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Cell-type specific DNA methylome signatures reveal epigenetic mechanisms for neuronal diversity and neurodevelopmental disorder

ABSTRACT: We report the application of an affinity purification approach (INTACT) to isolate tagged nuclei for GABAergic (GABA), Glutamatergic (Glu), and Purkinje neurons in mice. We performed whole-genome bisulfite sequencing (WGBS) to explore the cell type-specific DNA methylation signatures for the three neuronal types, together with matched transcriptomics. We found a substantial number of differentially methylated regions (DMRs) between cell types, characterized cell-type specific DMRs, and estimated the effects of DNA methylation on gene expression. Finally, we revealed that DNA methylation altered in a cell-type specific manner in a mouse model of Rett syndrome, a neurodevelopmental disorder caused by Mecp2 loss of function. The presented data emphasize the important role of DNA methylation-mediated epigenetic regulation in neuronal diversity and disease.

ORGANISM(S): Mus musculus

PROVIDER: GSE191211 | GEO | 2022/08/24

REPOSITORIES: GEO

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Project description:Background: Down syndrome (DS) is characterized by a genome-wide profile of differential DNA methylation that is skewed towards hypermethylation in most tissues, including brain, and includes pan-tissue differential methylation. The molecular mechanisms involve the overexpression of genes related to DNA methylation on chromosome 21. Here, we stably overexpressed the chromosome 21 gene DNA methyltransferase 3L (DNMT3L) in the human SH-SY5Y neuroblastoma cell line and assayed DNA methylation at over 26 million CpGs by whole genome bisulfite sequencing (WGBS) at three different developmental phases (undifferentiated, differentiating, and differentiated). Results: DNMT3L overexpression resulted in global CpG and CpG island hypermethylation as well as thousands of differentially methylated regions (DMRs). The DNMT3L DMRs were skewed towards hypermethylation and mapped to genes involved in neurodevelopment, cellular signaling, and gene regulation. Consensus DNMT3L DMRs showed that cell lines clustered by genotype and then differentiation phase, demonstrating sets of common genes affected across neuronal differentiation. The hypermethylated DNMT3L DMRs from all pairwise comparisons were enriched for regions of bivalent chromatin marked by H3K4me3 as well as differentially methylated sites from previous DS studies of diverse tissues. In contrast, the hypomethylated DNMT3L DMRs from all pairwise comparisons displayed a tissue-specific profile enriched for regions of heterochromatin marked by H3K9me3 during embryonic development. Conclusions: Taken together, these results support a mechanism whereby regions of bivalent chromatin that lose H3K4me3 during neuronal differentiation are targeted by excess DNMT3L and become hypermethylated. Overall, these findings demonstrate that DNMT3L overexpression during neurodevelopment recreates a facet of the genome-wide DS DNA methylation signature by targeting known genes and gene clusters that display pan-tissue differential methylation in DS.

Project description:To further our understanding of the role of DNA methylation in development, Methylated DNA Immunoprecipitation (MeDIP) was used in conjunction with a NimbleGen promoter plus CpG island array to identify Tissue and Developmental Stage specific Differentially Methylated DNA Regions (T-DMRs and DS-DMRs) on a genome-wide basis. Four tissues (brain, heart, liver, and testis) from C57BL/6J mice were analyzed at three developmental stages (15 day embryo, E15; new born, NB; 12 week adult, AD). Almost 5,000 adult T-DMRs and 10,000 DS-DMRs were identified. Surprisingly, almost all DS-DMRs were tissue specific (i.e., methylated and ummenthylated in one or more non-overlapping tissues), indicating that the vast majority of unique sequence DNA methylation has tissue specificity. Also, many DS-DMRs were methylated at early development stages (E15 and NB) but unmethylated in adult, indicating “demethylation” has a prominent role in tissue differentiation. The pattern of DNA methylation in adult testis was dramatically different from somatic tissues in many aspects, mostly notably with a very strong bias of methylation in non-CpGi (CpG island) promoter regions (94%). Although the majority of T-DMRs and DS-DMRs tended to be in non-CpGi promoter regions, a relatively large number were also located in CpGi in promoter, intra-genic and inter-genic regions (>15% of all CpGi). Gene Ontology analysis of genes with methylation in non-CpGi promoters indicates enrichment of genes related to membrane proteins and G-protein coupled receptors. Our data also suggest regulatory roles of DNA methylation outside of promoter regions and in alternative promoter selection. Overall, our studies indicate that change in DNA methylation during development is a dynamic, widespread and tissue-specific process involving both DNA methylation and demethylation. Comparison of DNA methylation across 3 developmental stages (15 day embryo, newborn, and adult) for four tissues (brain, heart, liver and testis)

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Cell-type specific DNA methylome signatures reveal epigenetic mechanisms for neuronal diversity and neurodevelopmental disorder

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