Global DNA methylation analysis of mouse totipotent potential stem cells, extended pluripotent stem cells and mouse embryonic stem cells

It is challenging to derive totipotent stem cells in vitro that functionally and molecularly resemble cells from totipotent embryos. Here, we report that a chemical cocktail enables the derivation of totipotent-like stem cells, designated as totipotent potential stem (TPS) cells, from 2-cell mouse embryos and extended pluripotent stem cells that can be stably maintained long-term in vitro. TPS cells shared features with 2-cell mouse embryos in terms of totipotency markers, transcriptome, chromatin accessibility and DNA methylation patterns. In vivo chimeric assays show that these cells have embryonic and extraembryonic developmental potentials at the single cell level. Moreover, TPS cells can be induced into blastocyst-like structures resembling preimplantation mouse blastocysts. Mechanistically, inhibition of HDAC1/2 and DOT1L activity and activation of RARγ signaling are important for inducing and maintaining totipotent features of TPS cells. Our study opens up a path toward fully capturing totipotent stem cells in vitro. Total of 10 samples were analyzed, which include mouse totipotent potential stem (TPS) cells, extended pluripotent stem (EPS) cells and embryonic stem (ES) cells. For mouse TPS cells, 2 cell lines were analyzed, which included 1 cell line converted from mouse EPS cells and 1 cell line derived from 2-cell embryos. For mouse EPS cells, 2 cell lines were analyzed. For mouse ES cells, 1 cell line was analyzed. For each sample, two technical replicates were included.