Fig. 2: RNA sequestration in P-bodies is cell type specific.

a, Schematic highlighting developmental stages profiled by P-body-seq. Prog., progenitor. b, Representative flow cytometry plots showing gating for GFP-LSM14A⁺ P-bodies in the indicated samples. c, Principal-component (PC) analysis of RNA-seq data for the indicated samples. d, Heatmap showing expression levels of differentially enriched mRNAs between purified P-body fractions of the indicated samples, with manual clustering based on P-body-enriched genes (log₂ (FC) > 0, P value < 0.05) that are specific to each cell type or enriched in all cell types (cluster I). Gene number in each cluster is indicated in the figure. Differential enrichment was assessed using DESeq2 (two-sided Wald test, Benjamini–Hochberg P_adj values, P < 0.05; n = 2 biologically independent samples per group). e, GO pathway analysis using expressed genes as a background for P-body-enriched mRNA in the indicated samples. Enrichment was tested using two-sided Fisher’s exact test with multiple-testing correction (Benjamini–Hochberg FDR). f, Gene tracks showing 8C-related genes from RNA-seq data of P-bodies and cytoplasm fractions of naive ES cells. g, log₂ (FC) of current cell fate markers in the current and downstream cell fate, where log₂ (FC) > 0 indicates P-body enrichment and log₂ (FC) < 0 indicates P-body depletion. Endo, endoderm; meso, mesoderm; NPC, neural progenitor cell. h, Venn diagrams showing the overlap of P-body-enriched genes in the downstream cell type (log₂ (FC) > 0, P < 0.05) with genes that are more highly expressed in the current cell type than in the downstream cell type (current/downstream log₂ (FC) > 0, P < 0.05). Differential enrichment was assessed using DESeq2 (two-sided Wald test, Benjamini–Hochberg P_adj values, P < 0.05). n = 2 biologically independent samples per group. i, Odds ratio of the overlap between the comparisons in h. Center points represent the odds ratio, and error bars represent 95% confidence intervals; n of gene sets is reflected in h. Dashed line indicates an odds ratio of 1 (odds ratios >1 indicate positive association between groups). j, log₂ (FC) of current fate lineage-specific genes (current/downstream log₂ (FC) > 2.5, P < 0.05) in the current and downstream cell state; box plots show the median (center line), interquartile range (IQR; box, 25th–75th percentiles) and whiskers extending to 1.5× the IQR). Differential enrichment was assessed using DESeq2 (two-sided Wald test, Benjamini–Hochberg P_adj values, P = 3.9 × 10⁻²³, P = 6.4 × 10⁻⁵⁴, P = 2.3 × 10⁻²⁹, P = 1.1 × 10⁻³⁷). n = 2 biologically independent samples per group (n of gene sets is reflected in h). k, GSEA performed on P-body-versus-cytoplasm differential expression in neurons for the neural progenitor-related gene expression signature and the neuron-related gene expression signature. Enrichment significance was calculated with the permutation test (two sided), with multiple-testing correction using the Benjamini–Hochberg method (FDR < 0.05). l, GSEA performed on P-body-versus-cytoplasm differential expression in human naive ES cells for human blastomere and naive ES cell-related gene sets from refs. ^58,103. Enrichment significance was calculated with the permutation test (two sided), with multiple-testing correction using the Benjamini–Hochberg method (FDR < 0.05).