Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Integrating 5 Hydroxymethylcytosine into the Epigenetic Landscape of Human Embryonic Stem Cells

ABSTRACT: Covalent modification of DNA distinguishes cellular identities and is crucial for regulating the pluripotency and differentiation of embryonic stem (ES) cells. The recent demonstration that 5-methylcytosine (5-mC) may be further modified to 5-hydroxymethylcytosine (5-hmC) in ES cells has revealed a novel regulatory paradigm to modulate the epigenetic landscape of pluripotency. To understand the role of 5-hmC in the epigenomic landscape of pluripotent cells, here we profile the genome-wide 5-hmC distribution and correlate it with the genomic profiles of 11 diverse histone modifications and six transcription factors in human ES cells. By integrating genomic 5-hmC signals with maps of histone enrichment, we link particular pluripotency-associated chromatin contexts with 5-hmC. Intriguingly, through additional correlations with defined chromatin signatures at promoter and enhancer subtypes, we show distinct enrichment of 5-hmC at enhancers marked with H3K4me1 and H3K27ac. These results suggest potential role(s) for 5-hmC in the regulation of specific promoters and enhancers. In addition, our results provide a detailed epigenomic map of 5-hmC from which to pursue future functional studies on the diverse regulatory roles associated with 5-hmC. Genome wide enrichment profile of 5-hmC in H1 human embryonic stem cells

ORGANISM(S): Homo sapiens

SUBMITTER: Keith Szulwach

PROVIDER: E-GEOD-30173 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Integrating 5-hydroxymethylcytosine into the epigenomic landscape of human embryonic stem cells.

Szulwach Keith E KE Li Xuekun X Li Yujing Y Song Chun-Xiao CX Han Ji Woong JW Kim SangSung S Namburi Sandeep S Hermetz Karen K Kim Julie J JJ Rudd M Katharine MK Yoon Young-Sup YS Ren Bing B He Chuan C Jin Peng P

PLoS genetics 20110623 6

Covalent modification of DNA distinguishes cellular identities and is crucial for regulating the pluripotency and differentiation of embryonic stem (ES) cells. The recent demonstration that 5-methylcytosine (5-mC) may be further modified to 5-hydroxymethylcytosine (5-hmC) in ES cells has revealed a novel regulatory paradigm to modulate the epigenetic landscape of pluripotency. To understand the role of 5-hmC in the epigenomic landscape of pluripotent cells, here we profile the genome-wide 5-hmC ...[more]

PMID: 21731508

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Project description:Gene-environment interactions mediated at the epigenetic level may provide an initial step in delivering an appropriate response to environmental changes. 5-hydroxymethylcytosine (5hmC), a DNA base derived from 5-methylcytosine (5mC), accounts for ~40% of modified cytosine in brain and has been implicated in DNA methylation-related plasticity. To identify the role of 5hmC in gene-environment interactions, we exposed both young (6-week-old) and aged (18-month-old) mice to both an enriched environment and a standard environment. Exposure to EE significantly improves learning and memory in aged mice and reduces 5hmC abundance in mouse hippocampus. Furthermore, we mapped the genome-wide distribution of 5hmC and found that the alteration of 5hmC modification occurred mainly at gene bodies. In particular, genes involved in axon guidance are enriched among the genes with altered 5hmC modification. These results together suggest that environmental enrichment could modulate the dynamics of 5hmC in hippocampus, which could potentially contribute to improved learning and memory in aged animals. To identify the role of 5hmC in gene-environment interactions, we exposed both young (6-week-old) and aged (18-month-old) C57B/6 mice to both an enriched environment (EE) and a standard environment. Exposure of mice to EE was achieved by keeping a group of mice in the EE chamber, which is larger than the standard cage, includes novel objects, such as toys of varied color and texture, tunnels, an exercise wheel for voluntary exercise, and extra bedding material, along with free access to food and water. Daily exposure to EE was kept at 5 hours during the daylight cycle for 4 consecutive weeks. Control animals were kept in their small cages that are used as standard housing cages (CE) containing bedding, food and access to water. YC, young mice exposing to control environment, YE, young mice exposing to enriched environment, AC, aged mice exposing to control environment, AE, aged mice exposed to enriched environment. Please note that 5hmC-containing DNA enrichment method was inspired by a unique character of beta-glucotransferase that can specifically add glucose to 5hmC modification. With a modified glucose conjugated with biotin, we are able to purify the 5hmC-containing DNA by biotin-streptavidin-based immunoprecipitation.

