DeepH&M: Simultaneous determination of single-CpG hydroxymethylation and methylation levels from enrichment and restriction enzyme sequencing methods
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ABSTRACT:
PROVIDER: PRJNA588377 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10421024_1.fastq.gz | Fastqsanger.gz | |||
| SRR10421024_2.fastq.gz | Fastqsanger.gz | |||
| SRR10421025_1.fastq.gz | Fastqsanger.gz | |||
| SRR10421025_2.fastq.gz | Fastqsanger.gz | |||
| SRR10421026_1.fastq.gz | Fastqsanger.gz |
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Project description:Increased appreciation of 5-hydroxymethylation (5hmC) as a stable epigenetic mark, which defines cell identity and disease progress, has engendered a need for cost-effective, but high-resolution 5hmC mapping technology. Current enrichment-based technologies provide cheap, but low-resolution and relative enrichment of 5hmC levels while single base-resolution methods can be prohibitively expensive to scale up to large experiments. To address this problem, we developed a deep learning-based method “DeepH&M”, which integrates enrichment and restriction enzyme sequencing methods to simultaneously estimate absolute hydroxymethylation and methylation levels at single CpG resolution. Using 7-week-old mouse cerebellum data for training DeepH&M model, we demonstrated that the 5hmC and 5mC levels predicted by DeepH&M were in high concordance with gold standard data predicted by TAB-seq and WGBS. The DeepH&M model can be applied to 7-week old frontal cortex and 79-week cerebellum revealing the robust generalizability of this method to other tissues from various biological time points.
2020-09-10 | GSE140125 | GEO
Project description:Increased appreciation of 5-hydroxymethylcytosine (5hmC) as a stable epigenetic mark, which defines cell identity and disease progress, has engendered a need for cost-effective, but high-resolution, 5hmC mapping technology. Current enrichment-based technologies provide cheap but low-resolution and relative enrichment of 5hmC levels, while single-base resolution methods can be prohibitively expensive to scale up to large experiments. To address this problem, we developed a deep learning-based method, "DeepH&M," which integrates enrichment and restriction enzyme sequencing methods to simultaneously estimate absolute hydroxymethylation and methylation levels at single-CpG resolution. Using 7-week-old mouse cerebellum data for training the DeepH&M model, we demonstrated that the 5hmC and 5mC levels predicted by DeepH&M were in high concordance with whole-genome bisulfite-based approaches. The DeepH&M model can be applied to 7-week-old frontal cortex and 79-week-old cerebellum, revealing the robust generalizability of this method to other tissues from various biological time points.
| S-EPMC7458459 | biostudies-literature
Project description:Simultaneous in vivo enzyme and host engineering through laboratory evolution
| PRJEB26704 | ENA
Project description:In this study we addressed the question whether changes of DNA hydroxymethylation after X-ray irradiation occur in a random fashion or if they show preferences for specific regions or features. We did not only focus on gene loci and promotor sights but also on gene body hydroxymethylation alterations and associated changes in gene expression. A more profound insight into hydroxymethylation characteristics triggered by radiation could provide clues to mechanisms and consequences of cellular response to irradiation including the immediate consequence of DNA strand breaks associated with active DNA demethylation as well as the long term risk of tumorigenesis or other associated diseases.
