Fig. 3: Both early and late-stage TFs are required for EVT differentiation.
a Western blot analysis showing KD efficiency and expression level of EVT marker genes in EVTs following KD of each candidate EVT regulator. EVTs differentiated from TSCs infected with lentivirus expressing non-targeting sequence were used as control. Two independent repeats were carried out, resulting in similar results. b Invasion ability of EVTs after KD of each candidate EVT regulator, compared to control cells. Scale bar: 100 µm. The invasion area was quantified using Image J software. Measurements were obtained from images taken in three independent biological replicates. Error bars: mean ± SD (n = 3, independent repeats). c Heatmap showing relative expression of class 1–4 genes in EVT factor KD cells compared to non-targeting control. Color represents log2 mRNA expression relative to control KD. d GSEA using EVT marker genes defined in scRNA-seq data of the human first-trimester placenta19,102 and fetal placenta EVT gene set obtained from the Molecular Signatures Database, comparing EVT factor KD cells with control. NES, normalized enrichment score. FDR, false discovery rate. e GO analysis was performed on genes exhibiting lower expression in EVTs after KD of each EVT factor compared to the control. The analysis was conducted using DAVID88, and p-values were calculated. Color representation indicates significance, with -log(p-value). f Principal component analysis of EVT factor KD and control cell transcriptome. g GO analysis was performed on genes exhibiting higher expression in EVTs after KD of each EVT factor compared to the control. The analysis was conducted using DAVID88, and p-values were calculated. Color representation indicates significance, with -log(p-value).