Project description:Alzheimer's disease (AD) is a chronic neurodegenerative disorder characterized by progressive deterioration of cognitive function. Evidence suggests a role for epigenetic regulation, in particular the cytosine modifications 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC,) in AD. 5hmC is highly enriched in the nervous system and displays neurodevelopment and age-related changes. To determine the role of 5hmC in AD, we performed genome-wide analyses of 5hmC in DNA from prefrontal cortex of post-mortem AD as well as RNA-Seq to correlate changes in methylation status with transcriptional changes. We also utilized the existing AD fly model to further test the functional significance of these epigenetically altered loci. We identified 325 genes containing differentially hydroxymethylated loci (DhMLs) in both the discovery and replication datasets, and these are enriched for pathways involved in neuron projection development and neurogenesis. Of the 325 genes identified, 140 also showed changes in gene expression by RNA-Seq. Proteins encoded by genes identified in the current analysis form direct protein-protein interactions with AD-associated genes, expanding the network of genes implicated in AD. Furthermore, we identified AD-associated single nucleotide polymorphisms (SNPs) located within or near DhMLs, suggesting that these SNPs may identify regions of epigenetic gene regulation that play a role in AD pathogenesis. Finally using the existing AD fly model we showed that some of these genes could modulate the toxicity associated with AD. Our data implicate neuron projection development and neurogenesis pathways as potential targets in AD. These results indicate that incorporating epigenomic and transcriptomic data with GWAS data can expand the known network of genes involved in disease pathogenesis. Combination of epigenome profiling and Drosophila model enables us to identify the epigenetic modifiers of Alzheimer's disease. University of Kentucky Alzheimer's Disease Research Center (3 control, 3 Alzheimer's) and Emory University Alzheimer's Disease Research Center (2 control, 2 Alzheimer's)

Project description:5-methylcytosine (5-mC) can be oxidized to 5-hydroxymethylcytosine (5-hmC). Genome-wide profiling of 5-hmC thus far indicated 5-hmC may not only be an intermediate form of DNA demethylation but could also constitute an epigenetic mark per se. We describe a cost-effective and selective method to detect both the hydroxymethylation and methylation status of cytosines in more than 1.8 million MspI sites in the human genome. This method involves the selective glucosylation of 5-hmC residues, short-sequence tag generation and high-throughput sequencing. We tested this method by screening H9 human embryonic stem cells and their differentiated embroid body cells, and found that differential hydroxymethylation preferentially occur in bivalent genes during cellular differentiation. Especially, our results support hydroxymethylation can regulate key transcription regulators with bivalent marks through demethylation and affect cellular decision on choosing active or inactive state of these genes upon cellular differentiation. We developed a cost-effective and selective method to detect both the hydroxymethylation and methylation status of cytosines in more than 1.8 million MspI sites in the human genome. In this method, we took advantage of the differential enzymatic sensitivities of the isoschizomers MspI and HpaII. HpaII cleaves only a completely unmodified site, any modification at either cytosine blocks the cleavage, while MspI recognizes and cleaves both 5-mC and 5-hmC, but not the newly discovered 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC). Furthermore, beta-glucosyltransferase (beta-GT) can transfer a glucose to the hydroxyl group of 5-hmC and generate beta-glucosyl-5-hydroxymethylcytosine (5-ghmC) that blocks MspI digestion. Thus, either by combining beta-GT treatment with MspI digestion or simply applying MspI/HpaII digestion, short sequence tags generated can be used for inferring hydroxymethylation or methylation status in around 1.8 million cytosine sites in the human genome. We tested this method by screening H9 human embryonic stem cells and their differentiated embroid body cells.

Dataset Information

Integrating 5 Hydroxymethylcytosine into the Epigenetic Landscape of Human Embryonic Stem Cells

Publications

Integrating 5-hydroxymethylcytosine into the epigenomic landscape of human embryonic stem cells.

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