We used MeDIP-seq to undertake the first genomic characterization of intra- and inter-individual variation in DNA methylation across multiple regions of the human brain and an easily accessible peripheral tissue (whole blood). In summary, we show that between-tissue variation in DNA methylation greatly exceeds between-individual differences within any one tissue, with clear hierarchical differences in DNA methylation across specific brain areas, and between brain and blood. Interestingly, we observe that TS-DMRs are strikingly under-represented in classic promoter CGIs, being located primarily in intragenic CGIs and LCPs. These TS-DMRs are dramatically enriched for genes known to be involved in brain development and neurobiological function, forming networks of co-methylated loci that define the cellular phenotype. Finally, although inter-individual differences in DNA methylation are dwarfed by tissue-specific variation, and absolute levels of DNA methylation at specific loci clearly differs between cortex, cerebellum and blood, we observe that some between-individual variation in DNA methylation is correlated between brain regions and blood.
To our knowledge, this study represents the most comprehensive cross-tissue inter-individual assessment of the methylomic landscape of the brain yet undertaken. The data generated in this project are available as a resource to the genomics research community; annotated UCSC tracks can be downloaded from our laboratory website [15
], and the raw data are being integrated into the Human Epigenome Atlas [16
] as part of the regular data release by NIH Epigenomics Roadmap Initiative. A primary goal of global research initiatives such as the International Human Epigenome Consortium and the NIH Epigenomics Roadmap is to create high-resolution reference epigenome maps across multiple human tissue and cell types that will expedite the application of epigenomic technologies to studies of human health and disease [18
]. In this regard, and given increasing evidence supporting a role for epigenetic disruption in neuropsychiatric disease, a unique aspect of this study was our use of blood and multiple brain regions from the same individuals, enabling us to address questions about the extent to which easily accessible peripheral tissues can be used to ask questions about inter-individual phenotypic variation manifest in inaccessible tissues such as the brain.
In addition to creating reference maps of DNA methylation across multiple brain regions and blood, we have identified key regions of the genome that are characterized by tissue-specific patterns of DNA methylation. Of note, TS-DMRs were significantly enriched near genes involved in functional pathways related to neurodevelopment and neuronal differentiation, including BDNF
, and SHANK3
. Although DNA methylation at promoter CGIs is largely conserved across tissues, reflecting data from other studies, we find striking evidence that intragenic CGIs are a primary location for TS-DMRs. This builds on recent data highlighting cell type-specific DNA methylation at intragenic CGIs in the immune system [37
], and evidence that these features may regulate transcription from alternative promoters across specific cell types [19
]. Our data highlight LCPs as another region characterized by considerable cross-tissue epigenetic variation; again these data support recent methylomic analyses of other tissues, indicating that differential DNA methylation across LCPs is associated with tissue-specific gene expression in somatic cells [20
]. Given the striking enrichment of tissue-relevant pathways and gene sets amongst the loci associated with the TS-DMRs we have identified, and the observation that many of these genes are differentially expressed across brain regions (Supplementary Table 7 and Supplementary Figure 6 in Additional file 1
), it is likely that they mediate functionally relevant differences in the cellular transcriptome.
There are several limitations to this study. First, although it represents one of the largest cross-tissue methylomic analyses performed to date, the number of individuals profiled in our initial MeDIP-seq screen was relatively small. Our successful verification and replication experiments (using bisulfite pyrosequencing and the Illumina 450 K array), combined with our gene expression data, in larger number of samples, however, highlights the validity of the between-tissue differences we observe. Second, the samples used in our initial MeDIP-seq screen were all obtained from donors >75 years old and may not fully represent patterns of DNA methylation earlier in life. However, our observation of highly significant gene expression differences corresponding to the top 50 cortex-cerebellum DMRs in a younger cohort of samples (average age = 62 ± 18 years) suggests that many of the tissue-specific differences observed are developmentally stable, resulting in functional differences at an earlier age. Future work should focus on identifying developmental trajectories of epigenetic change across multiple tissues.
In summary, this study reinforces the importance of DNA methylation in regulating cellular phenotype across tissues, and highlights genomic patterns of epigenetic variation across functionally distinct regions of the brain.