Fig. 2: Single-cell multiomic profiling reveals enhancer-guided early cell fate commitment in ZGA embryos.

a Experimental design and schematic representation of collection strategy and manual selections of ZGA embryos. Nuclei are isolated and processed for 10× Multiome (scATAC-seq + scRNA-seq). Adapted from an image created in BioRender. Cardamone, F. (2025) https://BioRender.com/a06c492. b UMAP embedding of scRNA-seq data of two integrated biological replicates of the ZGA embryo. Cluster identities were assigned by expression of marker genes. A-P ectoderm, anterior-posterior ectoderm. c UMAP embedding of scATAC-seq based on either promoter peaks or enhancer peaks associated to the highly variable genes, highlighting the contribution of enhancer regions in driving germ layers definition. d Distribution of spearman correlation between gene expression (GEX) and accessibility (ATAC) of highly variable genes (HGVs) at their most accessible promoter peak or at their highest score linked peak (enhancer), in each single-cell. Boxes center refers to mean, lower and upper quartiles (Q1 and Q3, respectively). Whiskers, 1.5 × IQR below Q1 and above Q3. Outliers are shown. Two-sided Mann–Whitney U test. P = 3.45 × 10⁻²². e, f Gene expression (GEX) or chromatin accessibility of most accessible promoter peak or highest score linked peak (enhancer) of Mes2 (mesoderm marker gene) and Lim1 (anterior endoderm marker gene). Dashed boxes highlight the germ layer where the gene is expressed. g, h Genome browser snapshot of Mes2 and Lim1 loci. Top, aggregated ATAC reads of each germ layer and violin plot of the respective Mes2 or Lim1 gene expression (GEX). Promoter is highlighted by blue dashed box, enhancer is highlighted by green dashed box. Bottom, ChIP-seq signal of H3K27me3 (this study), H3K27ac and RNA polymerase II (RNAPII) from mesoderm sorted nuclei⁴⁴. MESO, mesoderm. AP-ECTO, anterior-posterior ectoderm. D-ECTO, dorsal ectoderm. N-ECTO, neural ectoderm. V-ECTO, ventral ectoderm. A-ENDO, anterior endoderm. P-ENDO, posterior endoderm. YLK, yolk. i Volcano plot showing differential gene expression analysis in ZGA versus before ZGA embryos. Dot colour represents the marker genes used to annotate the 10× Multiome dataset or significance. Two-sided Wald test. Source data are provided as Source Data file. j Gene regulatory network of the wild-type ZGA embryo inferred by Pando⁸⁴ on the 10× Multiome dataset. Dot colour represents the annotated germ layer of the cell with the highest gene expression value for each gene.