2019-01-29 | GSE111437 | GEO
Project description:Background: 5-hydroxymethylcytosine (5-hmC) is a recently discovered epigenetic modification that is altered in cancers. Genome wide assays for 5-hmC determination are needed as many of the techniques commonly used to assay 5-methylcytosine (5-mC), including conventional methyl-sensitive restriction digest and bisulfite sequencing, are incapable of distinguishing between 5-mC and 5-hmC. Results: Glycosylation of 5-hmC residues by beta-Glucosyl Transferase (beta-GT) can make CCGG residues insensitive to digestion by MspI. We used this premise to modify the HELP-tagging assay to identify both 5-mC and 5-hmC loci in the genome. Comparison of sequencing libraries after HpaII, MspI and MspI+ beta-GT conversion resulted in locus specific 5-mC and 5-hmC determination. A custom bioinformatics pipeline was created to identify 5-hmC sites that were validated at global level by LS-MS and the locus specific level by qRT-PCR of 5-hmC pulldown DNA. Hydroxymethylation at both promoter and intragenic locations correlated positively with gene expression. Analysis of pancreatic cancer samples revealed striking redistribution of 5-hmC sites in cancer cells and demonstrated enrichment of this modification at many oncogenic promoters such as GATA6. Conclusions: The HELP-GT assay allows a high resolution, simultaneous determination of 5-hmC and 5-mC loci from small amounts of DNA with the utilisation of modest sequencing resources. Redistribution of 5-hmC seen in cancer highlights the importance of examining this modification in conjugation with conventional methylome analysis. We did methylation and hydroxymethylation tests for one control and two pancreatic cancer cases
2013-07-30 | E-GEOD-42723 | biostudies-arrayexpress
Project description:Peripheral blood leukocytes are the most commonly used surrogates to study epigenome-induced risk and epigenomic response to disease related stress. We considered the hypothesis that the TET enzyme catalyzed hydroxymethylation of 5mC to 5hmC might vary among peripheral blood leukocytes and reflect their responsiveness to environment. Reduction in TET1 and/or TET2 activity leads to the over-proliferation of various leukocyte precursors in bone marrow and acute leukemia, yet, the role of 5mC hydroxymethylation in peripheral blood is less well studied. We developed simplified protocols to rapidly and reiteratively isolate mostly non-overlapping leukocyte populations from a single small sample of fresh or frozen whole blood. Among peripheral leukocyte types we found extreme variation in the levels of transcripts encoding proteins involved in cytosine methylation (DNMT1, 3A, 3B) and turnover by de-methylation (TET1, 2, 3) and DNA repair (GADD45a, b, g) and in the gene-region-specific levels of DNA 5hmC (CD4 T cells >> CD14 monocytes > CD16 neutrophils > CD19 B cells > CD56 NK cells > Siglec 8 eosinophils > CD8 T cells). Taken together our results suggest a hierarchy of responsiveness among classes of leukocytes with CD4+ and CD8+ T cells and CD14 monocytes being the most distinctly potentiated for a rapid methylome response to physiological stress and disease. TAB-seq data on 5-hydroxymehtylcytosine (Yu, M. et al. 2012. Cell 149, 1368-1380.) was collected from seven leukocyte types (CD4+ T cells, CD8+ T cells, CD14+ monocytes, CD16+ neutrophils, CD19+ B cells, CD56+ natural killer cells, and Siglec-8+ eosinophils) reiteratively isolated from peripheral blood collected from a healthy male.
2016-07-05 | E-GEOD-70519 | biostudies-arrayexpress
Project description:Background: Despite the significant global loss of DNA hydroxymethylation marks in prostate cancer tissues, the locus-specific role of hydroxymethylation in prostate tumorigenesis is unknown. We characterized hydroxymethylation and methylation marks by performing whole-genome next generation sequencing in representative normal and prostate cancer-derived cell lines in order to determine functional pathways and key genes regulated by these epigenomic modifications in cancer. Results: Our cell line model shows disruption of hydroxymethylation distribution in cancer, with global loss and highly specific gain in promoter and CpG island regions. Significantly, we observed locus-specific retention of hydroxymethylation marks in specific intronic and intergenic regions which may play a novel role in the regulation of gene expression in critical functional pathways, such as BARD1 signaling and steroid hormone receptor signaling in cancer. We confirm a modest correlation of hydroxymethylation with expression in intragenic regions in prostate cancer, while identifying an original role for intergenic hydroxymethylation in differentially expressed regulatory pathways in cancer. We also demonstrate a successful strategy for the identification and validation of key candidate genes from differentially regulated biological pathways in prostate cancer. Conclusion: Our results indicate a distinct function for aberrant hydroxymethylation within each genomic feature in cancer, suggesting a specific and complex role for the deregulation of hydroxymethylation in tumorigenesis, similar to methylation. Subsequently, our characterization of key cellular pathways exhibiting dynamic enrichment patterns for methylation and hydroxymethylation marks may allow us to identify differentially epigenetically modified target genes implicated in prostate cancer tumorigenesis. Methylation profiles of representative normal prostate cell line RWPE-1 and prostate adenocarcinoma cell line 22Rv1 were generated by MBD capture followed by high-throughput sequencing on the HiSeq 2500 (Illumina), in triplicate. Hydroxymethylation profiles of RWPE-1 and 22Rv1 were generated by hMeSeal followed by high-throughput sequencing on the HiSeq 2500 (one replicate each). Additional hydroxymethylation profiling for RWPE-1 was generated by hMeDIP followed by high-throughput sequencing on the HiSeq 2000 (Illumina).
