Fig. 3: Phc1 maintains pluripotency of mESCs partly through Nanog.
From: PHC1 maintains pluripotency by organizing genome-wide chromatin interactions of the Nanog locus
a Designing sgRNAs targeting the 2nd exon and 3rd intron of mouse Phc1, and the morphology of the Phc1+/+ and Phc1−/− mESCs. Scale bars, 300 μm. b WB analysis of Phc1, Nanog, Oct4, Sox2, and Actin protein levels in the Phc1+/+ and Phc1−/− mESCs. c qPCR analysis of transcript levels of Pou5f1, Sox2 Nanog, and its known direct target genes including Klf4 and Esrrb. Mean ± s.d. of n = 4 independent experiments. Two-tailed t-tests were used (p < 0.0001 for Nanog; p = 0.0023 for Pou5f1; p = 0.0216 for Sox2; p < 0.0001 for Klf4; p < 0.0001 for Esrrb). d Flow cytometry analysis quantifying GFP signal after Phc1 knockdown in a mESC line carrying the Nanog-GFP reporter. Nanog suppression was used as the positive control. Mean ± s.d. of n = 3 independent experiments. e Morphology of Phc1+/+ and Phc1−/− mESCs transfected with an empty or Flag-Nanog vector. Scale bars, 600 μm. f Immunoblotting of Flag, Nanog, and Tubulin in extracts of mESCs in (e). g Alkaline phosphatase staining of mESCs in (e). h Quantification of colony-forming efficiencies of mESCs relative to Phc1+/+ + vector as the control in (e). Mean ± s.e.m. of n = 4 independent experiments. One-way ANOVA test with Bonferroni’s multiple comparison was used (**p = 0.0024, ***p = 0.0003, *p = 0.0319). Source data are provided as a Source Data file.
