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CpG methylation of half-CRE sequences creates C/EBPalpha binding sites that activate some tissue-specific genes.

ABSTRACT: DNA methylation of the cytosine in the CpG dinucleotide is typically associated with gene silencing. Genomic analyses have identified low CpG promoters that are both methylated and transcriptionally active, but the mechanism underlying the activation of these methylated promoters remains unclear. Here we show that CpG methylation of the CRE sequence (TGACGTCA) enhances the DNA binding of the C/EBP? transcription factor, a protein critical for activation of differentiation in various cell types. Transfection assays also show that C/EBP? activates the CRE sequence only when it is methylated. The biological significance of this observation was seen in differentiating primary keratinocyte cultures from newborn mice where certain methylated promoters are both bound by C/EBP? and activated upon differentiation. Experimental demethylation by either 5-azacytidine treatment or DNMT1 depletion diminished both C/EBP? binding and activation of the same methylated promoters upon differentiation suggesting that CpG methylation can localize C/EBP?. Transfection studies in cell cultures using methylated tissue-specific proximal promoters identified half-CRE (CGTCA) and half-C/EBP (CGCAA) sequences that need to be methylated for C/EBP? mediated activation. In primary dermal fibroblasts, C/EBP? activates a different set of methylated tissue-specific promoters upon differentiation into adipocytes. These data identify a new function for methyl CpGs: producing DNA binding sites at half-CRE and half-C/EBP sequences for C/EBP? that are needed to activate tissue-specific genes.

SUBMITTER: Rishi V

PROVIDER: S-EPMC2996703 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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CpG methylation of half-CRE sequences creates C/EBPalpha binding sites that activate some tissue-specific genes.

Rishi Vikas V Bhattacharya Paramita P Chatterjee Raghunath R Rozenberg Julian J Zhao Jianfei J Glass Kimberly K Fitzgerald Peter P Vinson Charles C

Proceedings of the National Academy of Sciences of the United States of America 20101108 47

DNA methylation of the cytosine in the CpG dinucleotide is typically associated with gene silencing. Genomic analyses have identified low CpG promoters that are both methylated and transcriptionally active, but the mechanism underlying the activation of these methylated promoters remains unclear. Here we show that CpG methylation of the CRE sequence (TGACGTCA) enhances the DNA binding of the C/EBPα transcription factor, a protein critical for activation of differentiation in various cell types. ...[more]

PMID: 21059933

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Project description:BackgroundThe complex composition of different cell types within a tissue can be estimated by deconvolution of bulk gene expression profiles or with various single-cell sequencing approaches. Alternatively, DNA methylation (DNAm) profiles have been used to establish an atlas for multiple human tissues and cell types. DNAm is particularly suitable for deconvolution of cell types because each CG dinucleotide (CpG site) has only two states per DNA strand-methylated or non-methylated-and these epigenetic modifications are very consistent during cellular differentiation. So far, deconvolution of DNAm profiles implies complex signatures of many CpGs that are often measured by genome-wide analysis with Illumina BeadChip microarrays. In this study, we investigated if the characterization of cell types in tissue is also feasible with individual cell type-specific CpG sites, which can be addressed by targeted analysis, such as pyrosequencing.ResultsWe compiled and curated 579 Illumina 450k BeadChip DNAm profiles of 14 different non-malignant human cell types. A training and validation strategy was applied to identify and test for cell type-specific CpGs. We initially focused on estimating the relative amount of fibroblasts using two CpGs that were either hypermethylated or hypomethylated in fibroblasts. The combination of these two DNAm levels into a "FibroScore" correlated with the state of fibrosis and was associated with overall survival in various types of cancer. Furthermore, we identified hypomethylated CpGs for leukocytes, endothelial cells, epithelial cells, hepatocytes, glia, neurons, fibroblasts, and induced pluripotent stem cells. The accuracy of this eight CpG signature was tested in additional BeadChip datasets of defined cell mixtures and the results were comparable to previously published signatures based on several thousand CpGs. Finally, we established and validated pyrosequencing assays for the relevant CpGs that can be utilized for classification and deconvolution of cell types.ConclusionThis proof of concept study demonstrates that DNAm analysis at individual CpGs reflects the cellular composition of cellular mixtures and different tissues. Targeted analysis of these genomic regions facilitates robust methods for application in basic research and clinical settings.

Dataset Information

CpG methylation of half-CRE sequences creates C/EBPalpha binding sites that activate some tissue-specific genes.

Publications

CpG methylation of half-CRE sequences creates C/EBPalpha binding sites that activate some tissue-specific genes.

Similar Datasets

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