Fig. 1: Chemically induced totipotent-like cells capture key features of mouse two-cell embryos.

a, Schematic of the stepwise protocol for using chemically induced totipotent-like stem cells to generate an embryo model mimicking different stages of mouse embryogenesis (E1.5 to E7.5 and beyond). PS, primitive streak; DE, definitive endoderm; ExM, extraembryonic mesoderm. b,c, Bright-field images of totipotent-like cell-derived embryo-like structures (b) at different stages and staged-matched natural embryos (c). Scale bars, 100 μm. NF, neural fold; H, heart; NT, neural tube; TB, tail bud. n = 9 embryo-like structures from 9 experiments, n = 3 natural embryo from 3 experiments. d,e, Immunofluorescent image showing the expression of MuERV-L-Gag in induced totipotent-like cells at S1 (c) and E1.5 2-cell embryos (d). n = 5 S1 cell aggregates from 2 experiments, n = 3 E1.5 2-cell embryo from 2 experiments. Scale bars, 20 μm. f, Bright-field and immunofluorescent image showing the presence of derivatives of induced totipotent-like cells in E4.5 mouse blastocysts. Td, tdTomato. Td⁺ cells were found to be presented in the EPI (dashed white line), PrE (white arrowhead) and TE (blue arrowhead) region. n = 5 from 3 experiments. Scale bars, 20 μm. g, Feature plots showing the expression of ZGA genes in EPS cells and cell aggregates at S1. h, Dot plots showing the average levels and proportion of cells expressing specific marker genes for early, middle, and late two-cell embryos in subpopulations 2 and 3 from cell aggregates at S1. Specific marker genes for early, middle and late two-cell embryos were identified using published single-cell transcriptomic datasets (GSE45719 and GSE136714). Expressions of these marker genes in zygotes, early to late 2-cell, 4-cell, 8-cell and 16-cell embryos are shown as the control. i, Pseudotime trajectory reconstruction of single cells from the three subpopulations at S1. j, UMAP analysis of transcriptome of subpopulations from induced totipotent-like cells at S1. These samples were integrated with published single-cell transcriptomic datasets of early mouse embryo stages from zygotes to blastocysts (GSE45719 and GSE136714). EPS cells were also integrated. k, Hierarchical clustering of subpopulations 2 and 3 from induced totipotent-like cells at S1, EPS cells and pre-implantation embryos (GSE45719 and GSE136714).