Germline epigenome editing identifies H3K9me3 as a mediator of intergenerational DNA methylation recovery in mice.

Inter/transgenerational epigenetic inheritance is a crucial and controversial theory that could reshape the concept of genetics. To investigate this theory directly, we invent a system for targeted reprogramming of epigenetic memory in mouse sperm. Using this system, we erase DNA methylation at the differentially methylated region of the H19 gene (H19-DMR) in sperm, which causes Silver-Russell syndrome-like phenotypes in F1 offspring. Although DNA methylation is fully lost in the sperm, it is partially restored during pre-implantation development, suggesting the existence of epigenetic memory that instructs de novo DNA methylation. Importantly, targeted removal of histone modifications in zygotes reveals that tri-methylation at lysine 9 of histone H3 (H3K9me3), which is deposited shortly after fertilization, is required for the subsequent de novo DNA methylation at the H19-DMR. Thus, our study provides a robust germline editing tool, which reveals partial intergenerational inheritance and no transgenerational inheritance at the model locus. Furthermore, we identify H3K9me3 as a mediator for DNA methylation recovery also acting at imprinted loci.