Folate deficiency reduced aberrant level of DOT1L-mediated histone H3K79 methylation causes disruptive SHH gene expression involved in neural tube defects

Neural tube defects (NTDs) are one of the most severe congenital abnormalities. Maternal folate deficiency could impact the occurrence of NTDs. Histone H3 methyltransferase disruptor of telomeric silencing 1-like (DOT1L) expression was significantly downregulated, and low levels of H3K79me2 were found in the corresponding NTDs samples with their maternal serum folate under low levels. Using ChIP-seq assays, we found that a decrease of H3K79me2 downregulates the expression of Shh and Sufu in mouse embryonic stem cells (mESC) under folate deficiency. Our results indicate that abnormal Shh and Sufu genes expression reduced by aberrant Dot1l-mediated H3K79me2 levels could be the cause of NTDs occurrence. Sv/129 mouse embryonic stem cells (mESCs) were cultured under normal folate medium for at least 2 passages before 1uM methotrexate(MTX) was applied for 24hours. 10^7 mESC cells from normal folate group and MTX group were used to obtain DNA with a concentration of 1 ng / ul and fragments were digested into100-500bp for CHIP(chromatin immunoprecipitation) sequencing. SimpleChip KIT (Cell Signaling Technology, America). CHIP-Seq libraries were prepared according to the protocol. Two IPs( Normal folate group and MTX group)precipitated with H3K79me2 antibody and two inputs(normal folate group and MTXgroup)were sequenced using Illumina NovaSeq 6000.