Extensive evaluation of DNA methylation of functional elements in the murine Fkbp5 locus using high‐accuracy DNA methylation measurement via targeted bisulfite sequencing

Abstract

AbstractFKBP5 is an important stress‐regulatory gene implicated in stress‐related psychiatric diseases. Single nucleotide polymorphisms of the FKBP5 gene were shown to interact with early life stress to alter the glucocorticoid‐related stress response and moderate disease risk. Demethylation of cytosine‐phosphate‐guanine‐dinucleotides (CpGs) in regulatory glucocorticoid‐responsive elements was suggested to be the mediating epigenetic mechanism for long‐term stress effects, but studies on Fkbp5 DNA methylation (DNAm) in rodents are so far limited. We evaluated the applicability of high‐accuracy DNA methylation measurement via targeted bisulfite sequencing (HAM‐TBS), a next‐generation sequencing‐based technology, to allow a more in‐depth characterisation of the DNA methylation of the murine Fkbp5 locus in three different tissues (blood, frontal cortex and hippocampus). In this study, we not only increased the number of evaluated sites in previously described regulatory regions (in introns 1 and 5), but also extended the evaluation to novel, possibly relevant regulatory regions of the gene (in intron 8, the transcriptional start site, the proximal enhancer and CTCF‐binding sites within the 5'UTR). We here describe the assessment of HAM‐TBS assays for a panel of 157 CpGs with possible functional relevance in the murine Fkbp5 gene. DNAm profiles were tissue‐specific, with lesser differences between the two brain regions than between the brain and blood. Moreover, we identified DNAm changes in the Fkbp5 locus after early life stress exposure in the frontal cortex and blood. Our findings indicate that HAM‐TBS is a valuable tool for broader exploration of the DNAm of the murine Fkbp5 locus and its involvement in the stress response.