarkers.
2013-04-03 | GSE38983 | GEO
Project description:Methylated-DNA sequencing technologies are producing vast amounts of methylome data from cancer samples, from which cancer-associated differentially methylated CpG sites (cDMCs) are continuously identified and filed. The inclusion of as many cDMCs as possible helps improve the accuracy of cancer diagnosis and sometimes identify cancer subtypes. However, the lack of an established method for the analysis of hundreds of cDMCs practically impedes their robust use in clinical medicine. Here, we tested the availability of targeted bisulfite-PCR-sequencing (TBPseq) technology for the assessment of methylation levels of a myriad of CpGs scattered over the genome. In randomly selected 46 cancer cell lines, multiplexed PCR yielded a variety of amplicons harboring 250 CpGs residing at promoters of 97 cancer-associated genes, all of which were sequenced in the same flow cell. Clustering analysis of the TBPseq-assessed methylation levels of target CpGs showed that the lung and liver cancer cell lines correlated relatively strongly with each other while they weakly correlated with colon cancer cells. CpGs at the MLH1 gene promoter, which are known to be hypermethylated in colon cancers, indeed were heavily methylated in the tested colon cancer cells. LIFR CpGs, also known to be methylated in colon cancers, were hypermethylated in the colon cancer cells. Moreover, MLH1 and LIFR promoter hypermethylations were found in colon cancer cells only, but not in biliary tract, liver, lung and stomach cancers cell lines. A meta-analysis with public cancer methylome datasets verified the colorectal cancer specificity of MLH1 and LIFR promoter methylation. These results demonstrate that our TBPseq-based methylation assessment could be considered an effective, accurate, and competitive method to simultaneously examine a large number of target cDMCs and patient samples.
2018-05-01 | GSE98921 | GEO
Project description:Many DNA-hypermethylated cancer genes are occupied by the polycomb (PcG) repressor complex in embryonic stem cells (ESCs). Their prevalence in the full spectrum of cancers, the exact context of chromatin involved, and their status in adult cell renewal systems are unknown. Using a genome-wide analysis, we demonstrate that approximately 75% of hypermethylated genes are marked by PcG in the context of bivalent chromatin in both ESC and adult stem/progenitor cells. A large number of these genes are key developmental regulators and a subset, which we call the "DNA hypermethylation module", comprise a portion of the PcG target genes that are downregulated in cancer. Genes with bivalent chromatin have a low, poised gene transcription state that has been shown to maintain stemness and self-renewal in normal stem cells. However, when DNA-hypermethylated in tumors, we find these genes are further repressed. We also show that the methylation status of these genes can cluster important subtypes of colon and breast cancers. By evaluating the subsets of genes that are methylated in different cancers with consideration of their chromatin status in ESCs, we provide evidence that DNA-hypermethylation preferentially targets the subset of PcG genes that are developmental regulators, and this may contribute to the stem-like state of cancer. Additionally, the capacity for global methylation profiling to cluster tumors by phenotype may have important implications for further refining tumor behavior patterns that may ultimately aid therapeutic interventions. The goal was to analyze the chromatin in MSCs, osteoblasts and U2OS cells of genes that are methylated in the osteosarcoma cell line U2OS. Cell culture: MSCs derived from adult bone marrow were cultured and differentiated to osteoblasts as described by Jaiswal et al., 1997 (J Cell Biochem 64, 295-312). U2OS cells were cultured in McCoy's 5A medium supplemented with 10% FBS. ChIP-seq: Cells were crosslinked in 3.7% formaldehyde, lysed and sonicated to obtain chromatin fragments in the range of 200 to 600 base pairs. Sonicated chromatin was incubated with primary antibody, H3K4Me3 (Millipore, 07-473) or H3K27Me3 (kind gift from Thomas Jenuwein/ Nicholas Shukeir), overnight at 4°C, followed by capturing primary antibodies by adding ProteinA/G magnetic beads (DynaBeads) and incubating at room temperature for 3-4 hrs. Captured chromatin was washed in low- and high-salt buffers, as well as TE. Chromatin fragments were subjected to simultaneous elution, de-crosslinking and Proteinase-K treatment at 65°C in elution buffer followed by phenol-chloroform-isoamyl alcohol extraction and ethanol precipitation. Sequencing and alignment: Purified DNA was used to prepare the sequencing library and sequenced on Applied Biosystems SOLiD (V3). Sequencing reads were aligned to hg18 (NCBI 36) using Bioscope 1.2.1. The software guarantees finding all alignments between the first 25 base pairs of the read (seed) and the reference sequence with up to 2 mismatches. Each match is extended to the full length of the read, scoring 1 point for matching and -2 points for mismatching bases. The read is trimmed to the length with the highest score. If there is only one alignment or if an alignment scores significantly higher than the others for the same read, it is considered unique and reported. Peak identification: Aligned reads were analyzed using Model-based Analysis of ChIP-Seq (MACS) to detect regions enriched for the histone marks (called peaks) with default settings. Input was used as background for peak identification.
2012-03-02 | E-GEOD-35573 | biostudies-arrayexpress