(A) Zbtb10Δ, Zbtb11Δ, and Zfp131Δ colony morphology in +2i + LIF and −2i + LIF conditions 3 days after doxycycline removal. Zbtb11Δ and Zfp131Δ cells lose typical ESC colony formation ability and adopt morphologies associated with differentiation. Scale bar, 200 μm.
(B) Volcano plots of log2 fold change of transcripts expressed in Zbtb10Δ, Zbtb11Δ, and Zfp131Δ cells versus wild-type (WT) cells grown in the absence of doxycycline for 3 days in +2i + LIF media (n = 3). Significant changes (*p < 0.05) were marked in color for each genotype. Zbtb11 and Zfp131 mutations in ESCs induce aberrant gene expression.
(C and D) GO-enrichment analysis of significantly upregulated genes in (C) Zbtb11Δ (693) and (D) Zfp131Δ (916) in +2i + LIF media (n = 3). Zbtb11 and Zfp131 mutations induce ESCs to express genes with a strong developmental signature.
(E) The log2 fold change of developmentally regulated TFs obtained from bulkRNA-seq experiment in Zbtb11Δ and Zfp131Δ cells versus WT in +2i + LIF and −2 + LIF conditions. Zbtb11 and Zfp131 mutations induce the expression of TFs associated with all three embryonic layers while downregulating pluripotency TFs.
See also Figures S6 and S7.