Whole genome bisulfite sequencing of E13.0 and E13.5 Tex14 mutant, heterozygote and wild-type murine female germ cells

Meiosis is critical to generating oocytes however, the mechanisms regulating the switch from mitotic primordial germ cells (PGCs) to meiotic germ cells are poorly understood in females. We showed that the onset of meiosis in the fetal murine ovary propagates via intercellular bridges between developing germ cells by using Tex14 mutant (Tex14-/-) mice. Tex14-/- germ cells, which lack intercellular bridges, initiates meiosis prematurely and they more rapidly extinguish pluripotency-associated transcripts (such as Dppa3) upon entering meiosis. Considering the involvement of Dppa3 in methylation process as well as the importance of epigenetic reprogramming for gamete formation and meiosis, we isolated Tex14 wild-type, heterozygous and mutant germ cells from E13.0 and E13.5 ovaries for whole genome bisulfite sequencing. We compared how methylation differs between age-matched wild-type/heterozygous controls and mutants. We find that DNA methylation across the genome is not affected in Tex14 mutant germ cells however, promoters of Germline Reprogramming Responsive (GRR) genes and younger L1 genes are hypomethylated. These results suggest that cytoplasmic sharing via intercellular bridges slows and coordinates the cell state transition from pluripotency to meiosis potentially by diluting regulatory factors of epigenetic reprogramming. Overall design: E13.0 and E13.5 Oct4-GFP+ female germ cells from single Tex14 mutant, heterozygote or wild-type embryos were purified by fluorescence-activated flow cytometry (8,611 cells from wild-type; 9,944 cells from heterozygote; 4,371 cells from mutant at E13.0 and 3,224 and 8,606 cells from heterozygotes; 3,710 and 10,371 cells from mutants at E13.5). Sequencing was completed for whole-genome analysis as well as specifically for GRR genes and LINE1 elements.