Stable Transmission of DNA Methylation Epimutations from Germlines to the Liver and Their Association with Fatty Liver Disease in Medaka.

Background Environmental stressors can induce heritable traits in organisms across phyla, with distinct epigenetic alterations in gametes and phenotypic outcomes across several generations. However, the mechanisms underlying such intergenerational inheritance, mainly from the germline to the germline and from the germline to the soma, are enigmatic, given that postfertilization embryos and germline cells reprogram the epigenome in each generation to gain their cellular identity. Here, we report stable germline transmission of differential DNA methylation alterations (epimutations) and their associations with nonalcoholic fatty liver disease (NAFLD) in medaka exposed to a model estrogenic chemical but a ubiquitous environmental contaminant, bisphenol A (BPA). Results Ancestral BPA exposure in the F0 generation led to advanced NAFLD in the unexposed grandchildren generation (F2) of medaka. The F2 liver transcriptome and histopathology revealed a severe NAFLD phenotype in females. Whole-genome bisulfite sequencing of the sperm and liver revealed a gradual shift in promoter methylation from F0 sperm (hypomethylated) to F1 sperm (mix of hypo- and hypermethylated) and F2 liver (predominantly hypermethylated). Many differentially methylated promoters (DMPs) overlapped in F0 sperm, F1 sperm, and F2 liver, regardless of sex. In females, stable transmission of 1511 DMPs was found across three generations, which are associated with protein-coding genes, miRNAs, and others and linked to NAFLD and nonalcoholic steatohepatitis (NASH). Among them, 27 canonical genes maintained consistently hypermethylated promoters across three generations, with significant downregulation of their expression and enrichment in NAFLD-related pathways, mainly fat digestion, glycerolipid metabolism, and steroid biosynthesis. Conclusions The present results demonstrate stable inter- and transgenerational germline-to-germline and germline-to-soma transmission of environmentally induced DNA epimutations with F0 and F1 gametic epimutations, predicting the F2 liver phenotype-a clear transgenerational passage of the disease phenotype in medaka.