2016-03-15 | E-GEOD-74464 | biostudies-arrayexpress
Project description:The newly discovered 5-hydroxymethylcytosine (5hmC) may complicate previous observations of abnormal cytosine methylation statuses used for the identification of new tumor suppressor gene candidates relevant to human hepatocarcinogenesis. The simultaneous detection of 5mC and 5hmC will stimulate the discovery of aberrantly methylated genes with increased accuracy in human hepatocellular carcinoma (HCC). Here, we performed a newly developed single-base high-throughput sequencing approach (hydroxymethylation and methylation Sensitive Tag sequencing, HMST-seq) to synchronously measure these two modifications in HCC samples. After identifying the differentially methylated and hydroxymethylated genes in HCC, we integrated the DNA copy-number alterations as determined using array-based comparative genomic hybridization (aCGH) data with gene expression to identify genes potentially silenced by promoter hypermethylation. As a result, we report a high enrichment of genes with epigenetic aberrations in cancer signaling pathways. Six genes were selected as tumor suppressor gene (TSG) candidates, among which, ECM1, ATF5 and EOMES were confirmed to have potential anti-cancer function via siRNA experiments. To fully examine 5mC and 5hmC status in HCC, we used a newly developed single-base high-throughput sequencing approach (hydroxymethylation and methylation sensitive tag sequencing, HMST-seq) to synchronously measure these two modifications in HCC samples and their adjacent non-cancerous liver tissues (non-HCCs).
2014-09-01 | E-GEOD-54141 | biostudies-arrayexpress
Project description:We introduce methylation-sensitive restriction enzyme bisulfite sequencing (MREBS) which has the reduced sequencing requirements of RRBS, but significantly expands the coverage of CpG sites in the genome. Here, we compare MREBS to WGBS and RRBS.
2018-12-11 | GSE123566 | GEO
Project description:Digestion with restriction enzymes is a classical approach for probing DNA accessibility in chromatin. It allows to monitor both the cut and the uncut fraction and thereby the determination of accessibility or occupancy (= 1- accessibility) in absolute terms as the percentage of cut or uncut out of the total molecules. The here presented protocol takes this classical approach to the genome-wide level. After exhaustive restriction enzyme digestion of chromatin, DNA is purified, sheared and converted into libraries for high throughput sequencing. Bioinformatic analysis counts DNA fragments cut by the restriction enzyme as well as DNA ends generated by restriction enzyme digest and derives thereof the fraction of accessible DNA. This straight forward principle is technically challenged as preparation and sequencing of the libraries leads to biased scoring of DNA fragments with ends generated by restriction enzymes versus by shearing. Our protocol includes two orthogonal approaches to correct for this bias, the “corrected cut-uncut” and the “cut-all cut” method, so that accurate measurements of absolute accessibility/occupancy at restriction sites throughout a genome are possible. The protocol is presented for the example of S. cerevisiae chromatin but may be adapted for any other species.
2023-01-01 | GSE189142 | GEO