A Ternary-code DNA Methylome Atlas of Mouse Tissues

DNA cytosine modifications, including 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), are key epigenetic regulators with distinct functions. To better understand the ternary code (C, 5mC, 5hmC) across different tissues and cell types, we used bACE conversion with scalable assays to map these modifications at base resolution in 29 mouse tissues across various ages and sexes. Our atlas revealed complex patterns of 5hmC distribution, shaped by mitotic activity, chromatin states, and interactions with 5mC. Notably, 5hmC complements 5mC in identifying cell types, extending beyond the brain. Each modification, including 5hmC alone, distinguishes tissue types, enabling high-precision classification of epigenetic identity. Additionally, changes in the methylome correlate with gene transcription, with age-related effects varying by tissue type, offering new insights into cytosine modification biology and potential biomarkers. We examined 5mC and 5hmC in 267 tissue samples from 29 tissue types obtained from 32 C75BL/6 mice.